Natural Hydrogen Sulfide Donors from Allium sp. as a Nutraceutical Approach in Type 2 Diabetes Prevention and Therapy

Type 2 diabetes mellitus (DM) is a socially relevant chronic disease with high prevalence worldwide. DM may lead to several vascular, macrovascular, and microvascular complications (cerebrovascular, coronary artery, and peripheral arterial diseases, retinopathy, neuropathy, and nephropathy), often accelerating the progression of atherosclerosis. Dietary therapy is generally considered to be the first step in the treatment of diabetic patients. Among the current therapeutic options, such as insulin therapy and hypoglycemic drugs, in recent years, attention has been shifting to the effects and properties—that are still not completely known—of medicinal plants as valid and inexpensive therapeutic supports with limited side effects. In this review, we report the relevant effects of medicinal plants and nutraceuticals in diabetes. In particular, we paid attention to the organosulfur compounds (OSCs) present in plant extracts that due to their antioxidant, hypoglycemic, anti-inflammatory, and immunomodulatory effects, can contribute as cardioprotective agents in type 2 DM. OSCs derived from garlic (Allium sp.), due to their properties, can represent a valuable support to the diet in type 2 DM, as outlined in this manuscript based on both in vitro and in vivo studies. Moreover, a relevant characteristic of garlic OSCs is their ability to produce the gasotransmitter H2S, and many of their effects can be explained by this property. Indeed, in recent years, several studies have demonstrated the relevant effects of endogenous and exogenous H2S in human DM, including by in vitro and in vivo experiments and clinical trials; therefore, here, we summarize the effects and the underlying molecular mechanisms of H2S and natural H2S donors.


Introduction
Diabetes mellitus (DM), as reported in the WHO 2016 global report, is a chronic disease with high incidence worldwide, creating a crucial social issue that represents one of four major noncommunicable diseases as outlined in world forums. The International Diabetes Federation recently estimated that DM affects about 425 million people and that this number will increase to 629 million in 2045. Type 2 DM is the most frequent form of the disease (over 90% of DM patients), characterized by hyperglycemia due to insulin resistance or inadequate insulin secretion [1]. Progressive hyperglycemia is one of the main causes of oxidative stress and is recognized to be principally responsible for type 2 DM complications [2][3][4]. Inflammation and oxidative stress are determinants for the loss of endothelial function and dysfunction of the vascular endothelium leading to macrovascular (cerebrovascular or

OSCs from Garlic as Nutraceuticals for Prevention and Therapy in Type 2 DM
Among the nutraceuticals described above, the plant extracts with organosulfur compounds (OSCs) deserve particular interest. Several studies have shown that OSCs and their different formulations inhibit insulin resistance and hyperglycemia, and they subsequently protect DM patients from several clinical effects, including cardiovascular complications. There are two main groups of vegetables characterized by the presence of OSCs with special properties: Brassicaceae and Amaryllidaceae. The first family includes cabbage, cauliflower, and Brussels sprouts, and kale and rucola (also known as rocket salad) are part of the Eruca genus of the mustard or cruciferous family; all of these produce S-methyl cysteine-l-sulfoxide [113]. The second one includes shallot, garlic, leek, onion, and chives; they belong to the Allium genus and produce S-alk(en)yl-l-cysteine sulfoxides. OSCs, contained in both these vegetable families, can be used as nutraceuticals and the mechanisms of action of either original produced sulfoxides or their derivatives have been studied in detail for their therapeutic effects. According to these investigations, type 2 DM patients eating broccoli sprouts, containing sulforaphane (1-isothiocyanato-4-(methylsulfinyl)butane), show increased total antioxidant capacity in their blood, serum insulin, and insulin resistance, with reduced lipid peroxidation, serum triglycerides, oxidative stress index, oxidized low-density lipoprotein (LDL)/LDL cholesterol ratio, and blood high-sensitivity C-reactive protein (CRP) [114]. Therefore, sulforaphane seems to reduce nephropathy, diabetic fibrosis, and vascular complications. The underlying molecular mechanism of sulforaphane seems to involve the Nrf2-related antioxidant response, elevation of phase 2 enzymes and PPARs, reduction of oxidative stress, and NF-κB (nuclear factor kappa light chain enhancer of activated B cells) activity reduction (with reduction of its related inflammation). According to these investigations, sulforaphane, as a component of young broccoli sprouts, is an excellent food additive for diabetic patients [114]. One of the most important glycemic-controlled herbal medicines with OSCs is garlic (A. sativum L.) [115,116]. Epidemiological and preclinical studies support the effects of garlic extract and its OSCs as cardiovascular-protective agents [117][118][119][120][121][122][123][124][125][126][127][128][129][130][131][132][133], due to the properties of these compounds, which are summarized in Figure 1.

OSCs from Garlic as Nutraceuticals for Prevention and Therapy in Type 2 DM
Among the nutraceuticals described above, the plant extracts with organosulfur compounds (OSCs) deserve particular interest. Several studies have shown that OSCs and their different formulations inhibit insulin resistance and hyperglycemia, and they subsequently protect DM patients from several clinical effects, including cardiovascular complications. There are two main groups of vegetables characterized by the presence of OSCs with special properties: Brassicaceae and Amaryllidaceae. The first family includes cabbage, cauliflower, and Brussels sprouts, and kale and rucola (also known as rocket salad) are part of the Eruca genus of the mustard or cruciferous family; all of these produce S-methyl cysteine-l-sulfoxide [113]. The second one includes shallot, garlic, leek, onion, and chives; they belong to the Allium genus and produce S-alk(en)yl-l-cysteine sulfoxides. OSCs, contained in both these vegetable families, can be used as nutraceuticals and the mechanisms of action of either original produced sulfoxides or their derivatives have been studied in detail for their therapeutic effects. According to these investigations, type 2 DM patients eating broccoli sprouts, containing sulforaphane (1-isothiocyanato-4-(methylsulfinyl)butane), show increased total antioxidant capacity in their blood, serum insulin, and insulin resistance, with reduced lipid peroxidation, serum triglycerides, oxidative stress index, oxidized low-density lipoprotein (LDL)/LDL cholesterol ratio, and blood high-sensitivity C-reactive protein (CRP) [114]. Therefore, sulforaphane seems to reduce nephropathy, diabetic fibrosis, and vascular complications. The underlying molecular mechanism of sulforaphane seems to involve the Nrf2-related antioxidant response, elevation of phase 2 enzymes and PPARs, reduction of oxidative stress, and NF-κB (nuclear factor kappa light chain enhancer of activated B cells) activity reduction (with reduction of its related inflammation). According to these investigations, sulforaphane, as a component of young broccoli sprouts, is an excellent food additive for diabetic patients [114]. One of the most important glycemic-controlled herbal medicines with OSCs is garlic (A. sativum L.) [115,116]. Epidemiological and preclinical studies support the effects of garlic extract and its OSCs as cardiovascular-protective agents [117][118][119][120][121][122][123][124][125][126][127][128][129][130][131][132][133], due to the properties of these compounds, which are summarized in Figure 1. Garlic shows powerful effects in DM, such as hypoglycemia, hyperinsulinemia, hypotriglyceridemia, anti-glycation, hypocholesterolemia, and anti-lipidperoxidation effects [116,117] (Figure 2). Garlic, either dried or fresh, and its derivatives show antihyperglycemic effects in genetic animal models of DM [116,[134][135][136][137] and clinically in humans [138,139]. Garlic improves insulin sensitivity and the associated metabolic syndrome in animal models [134], and its derivatives reduce both insulin resistance [67] and blood glucose in streptozotocin-induced and alloxan-induced DM mellitus in rats and mice [140,141]. These beneficial effects are attributed to the presence of OSCs, such as derivatives from alliin and sulfoxide amino acids. The effect of garlic homogenate in  Garlic shows powerful effects in DM, such as hypoglycemia, hyperinsulinemia, hypotriglyceridemia, anti-glycation, hypocholesterolemia, and anti-lipidperoxidation effects [116,117] ( Figure 2). Garlic, either dried or fresh, and its derivatives show antihyperglycemic effects in genetic animal models of DM [116,[134][135][136][137] and clinically in humans [138,139]. Garlic improves insulin sensitivity and the associated metabolic syndrome in animal models [134], and its derivatives reduce both insulin resistance [67] and blood glucose in streptozotocin-induced and alloxan-induced DM Nutrients 2019, 11, 1581 7 of 25 mellitus in rats and mice [140,141]. These beneficial effects are attributed to the presence of OSCs, such as derivatives from alliin and sulfoxide amino acids. The effect of garlic homogenate in reducing heart hypertrophy and fructose-induced myocardial oxidative stress is due to activation of the PI3K/Akt/Nrf2-Keap1-dependent pathway [142]. Diabetic erectile dysfunction may be associated with an elevated level of ROS in penile tissue [143] and ROS formation prevention and the restoration of the erectile function by S-allyl cysteine (SAC), the main OSC in aged garlic extract, in diabetic rats was obtained by modulation of NADPH oxidase expression. Recent studies on SAC demonstrated its antidiabetic, antioxidant, anti-inflammatory, and neuroprotective properties [144,145]. Trials using raw garlic on type 2 DM patients have reported a significant lowering of glycaemia and lipid metabolism with a concomitant amelioration of redox metabolism (SOD, catalase, and GPx in erythrocytes) [146]. Similar effects have been reported by other trials following administration of garlic or garlic compounds [147,148]. Although several investigations using garlic or its extracts, both in animal models and in clinical trials, have shown a clear beneficial effect in the treatment of patients with DM, nonetheless, additional investigations are needed to further explore the benefits of garlic for patients with type 2 DM.   Garlic OSCs are spontaneously derived from allicin after cutting of the garlic cloves ( Figure 3) and are the principal active ingredients that are responsible for the beneficial effects of the garlic extracts. The alliin (S-allyl-cysteine sulfoxide) is metabolized to allicin (diallyl thiosulfinate) by alliinase, a carbon-sulfur lyase enzyme that can be released only by breaking the garlic cells. Subsequently, allicin rapidly undergoes nonenzymatic decomposition that transforms into a series of OSCs, such as diallyl monosulfide (DAS) and oil-soluble polysulfides, including diallyl disulfide (DADS) as a main product and diallyl trisulfide (DATS) (Figure 3).  These first compounds are responsible for the characteristic pungency of garlic [152]. Among the compounds with notable current interest, there are the so-called polysulfites, which are abundant constituents, especially in the essential oils of garlic. Zhao and colleagues [153] identified 16 compounds as the main components of commercial garlic essential oil, accounting for 97.44% of the total oil of A. sativum. These were diallyl trisulfide (DATS; 50.43%), diallyl disulfide (DADS; 25.30%), diallyl sulfide (DAS; 6.25%), diallyl tetrasulfide (DATES; 4.03%), 1,2-dithiolane (3.12%), allyl methyl disulfide (3.07%), 1,3-dithiane (2.12%), and allyl methyl trisulfide (2.08%).
Originally, the antidiabetic properties of allicin were demonstrated in rabbits by a reduction of fasting blood glucose, with comparable efficacy to the standard drug tolbutamide [154]. The heart-related complications in DM, such as suppression of myocardial fibrosis progression in streptozotocin-induced diabetic rats, can be reduced by allicin administration. The attenuation of apoptosis and fibrosis after allicin treatment was related to the inhibition of Bcl-2, CD95, connective tissue growth factor (CTGF), and transforming growth factor β1 (TGF-β1) protein expression, eventually preventing DM-induced cardiac complication progression [155]. Bcl-2 and CD95 drive the cell fate, while CTGF and TGF-β1 are highly sensitive myocardial fibrosis markers. Allicin substantially down-regulates both Bcl2 and CD95 in diabetic rats, and thereby reverses myocardial apoptosis remodeling [155]. Moreover, ventricular arrhythmias activated by Bcl-2 treatment in diabetic rats can be considerably suppressed by allicin. Electrophysiology experiments have also demonstrated that allicin attenuated the action potential duration by inhibiting the L-type calcium current (ICa-L) and improving the inward rectifier potassium current (IK1) [156]. Nephropathy is also a disease linked to DM and it is typically related to a high kidney weight/body ratio, blood urea, and creatinine, with a reduced creatinine clearance rate. Allicin treatment efficiently ameliorated the diabetic nephropathy in rats by preventing the effects on the TGF-1/p-ERK1/2 signaling pathway [157].
In order to ameliorate the efficacy and stability of allicin, it was conjugated with captopril to produce S-allyl-mercapto-captopril (CPSSA). Prolonged CPSSA administration reduces body weight gain, blood pressure, and blood glucose levels in Cohen-Rosenthal Diabetic Hypertensive mice.
All these data demonstrate that allicin can provide an important contribution to reducing obesity, hypertension, and diabetes, which are also important risk factors for cardiac and metabolic disease [158]. Therefore, allicin can prevent insulin resistance and other complications [159]. The molecular mechanism by which allicin reduces the pathologies related to the DM was also investigated, and it was related to its ability to scavenge ROS. Free radical generation, which is also due to hyperglycemia, could in fact be one of the primary causes of insulin resistance in DM and its related complications [160].
In vitro studies have demonstrated that allicin attenuated nicotinamide adenine dinucleotide phosphate oxidase (NOX) activation and ROS production when oxidized LDL-cholesterol was exposed to endothelial cells [159,161].
Other derivatives from allicin have been studied for their properties as adjuvants in diabetic pathologies. One of these derivatives is allyl methyl sulfide (AMS), which is one of the major bioactive components in garlic present in the volatile garlic fraction with antibacterial [162], antioxidant [163], and anticancer properties [164]. The administration of AMS to experimental hyperglycemic rats considerably enhanced glutathione (GSH) and vitamin C and E levels [165]. Moreover, AMS treatment, by way of its free radical scavenging property and control of free radicals in the liver, is able to increase the activities of antioxidant enzymes.
Antioxidants and anti-inflammatory phytochemicals have a crucial role in the prevention of acute liver damage [166]. Due to the hepatoprotective effects of AMS, its administration can reduce the elevation of hepatic injury enzymes [165]. Several studies have shown that AMS treatment improves hepatic cellular damage, thereby conquering diabetic complications. Beneficial effects in alleviating diabetic liver damage and improving the hepatic function were obtained, in addition to exertion of a better glycemic control through stimulating insulin secretion in the remnant β-cells and ameliorating inflammatory markers. Dietary administration of AMS exhibited significant preservation of the structural and functional integrity of hepatocytes, probably due to the attenuation of hyperglycemia-mediated oxidative stress [165]. Further in vivo and clinical studies are necessary to confirm the possibility of using this phytochemical for dietary treatment in DM.
Garlic OSCs and their conjugates are also optimal slow H 2 S-releasing agents [167][168][169] and are able to release H 2 S in a non-enzymatic reaction with intracellular GSH ( Figure 4B) [169]. A growing body of evidence has shown that H 2 S plays an important role in the disordered glucose metabolism [170,171] that is the most important features of DM. Therefore, garlic-derived OSC supplementation could increase H 2 S levels, help to restore kidney function, and represent a natural therapeutic strategy.  Basal CSE expression is quite low in islet β cells, but can be increased by high concentrations of glucose. H2S can affect insulin-secreting β cells, both inhibiting secretion of insulin from the cells [195,196] and protecting them against cellular apoptosis induced by various stimuli [193,197]. Deregulated production of insulin is the major reason for glycometabolism disorder, and therefore DM; this is because insulin is the only hormone that is able to decrease blood glucose. Some studies have demonstrated that the expression of CSE and CBS is significantly upregulated in both liver and pancreas in streptozotocin-induced diabetic rats compared to the control [198]. H2S administration to beta cell lines INS-1E and HIT-T15 cells attenuated insulin secretion triggered by a high concentration of glucose [195]. This inhibitory effect of H2S on insulin secretion is related to the opening of KATP channels [199]. During diabetic hyperglycemia, high levels of endogenous H2S can open KATP channels in islet β cell membrane, causing elevated polarization of the membrane potential and lower insulin secretion [195] ( Figure 5B). Moreover, exogenous H2S by NaHS inhibits L-type voltage-dependent Ca 2 C channels, further lowering insulin secretion in a KATP channel-independent manner [196]. Therefore, H2S inhibits insulin secretion by targeting several biochemical processes, such as activation of KATP channels, inhibition of ATP synthesis, and inactivation of L-type voltage-dependent Ca 2 C channels [170]. Of note is that the physiological synthesis of H2S inhibition in Zucker DM rats increased insulin release and reduced hyperglycemia [200]. Altogether, the above strongly support that the downregulation of the H2S system apparently promotes diabetic prevention and treatment. The regulation of endogenous H2S production can be very relevant in DM, i.e., at the early phases of diabetes, the administration of H2S may be beneficial, while at its late stage, inhibiting H2S generation may be a possible therapeutic strategy. Basal CSE expression is quite low in islet β cells, but can be increased by high concentrations of glucose. H2S can affect insulin-secreting β cells, both inhibiting secretion of insulin from the cells [195,196] and protecting them against cellular apoptosis induced by various stimuli [193,197]. Deregulated production of insulin is the major reason for glycometabolism disorder, and therefore DM; this is because insulin is the only hormone that is able to decrease blood glucose. Some studies have demonstrated that the expression of CSE and CBS is significantly upregulated in both liver and pancreas in streptozotocin-induced diabetic rats compared to the control [198]. H2S administration to beta cell lines INS-1E and HIT-T15 cells attenuated insulin secretion triggered by a high concentration of glucose [195]. This inhibitory effect of H2S on insulin secretion is related to the opening of KATP channels [199]. During diabetic hyperglycemia, high levels of endogenous H2S can open KATP channels in islet β cell membrane, causing elevated polarization of the membrane potential and lower insulin secretion [195] ( Figure 5B). Moreover, exogenous H2S by NaHS inhibits L-type voltage-dependent Ca 2 C channels, further lowering insulin secretion in a KATP channel-independent manner [196]. Therefore, H2S inhibits insulin secretion by targeting several biochemical processes, such as activation of KATP channels, inhibition of ATP synthesis, and inactivation of L-type voltage-dependent Ca 2 C channels [170]. Of note is that the physiological synthesis of H2S inhibition in Zucker DM rats increased insulin release and reduced hyperglycemia [200]. Altogether, the above strongly support that the downregulation of the H2S system apparently promotes diabetic prevention and treatment. The regulation of endogenous H2S production can be very relevant in DM, i.e., at the early phases of diabetes, the administration of H2S may be beneficial, while at its late stage, inhibiting H2S generation may be a possible therapeutic strategy. Basal CSE expression is quite low in islet β cells, but can be increased by high concentrations of glucose. H2S can affect insulin-secreting β cells, both inhibiting secretion of insulin from the cells [195,196] and protecting them against cellular apoptosis induced by various stimuli [193,197]. Deregulated production of insulin is the major reason for glycometabolism disorder, and therefore DM; this is because insulin is the only hormone that is able to decrease blood glucose. Some studies have demonstrated that the expression of CSE and CBS is significantly upregulated in both liver and pancreas in streptozotocin-induced diabetic rats compared to the control [198]. H2S administration to beta cell lines INS-1E and HIT-T15 cells attenuated insulin secretion triggered by a high concentration of glucose [195]. This inhibitory effect of H2S on insulin secretion is related to the opening of KATP channels [199]. During diabetic hyperglycemia, high levels of endogenous H2S can open KATP channels in islet β cell membrane, causing elevated polarization of the membrane potential and lower insulin secretion [195] ( Figure 5B). Moreover, exogenous H2S by NaHS inhibits L-type voltage-dependent Ca 2 C channels, further lowering insulin secretion in a KATP channel-independent manner [196]. Therefore, H2S inhibits insulin secretion by targeting several biochemical processes, such as activation of KATP channels, inhibition of ATP synthesis, and inactivation of L-type voltage-dependent Ca 2 C channels [170]. Of note is that the physiological synthesis of H2S inhibition in Zucker DM rats increased insulin release and reduced hyperglycemia [200]. Altogether, the above strongly support that the downregulation of the H2S system apparently promotes diabetic prevention and treatment. The regulation of endogenous H2S production can be very relevant in DM, i.e., at the early phases of diabetes, the administration of H2S may be beneficial, while at its late stage, inhibiting H2S generation may be a possible therapeutic strategy.

H 2 S-Releasing Agents for Prevention and a Therapeutic Approach in Type 2 DM
Hydrogen sulfide is one of three important endogenous gasotransmitters and is released in tissues from the metabolism of L-cysteine or polysulfides [170] (Figure 4A,B). Principally, the enzymatic production of H 2 S in mammalian cells is due to the cytosolic pyridoxal 5'-phosphate (PLP)-dependent enzymes cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE) and to the 3-mercaptopyruvate sulfurtransferase (MST) that is present in both the cytosol and mitochondria. H 2 S exerts relevant protective effects and has essential roles in the central nervous, respiratory, and cardiovascular systems. H 2 S is a physiological mediator able to limit inflammation and free radical damage by reacting with multiple oxidant stressors, including peroxynitrite [172], superoxide radical anion [173], and hydrogen peroxide [174], and by producing glutathione persulfide (GSSH) in mitochondria [175][176][177], a more efficient H 2 O 2 scavenging molecule than GSH. Its antioxidant activity is also due to activation of the Nrf2-ARE pathway [178] (Figure 5A). In the last few years, H 2 S donors have shown great therapeutic potential for widely diffused pathologies, such as cardiovascular [179][180][181], neurodegenerative [182][183][184], and gastrointestinal diseases [185,186]. Moreover, H 2 S seems to be able to protect islet beta cells from damage elicited by distinct toxic or stress events [187,188]. Both exogenous administration of H 2 S (NaHS) and stimulating endogenous H 2 S generation with L-cysteine seem to reduce programmed cell death [187]. The pathological loss of beta cells, but also the chronic inflammation of damaged islet cells, is the primary cause of several DM complications. DM can be, therefore, considered an inflammatory response disease, suggesting a possible anti-inflammatory therapy [189][190][191]. Of note is that the NaHS administration can significantly inhibit pro-inflammatory-factor-induced injury in primary cultured pancreatic beta cells and MIN6 cells [192]. The reduction of H 2 S by knockout of CSE in high-fat-diet-induced type 2 DM mice exhacerbates oxidative insults without insulin secretion or reduction of blood glucose levels [193]. Actually, endogenous H 2 S does not always work as a friend. Conversely, other authors demonstrated that H 2 S contributes to ER stress-mediated cellular apoptosis through activation of the p38 MAPK pathway [194], not fully in keeping with the work by Taniguchi et al. [192].   On these bases, we can conclude that the effects of H2S on insulin secretion can change at different phases of diabetic development. Therefore, in the early phases of the disease, hyperglycemia-induced CSE upregulation seems a beneficial mechanism for the patients and the increased H2S levels protect islet β cells by reducing oxidation and inflammation, and by inhibiting the autoimmune response. The development of diabetes leads to a further increase of H2S that can inhibit insulin secretion and reduce the overload of diabetic beta cells by the reduction of the ATP content, activation of KATP channels, or inhibition of L-type voltage-dependent calcium channels [201]. In persistent hyperglycemia conditions, an increase in endogenous H2S can trigger an ER stress response, and consequently apoptosis [194]. On these bases, we can conclude that the effects of H2S on insulin secretion can change at different phases of diabetic development. Therefore, in the early phases of the disease, hyperglycemia-induced CSE upregulation seems a beneficial mechanism for the patients and the increased H2S levels protect islet β cells by reducing oxidation and inflammation, and by inhibiting the autoimmune response. The development of diabetes leads to a further increase of H2S that can inhibit insulin secretion and reduce the overload of diabetic beta cells by the reduction of the ATP content, activation of KATP channels, or inhibition of L-type voltage-dependent calcium channels [201]. In persistent hyperglycemia conditions, an increase in endogenous H2S can trigger an ER stress response, and consequently apoptosis [194]. On these bases, we can conclude that the effects of H2S on insulin secretion can change at different phases of diabetic development. Therefore, in the early phases of the disease, hyperglycemia-induced CSE upregulation seems a beneficial mechanism for the patients and the increased H2S levels protect islet β cells by reducing oxidation and inflammation, and by inhibiting the autoimmune response. The development of diabetes leads to a further increase of H2S that can inhibit insulin secretion and reduce the overload of diabetic beta cells by the reduction of the ATP content, activation of KATP channels, or inhibition of L-type voltage-dependent calcium channels [201]. In persistent hyperglycemia conditions, an increase in endogenous H2S can trigger an ER stress response, and consequently apoptosis [194]. On these bases, we can conclude that the effects of H2S on insulin secretion can change at different phases of diabetic development. Therefore, in the early phases of the disease, hyperglycemia-induced CSE upregulation seems a beneficial mechanism for the patients and the increased H2S levels protect islet β cells by reducing oxidation and inflammation, and by inhibiting the autoimmune response. The development of diabetes leads to a further increase of H2S that can inhibit insulin secretion and reduce the overload of diabetic beta cells by the reduction of the ATP content, activation of KATP channels, or inhibition of L-type voltage-dependent calcium channels [201]. In persistent hyperglycemia conditions, an increase in endogenous H2S can trigger an ER stress response, and consequently apoptosis [194]. Although endogenous H2S production could have different effects in the stages of DM, several studies have demonstrated that the treatment with H2S-releasing agents can be important in reducing the damage induced by DM. Oxidative stress in DM leads to excessive autophagy with consequent vascular endothelial cell (EC) dysfunction.      Basal CSE expression is quite low in islet β cells, but can be increased by high concentrations of glucose. H 2 S can affect insulin-secreting β cells, both inhibiting secretion of insulin from the cells [195,196] and protecting them against cellular apoptosis induced by various stimuli [193,197]. Deregulated production of insulin is the major reason for glycometabolism disorder, and therefore DM; this is because insulin is the only hormone that is able to decrease blood glucose. Some studies have demonstrated that the expression of CSE and CBS is significantly upregulated in both liver and pancreas in streptozotocin-induced diabetic rats compared to the control [198]. H 2 S administration to beta cell lines INS-1E and HIT-T15 cells attenuated insulin secretion triggered by a high concentration of glucose [195]. This inhibitory effect of H 2 S on insulin secretion is related to the opening of K ATP channels [199]. During diabetic hyperglycemia, high levels of endogenous H 2 S can open K ATP channels in islet β cell membrane, causing elevated polarization of the membrane potential and lower insulin secretion [195] ( Figure 5B). Moreover, exogenous H 2 S by NaHS inhibits L-type voltage-dependent Ca 2 C channels, further lowering insulin secretion in a K ATP channel-independent manner [196]. Therefore, H 2 S inhibits insulin secretion by targeting several biochemical processes, such as activation of K ATP channels, inhibition of ATP synthesis, and inactivation of L-type voltage-dependent Ca 2 C channels [170]. Of note is that the physiological synthesis of H 2 S inhibition in Zucker DM rats increased insulin release and reduced hyperglycemia [200]. Altogether, the above strongly support that the downregulation of the H 2 S system apparently promotes diabetic prevention and treatment. The regulation of endogenous H 2 S production can be very relevant in DM, i.e., at the early phases of diabetes, the administration of H 2 S may be beneficial, while at its late stage, inhibiting H 2 S generation may be a possible therapeutic strategy.
On these bases, we can conclude that the effects of H 2 S on insulin secretion can change at different phases of diabetic development. Therefore, in the early phases of the disease, hyperglycemia-induced CSE upregulation seems a beneficial mechanism for the patients and the increased H 2 S levels protect islet β cells by reducing oxidation and inflammation, and by inhibiting the autoimmune response. The development of diabetes leads to a further increase of H 2 S that can inhibit insulin secretion and reduce the overload of diabetic beta cells by the reduction of the ATP content, activation of K ATP channels, or inhibition of L-type voltage-dependent calcium channels [201]. In persistent hyperglycemia conditions, an increase in endogenous H 2 S can trigger an ER stress response, and consequently apoptosis [194]. Although endogenous H 2 S production could have different effects in the stages of DM, several studies have demonstrated that the treatment with H 2 S-releasing agents can be important in reducing the damage induced by DM. Oxidative stress in DM leads to excessive autophagy with consequent vascular endothelial cell (EC) dysfunction. Several studies have shown that exogenous H 2 S administration is able to prevent arterial EC dysfunction by inhibition of excessive autophagy through the Nrf2-ROS-AMPK signaling pathway [202]. NaHS treatment ameliorated myocardial autophagy, and more generally, the myocardial fibrosis, which is a predominant pathological characteristic of diabetic myocardial damage, by PI3K/Akt1 pathway activation [203]. High blood glucose levels and DM are implicated in neurodegeneration, and one of the hallmarks of this pathology is protein aggregation; H 2 S treatment could represent a novel strategy against protein aggregation in the diabetic brain [204]. Other common complications of DM are reduced angiogenesis and intractable wound lesions. H 2 S has been reported to have pro-angiogenic effects and H 2 S donors are able to promote diabetic wound healing by restoring endothelial progenitor cell (EPC) function and inducing an upregulation of in-wound skin tissue and EPCs [205]. In diabetic skin complications, H 2 S provided by NAC or NSHD-1, a synthetic slow H 2 S-releasing donor, can exert protective effects against DM-like injury [206,207]. More recently, several groups have produced slow H 2 S-releasing materials able to promote cell proliferation and migration and tissue repair, also reducing oxidative stress due to ROS [208,209]. A microparticle system comprising hydrophobic phase-change materials able to release H 2 S, termed NaHS@MPs, was produced for wound healing applications in DM [210]. In this study, significantly accelerated re-epithelialization and wound closure in diabetic mice was obtained using Tegaderm integrated with NaHS@MPs. Other H 2 S-releasing biomaterials for potential application in wound healing in DM have been obtained using OSCs derived from garlic. Wang et al. [211] demonstrated that mesoporous silica nanoparticles (MSNs) loaded with DATS, named DATS-MSN, and able to release H 2 S can stimulate endothelial cells proliferation and migration and have cytoprotective effects, reducing the inflammatory cytokines production and adhesion molecule expression. Other formulations of slow H 2 S-releasing microfibrous mats were produced by functionalization or doping with OSCs or oil-soluble extracts derived from garlic, named DADS-PFM/PFM+DADS and GaOS-PFM/ PFM+GaOS [212], and were shown to scavenge hydrogen peroxide, increasing pro-cell survival signaling, and at the same time, decreasing pro-apoptotic signaling. The development of slow H 2 S-releasing biomaterials opens new perspectives for applications of OSC H 2 S-releasing donors for the fabrication of biomedical devices. Functionalized biomaterials could then be used inside or outside the body for both non-implantable devices and patches for wound dressing and implantable vascular grafts and implants in order to reduce damage due to DM, improving the patient's health.

Conclusions
Currently the worldwide attention is focused on the development of prevention and treatment of diseases by daily consume of nutraceuticals, which can have a supportive role in preserving the life quality of the public. In this review we revised the state of the art on the use of nutraceuticals for prevention and adjuvant therapy of type 2 DM and its complications, focusing our attention in particular on nutraceuticals with sulfur and derived from Allium spp. The peculiarity of these nutraceuticals is their ability to release the gasotrasmitter H 2 S. Although endogenous H 2 S, as a signaling molecule, can show different effects at different stages of DM, several in vitro and in vivo studies have demonstrated that H 2 S donors can reduce the onset of DM and the damage it causes. However, more clinical studies are requested to support the validity of OSCs administration in the prevention and therapy of DM. In general, although several studied have demonstrated the beneficial effects of nutraceuticals in DM, one of the most important problems with natural compounds, including garlic OSCs, is their stability over time. Indeed, many garlic OSCs, such as allicin and its derivatives, can rapidly degrade even at low temperatures. Accordingly, several groups are developing promising strategies of administration of these natural compounds, such as capsules containing garlic oil self-nanoemulsifying systems [213] or nano-emulsions obtained in combination with other nutraceuticals, as we previously have shown with omega 3 and proteins [214], for improving their stability and bioavailability. Therefore, the production of new formulations with other nutraceuticals, having synergistic effects, may be relevant to obtaining good administration and reproducibility in clinical trials. Moreover, the variability of the chemical composition of the vegetables, which can vary with the environmental conditions and countries where they are produced, represents another relevant problem. Therefore, trials on the use of vegetables containing H 2 S donors should include information on their chemical composition and standardized preparations. In this context, the possibility to increase the production of the optimal H 2 S-releasing agents in the OSC-rich-vegetables should be explored in order to produce optimized food for daily usage as a prevention strategy and adjuvant cure for type 2 DM.