Advertisement

SGLT2-inhibitors reduce the cardiac autonomic neuropathy dysfunction and vaso-vagal syncope recurrence in patients with type 2 diabetes mellitus: the SCAN study

Open AccessPublished:June 19, 2022DOI:https://doi.org/10.1016/j.metabol.2022.155243

      Highlights

      • Type 2 diabetes mellitus (T2DM) causes higher rate of vaso-vagal syncope (VVS) recurrence.
      • VVS recurrence could be due to alteration of autonomic system function.
      • heart rate variability and 123I-mIBG myocardial scintigraphy could evaluate these alterations.
      • SGLT2-I could reduce autonomic dysfunction and the VVS recurrence in T2DM patients.

      Abstract

      Background

      In patients with type 2 diabetes mellitus (T2DM) the vaso-vagal syncope (VVS) recurrence could be due to the alteration of autonomic system function, evaluated by heart rate variability (HRV), and by 123I-metaiodobenzylguanidine (123I-mIBG) myocardial scintigraphy indexes: Heart to Mediastinum ratio (H/Mlate), and Washout rate (WR). The SGLT2-I could modulate/reduce autonomic dysfunction in T2DM patients with VVS. This effect could reduce the VVS recurrence in T2DM patients.

      Methods

      In a prospective multicenter study, after propensity score matching, we studied a population of 324 T2DM patients with VVS, divided into 161 SGLT2-I-users vs. 163 Non-SGLT2-I users. In these patients as SGLT2-I-users vs. Non-SGLT2-I users, we investigated the HRV and 123I-MIBG modifications and VVS recurrence at 12 months of follow-up.

      Results

      At follow-up end, the SGLT2-I-users vs. Non-SGLT2-I users had best glucose homeostasis and lower values of inflammatory markers, and resting heart rate (p < 0.05). The SGLT2-I-users vs. Non-SGLT2-I users evidenced the lowest low frequency/high frequency ratio (LF/HFr), a significant difference for all the indexes of autonomic dysfunction via ECG Holter analysis, and higher values of H/Mlate (p < 0.05). Finally, comparing SGLT2-I-users vs. Non-SGLT2-I users, we found a higher rate of VVS recurrence events, specifically of the vasodepressor VVS recurrence at 1-year follow-up (p < 0.05). We did not find a significant difference of mixed and cardio-inhibitory VVS recurrence events at 1 year of follow-up in the study cohorts (p > 0.05). At the Cox regression analysis H/Mlate (0.710, [0.481–0.985]), and SGLT2-I therapy (0.550, [0.324–0.934]) predicted all causes of syncope recurrence at 1 year of follow-up.

      Conclusions

      Non-SGLT2-I users vs. SGLT2-I-users had alterations of the autonomic nervous system, with a higher rate of VVS recurrence at 1 year of follow-up. The indexes of cardiac denervation predicted the VVS recurrence, while the SGLT2-I reduced the risk of VVS recurrence.
      Clinical trial registration number: NCT03717207.

      Graphical abstract

      Keywords

      Abbreviations:

      123I-mIBG (123I-metaiodobenzylguanidine), BNP (B type natriuretic peptide), CAN (cardiac autonomic dysfunction), CRP (C reactive protein), DBP (diastolic blood pressure), ECG (electrocardiogram), eGFR (glomerular filtration rate), HR (heart rate), HRV (heart rate variability), HF (high frequency), LF/HFr (LF to HF ratio), H/Mlate (late heart-to-mediastinum ratio), Hb1Ac (glycated hemoglobin.), HR (Hazard Ratios), HUT (Head-Up Tilt Test), LF (low frequency), LVEF (left ventricular ejection fraction), SGLT2-I (sodium-glucose transporter inhibitors), T2DM (type 2 diabetes mellitus), VVS (Vasovagal syncope), WR (washout rate)

      1. Introduction

      Vasovagal syncope (VVS) is caused by a transient loss of consciousness due to transient global hypo-perfusion, with a rapid onset, short duration, and complete spontaneous recovery [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ]. The VVS could recur until 35 % of the cases, leading to adverse quality of life and worse prognosis [
      • Khera S.
      • Palaniswamy C.
      • Aronow W.S.
      • et al.
      Predictors of mortality, rehospitalization for syncope, and cardiac syncope in 352 consecutive elderly patients with syncope.
      ]. Notably, the VVS recurrence is higher in patients with type 2 diabetes mellitus (T2DM), and T2DM is an independent predictor of VVS recurrence [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Intriguingly, T2DM patients with VVS have a significant autonomic nervous system dysfunction with excessive vagal tone and sympathetic tone withdrawal [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. This could cause significant heart rate (HR) alterations with resting tachycardia, exercise intolerance, abnormal blood pressure regulation, and orthostatic hypotension [
      • Sardu C.
      • Marfella R.
      • Testa G.
      • Santamaria M.
      • Sacra C.
      • Ranauro A.
      • Paolisso G.
      • Rizzo M.R.
      • Barbieri M.
      ELectrophysiological mechanisms underlying the Inhibitory CArdiac syncope without asystolic significant pause: therapeutic and prognostic implications. The ELICA randomized trial.
      ]. The excessive vagal tone could be evaluated by ECG Holter monitoring, via measurements of heart rate variability (HRV) parameters, such as the low frequency (LF), the high frequency (HF), and the LF to HF ratio (LF/HFr), [
      • Pfeifer M.A.
      • Weinberg C.R.
      • Cook D.L.
      • et al.
      Autonomic neural dysfunction in recently diagnosed diabetic subjects.
      ]. The sympathetic system over-activity, and the cardiac autonomic dysfunction (CAN) in patients with VVS, could be evaluated by values of serum norepinephrine, and by 123I-metaiodobenzylguanidine (123I-mIBG) myocardial scintigraphy [
      • Paolisso P.
      • Bergamaschi L.
      • Rambaldi P.
      • Gatta G.
      • Foà A.
      • Angeli F.
      • Fabrizio M.
      • Casella G.
      • Barbieri M.
      • Galiè N.
      • Marfella R.
      • Pizzi C.
      • Sardu C.
      Impact of admission hyperglycemia on heart failure events and mortality in patients with takotsubo syndrome at long-term follow-up: data from HIGH-GLUCOTAKO investigators.
      ,
      • Kochiadakis G.
      • Marketou M.
      • Koukouraki S.
      • Parthenakis F.
      • Chlouverakis G.
      • Karkavitsas N.
      • Vardas P.
      Cardiac autonomic disturbances in patients with vasovagal syndrome: comparison between iodine-123-metaiodobenzylguanidine myocardial scintigraphy and heart rate variability.
      ]. On the other hand, it should be pointed out that there are no conclusive data about the evaluation of CAN by the analysis of HRV and of cardiac MIBG scintigraphy indexes in T2DM patients with VVS, and that less is known about the effects of anti-diabetic therapies on CAN, and their prognostic implications in T2DM patients with VVS. Concerning this, sodium-glucose transporter inhibitors (SGLT2-I) are drugs with direct effects on the glucose-control, which reduce cardiovascular outcomes in T2DM patients [
      • Zinman B.
      • Wanner C.
      • Lachin J.M.
      • Fitchett D.
      • Bluhmki E.
      • Hantel S.
      • Mattheus M.
      • et al.
      Empagliflozin, cardiovascular outcomes, and mortality in type 2 diabetes.
      ]. Intriguingly, the SGLT2-I could exert cardiovascular protective effects also via the modulation of sympathetic nerve activity which is an independent effect of glucose-control efficacy [
      • Hussein A.M.
      • Eid E.A.
      • Taha M.
      • Elshazli R.M.
      • Bedir R.F.
      • Lashin L.S.
      Comparative study of the effects of GLP1 analog and SGLT2 inhibitor against diabetic cardiomyopathy in type 2 diabetic rats: possible underlying mechanisms.
      ]. Therefore, we might hypothesize that the SGLT2-I might significantly modulate the CAN in T2DM with VVS, leading to the reduction of VVS recurrence in T2DM under SGLT2-I (SGLT2-I users) as compared to T2DM patients without SGLT2-I therapy (Non-SGLT2-I users). On the other hand, the chronic use of SGLT2-I to reduce VVS recurrence in T2DM has never been investigated before, and it represents the novelty of the current study. Moreover, here we aimed to evaluate the CAN by HRV parameters and cardiac MIBG scintigraphy indexes at baseline and at 1 year of follow-up, and the rate of VVS recurrence in SGLT2-I users vs. Non-SGLT2-I users' T2DM patients at 1 year of follow up.

      2. Methods

      In a prospective multicenter study conducted at University of Campania “Luigi Vanvitelli”, Naples, at “S. Maria della Pietà Hospital”, Naples, at “Vecchio Pellegrini Hospital”, Naples, at “Catholic University of the Sacred Heart”, Rome, Italy, at Catholic University of Sacred Heart, Campobasso, and at Gemelli Molise, Campobasso, Italy, we investigated from June 2018 to March 2021 a population of 15,794 consecutive patients who had reported at least two syncopal episodes of unknown origin during the previous 6 months, and who had a recurrence of syncope during Head-Up Tilt Test (HUT). We defined the VVS as a transient loss of consciousness whose cause was not determined by the following series of examinations performed in affected patients: clinical history, clinical examination (auscultation, carotid sinus massage, blood pressure measurement in supine, and upright position), electrocardiogram (ECG), chest X-ray, echocardiography, 24 h Holter ambulatory monitoring, late potentials, and a complete neurological examination (head imaging and electroencephalograms), according to last international guidelines [
      • Stewart J.M.
      • Medow M.S.
      • Sutton R.
      • Visintainer P.
      • Jardine D.L.
      • Wieling W.
      Mechanisms of vasovagal syncope in the young: reduced systemic vascular resistance versus reduced cardiac output.
      ]. However, all enrolled patients performed a HUT. These patients were in stable sinus rate before performing the HUT, and they performed a 24 h ECG Holter to assess sinus rhythm, HR, HRV, and the MIBG myocardial scintigraphy before receiving a HUT. Thus, we included only T2DM patients (n 2621) from the study populations, according to diagnostic criteria for T2DM [
      • Stewart J.M.
      • Medow M.S.
      • Sutton R.
      • Visintainer P.
      • Jardine D.L.
      • Wieling W.
      Mechanisms of vasovagal syncope in the young: reduced systemic vascular resistance versus reduced cardiac output.
      ]. Fig. 1. From this cohort, we had the T2DM patients under SGLT2-I therapy (n 426) vs. those that did not receive the SGLT2-I therapy (n 2195). Then, we calculated a propensity score matching (PSM) of 160 patients for each cohort of the study. The study complies with the Declaration of Helsinki, and the locally appointed Ethics Committee approved the research protocol (n 06062.18), and informed consent was obtained from all the subjects. The study population respected the following inclusion/exclusion criteria.
      • Inclusion criteria: eligible adults' patients with T2DM (≥18 years of age); previous VVS event; indication to receive a HUT; indication to practice cardiac MIBG scintigraphy; patients without previous cardiovascular disease; patients with an estimated glomerular filtration rate (eGFR) of at least 30 ml per minute per 1.73 m2 of body-surface area.
      • Exclusion criteria: patients with a diagnosis of type 1 diabetes, chronic neurological disorders, heart failure and coronary heart disease or depression of left ventricular ejection fraction (LVEF <55 %); patients with severe anemia, thyroid dysfunction, concomitant treatment with anticholinergic agents, adrenergic antagonists, and vaso-constrictive agents that can affect the results of autonomic function test; patients with uncontrolled blood pressure (blood pressure ≥ 140/90 mmHg on two occasions 2 weeks apart).
      Fig. 1
      Fig. 1Study flow chart. CHD: coronary heart disease; eGFR: estimated glomerular filtration rate; HUT: Head-Up Tilt test; MIBG: metaiodobenzylguanidine; SGLT2-I: Sodium-Glucose Transporter 2 inhibitor; T2DM: type 2 diabetes mellitus; VVS: vaso-vagal syncope.

      2.1 Patients' monitoring

      The enrolled patients were regularly followed by clinical visits 10 days, and at 3rd, 6th, and 12th months after clinical discharge by the treating physician. These patients had a diagnosis of VVS in accordance with the HUT result [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ]. All the patients gave their written informed to participate in the study. Clinical evaluations included physical examination, vital signs, and review of adverse events. We performed fasting blood (at least 12 h from last meal) for glycemia, lipid profile (total cholesterol (TC), triglycerides, HDL-C, and LDL-C) and B type natriuretic peptide (BNP) at every visit. Syncope recurrence and other clinical events were collected during patients' interviews, visits, and hospital discharge schedules. The diagnosis of T2DM was made according to international recommendations, by plasma glucose values as fasting plasma glucose level ≥ 7.0 mmol/l (126 mg/dl), plasma glucose ≥11.1 mmol/l (200 mg/dl) either while fasting or not fasting, glycated hemoglobin ≥48 mmol/mol, and by the known clinical history of diabetes and by the current use of anti-diabetic medications [
      American Diabetes Association
      2. Classification and diagnosis of diabetes: standards of medical care in diabetes-2021.
      ]. In the T2DM patients, we defined SGLT2-I users as those receiving a SGLT2-I therapy before starting the study. We defined the rest of the T2DM patients as Non-SGLT2-I users. The SGLT2-I users received either 10 mg or 25 mg of empagliflozin once daily and/or canaglifozin 100 mg daily in the last month before starting the study (mean duration of SGLT2-I therapy 16 ± 4.8 days at study beginning).

      2.2 Cardiovascular autonomic neuropathy evaluation

      We evaluated the autonomic function of the study population by classical Ewing cardiovascular autonomic function tests, heart and pulse rate variability [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. In the morning (08:00–10:30), we performed the test in a room with a quiet ambiance and temperature of 19–22 °C. The patients were fasting from midnight and refrained from smoking and caffeine-containing beverages for almost 12 h before the evaluations. Two experienced physicians blinded to study protocol evaluated the cardiovascular autonomic reflex function tests as parasympathetic and sympathetic tests [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ,
      • Sardu C.
      • Marfella R.
      • Testa G.
      • Santamaria M.
      • Sacra C.
      • Ranauro A.
      • Paolisso G.
      • Rizzo M.R.
      • Barbieri M.
      ELectrophysiological mechanisms underlying the Inhibitory CArdiac syncope without asystolic significant pause: therapeutic and prognostic implications. The ELICA randomized trial.
      ,
      • Pfeifer M.A.
      • Weinberg C.R.
      • Cook D.L.
      • et al.
      Autonomic neural dysfunction in recently diagnosed diabetic subjects.
      ]. The parasympathetic tests evaluated the HR response during deep breathing, the Valsalva maneuver, and quick standing [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ,
      • Sardu C.
      • Marfella R.
      • Testa G.
      • Santamaria M.
      • Sacra C.
      • Ranauro A.
      • Paolisso G.
      • Rizzo M.R.
      • Barbieri M.
      ELectrophysiological mechanisms underlying the Inhibitory CArdiac syncope without asystolic significant pause: therapeutic and prognostic implications. The ELICA randomized trial.
      ,
      • Pfeifer M.A.
      • Weinberg C.R.
      • Cook D.L.
      • et al.
      Autonomic neural dysfunction in recently diagnosed diabetic subjects.
      ]. The sympathetic tests evaluated the blood pressure response during the sustained handgrip test and quick standing [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ,
      • Sardu C.
      • Marfella R.
      • Testa G.
      • Santamaria M.
      • Sacra C.
      • Ranauro A.
      • Paolisso G.
      • Rizzo M.R.
      • Barbieri M.
      ELectrophysiological mechanisms underlying the Inhibitory CArdiac syncope without asystolic significant pause: therapeutic and prognostic implications. The ELICA randomized trial.
      ,
      • Pfeifer M.A.
      • Weinberg C.R.
      • Cook D.L.
      • et al.
      Autonomic neural dysfunction in recently diagnosed diabetic subjects.
      ]. We evaluated the deep breathing test to determine the maximum and minimum R–R intervals during each breathing cycle during six deep breaths in 1 min [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. However, we expressed the R–R intervals during inspiration and expiration as the R–R inspiration/R–R expiration ratio [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Again, we evaluated the Valsalva maneuver as a forced expiration in a manometer against 40 mmHg for 15 s. The Valsalva ratio is divided into the longest R–R interval by the shortest R–R interval [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Moreover, during the quick standing test, we measured the HR response after standing from the R–R intervals calculation at 15 and 30 beats after standing and reported as the ratio of the longest vs. the shortest R–R interval [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Thus, we calculated the sympathetic component of the standing test by the values of the systolic blood pressure response 2 min after standing [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Finally, we evaluated the diastolic blood pressure response during the sustained handgrip by a dynamometer to establish a maximum developed force, followed by a handgrip squeeze of 30 % of the maximum force for 5 min [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Furthermore, we defined Cardiovascular autonomic neuropathy (CAN) as the composite CAN index including the HR ratio < 1.30 plus Valsalva ratio < 1.5 or a decrease of >10 mmHg in diastolic blood pressure (DBP) upon standing (CAN+) [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ].

      2.3 Head-Up Tilt Test (HUT)

      In the overall study population, and selectively in SGLT2-I-users vs. Non-SGLT2-I users, we performed the HUT in the morning in a quiet room with lights slightly dimmed after overnight fasting [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ,
      • Khera S.
      • Palaniswamy C.
      • Aronow W.S.
      • et al.
      Predictors of mortality, rehospitalization for syncope, and cardiac syncope in 352 consecutive elderly patients with syncope.
      ,
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Using a motorized tilt table with foot support, we had a first 5-minute supine control phase, and then we moved the patients to the 60° upright position for a maximum duration of 45 min or until syncope developed [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ,
      • Khera S.
      • Palaniswamy C.
      • Aronow W.S.
      • et al.
      Predictors of mortality, rehospitalization for syncope, and cardiac syncope in 352 consecutive elderly patients with syncope.
      ,
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. However, at 20 min, we administered to the patients 400 mg of nitroglycerin spray sublingually [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ,
      • Khera S.
      • Palaniswamy C.
      • Aronow W.S.
      • et al.
      Predictors of mortality, rehospitalization for syncope, and cardiac syncope in 352 consecutive elderly patients with syncope.
      ,
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. To date, at the time of syncope, we immediately tilted back the patients to the horizontal position [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ,
      • Khera S.
      • Palaniswamy C.
      • Aronow W.S.
      • et al.
      Predictors of mortality, rehospitalization for syncope, and cardiac syncope in 352 consecutive elderly patients with syncope.
      ,
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. We defined the syncope as an abrupt, transient loss of consciousness and a loss of postural tone associated with bradycardia, hypotension, or both [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ,
      • Khera S.
      • Palaniswamy C.
      • Aronow W.S.
      • et al.
      Predictors of mortality, rehospitalization for syncope, and cardiac syncope in 352 consecutive elderly patients with syncope.
      ,
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Therefore, we classified the VVS into three groups according to HUT response during the onset of syncope: i) vasodepressor VVS by evidence of a decrease in systolic blood pressure to <60 mmHg without a decrease in HR during symptoms; ii) cardioinhibitory by evidence of an abrupt decrease in HR by ≥20 % without any antecedent decrease in systolic blood pressure; iii) mixed response by evidence of a concurrent decrease in systolic blood pressure to <60 mmHg and a decrease in HR by ≥20 % compared with averages 4 min before the onset of symptoms [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ,
      • Khera S.
      • Palaniswamy C.
      • Aronow W.S.
      • et al.
      Predictors of mortality, rehospitalization for syncope, and cardiac syncope in 352 consecutive elderly patients with syncope.
      ,
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ].

      2.4 MIBG scintigraphy

      At hospital admission for the VVS clinical event and before performing the HUT, the authors practiced the 123I-MIBG myocardial scintigraphy. We used a standardized protocol at the time of hospitalization and at 1 year of follow-up [
      • Paolisso P.
      • Bergamaschi L.
      • Rambaldi P.
      • Gatta G.
      • Foà A.
      • Angeli F.
      • Fabrizio M.
      • Casella G.
      • Barbieri M.
      • Galiè N.
      • Marfella R.
      • Pizzi C.
      • Sardu C.
      Impact of admission hyperglycemia on heart failure events and mortality in patients with takotsubo syndrome at long-term follow-up: data from HIGH-GLUCOTAKO investigators.
      ]. The 123I-MIBG myocardial scintigraphy was performed in VVS patients to assess the cardiac sympathetic nerve activity [
      • Kochiadakis G.
      • Marketou M.
      • Koukouraki S.
      • Parthenakis F.
      • Chlouverakis G.
      • Karkavitsas N.
      • Vardas P.
      Cardiac autonomic disturbances in patients with vasovagal syndrome: comparison between iodine-123-metaiodobenzylguanidine myocardial scintigraphy and heart rate variability.
      ]. Moreover, we used the 123I-MIBG, a norepinephrine analogous, to calculate the late heart-to-mediastinum ratio (H/Mlate) and washout rate (WR). Thus, we evaluated the H/Mlate as the index of global neuronal function due to norepinephrine uptake and the WR as the index of sympathetic tone. We withheld the drugs interfering with 123I-MIBG uptake, and thyroid uptake of unbound 123I was blocked with 500 mg of potassium perchlorate given orally 30 min before 123I-MIBG injection [
      • Paolisso P.
      • Bergamaschi L.
      • Rambaldi P.
      • Gatta G.
      • Foà A.
      • Angeli F.
      • Fabrizio M.
      • Casella G.
      • Barbieri M.
      • Galiè N.
      • Marfella R.
      • Pizzi C.
      • Sardu C.
      Impact of admission hyperglycemia on heart failure events and mortality in patients with takotsubo syndrome at long-term follow-up: data from HIGH-GLUCOTAKO investigators.
      ,
      • Kochiadakis G.
      • Marketou M.
      • Koukouraki S.
      • Parthenakis F.
      • Chlouverakis G.
      • Karkavitsas N.
      • Vardas P.
      Cardiac autonomic disturbances in patients with vasovagal syndrome: comparison between iodine-123-metaiodobenzylguanidine myocardial scintigraphy and heart rate variability.
      ]. We injected intravenously at rest, the dose ranging from 148 MBq to 370 MBq of 123I-MIBG. Then, we acquired both planar and SPECT images at 15 min after injection (early) and 4 h after injection (delayed, by the use of a dual-head gamma camera - ECAM Siemens, Erlangen, Germany) equipped with a low-energy, high-resolution collimator. A 20 % window was usually centered over the 159-keV photopeak of 123I for imaging. Anterior planar images of the chest for global assessment of cardiac innervation were acquired using a 256 × 256 matrix. Thereafter, we acquired the SPECT images for the regional evaluation, using a 64 × 64 matrix over 180°, from the right anterior oblique position to the left posterior diagonal position. Thus, we performed the quantitative evaluations with a standard protocol from this imaging [
      • Paolisso P.
      • Bergamaschi L.
      • Rambaldi P.
      • Gatta G.
      • Foà A.
      • Angeli F.
      • Fabrizio M.
      • Casella G.
      • Barbieri M.
      • Galiè N.
      • Marfella R.
      • Pizzi C.
      • Sardu C.
      Impact of admission hyperglycemia on heart failure events and mortality in patients with takotsubo syndrome at long-term follow-up: data from HIGH-GLUCOTAKO investigators.
      ]. Moreover, we evaluated at baseline and at 1 year of follow-up the H/Mlate and WR in the study cohorts.
      The formula to calculate WR was: WR BKG corrected = (He-Me) – [(Hl-Ml) × 1.21] (He-Me)
      The BKG is background; H is heart mean counts per pixel; M is mediastinum mean counts per pixel; “e” is early; “l” is late; and 1.21 is the correction factor for 123I decay at 3 h and 45 min [
      • Paolisso P.
      • Bergamaschi L.
      • Rambaldi P.
      • Gatta G.
      • Foà A.
      • Angeli F.
      • Fabrizio M.
      • Casella G.
      • Barbieri M.
      • Galiè N.
      • Marfella R.
      • Pizzi C.
      • Sardu C.
      Impact of admission hyperglycemia on heart failure events and mortality in patients with takotsubo syndrome at long-term follow-up: data from HIGH-GLUCOTAKO investigators.
      ,
      • Kochiadakis G.
      • Marketou M.
      • Koukouraki S.
      • Parthenakis F.
      • Chlouverakis G.
      • Karkavitsas N.
      • Vardas P.
      Cardiac autonomic disturbances in patients with vasovagal syndrome: comparison between iodine-123-metaiodobenzylguanidine myocardial scintigraphy and heart rate variability.
      ].

      2.5 Study endpoints

      The study endpoints were: 1) the evaluation of the HRV parameters in SGLT2-I-users vs. Non-SGLT2-I users at baseline and at 1 year of follow-up; 2) the evaluation of cardiac 123I-MIBG indexes in SGLT2-I-users vs. Non-SGLT2-I users at baseline and at 1 year of follow-up; 3) the rate of VVS recurrence after HUT diagnosis in SGLT2-I-users vs. Non-SGLT2-I users at 1 year of follow-up.

      3. Statistical analysis

      We made the descriptive statistical analysis using correct measurements on the collected data. However, before the HUT we divided the VVS patients into SGLT2-I-users vs. Non-SGLT2-I users. Then, at follow-up after HUT, and during follow-up visits and controls, we divided patients with VVS recurrence vs. patients without VVS recurrence. Thus, as appropriate, we tested the study variables by parametric and non-parametric tests. The normality of the distribution was tested with the Kolmogorov-Smirnov test. We compared the categorical variables by chi-square or Fisher exact test where appropriate. The statistical significance was set at p < 0.05 (two-sided tests), and for multiple testing, we used a statistical significance of p < 0.01.
      The PSM was constructed as a 2-category multinomial logistic regression model for the probability of study cohort to have VVS recurrence. Group 1 of patients (SGLT2-I users) were matched to group 2 (Non-SGLT2-I users) on PSM (group 1 vs. group 2); group 1 was the common referent for PSM. For the propensity score estimation, the following clinical characteristics were included as model predictors: age, BMI, current smoking, history of hypertension, dyslipidemia, smoking history, baseline therapies, metabolic characteristics, and other clinical variables. A nearest-neighbor algorithm on a 1:1 basis without replacement within a caliper of 0.01 was used to match patients. We checked covariates balance using the standardized difference of mean between groups with a threshold of 10 % to determine substantial imbalance. The Kaplan curves were made at 1 year of follow-up to show the cumulative risk to have VVS recurrence (all causes, mixed, cardio-inhibitory, and vasodepressor VVS recurrence). The Log Rank test evaluated the comparison between SGLT2-I users and Non-SGLT2-I users regarding the cumulative risk of VVS recurrence (all causes, mixed, cardio-inhibitory, and vasodepressor) VVS recurrence) at 1 year of follow-up. We made the multivariate Cox multivariate regression analysis to predict all causes of VVS recurrence at 1 year of follow-up in the study population. Among all risk factors and all clinical and angiographic parameters evaluated (age, smoking, resting HR, systolic and diastolic blood pressure, etc.), only the variables presenting a p ≤ 0.25 at univariate analysis were included in the model, and we used Hazard Ratios (HR) with 95 % confidence intervals was calculated. We performed the statistical analysis using the SPSS software package for Windows 17.0 (SPSS Inc., Chicago, Illinois).

      4. Results

      In the current prospective multicenter study, we included 324 T2DM patients with VVS, divided into 161 SGLT2-I-users vs. 163 Non-SGLT2-I users. The SGLT2-I users received either 10 mg (n 64), or 25 mg (n 45) of empagliflozin once daily, and/or canaglifozin 100 mg daily (n 52) in this study.
      The baseline characteristics, tests of autonomic dysfunction, echocardiographic parameters, and MIBG index of autonomic dysfunction of the overall study population and of SGLT2-I-users vs. Non-SGLT2-I users are reported in Table 1. Notably, the cohorts of the study did not differ for investigated parameters at baseline. Table 1.
      Table 1Characteristics of the study population at baseline, and at follow-up end as SGLT2-I users (n 161) vs. Non-SGLT2-I users (n 163).
      Study variablesAt baselineAt 1 year of follow-up
      SGLT2-I users

      (n 161)
      Non-SGLT2-I users

      (n 163)
      PSGLT2-I users

      (n 161)
      Non-SGLT2-I users (n 163)P
      Clinical characteristics
      Age, median IQR (years)61 (41–74)57 (34–73)0.200//
      Gender (male %)81 (50.3)234 (52.5)0.681//
      Smokers19 (11.8)74 (16.6)0.24124 (14.9)26 (15.9)0.887
      BMI > 30 kg/m212 (7.4)38 (8.5)0.12112 (7.5)12 (7.4)0.850
      Dyslipidemia (%)53 (32.9)162 (36.3)0.51652 (32.3)53 (32.5)0.514
      Hypothyroidism (%)18 (11.2)52 (11.7)0.17410 (6.2)10 (6.1)0.078
      Systolic BP (mmHg)123 ± 17126 ± 190.134120 ± 16122 ± 170.136
      Diastolic BP (mmHg)72 ± 974 ± 80.43173 ± 976 ± 80.061
      Systemic Hypertension92 (57.1)266 (59.6)0.21481 (50.3)83 (50.9)0.148
      Prodromes (%)84 (52.2)236 (52.9)0.718//
      Episodes/year (n)2.5 ± 1.22.2 ± 1.50.128//
      Laboratory data
      White blood cells, median IQR × 1039.0 (7.0–9.2)8.9 (6.5–9.4)0.1249.3 (8.2–10.0)8.8 (6.6–9.5)0.024
      Statistical significant (p < 0.05).
      Platelets, ×103244.16 ± 74.21269.45 ± 78.150.882250.26 ± 78.05210.20 ± 74.150.621
      C reactive protein, mg4.71 ± 0.225.43 ± 0.370.2311.56 ± 0.172.47 ± 0.200.001
      Statistical significant (p < 0.05).
      Fasting glucose, median IQR (mmol)6.82 ± 1.867.11 ± 2.170.4026.0 ± 0.416.38 ± 0.520.001
      Statistical significant (p < 0.05).
      HB1Ac (%)6.6 (5.5–6.9)6.7 (5.4–7.0)0.2356.1 ± 0.816.4 ± 0.930.001
      Statistical significant (p < 0.05).
      Norepinephrine, median IQR (pg/ml)1765.12

      (1517.52–1920.18)
      1890.21

      (1630.09–2250.15)
      0.0981265.12 (1017.52–1414.36)1972.41 (1702.40–2323.24)0.001
      Statistical significant (p < 0.05).
      eGFR (ml/min/1.73 m2)61.8 ± 20.161.9 ± 19.90.81261.5 ± 19.961.3 ± 19.80.893
      BNP, pg/ml53.02 ± 23.8851.42 ± 1951 20.50928.78 ± 19.2839.86 ± 20.870.002
      Statistical significant (p < 0.05).
      Autonomic dysfunction tests
      Resting heart rate (bpm)70 ± 1472 ± 150.10865 ± 1270 ± 110.001
      Statistical significant (p < 0.05).
      Deep breathing; HR (ratio)1.23 ± 0.121.26 ± 0.090.209//
      Valsalva maneuver (ratio)1.13 ± 0.121.16 ± 0.150.184//
      Lying to standing

      (30:15 ratio)
      1.06 ± 0.111.11 ± 0.080.125//
      Postural BP changes (mmHg)9.2 ± 3.18.9 ± 2.50.227//
      Sustained handgrip test (mmHg)15.2 ± 3.015.0 ± 4.00.972//
      LF, normalized units84.21 ± 4.8783.88 ± 4.800.43183.76 ± 4.7087.15 ± 4.570.001
      Statistical significant (p < 0.05).
      HF, normalized units18.28 ± 1.5318.45 ± 1.420.19719.22 ± 0.7915.97 ± 1.620.001
      Statistical significant (p < 0.05).
      LF/HF ratio4.64 ± 0.464.58 ± 0.490.1754.48 ± 0.325.52 ± 0.660.001
      Statistical significant (p < 0.05).
      ECG Holter parameters
      Mean NN802.71 ± 68.52806.35 ± 68.860.556805.06 ± 70.51781.97 ± 61.900.010
      Statistical significant (p < 0.05).
      SDNN173,67 ± 30.96174.49 ± 31.390.774184,12 ± 30.63166.12 ± 25.820.001
      Statistical significant (p < 0.05).
      SDANN166.26 ± 38.39170.35 ± 38.190.245173.27 ± 35.16160.92 ± 36.030.001
      Statistical significant (p < 0.05).
      SD81.62 ± 17.3882.31 ± 17.500.89887.32 ± 16.3480.51 ± 16.890.001
      Statistical significant (p < 0.05).
      RMSSD62.03 ± 15.3263.11 ± 15.220.59168.84 ± 16.0857.25 ± 16.300.001
      Statistical significant (p < 0.05).
      pNN5023.06 ± 7.6023.93 ± 7.880.22724.85 ± 8.7420.32 ± 6.500.001
      Statistical significant (p < 0.05).
      123I-MIBG myocardial scintigraphy parameters
      Heart to mediastinum ratio2.03 ± 0.571.96 ± 0.510.2152.53 ± 0.342.05 ± 0.520.001
      Statistical significant (p < 0.05).
      Washout rate (%)49.10 ± 7.6748.62 ± 9.88023639.79 ± 6.1536.38 ± 9.310.132
      Echocardiographic parameters
      IVS (mm)10.6 ± 1.410.7 ± 1.40.52710.5 ± 1.510.8 ± 1.30.482
      LVEDv (ml/m2)50.2 ± 11.851.1 ± 11.20.22351.6 ± 12.152.3 ± 11.50.261
      LVESv (ml/m2)19.1 ± 5.619.7 ± 5.90.30719.7 ± 5.919.9 ± 6.10.342
      LVEF (%)58 ± 660 ± 50.45362 ± 661 ± 40.582
      LAV (ml/m2)26.1 ± 2.626.3 ± 2.80.57126.5 ± 2.826.9 ± 2.90.307
      Cardiovascular medications
      Beta-blockers (%)42 (26.1)124 (27.8)0.20136 (22.4)44 (27.0)0.201
      Calcium blockers (%)27 (16.8)76 (17.0)0.54720 (12.4)21 (12.9)0.606
      ACE inhibitors (%)67 (41.6)190 (42.6)0.62560 (37.3)65 (39.9)0.341
      ARS blockers (%)22 (13.7)62 (13.9)0.51820 (12.4)22 (13.5)0.499
      Loop diuretics (%)24 (14.9)86 (19.3)0.21819 (11.8)30 (18.4)0.258
      Thiazides diuretics (%)30 (18.6)100 (22.4)0.36729 (18.0)36 (22.1)0.309
      Statins (%)66 (41.0)184 (41.2)0.42356 (34.8)55 (33.7)0.389
      Class 1 antiarrhythmic drugs (%)16 (9.9)46 (10.3)0.82516 (9.9)16 (9.8)0.825
      Class 3 antiarrhythmic drugs (%)4 (2.5)14 (3.1)0.3624 (2.5)5 (3.1)0.362
      Digitalis (%)11 (6.9)36 (8.1)0.41011 (6.9)13 (8.0)0.410
      Anti platelets (%)52 (32.3)136 (30.5)0.70752 (32.3)50 (30.7)0.707
      Anticoagulants (%)10 (6.2)44 (9.9)0.20110 (6.2)13 (7.9)0.201
      Hypoglycemic drugs
       Metformin81 (50.3)232 (52.0)0.27585 (52.8)87 (53.4)0.568
       Sulfonylureas18 (11.2)54 (12.1)0.48418 (11.2)19 (11.6)0.484
       Glinides7 (4.3)20 (4.5)0.1947 (4.3)8 (4.9)0.194
       Glitazones8 (5)22 (4.9)0.2028 (5)9 (5.5)0.202
       Incretins21 (13.0)58 (13.0)0.82124 (14.9)026 (15.9)0.342
       Insulin therapy32 (19.9)90 (20.2)0.26235 (21.7)38 (23.3)0.318
      Syncope recurrence events (%)
      Total number of events32780.001
      Mixed (%)15 (42.8)31 (39.8)0.139
      Cardio-inhibitory (%)4 (12.5)8 (10.2)0.224
      Vasodepressor (%)13 (40.6)39 (50.0)0.004
      Statistical significant (p < 0.05).
      ACE: angiotensin-converting enzyme; ARS: angiotensin receptors; BMI: body mass index; BNP: B type natriuretic peptide; BP: blood pressure; CRP: C reactive protein; eGFR: estimated Glomerular Filtration Rate; HB1Ac: glycated hemoglobin 1Ac; HF: high frequency; HR: heart rate; IQR: interquartile range; IVS: inter-ventricular septum; LF: low frequency; LF/HF ratio: low frequency/high frequency ratio; LAV: left atrium volume; LVEDv: left ventricle end-diastolic volume; LVESv: left ventricle end-systolic volume; LVEF: left ventricular ejection fraction; pNN50: the proportion of adjacent normal RR intervals differing by 50 ms; n: number; NN: mean of all coupling intervals between normal beats; RMSSD: the root-mean-square of the difference between successive RRs; SDNN: the standard deviation about the mean of all coupling intervals between normal beats; SDANN: the standard deviation of 5 min mean NN intervals; SD: the mean of all 5 min standard deviations of NNs; SGLT2-I: sodium-glucose transporter 2 inhibitors.
      The bold character expressed the word used for indicatibg hte exams used as example "laboratory data; autonomic dysfunction texts etc.".
      low asterisk Statistical significant (p < 0.05).

      4.1 At 1 year of follow-up

      the SGLT2-I-users vs. Non-SGLT2-I users had significantly lower values of the mean values of fasting glucose, Hb1Ac, CRP, WBC, norepinephrine, resting HR, and BNP (p < 0.05). Table 1. The SGLT2-I-users vs. Non-SGLT2-I users evidenced significant LF and HF values differences, with the lowest LF/HFr (p < 0.05). Table 1. Conversely, SGLT2-I-users vs. Non-SGLT2-I users had a significant difference for all the indexes of autonomic dysfunction via ECG Holter analysis (p < 0.05). Table 1. The SGLT2-I-users vs. Non-SGLT2-I users had significantly higher values of the Heart to Mediastinum ratio (p < 0.05). Table 1.
      Finally, SGLT2-I-users had lower rate of VVS recurrence events vs. Non-SGLT2-I users, specifically of the vasodepressor VVS recurrence at 1-year follow-up (p < 0.05). Table 1. We did not find a significant difference of mixed and cardio-inhibitory VVS recurrence events at 1 year of follow-up in the study cohorts (p > 0.05). Table 1.
      Notably, comparing the baseline vs. follow-up data in the SGLT2-I users and Non-SGLT2-I users, we noted in SGLT2-I users a significant reduction of CRP, serum norepinephrine, HR, LF/HFr, and WHR (p < 0.05). Fig. 2. Conversely, these patients evidenced a significant increase of H/Mlate (p < 0.05). Fig. 2. In the Non-SGLT2-I users, we found a significant reduction of CRP and WHR, with a significant increase of LF/HFr (p < 0.05). Fig. 2.
      Fig. 2
      Fig. 2C reactive protein, norepinephrine values, heart rate, LF/HF ratio, Heart to Mediastinum ratio, and Washout rate (%) at baseline vs. 1 year of follow-up in SGLT2-I users (red color) and Non-SGLT2-I users (blu color). LF: low frequency; HF: high frequency; SGLT2-I: sodium-glucose transporter 2 inhibitors. *is for statistical significant vs. follow-up in SGLT2-I users (p < 0.05); **is for statistical significant vs. follow-up in Non-SGLT2-I users (p < 0.05). (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)
      Finally, the number hypoglycemic attacks were not different comparing the SGLT2-I users (8 patients, 5 %) vs. Non-SGLT2-I users (6 patients, 4.3 %; p > 0.05).

      4.2 The Cox regression analysis evaluated the risk factors to predict all causes of VVS recurrence at 1 year of follow-up in the study population (Table 3)

      To date, Heart to Mediastinum rate (0.710, CI 95 % [0.481–0.985]; p 0.024), and SGLT2-I therapy (0.550, CI 95 % [0.324–0.934]; p 0.027) predicted all causes of syncope recurrence at 1 year of follow-up (Table 2).
      Table 2Multivariate Cox regression analysis for all causes of syncope recurrence at 1 year of follow-up. BMI: body mass index; CRP: C reactive protein; HR: Hazard ratio; LVEF: left ventricular ejection fraction; LF/HF ratio: low frequency/high frequency ratio; SGLT2-I: sodium-glucose transporter 2 inhibitors.
      All causes of vasovagal syncope recurrenceMultivariate analysis

      HR (95 % CI)
      P-value
      Age0.763 [0.125–1.018]0.135
      Smoking1.271 [0.839–1.927]0.258
      BMI1.024 [0.506–2.070]0.158
      Prodromes0.992 [0.944–1.043]0.075
      Hypertension0.751 [0.471–1.198]0.229
      Systolic blood pressure0.999 [0.950–1.050]0.960
      Glycemia0.934 [0.677–1.289]0.677
      CRP1.043 [0.747–1.455]0.805
      Noradrenaline1.001 [0.998–1.003]0.618
      LVEF1.005 [0.973–1.038]0.749
      Heart rate1.007 [0.993–1.021]0.323
      LF/HF ratio0.708 [0.458–1.096]0.122
      Heart to Mediastinum rate0.710 [0.481–0.985]0.024
      Confidence interval (CI) 95 %, p < 0.05.
      Washout rate0.994 [0.972–1.016]0.568
      Beta blockers1.007 [0.607–1.670]0.980
      SGLT2-I0.550 [0.324–0.934]0.027
      Confidence interval (CI) 95 %, p < 0.05.
      low asterisk Confidence interval (CI) 95 %, p < 0.05.
      Finally, the Kaplan curves showed the cumulative risk to have all causes of syncope (A), vaso-depressor syncope (B), cardio-inhibitory syncope (C), and mixed syncope (D) recurrence in SGLT2-I-users vs. Non-SGLT2-I users at 1 year of follow-up (Fig. 3).
      Fig. 3
      Fig. 3Kaplan curve for syncope recurrence (upper on the left), mixed syncope recurrence (upper right), cardio-inhibitory syncope recurrence (lower left), and vaso-depressor syncope recurrence (lower right) at 1 year of follow-up comparing SGLT2-I users (red color) vs. Non-SGLT2-I users (blu color). SGLT2-i: sodium-glucose transporter 2 inhibitors. *is for statistical significant (p < 0.05). (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)
      In Fig. 4, we showed the representative cases of 123I-mIBG myocardial scintigraphy with SGLT2-I-users of no-VVS recurrence and Non-SGLT2-I users of VVS recurrence (pre and 1 year follow up).
      Fig. 4
      Fig. 4The planar images of 123I-metaiodobenzylguanidine (MIBG) myocardial scintigraphy for representative cases of SGLT2-I-users without VVS recurrence and Non-SGLT2-I users with VVS recurrence at baseline and 1 year follow up. The images are reported as early and late 123I-MIBG values at baseline and follow-up end for SGLT2-I-users without VVS recurrence (panel A and B, respectively), and for Non-SGLT2-I users with VVS recurrence (panel C and D, respectively). In green color and with rectangular shape the mediastinum capitation of the 123I-MIBG; in red color and with round shape the heart capitation of 123I-MIBG. In the superior part of any panel (A, B, C, and D) the representative values of H/M and WHR (%). H/M: Heart to Mediastinum ratio; WHR: and Washout rate (%); SGLT2-I-users: sodium-glucose transporter 2 inhibitors; VVS: vaso vagal syncope. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

      5. Discussion

      The principal findings of the current prospective multicenter study are that the SGLT2-I could modulate the autonomic system and reduce the VVS recurrence and the vasodepressor VVS recurrence in T2DM patients at 1 year of follow-up. Notably, Non-SGLT2-I users vs. SGLT2-I users had over-inflammation, worse glycemic control, higher blood values of catecholamines, and more severe dysfunction of the autonomic system at follow-up end. Finally, the Non-SGLT2-I users vs. SGLT2-I users showed a higher recurrence rate of all causes of VVS and vasodepressor VVS at 1 year of follow-up. Notably, we noted that, compared to the baseline vs. follow-up data, the SGLT2-I users had a significant reduction of CRP, serum norepinephrine, HR, LF/HFr, and WHR, with a significant increase H/Mlate. Moreover, the SGLT2-I treatment could induce an ameliorative regulation of sympathetic systemic tone. Moreover, this effect could reduce cardiac autonomic dysfunction and cardiac denervation. These effects were lost in the Non-SGLT2-I users, those patients with worse prognoses (higher VVS recurrence).
      Conversely, we would point out that the SGLT2-I users' patients differed regards those receiving 10 mg vs. 25 mg daily of empagliflozin, and a third group receiving canaglifozin 100 mg daily. On the other hand, the two-dose groups for empaglifozin, and the group under canaglifozin had a similar hazard ratio for cardiovascular outcomes. In any way, we might confirm the cardio-protective effects of the SGLT2-I in the T2DM with VVS recurrence.
      The SGLT2-I could significantly reduce the volume overload and the blood pressure [
      • Matthews V.B.
      • Elliot R.H.
      • Rudnicka C.
      • Hricova J.
      • Herat L.
      • Schlaich M.P.
      Role of the sympathetic nervous system in regulation of the sodium-glucose cotransporter2.
      ], with anti-inflammatory properties and the modulation of the autonomic system [
      • D'Onofrio N.
      • Sardu C.
      • Trotta M.C.
      • Scisciola L.
      • Turriziani F.
      • Ferraraccio F.
      • Panarese I.
      • Petrella L.
      • Fanelli M.
      • Modugno P.
      • Massetti M.
      • Marfella L.V.
      • Sasso F.C.
      • Rizzo M.R.
      • Barbieri M.
      • Furbatto F.
      • Minicucci F.
      • Mauro C.
      • Federici M.
      • Balestrieri M.L.
      • Paolisso G.
      • Marfella R.
      Sodium-glucose co-transporter2 expression and inflammatory activity in diabetic atherosclerotic plaques: effects of sodium-glucose co-transporter2 inhibitor treatment.
      ]. In our study, the SGLT2-I users vs. Non-SGLT2-I users had a significant down-regulation of inflammatory markers, and norepinephrine serum values at the follow-up end. Intriguingly, the Non-SGLT2-I vs. SGLT2-I users' patients evidenced the significant increase of LF/HFr, which is an index of cardiac autonomic dysfunction in patients with VVS [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Therefore, the increase of LF/HFr in the Non-SGLT2-I users could be due to an increase in the sympathetic component and a decrease of the vagal component [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Indeed, the LF is modulated by both the sympathetic and parasympathetic nervous system, and associated with baroreceptor activity. In contrast, HF is modulated by the parasympathetic nervous system and connected with respiration and blood pressure changes [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Therefore, the LF/HFr is an index reflecting the interactions of both types of autonomic modulation with the regulation of heart rate and blood pressure [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. To date, the SGLT2-I could reduce autonomic dysfunction, with the best balance between vaso-vagal and sympathetic tone axis, and baroreceptor activity in patients with VVS, then leading to a reduction of VVS recurrence.
      In this setting, the cardiac 123I-MIBG could evaluate the cardiac denervation and the over-sympathetic tone in T2DM patients [
      • Paolisso P.
      • Bergamaschi L.
      • Rambaldi P.
      • Gatta G.
      • Foà A.
      • Angeli F.
      • Fabrizio M.
      • Casella G.
      • Barbieri M.
      • Galiè N.
      • Marfella R.
      • Pizzi C.
      • Sardu C.
      Impact of admission hyperglycemia on heart failure events and mortality in patients with takotsubo syndrome at long-term follow-up: data from HIGH-GLUCOTAKO investigators.
      ] and patients with VVS recurrence [
      • Kochiadakis G.
      • Marketou M.
      • Koukouraki S.
      • Parthenakis F.
      • Chlouverakis G.
      • Karkavitsas N.
      • Vardas P.
      Cardiac autonomic disturbances in patients with vasovagal syndrome: comparison between iodine-123-metaiodobenzylguanidine myocardial scintigraphy and heart rate variability.
      ,
      • Garg V.
      • Verma S.
      • Connelly K.A.
      • Yan A.T.
      • Sikand A.
      • Garg A.
      Does empagliflozin modulate the autonomic nervous system among individuals with type 2 diabetes and coronary artery disease? The EMPA-HEART CardioLink-6 Holter analysis.
      ,

      W Shimizu Y Kubota Y Hoshika K Mozawa S Tara Y Tokita et al. EMBODY trial investigators. Effects of empagliflozin versus placebo on cardiac sympathetic activity in acute myocardial.

      ]. Intriguingly, in our investigation, the SGLT2-I-users showed the significant increase of H/Mlate at follow-up end (vs. baseline condition), and highest values of H/Mlate compared to Non-SGLT2-I users. The H/Mlate is an index of cardiac denervation and a marker of autonomic system dysfunction in VVS patients [
      • Garg V.
      • Verma S.
      • Connelly K.A.
      • Yan A.T.
      • Sikand A.
      • Garg A.
      Does empagliflozin modulate the autonomic nervous system among individuals with type 2 diabetes and coronary artery disease? The EMPA-HEART CardioLink-6 Holter analysis.
      ,

      W Shimizu Y Kubota Y Hoshika K Mozawa S Tara Y Tokita et al. EMBODY trial investigators. Effects of empagliflozin versus placebo on cardiac sympathetic activity in acute myocardial.

      ]. Indeed, the increase of H/Mlate could indicate a higher heart uptake of 123I, which is analogous of norepinephrine [
      • Kochiadakis G.
      • Marketou M.
      • Koukouraki S.
      • Parthenakis F.
      • Chlouverakis G.
      • Karkavitsas N.
      • Vardas P.
      Cardiac autonomic disturbances in patients with vasovagal syndrome: comparison between iodine-123-metaiodobenzylguanidine myocardial scintigraphy and heart rate variability.
      ]. Therefore, the increase of H/Mlate could indirectly reflect an amelioration of cardiac denervation in patients with VVS [
      • Garg V.
      • Verma S.
      • Connelly K.A.
      • Yan A.T.
      • Sikand A.
      • Garg A.
      Does empagliflozin modulate the autonomic nervous system among individuals with type 2 diabetes and coronary artery disease? The EMPA-HEART CardioLink-6 Holter analysis.
      ,

      W Shimizu Y Kubota Y Hoshika K Mozawa S Tara Y Tokita et al. EMBODY trial investigators. Effects of empagliflozin versus placebo on cardiac sympathetic activity in acute myocardial.

      ]. In our setting, its increase has been seen in the arm of SGLT2-I treatment. Moreover, after 12 months of treatment with SGLT2-I, we might speculate about observing an increase of the cardiac norepinephrine uptake. Notably, at the clinical level, the SGLT2-I-users vs. Non-SGLT2-I users had a lower rate of VVS recurrence at the follow-up end. Thus, we might speculate that the SGLT2-I might regulate the sympathetic nervous system and autonomic dysfunction in patients with VVS, inducing a significant reduction of VVS recurrence. Conversely, the over-activation of the sympathetic system might cause the over-expression of SGLT2 pathways, while on the contrary, the SGLT2-I (block of SGLT2 pathways) might mediate inhibitory effects on the sympathetic system [
      • Sardu C.
      • D'Onofrio N.
      • Torella M.
      • Portoghese M.
      • Mureddu S.
      • Loreni F.
      • Ferraraccio F.
      • Panarese I.
      • Trotta M.C.
      • Gatta G.
      • Galdiero M.
      • Sasso F.C.
      • D'Amico M.
      • De Feo M.
      • Balestrieri M.L.
      • Paolisso G.
      • Marfella R.
      Metformin therapy effects on the expression of sodium-glucose cotransporter 2, leptin, and SIRT6 levels in pericoronary fat excised from pre-diabetic patients with acute myocardial infarction.
      ,
      • Mahaffey K.W.
      • Jardine M.J.
      • Bompoint S.
      • Cannon C.P.
      • Neal B.
      • Heerspink H.J.L.
      • Charytan D.M.
      • Edwards R.
      • Agarwal R.
      • Bakris G.
      • Bull S.
      • Capuano G.
      • de Zeeuw D.
      • Greene T.
      • Levin A.
      • Pollock C.
      • Sun T.
      • Wheeler D.C.
      • Yavin Y.
      • Zhang H.
      • Zinman B.
      • Rosenthal N.
      • Brenner B.M.
      • Perkovic V.
      Canagliflozin and cardiovascular and renal outcomes in type 2 diabetes mellitus and chronic kidney disease in primary and secondary cardiovascular prevention groups.
      ]. However, we might confirm the H/Mlate as an index of cardiac sympathetic innervation and adrenergic nervous system integrity and function [
      • Sisson J.C.
      • Wieland D.M.
      • Sherman P.
      • Mangner T.J.
      • Tobes M.C.
      • et al.
      Metaiodobenzylguanidine as an index of the adrenergic nervous system integrity and function.
      ]. However, we found that the highest values of H/Mlate could lead to the lowest risk of having VVS recurrence at follow-up end.
      On the other hand, the SGLT2-I therapy could inversely and independently predict the risk of having VVS recurrence at follow-up. Indeed, we reported that the SGLT2-I therapy could reduce about the ~45 % risk to have all causes of VVS recurrence. Therefore, the SGLT2-I might positively regulate/ameliorate the autonomic dysfunction in the patients with all causes of VVS, and with the vasodepressor VVS. This could consequently result in a lower rate of VVS recurrence in the SGLT2-I-users vs. Non-SGLT2-I users at 1 year of follow-up. In this context, the empagliflozin (SGLT2-I) at a 10-mg dosage significantly improved the HRV via modifying the HF, LF, and LF/HFr parameters [

      W Shimizu Y Kubota Y Hoshika K Mozawa S Tara Y Tokita et al. EMBODY trial investigators. Effects of empagliflozin versus placebo on cardiac sympathetic activity in acute myocardial.

      ]. Intriguingly, from a previous study, the LF/HFr is an independent predictor of VVS recurrence in T2DM patients [
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Therefore, here we might underline the SGLT2-I could act on autonomic dysfunction via the modulation of the LF, HF and LF/HFr parameters.
      Furthermore, the lowest values of LF/HRr, and the significant increase in the H/Mlate, could evidence the significant amelioration of autonomic function in SGLT2-I vs. Non-SGLT2-I users' patients with VVS. On the opposite, these parameters could evidence a more severe CAN in Non-SGLT2-I users vs. SGLT2-I-users' patients with VVS. Thus, we might suppose that the SGLT2-I therapy might cause a significant modification of vasovagal tone (HRV indexes) and of the cardiac sympathetic dysfunction (123I-MIBG parameters) in T2DM patients with VVS. Conversely, this might lead to the significant reduction of VVS recurrence in the SGLT2-I users vs. Non-SGLT2-I users at 1 year of follow-up. Moreover, the SGLT2-I might reduce inflammatory burden and ameliorate glucose homeostasis, hemodynamic, and CAN in T2DM patients with VVS. Indeed, the best control of glycemia, blood pressure, and hemodynamic could reduce the VVS recurrence in T2DM patients with VVS [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ,
      • Sardu C.
      • Paolisso P.
      • Santamaria M.
      • Sacra C.
      • Pieretti G.
      • Rizzo M.R.
      • Barbieri M.
      • Scisciola L.
      • Nicoletti G.
      • Paolisso G.
      • Marfella R.
      Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
      ]. Accordingly, the SGLT2-I users vs. Non-SGLT2-I users showed a significant reduction of BNP values at follow-up end. The BNP could contribute to VVS induced by HUT [
      • Jardine D.L.
      • Melton I.C.
      • Crozier I.G.
      • Bennett S.I.
      • Donald R.A.
      • Ikram H.
      Neurohormonal response to head-up tilt and its role in vasovagal syncope.
      ], exerting multiple actions on target tissues and binding to receptors implied on the heart, vasculature, and nervous system control [
      • Nilsson D.
      • Sutton R.
      • Melander O.
      • Fedorowski A.
      Spontaneous vs nitroglycerin-induced vasovagal reflex on head-up tilt: are there neuroendocrine differences?.
      ]. Thus, the modifications of BNP might markedly change in common forms of syncope, suggesting the involvement of the neurohormonal mechanisms in syncopal attacks [
      • Fedorowski A.
      • Burri P.
      • Struck J.
      • Juul-Moller S.
      • Melander O.
      Novel cardiovascular biomarkers in unexplained syncopal attacks: the SYSTEMA cohort.
      ]. Conversely, the effects of SGLT2-I on the BNP expression are somewhat inconsistent, showing both improvements [
      • Sezai A.
      • Sekino H.
      • Unosawa S.
      • Taoka M.
      • Osaka S.
      • Tanaka M.
      Canagliflozin for japanese patients with chronic heart failure and type II diabetes.
      ,
      • Lee M.M.Y.
      • Brooksbank K.J.M.
      • Wetherall K.
      • et al.
      Effect of empagliflozin on left ventricular volumes in patients with type 2 diabetes, or prediabetes, and heart failure with reduced ejection fraction (SUGAR-DM-HF).
      ], and demonstrating no substantial changes [
      • Jensen J.
      • Omar M.
      • Kistorp C.
      • et al.
      Empagliflozin in heart failure patients with reduced ejection fraction: a randomized clinical trial (Empire HF).
      ,
      • Jensen J.
      • Omar M.
      • Kistorp C.
      • et al.
      Empagliflozin in heart failure patients with reduced ejection fraction: a randomized clinical trial (Empire HF).
      ,
      • Yu Y.-W.
      • Zhao X.-M.
      • Wang Y.-H.
      • Zhou Q.
      • Huang Y.
      • Zhai M.
      • Zhang J.
      Effect of sodium–glucose cotransporter 2 inhibitors on cardiac structure and function in type 2 diabetes mellitus patients with or without chronic heart failure: a meta-analysis.
      ]. Intriguingly, in our study, the SGLT2-I could significantly reduce the BNP values. This might probably be due to a diuretic effect of SGLT2i, and switching of myocardial metabolism from glucose to ketone bodies and free fatty acids, with a more efficient ATP production [
      • Mudaliar S.
      • Polidori D.
      • Zambrowicz B.
      • Henry R.R.
      Sodium–glucose cotransporter inhibitors: effects on renal and intestinal glucose transport: from bench to bedside.
      ], and inhibition of cardiac sodium exchanger, thereby reducing intracellular calcium and increasing the mitochondrial calcium [
      • Baartscheer A.
      • Schumacher C.A.
      • Wüst R.C.
      • Fiolet J.W.
      • Stienen G.J.
      • Coronel R.
      • Zuurbier C.J.
      Empagliflozin decreases myocardial cytoplasmic Na+ through inhibition of the cardiac Na+/H+ exchanger in rats and rabbits.
      ]. However, these effects might modulate the autonomic system dysfunction and the blood pressure regulation, which are adaptive mechanisms leading to reflex syncope events [
      • Brignole M.
      • Rivasi G.
      • Sutton R.
      • et al.
      Low-blood pressure phenotype underpins the tendency to reflex syncope.
      ].
      Furthermore, the SGLT2-I might ameliorate hemodynamic and electrical function in patients with VVS via BNP pathways, and leading to counter-regulation and compensation of the heart rate and blood pressure abnormalities, and reduced blood supply to the brain during a syncope event [
      • Brignole M.
      • Rivasi G.
      • Sutton R.
      • et al.
      Low-blood pressure phenotype underpins the tendency to reflex syncope.
      ]. In this setting, an in vivo study showed that SGLT2i stabilized the circadian rhythms of blood pressure and sympathetic nervous system activity, reducing the blood pressure [
      • Takeshige Y.
      • Fujisawa Y.
      • Rahman A.
      • Kittikulsuth W.
      • Nakano D.
      • Mori H.
      • Masaki T.
      • Ohmori K.
      • Kohno M.
      • Ogata H.
      • et al.
      A sodium-glucose co-transporter 2 inhibitor empagliflozin prevents abnormality of circadian rhythm of blood pressure in salt-treated obese rats.
      ]. To date, the SGLT2-I could suppress the norepinephrine turnover and improve the baroreflex sensitivity in diabetic rats, restoring arterial pressure stability [
      • Yoshikawa T.
      • Kishi T.
      • Shinohara K.
      • Takesue K.
      • Shibata R.
      • Sonoda N.
      • Inoguchi T.
      • Sunagawa K.
      • Tsutsui H.
      • Hirooka Y.
      Arterial pressure lability is improved by sodium-glucose cotransporter 2 inhibitor in streptozotocin-induced diabetic rats.
      ]. Conversely, the SGLT2-I reduced the production of noradrenaline in the heart of high-fat-diet-fed mice by reducing inflammatory cytokines [
      • Matthews V.B.
      • Elliot R.H.
      • Rudnicka C.
      • Hricova J.
      • Herat L.
      • Schlaich M.P.
      Role of the sympathetic nervous system in regulation of the sodium glucose cotransporter 2.
      ]. In humans, SGLT2i mediated calorie loss and diuresis by sodium and glucose reabsorption blockade [
      • Heerspink H.J.L.
      • De Zeeuw D.
      • Wie L.
      • Leslie B.
      • List J.
      Dapagliflozin a glucose-regulating drug with diuretic properties in subjects with type 2 diabetes.
      ], with diuretic actions acting on the control of the baroreflex and sympathetic activity [
      • Scheen A.
      Effect of SGLT2 inhibitors on the sympathetic nervous system and blood pressure.
      ]. However, all these effects might suggest the regulative mechanisms of SGLT2i-on the human autonomic cardiovascular system [
      • Jordan J.
      • Jordan J.
      • Heusser K.
      • Heise T.
      • Wanner C.
      • Heer M.
      • Macha S.
      • Mattheus M.
      • Lund S.S.
      • Woerle H.J.
      • et al.
      The effect of empagliflozin on muscle sympathetic nerve activity in patients with type II diabetes mellitus.
      ]. By the way, the sympathetic nerves innervate the kidney's proximal tubules, regulating the expression of SGLT2 transporters [
      • Elliott R.H.
      • Matthews V.B.
      • Rudnicka C.
      • Schlaich M.P.
      Is it time to think about the sodium glucose co-transporter 2 sympathetically?.
      ]. Therefore, the effects of SGLT2-I on sympathetic nervous activity might be secondary to a reduction in renal stress with inhibition of renal afferent sympathetic activation [
      • Sano M.
      A new class of drugs for heart failure: SGLT2 inhibitors reduce sympathetic overactivity.
      ]. Finally, but no less relevant, in the central nervous system, SGLT2-I may act on the central nuclei-regulated cardiovascular system, then decreasing blood pressure and heart rate [
      • Nguyen T.
      • Wen S.
      • Gong M.
      • Yuan X.
      • Xu D.
      • Wang C.
      • Jin J.
      • Zhou L.
      Dapagliflozin activates neurons in the central nervous system and regulates cardiovascular activity by inhibiting SGLT-2 in mice.
      ]. Conversely, also the hypoglycemic attack could be responsible for the alteration of the sympathetic nervous system tone, thus favoring the VVS recurrence [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ]. On the other hand, the low risk of hypoglycemia under SGLT2-I therapy [
      • Zinman B.
      • Wanner C.
      • Lachin J.M.
      • Fitchett D.
      • Bluhmki E.
      • Hantel S.
      • Mattheus M.
      • et al.
      Empagliflozin, cardiovascular outcomes, and mortality in type 2 diabetes.
      ], could be due to insulin resistance and dysregulation of insulin secretion [
      Adverse events and their association with treatment regimens in the diabetes control and complications trial.
      ]. Notably, it could occur almost exclusively in older, long-term diabetic patients, especially with antidiabetic medication such as sulfonylurea or insulin [
      Adverse events and their association with treatment regimens in the diabetes control and complications trial.
      ]. Intriguingly, hypoglycemia could be the true cause of VVS, such as a possible false diagnostic factor for syncope [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ]. Indeed, the hypoglycemia could lead to impairment of consciousness instead of loss thereof, and cause much longer duration than the typically short in the syncope [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ]. Despite this, the hypoglycemia could lead to a short, self-limiting loss of consciousness, hence perfectly imitating a syncope [
      • Brignole M.
      • Moya A.
      • de Lange F.J.
      • et al.
      2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
      ]. However, the significant reduction of the cumulative risk to have VVS recurrence played by SGLT-I, added to the low risk of hypoglycemia, could be due to the chronic effect played by this class of anti-diabetic agents on the systemic and cardiac autonomic axis.

      5.1 Study limitations

      The current study evidenced few limitations. Firstly, we collected the data of VVS recurrence by hospital discharge schedules, visits of follow-up, and interrogation of patients' diaries. However, we revised the collected data and categorized the patients with vs. those without VVS recurrence. The duration of follow-up could affect long-term clinical outcomes. In addition, we did not implant internal loop recorders for continuous monitoring of syncope recurrence, and this could be limiting in the analysis of arrhythmic events in patients with VVS recurrence. Again, we did not investigate the effects of SGLT2-I at the molecular and cellular level in patients with VVS recurrence. Thus, we cannot report conclusive data about the cardio-protective and anti-inflammatory effects played by SGLT2-I [
      • Marfella R.
      • D'Onofrio N.
      • Trotta M.C.
      • Sardu C.
      • Scisciola L.
      • Amarelli C.
      • Balestrieri M.L.
      • Grimaldi V.
      • Mansueto G.
      • Esposito S.
      • D'Amico M.
      • Golino P.
      • Signoriello G.
      • De Feo M.
      • Maiello C.
      • Napoli C.
      • Paolisso G.
      Sodium/glucose cotransporter 2 (SGLT2) inhibitors improve cardiac function by reducing JunD expression in human diabetic hearts.
      ,
      • Sardu C.
      • Massetti M.
      • Testa N.
      • Di Martino L.
      • Castellano G.
      • Turriziani F.
      • Sasso F.C.
      • Torella M.
      • De Feo M.
      • Santulli G.
      • Paolisso G.
      • Marfella R.
      Effects of sodium-glucose transporter 2 inhibitors (SGLT2-I) in patients with ischemic heart disease (IHD) treated by coronary artery bypass grafting via MiECC: inflammatory burden, and clinical outcomes at 5 years of follow-up.
      ] in the patients with VVS recurrence. Again, in the current study, we measured the plasma norepinephrine for the evaluation of systemic sympathetic activity. This could be limiting and evidence intra-subject variability, but it is a valid and reproducible index of sympathetic activity, and used to evaluate the cardiac denervation by MIBG [
      • Paolisso P.
      • Bergamaschi L.
      • Rambaldi P.
      • Gatta G.
      • Foà A.
      • Angeli F.
      • Fabrizio M.
      • Casella G.
      • Barbieri M.
      • Galiè N.
      • Marfella R.
      • Pizzi C.
      • Sardu C.
      Impact of admission hyperglycemia on heart failure events and mortality in patients with takotsubo syndrome at long-term follow-up: data from HIGH-GLUCOTAKO investigators.
      ]. Indeed, the evaluations made are based on indirect assessment of the autonomic nervous system via serum norepinephrine and 123I-MIBG. In this setting, the 123I-MIBG does not measure the activity of the sympathetic nerves, nor does it take into consideration any change in postsynaptic receptor expression levels which we know change drastically in cardiovascular disease, while serum noradrenaline is affected not by release only but breakdown rates [
      • Kochiadakis G.
      • Marketou M.
      • Koukouraki S.
      • Parthenakis F.
      • Chlouverakis G.
      • Karkavitsas N.
      • Vardas P.
      Cardiac autonomic disturbances in patients with vasovagal syndrome: comparison between iodine-123-metaiodobenzylguanidine myocardial scintigraphy and heart rate variability.
      ]. Finally, the dimension of the study population could limit the importance of study results, and it cannot draw any definitive conclusion on the possible correlation between SGLT2-I therapy and VVS recurrence findings. Therefore, this remains a point to be demonstrated by an appropriately designed study conducted on a higher number of VVS patients with T2DM diagnosis under the SGLT2-I vs. other classes of hypoglycemic drugs.

      6. Conclusion

      In T2DM patients with VVS recurrence, the SLGT2i-users vs. Non-SLGT2i users evidenced a significant reduction of the inflammatory burden, with lowest LF/HFr and norepinephrine serum values, and highest (increase) H/Mlate values at the follow-up end. These effects SGLT2-I induced could evidence a more balanced autonomic system activity and an induced amelioration of CAN in T2DM patients with VVS. However, at the clinical level, the effects of SGLT2-I might cause the reduction of all causes of VVS and of vasodepressor VVS recurrence in SGLT2-I-users vs. Non-SGLT2-I users at follow-up end. Thus, the SGLT2-I might have positive implications in the treatment of VVS recurrence, and specifically of the vasodepressor VVS in T2DM patients.

      Ethics approval and consent to participate

      the study complies with the Declaration of Helsinki, and the locally appointed Ethics Committee approved the research protocol (n 06062.18), and informed consent was obtained from all the participants.

      Consent for publication

      The authors give full consent for publication.

      Availability of data and materials

      The study data are available on demand.

      Funding

      This research received no specific grant from any funding agency in the public, commercial or not-for-profit sectors.

      CRediT authorship contribution statement

      C.S.: wrote the research project, and wrote and edited the manuscript; P.R., G.G., M. S: data collection regarding autonomic dysfunction tests and VVS recurrence; M.V., V.D., G.S.: analyzed and interpreted the patient data regarding the VVS recurrence; M.M., S.C., F.C.S., G.P., R.M.: edited and revised the manuscript.

      Author agreement submission

      This original article has not been previously published. Upon acceptance, the Author assigns to the Metabolism the right to publish and distribute the manuscript in part or in its entirety. The Author's name will always be included with the publication of the manuscript. These exclusive rights run the full term of the copyright, and all renewals and extensions thereof. I hereby accept the terms of the above Author Agreement.

      Declaration of competing interest

      None to declare.

      Acknowledgments

      All the authors equally contributed to this research.

      References

        • Brignole M.
        • Moya A.
        • de Lange F.J.
        • et al.
        2018 ESC guidelines for the diagnosis and management of syncope the task force for the diagnosis and management of syncope of the european Society of Cardiology (ESC) developed with the special contribution of the european heart rhythm association (EHRA).
        Eur Heart J. 2018; 00: 1-69
        • Khera S.
        • Palaniswamy C.
        • Aronow W.S.
        • et al.
        Predictors of mortality, rehospitalization for syncope, and cardiac syncope in 352 consecutive elderly patients with syncope.
        J Am Med Dir Assoc. 2013 May; 14: 326-330
        • Sardu C.
        • Paolisso P.
        • Santamaria M.
        • Sacra C.
        • Pieretti G.
        • Rizzo M.R.
        • Barbieri M.
        • Scisciola L.
        • Nicoletti G.
        • Paolisso G.
        • Marfella R.
        Cardiac syncope recurrence in type 2 diabetes mellitus patients vs. normoglycemic patients: the CARVAS study.
        Diabetes Res Clin Pract. 2019 May; 151https://doi.org/10.1016/j.diabres.2019.04.015
        • Sardu C.
        • Marfella R.
        • Testa G.
        • Santamaria M.
        • Sacra C.
        • Ranauro A.
        • Paolisso G.
        • Rizzo M.R.
        • Barbieri M.
        ELectrophysiological mechanisms underlying the Inhibitory CArdiac syncope without asystolic significant pause: therapeutic and prognostic implications. The ELICA randomized trial.
        Medicine (Baltimore). 2018; 97e11757
        • Pfeifer M.A.
        • Weinberg C.R.
        • Cook D.L.
        • et al.
        Autonomic neural dysfunction in recently diagnosed diabetic subjects.
        Diabetes Care. 1984; 7: 447-453
        • Paolisso P.
        • Bergamaschi L.
        • Rambaldi P.
        • Gatta G.
        • Foà A.
        • Angeli F.
        • Fabrizio M.
        • Casella G.
        • Barbieri M.
        • Galiè N.
        • Marfella R.
        • Pizzi C.
        • Sardu C.
        Impact of admission hyperglycemia on heart failure events and mortality in patients with takotsubo syndrome at long-term follow-up: data from HIGH-GLUCOTAKO investigators.
        Diabetes Care. 2021 Sep; 44: 2158-2161https://doi.org/10.2337/dc21-0433
        • Kochiadakis G.
        • Marketou M.
        • Koukouraki S.
        • Parthenakis F.
        • Chlouverakis G.
        • Karkavitsas N.
        • Vardas P.
        Cardiac autonomic disturbances in patients with vasovagal syndrome: comparison between iodine-123-metaiodobenzylguanidine myocardial scintigraphy and heart rate variability.
        Europace. 2012 Sep; 14: 1352-1358https://doi.org/10.1093/europace/eus063
        • Zinman B.
        • Wanner C.
        • Lachin J.M.
        • Fitchett D.
        • Bluhmki E.
        • Hantel S.
        • Mattheus M.
        • et al.
        Empagliflozin, cardiovascular outcomes, and mortality in type 2 diabetes.
        N Engl J Med. 2015; 373: 2117-2128https://doi.org/10.1056/NEJMoa1504720
        • Hussein A.M.
        • Eid E.A.
        • Taha M.
        • Elshazli R.M.
        • Bedir R.F.
        • Lashin L.S.
        Comparative study of the effects of GLP1 analog and SGLT2 inhibitor against diabetic cardiomyopathy in type 2 diabetic rats: possible underlying mechanisms.
        Biomedicines. 2020; 8: 43
        • Stewart J.M.
        • Medow M.S.
        • Sutton R.
        • Visintainer P.
        • Jardine D.L.
        • Wieling W.
        Mechanisms of vasovagal syncope in the young: reduced systemic vascular resistance versus reduced cardiac output.
        J Am Heart Assoc. 2017 Jan 18; 6e004417https://doi.org/10.1161/JAHA.116.004417
        • American Diabetes Association
        2. Classification and diagnosis of diabetes: standards of medical care in diabetes-2021.
        Diabetes Care. 2021 Jan; 44 (S15-S33)
        • Matthews V.B.
        • Elliot R.H.
        • Rudnicka C.
        • Hricova J.
        • Herat L.
        • Schlaich M.P.
        Role of the sympathetic nervous system in regulation of the sodium-glucose cotransporter2.
        J Hypertens. 2017; 35: 2059-2068
        • D'Onofrio N.
        • Sardu C.
        • Trotta M.C.
        • Scisciola L.
        • Turriziani F.
        • Ferraraccio F.
        • Panarese I.
        • Petrella L.
        • Fanelli M.
        • Modugno P.
        • Massetti M.
        • Marfella L.V.
        • Sasso F.C.
        • Rizzo M.R.
        • Barbieri M.
        • Furbatto F.
        • Minicucci F.
        • Mauro C.
        • Federici M.
        • Balestrieri M.L.
        • Paolisso G.
        • Marfella R.
        Sodium-glucose co-transporter2 expression and inflammatory activity in diabetic atherosclerotic plaques: effects of sodium-glucose co-transporter2 inhibitor treatment.
        Mol Metab. 2021 Sep; 7101337https://doi.org/10.1016/j.molmet.2021.101337
        • Garg V.
        • Verma S.
        • Connelly K.A.
        • Yan A.T.
        • Sikand A.
        • Garg A.
        Does empagliflozin modulate the autonomic nervous system among individuals with type 2 diabetes and coronary artery disease? The EMPA-HEART CardioLink-6 Holter analysis.
        Metabol Open. 2020; 7 (7100039): 10039-100039https://doi.org/10.1016/j.metop.2020.100039
      1. W Shimizu Y Kubota Y Hoshika K Mozawa S Tara Y Tokita et al. EMBODY trial investigators. Effects of empagliflozin versus placebo on cardiac sympathetic activity in acute myocardial.

        • Sardu C.
        • D'Onofrio N.
        • Torella M.
        • Portoghese M.
        • Mureddu S.
        • Loreni F.
        • Ferraraccio F.
        • Panarese I.
        • Trotta M.C.
        • Gatta G.
        • Galdiero M.
        • Sasso F.C.
        • D'Amico M.
        • De Feo M.
        • Balestrieri M.L.
        • Paolisso G.
        • Marfella R.
        Metformin therapy effects on the expression of sodium-glucose cotransporter 2, leptin, and SIRT6 levels in pericoronary fat excised from pre-diabetic patients with acute myocardial infarction.
        Biomedicines. 2021 Jul 28; 9: 904https://doi.org/10.3390/biomedicines9080904
        • Mahaffey K.W.
        • Jardine M.J.
        • Bompoint S.
        • Cannon C.P.
        • Neal B.
        • Heerspink H.J.L.
        • Charytan D.M.
        • Edwards R.
        • Agarwal R.
        • Bakris G.
        • Bull S.
        • Capuano G.
        • de Zeeuw D.
        • Greene T.
        • Levin A.
        • Pollock C.
        • Sun T.
        • Wheeler D.C.
        • Yavin Y.
        • Zhang H.
        • Zinman B.
        • Rosenthal N.
        • Brenner B.M.
        • Perkovic V.
        Canagliflozin and cardiovascular and renal outcomes in type 2 diabetes mellitus and chronic kidney disease in primary and secondary cardiovascular prevention groups.
        Circulation. 2019 Aug 27; 140: 739-750https://doi.org/10.1161/CIRCULATIONAHA.119.042007
        • Sisson J.C.
        • Wieland D.M.
        • Sherman P.
        • Mangner T.J.
        • Tobes M.C.
        • et al.
        Metaiodobenzylguanidine as an index of the adrenergic nervous system integrity and function.
        J Nucl Med. 1987; 28 (pmid:3655914): 1620-1624
        • Jardine D.L.
        • Melton I.C.
        • Crozier I.G.
        • Bennett S.I.
        • Donald R.A.
        • Ikram H.
        Neurohormonal response to head-up tilt and its role in vasovagal syncope.
        Am J Cardiol. 1997; 79: 1302-1306
        • Nilsson D.
        • Sutton R.
        • Melander O.
        • Fedorowski A.
        Spontaneous vs nitroglycerin-induced vasovagal reflex on head-up tilt: are there neuroendocrine differences?.
        Heart Rhythm. 2016; 13: 1674-1678
        • Fedorowski A.
        • Burri P.
        • Struck J.
        • Juul-Moller S.
        • Melander O.
        Novel cardiovascular biomarkers in unexplained syncopal attacks: the SYSTEMA cohort.
        J Intern Med. 2013; 273: 359-367
        • Sezai A.
        • Sekino H.
        • Unosawa S.
        • Taoka M.
        • Osaka S.
        • Tanaka M.
        Canagliflozin for japanese patients with chronic heart failure and type II diabetes.
        Cardiovasc Diabetol. 2019; 18 ([PMC free article] [PubMed] [CrossRef] [Google Scholar]): 1-13https://doi.org/10.1186/s12933-019-0877-2
        • Lee M.M.Y.
        • Brooksbank K.J.M.
        • Wetherall K.
        • et al.
        Effect of empagliflozin on left ventricular volumes in patients with type 2 diabetes, or prediabetes, and heart failure with reduced ejection fraction (SUGAR-DM-HF).
        Circulation. 2020; 143
        • Jensen J.
        • Omar M.
        • Kistorp C.
        • et al.
        Empagliflozin in heart failure patients with reduced ejection fraction: a randomized clinical trial (Empire HF).
        Trials. 2019; 20 (374-4)
        • Jensen J.
        • Omar M.
        • Kistorp C.
        • et al.
        Empagliflozin in heart failure patients with reduced ejection fraction: a randomized clinical trial (Empire HF).
        Trials. 2019; 20 (374-4)
        • Yu Y.-W.
        • Zhao X.-M.
        • Wang Y.-H.
        • Zhou Q.
        • Huang Y.
        • Zhai M.
        • Zhang J.
        Effect of sodium–glucose cotransporter 2 inhibitors on cardiac structure and function in type 2 diabetes mellitus patients with or without chronic heart failure: a meta-analysis.
        Cardiovasc Diabetol. 2021; 20: 1-17
        • Mudaliar S.
        • Polidori D.
        • Zambrowicz B.
        • Henry R.R.
        Sodium–glucose cotransporter inhibitors: effects on renal and intestinal glucose transport: from bench to bedside.
        Diabetes Care. 2015; 38: 2344-2353
        • Baartscheer A.
        • Schumacher C.A.
        • Wüst R.C.
        • Fiolet J.W.
        • Stienen G.J.
        • Coronel R.
        • Zuurbier C.J.
        Empagliflozin decreases myocardial cytoplasmic Na+ through inhibition of the cardiac Na+/H+ exchanger in rats and rabbits.
        Diabetologia. 2017; 60: 568-573
        • Brignole M.
        • Rivasi G.
        • Sutton R.
        • et al.
        Low-blood pressure phenotype underpins the tendency to reflex syncope.
        J Hypertens. 2021; 39: 1319-1325
        • Takeshige Y.
        • Fujisawa Y.
        • Rahman A.
        • Kittikulsuth W.
        • Nakano D.
        • Mori H.
        • Masaki T.
        • Ohmori K.
        • Kohno M.
        • Ogata H.
        • et al.
        A sodium-glucose co-transporter 2 inhibitor empagliflozin prevents abnormality of circadian rhythm of blood pressure in salt-treated obese rats.
        Hypertens Res. 2016; 39: 415-422
        • Yoshikawa T.
        • Kishi T.
        • Shinohara K.
        • Takesue K.
        • Shibata R.
        • Sonoda N.
        • Inoguchi T.
        • Sunagawa K.
        • Tsutsui H.
        • Hirooka Y.
        Arterial pressure lability is improved by sodium-glucose cotransporter 2 inhibitor in streptozotocin-induced diabetic rats.
        Hypertens Res. 2017; 40: 646-651
        • Matthews V.B.
        • Elliot R.H.
        • Rudnicka C.
        • Hricova J.
        • Herat L.
        • Schlaich M.P.
        Role of the sympathetic nervous system in regulation of the sodium glucose cotransporter 2.
        J Hypertens. 2017; 35: 2059-2068
        • Heerspink H.J.L.
        • De Zeeuw D.
        • Wie L.
        • Leslie B.
        • List J.
        Dapagliflozin a glucose-regulating drug with diuretic properties in subjects with type 2 diabetes.
        Diabetes Obes Metab. 2013; 15: 853-862
        • Scheen A.
        Effect of SGLT2 inhibitors on the sympathetic nervous system and blood pressure.
        Curr Cardiol Rep. 2019; 21: 70
        • Jordan J.
        • Jordan J.
        • Heusser K.
        • Heise T.
        • Wanner C.
        • Heer M.
        • Macha S.
        • Mattheus M.
        • Lund S.S.
        • Woerle H.J.
        • et al.
        The effect of empagliflozin on muscle sympathetic nerve activity in patients with type II diabetes mellitus.
        J Am Soc Hypertens. 2017; 11: 604-612
        • Elliott R.H.
        • Matthews V.B.
        • Rudnicka C.
        • Schlaich M.P.
        Is it time to think about the sodium glucose co-transporter 2 sympathetically?.
        Nephrology. 2016; 21: 286-294
        • Sano M.
        A new class of drugs for heart failure: SGLT2 inhibitors reduce sympathetic overactivity.
        J Cardiol. 2018; 71: 471-476
        • Nguyen T.
        • Wen S.
        • Gong M.
        • Yuan X.
        • Xu D.
        • Wang C.
        • Jin J.
        • Zhou L.
        Dapagliflozin activates neurons in the central nervous system and regulates cardiovascular activity by inhibiting SGLT-2 in mice.
        Diabetes Metab Syndr Obes Targets Ther. 2020; 13: 2781-2799
      2. Adverse events and their association with treatment regimens in the diabetes control and complications trial.
        Diabetes Care. 1995 Nov; 18: 1415-1427
        • Marfella R.
        • D'Onofrio N.
        • Trotta M.C.
        • Sardu C.
        • Scisciola L.
        • Amarelli C.
        • Balestrieri M.L.
        • Grimaldi V.
        • Mansueto G.
        • Esposito S.
        • D'Amico M.
        • Golino P.
        • Signoriello G.
        • De Feo M.
        • Maiello C.
        • Napoli C.
        • Paolisso G.
        Sodium/glucose cotransporter 2 (SGLT2) inhibitors improve cardiac function by reducing JunD expression in human diabetic hearts.
        Metabolism. 2021 Nov; 18154936https://doi.org/10.1016/j.metabol.2021.154936. EX 20
        • Sardu C.
        • Massetti M.
        • Testa N.
        • Di Martino L.
        • Castellano G.
        • Turriziani F.
        • Sasso F.C.
        • Torella M.
        • De Feo M.
        • Santulli G.
        • Paolisso G.
        • Marfella R.
        Effects of sodium-glucose transporter 2 inhibitors (SGLT2-I) in patients with ischemic heart disease (IHD) treated by coronary artery bypass grafting via MiECC: inflammatory burden, and clinical outcomes at 5 years of follow-up.
        Front Pharmacol. 2021 Nov; 12: 3141https://doi.org/10.3389/fphar.2021.777083