A novel dendritic mesoporous silica based sustained hydrogen sulfide donor for the alleviation of adjuvant-induced inflammation in rats

PurposeKetoprofen (KETO) is a traditional non-steroidal anti-inflammatory drug (NSAIDs) with good analgesic and antipyretic effects. However, as NASIDs, the toxicity of KETO towards gastrointestinal (GI) system might limit its clinical use. S-propargyl-cysteine (SPRC) is an excellent endogenous H2S donor showed wide application in the field of anti-inflammation, anti-oxidative stress, or even the protection of cardiovascular system through the elevation of endogenous H2S concentration. As recently studies reported, co-administration of H2S donor might potentially mitigate the GI toxicity and relevant side effects induced by series of NSAIDs.MethodsIn this study, we established a SPRC and KETO co-encapsulated poly (lactic-co-glycolic acid) microsphere (SK@MS), and its particle size, morphology, storage stability and in vitro release profile were firstly investigated. The elevation of endogenous H2S level of SK@MS was then calculated, and the pharmacodynamic study (anti-inflammation and analgesic effects) of SK@MS, SPRC, and KETO towards adjuvant induced arthritis (AIA) in rats were also studied. Finally, to test the potential side effect, the heart, liver, spleen, lung, kidney, stomach, small intestine, and large intestine were resected from rats and examined by H&E staining.ResultsA monodispersed SK@MS could be observed under the SEM, and particle size was calculated around 25.12 μm. The loading efficiency (LE) for SPRC and KETO were 6.67% and 2.64%, respectively, while the encapsulation efficiency (EE) for SPRC and KETO were 37.20% and 68.28%, respectively. SK@MS showed a sustained release of SPRC and KETO in vitro, which was up-to 15 days. SK@MS could achieve a long-term elevation of the H2S concentration in vivo, while SPRC showed an instant H2S elevation and metabolize within 6 h. Interestingly, the KETO did not show any influence on the H2S concentration in vivo. After establishment of AIA model, neither SPRC nor KETO showed scarcely anti-inflammation and anti-nociception effect, while conversely, SK@MS showed an obvious mitigation towards paw edema and pain in AIA rats, which indicated an improved anti-inflammation and anti-nociception effect when co-delivery of SRC and KETO. Besides, low stimulation towards major organs in rats observed in any experimental group.ConclusionA monodispersed was successfully prepared in this study, and SK@MS showed a sustained SPRC and KETO release in vitro and H2S release in vivo. In the pharmacodynamics study, SK@MS not only exhibited an excellent anti-inflammation and analgesic effects in AIA rats but also showed low stimulation towards rats.

We previously reported that S-propargyl-cysteine (SPRC) exerts cardioprotective effects by elevating H2S levels via the CSE/H2S pathway. In the present study, we investigated the cardioprotective effects and pharmacokinetic properties of a controlled release formulation of SPRC (CR-SPRC) in an in vivo rat model of myocardial infarction (MI). Rats were randomly assigned to seven groups that were pre-treated with CR-SPRC daily for 7 days prior to ligation of the left anterior descending coronary artery to induce MI. Cardiac function and infarct size were determined after MI, and we examined the activity of antioxidant enzymes, expression of anti-inflammation proteins and hydrogen sulfide levels. Mixed-mode, reversed-phase and cation-exchange HPLC-MS/MS were used to compare the pharmacokinetic properties of CR-SPRC and SPRC. CR-SPRC significantly reduced infarct size and creatine kinase (CK) and lactate dehydrogenase (LDH) leakage and it preserved cardiac function during MI. CR-SPRC displayed antioxidant properties, preserving glutathione (GSH), catalase (CAT) and superoxide dismutase (SOD) levels whereas reducing malondialdehyde (MDA) levels. Moreover, CR-SPRC significantly reduced the protein levels of inflammatory biomarkers (phospho-NF-κB p65/NF-κB p65, TNF-α) and increased cystathionine-γ-lyase (CSE) and Iκ-Bα protein levels. CR-SPRC had better pharmacokinetic properties than SPRC, with a reduced concentration peak (Cmax), prolonged time to reach peak concentration (Tmax), prolonged mean residence time (MRTinf) and increased AUC0-t. CR-SPRC showed protective effects against MI via the CSE/H2S pathway and demonstrated better cardioprotective effects than SPRC by prolonging the release of endogenous H2S.

BackgroundRheumatoid arthritis (RA) is a long-term autoimmune disorder characterized by chronic inflammation that results in swollen and painful joints and even cartilage and bone damage. The gut microbiota, a novel anti-inflammatory target, is considered an important environmental factor in the development of RA. S-propargyl-cysteine (SPRC), an amino acid analogue, exerts anti-inflammatory, cardioprotective effects, and neuroprotective effects on various diseases. In recent studies, an SPRC treatment exerted anti-inflammatory effects on RA. Meanwhile, gut microbiome dysbiosis in individuals with RA has also been reported by many researchers. However, the relationship between SPRC and gut microbiota in individuals with RA remains unclear.MethodsThirty male Sprague-Dawley (SD) rats were randomly divided into three groups of 10 each, including the Control, Model, and SPRC groups. Adjuvant-induced arthritis (AIA) rats in SPRC group were treated with SPRC. Measurement of paw volume and serum tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6) levels were applied to evaluate the inflammatory status. Fecal samples were collected on the 14th day and 28th day. Gut microbiota were analyzed using 16S ribosomal RNA (rRNA) gene amplicon sequencing. Untargeted metabolomics on plasma samples was applied to investigate the metabolic changes induced by the altered gut microbiota by using derivatization-UHPLC-Q-TOF/MS.FindingsUsing 16S rRNA amplicon sequencing, we found that SPRC significantly altered the gut microbiota structure in AIA rats. In particular, Bifidobacterium, a genus of BSH (Bile Salt Hydrolase)-producing microbes, was overrepresented in SPRC-treated AIA rats. Additionally, a subsequent metabolomics analysis indicated that bile acid metabolism was also altered by SPRC treatment. Interestingly, glycochenodeoxycholic acid (GCDCA) and glycocholic acid (GCA), which are formed with the participation of BSH-producing microbes in the intestine, were identified as crucial biomarkers responding to SPRC treatment with significantly lowered levels.InterpretationA mechanistic link between the gut microbiota and plasma metabolites was revealed in this study, which provides insights into the mechanism of SPRC treatment for RA from the perspective of the gut microbiota.

Aims: S-propargyl-cysteine (SPRC) is an endogenous hydrogen sulfide (H2S) donor obtained by modifying the structure of S-allyl cysteine in garlic. This study aims to investigate the effect of SPRC on mitigating cardiac aging and the involvement of jumonji domain-containing protein 3 (JMJD3), a histone demethylase, which represents the primary risk factor in major aging related diseases, in this process, elucidating the preliminary mechanism through which SPRC regulation of JMJD3 occurs. Results: In vitro, SPRC mitigated the elevated levels of reactive oxygen species, senescence-associated β-galactosidase, p53, and p21, reversing the decline in mitochondrial membrane potential, which represented a reduction in cellular senescence. In vivo, SPRC improved Dox-induced cardiac pathological structure and function. Overexpression of JMJD3 accelerated cardiomyocytes and cardiac senescence, whereas its knockdown in vitro reduced the senescence phenotype. The potential binding site of the upstream transcription factor of JMJD3, sheared X box binding protein 1 (XBP1s), was determined using online software. SPRC promoted the expression of cystathionine γ-lyase (CSE), which subsequently inhibited the IRE1α/XBP1s signaling pathway and decreased JMJD3 expression. Innovations: This study is the first to establish JMJD3 as a crucial regulator of cardiac aging. SPRC can alleviate cardiac aging by upregulating CSE and inhibiting endoplasmic reticulum stress pathways, which in turn suppress JMJD3 expression. Conclusions: JMJD3 plays an essential role in cardiac aging regulation, whereas SPRC can suppress the expression of JMJD3 by upregulating CSE, thus delaying cardiac aging, which suggests that SPRC may serve as an aging protective agent, and pharmacological targeting of JMJD3 may also be a promising therapeutic approach in age-related heart diseases. Antioxid. Redox Signal. 42, 301-320.

Designed synthesis of silica based nanocarriers for mRNA delivery

P24 Cardioprotective effects of S-propargyl-cysteine controlled release formulation to heart failure rats after myocardial infarction and the possible involved mechanism

Hydrogen sulfide, once considered as toxic gas, is now recognized as an important biological mediator. The deficiency of hydrogen sulfide could lead to various pathological changes, such as arterial and pulmonary hypertension, Alzheimer’s disease, gastric mucosal injury and liver cirrhosis. However, excessive production of hydrogen sulfide, by using inorganic hydrogen sulfide donors such as NaHS, may contribute to the pathogenesis of inflammatory diseases, septic shock, cerebral stroke and mental retardation in patients with Down syndrome. Therefore, an increasing interest in organic molecules that are capable of regulating the formation of hydrogen sulfide has extended in recent years. Allium vegetables are one natural source of organic sulfur-containing compounds and have been widely investigated regarding their therapeutic applications, and it has been proven that the ingredients of garlic, such as diallyl disulfide, diallyl trisulfide and S-ally cysteine act as hydrogen sulfide donors or mediators in pharmaceutical studies. In addition, S-propargyl cysteine (ZYZ-802) and S-propyl cysteine, two synthetic cysteine analogs, have been examined and could be used to treat ischemic heart disease via modulation of the hydrogen sulfide pathway. In addition, drugs containing hydrogen sulfide-releasing moieties have been synthesized and widely reported in recent years, such as S-nonsteroidal anti-inflammatory drugs and the derivative of Lawesson’s reagents, which exhibit varied biological effects in experiments. As cystathionine β-synthase and cystathionine γ-lyase are the enzymes that are able to catalyze the production of endogenous hydrogen sulfide from cysteine, their inhibitors, such as dl-propylargylglycine and β-cyanoalanine, have been frequently used in studies on the biological mechanism of hydrogen sulfide. All these hydrogen sulfide donors, mediators and inhibitors have provided useful tools in the research of a variety of biological effects and are promising drug candidates of hydrogen sulfide.

P19 S-propargyl-cysteine (SPRC) attenuates cognitive impairment in APP/PS1 mice and protects neurons against glial overactivation induced by Aβ1−40 via modulation of cystathionine-β-synthase

Mesoporous silica nanoparticles have drawn increasing attention as promising candidates in vaccine delivery. Previous studies evaluating silica-based vaccine delivery systems concentrated largely on macromolecular antigens, such as inactivated whole viruses. In this study, we synthesized dendritic mesoporous silica nanoparticles (DMSNs), and we evaluated their effectiveness as delivery platforms for peptide-based subunit vaccines. We encapsulated and tested in vivo an earlier reported foot-and-mouth disease virus (FMDV) peptide vaccine (B2T). The B2T@DMSNs formulation contained the peptide vaccine and the DMSNs without further need of other compounds neither adjuvants nor emulsions. We measured in vitro a sustained release up to 930 h. B2T@DMSNs-57 and B2T@DMSNs-156 released 23.7% (135 µg) and 22.8% (132 µg) of the total B2T. The formation of a corona of serum proteins around the DMSNs increased the B2T release up to 61% (348 µg/mg) and 80% (464 µg/mg) for B2T@DMSNs-57 and B2T@DMSNs-156. In vitro results point out to a longer sustained release, assisted by the formation of a protein corona around DMSNs, compared to the reference formulation (i.e., B2T emulsified in Montanide). We further confirmed in vivo immunogenicity of B2T@DMSNs in a particle size-dependent manner. Since B2T@DMSNs elicited specific immune responses in mice with high IgG production like the reference B2T@Montanide™, self-adjuvant properties of the DMSNs could be ascribed. Our results display DMSNs as efficacious nanocarriers for peptide-based vaccine administration.

Development of Ce/Cu co-doped dendritic mesoporous silica nanoparticles (DMSNs) as novel abrasive systems toward high-performance chemical mechanical polishing

EP04 Therapeutic modulators of endogenous H2S synthesis: Bench to bedside

To study the distribution, metabolism and excretion of S-propargyl-cysteine (SPRC), a novel hydrogen sulfide (H2S) donor, after oral administration in rats. Adult Sprague-Dawley rats were used. The tissue distribution of [(35)S] SPRC-derived radioactivity was measured using a liquid scintillation counter. The plasma protein binding of SPRC was examined using 96-well equilibrium dialysis. The excretion of SPRC in urine, bile and feces was analyzed using the LC-MS/MS method. The major metabolites in rat biomatrices were identified using MRM information-dependent, acquisition-enhanced product ion (MRM-IDA-EPI) scans on API 4000QTrap system. After oral administration of [(35)S]-SPRC at a dose of 75 mg/kg, [(35)S] SPRC-derived radioactivity displayed broad biological distribution in various tissues of rats, including its target organs (heart and brain) with the highest in kidney. On the other hand, the binding of SPRC to human, rat and dog plasma protein was low. Only 2.18% ± 0.61% and 0.77% ± 0.61% of the total SPRC administered was excreted unchanged in the bile and urine. However, neither intact SPRC nor its metabolites were detected in rat feces. The major metabolic pathway in vivo (rat bile, urine, and plasma) was N-acetylation. The preliminary results suggest that SPRC possesses acceptable pharmacokinetic properties in rats.

Histone deacetylase 6 (HDAC6) acts as a regulator of the nuclear factor kappa-B (NF-κB) signaling pathway by deacetylating the non-histone protein myeloid differentiation primary response 88 (MyD88) at lysine residues, which is an adapter protein for the Toll-like receptor (TLR) and interleukin (IL)-1β receptor. Over-activated immune responses, induced by infiltrated immune cells, excessively trigger the NF-κB signaling pathway in other effector cells and contribute to the development of rheumatoid arthritis (RA). It has also been reported that HDAC6 can promote the activation of the NF-κB signaling pathway. In the present study, we showed that HDAC6 protein level was increased in the synovium tissues of adjuvant-induced arthritis rats. In addition, hydrogen sulfide (H2S) donor S-propargyl-cysteine (SPRC) can inhibit HDAC6 expression and alleviate inflammatory response in vivo. In vitro study revealed that HDAC6 overexpression activated the NF-κB signaling pathway by deacetylating MyD88. Meanwhile, sodium hydrosulfide (NaHS) or HDAC6 inhibitor tubastatin A (tubA) suppressed the pro-inflammatory function of HDAC6. Furthermore, the reduced expression of HDAC6 appeared to result from transcriptional inhibition by S-sulfhydrating specificity protein 1 (Sp1), which is a transcription factor of HDAC6. Our results demonstrate that Sp1 can regulate HDAC6 expression, and S-sulfhydration of Sp1 by antioxidant molecular H2S ameliorates RA progression via the HDAC6/MyD88/NF-κB signaling pathway.

Hydrogen sulfide (H2S), a novel gaseous messenger, is synthesized endogenously from L-cysteine by two pyridoxal-5′-phosphate-dependent enzymes, cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE). S-propargyl-cysteine (SPRC) is a slow H2S releasing drug that provides cysteine, a substrate of CSE. The present study was aimed to investigate the effects of SPRC in an in vivo model of acute pancreatitis (AP) in mice. AP was induced in mice by hourly caerulein injections (50 µg/kg) for 10 hours. Mice were treated with SPRC (10 mg/kg) or vehicle (distilled water). SPRC was administered either 12 h before or 3 h before the induction of pancreatitis. Mice were sacrificed 1 h after the last caerulein injection. Blood, pancreas and lung tissues were collected and processed to measure the plasma amylase, plasma H2S, myeloperoxidase (MPO) activities and cytokine levels in pancreas and lung. The results revealed that significant reduction of inflammation, both in pancreas and lung was associated with SPRC given 3 h prior to the induction of AP. Furthermore, the beneficial effects of SPRC were associated with reduction of pancreatic and pulmonary pro-inflammatory cytokines and increase of anti-inflammatory cytokine. SPRC administered 12 h before AP induction did not cause significant improvement in pancreatic and lung inflammation. Plasma H2S concentration showed significant difference in H2S levels between control, vehicle and SPRC (administered 3 h before AP) treatment groups. In conclusion, these data provide evidence for protective effects of SPRC in AP possibly by virtue of its slow release of endogenous H2S.

Objective Sepsis is one of major reasons of cardiorenal syndrome type 5 (CRS-5), resulting in irreversible tissue damage and organ dysfunction. Macrophage has been demonstrated to play key role in the pathophysiology of sepsis, highlighting the need to identify therapeutic targets for modulating macrophage phenotype in sepsis. Methods and Results In this study, a rapid-releasing hydrogen sulfide (H2S) donor NaSH, and a slow-releasing H2S compound S-propargyl-cysteine (SPRC) which is derived from garlic, have been studied for the immune-regulatory effects on macrophages. The NaSH and SPRC showed the potential to protect the heart and kidney from tissue injury induced by LPS. The immunohistochemistry of F4/80+ revealed that the infiltration of macrophages in the heart and kidney tissues of LPS-treated mice was reduced by NaSH and SPRC. In addition, in the LPS-triggered inflammatory cascade of RAW264.7 macrophage cells, NaSH and SPRC exhibited significantly inhibitory effects on the secretion of inflammatory cytokines, production of reactive oxygen species (ROS), and regulation of the macrophage phenotype from M1-like to M2-like. Moreover, autophagy, a crucial process involved in the elimination of impaired proteins and organelles during oxidative stress and immune response, was induced by NaSH and SPRC in the presence of LPS stimulation. Consequently, there was an increase in the number of mitochondria and an improvement in mitochondrial membrane potential. This process was mainly mediated by PINK1/Parkin pathway mediated mitophagy. Discussion These results demonstrated that the immunoregulatory effects of H2S donors were through the PINK1/Parkin-mediated mitophagy pathway. Overall, our study provided a new therapeutic direction in LPS-induced cardiorenal injury.