Astragalin ameliorates renal injury in diabetic mice by modulating mitochondrial quality control via AMPK-dependent PGC1α pathway.

Diabetic kidney disease (DKD) is a common microvascular complication of diabetes mellitus, and oxidative stress and mitochondrial dysfunction play an important role in this process. It has been shown that aldose reductase (ALR2) catalyzes NADPH-dependent reduction of glucose to sorbitol, resulting in oxidative stress and mitochondrial dysfunction in diabetic patients. Astragalin (AG), a flavonoid extracted from Thesium chinense Turcz., shows an inhibitory activity on ALR2. In this study, we investigated the therapeutic effects of AG against renal injury in streptozocin (STZ)-induced diabetic mouse model. Diabetic mice were orally administered AG (5, 10 mg·kg-1·d-1) for 4 weeks. We showed that AG treatment greatly improved the proteinuria and ameliorated renal pathological damage without affecting the elevated blood glucose in diabetic mice. Furthermore, AG treatment significantly suppressed highly activated ALR2, and reduced oxidative stress in the kidney of diabetic mice and in high glucose and lipids-stimulated HK2 cells in vitro. We demonstrated that AG treatment modulated mitochondrial quality control and ameliorated apoptosis, boosting mitochondrial biogenesis, maintaining mitochondrial dynamic homeostasis, and improving energy metabolism disorder in vivo and in vitro. In high glucose and lipids-stimulated HK2 cells, we found that AG (20 µM) restored the phosphorylation level of AMPK, and upregulated the expression and transcriptional activity of PGC1α, whereas treatment with H2O2, blockade of AMPK with Compound C or knockdown of AMPKα with siRNA abolished the protective effect of AG on mitochondrial function, suggesting that antioxidant effects and activation of AMPK-dependent PGC1α pathway might be the molecular mechanisms underlying the protective effects of AG on mitochondrial quality control. We conclude that AG could be a promising drug candidate for the treatment of diabetic renal injury through activating AMPK.

Phloretin ameliorates diabetes-induced endothelial injury through AMPK-dependent anti-EndMT pathway.

Diabetic cardiovascular complications contribute more than half of diabetes mortality. Endothelial damage and subsequent pathological changes play a key role in this process. Phloretin, a plant-derived dihydrochalcone compound, was reported to have the activities in regulating metabolism homeostasis and anti-inflammation. However, its effects and the mechanism on early stage endothelial injury caused by diabetes are not clear yet. In our present study, human umbilical vein endothelial cells (HUVECs) were stimulated by high glucose or advanced glycation end products (AGEs) to induce endothelial damage, and streptozotocin (STZ) -induced diabetes mouse model was used for in vivo study. Our results showed that phloretin effectively reduced endothelial damage marker monocyte chemotactic protein-1 (MCP1) as well as pro-calcification factors bone morphogenetic protein-2 (BMP2) and receptor activator of NF-κB ligand (RANKL) expression, reversed the increased vimentin and decreased CD31 dose-dependently in vitro and in vivo. Phloretin had no effect on blood glucose level. However, it ameliorated endothelial injury and vascular fibrosis in diabetic mice. Further experiments revealed that phloretin could enhance AMP activated protein kinase (AMPK) activation and upregulate peroxidase proliferator activated receptor-gamma coactivator-lα (PGC1α) level, and inhibit the activation of TGFß-Smad2-Snail signalling pathway which was abrogated by AMPK inhibitor, providing a rational mechanism that AMPK activation was required for the effects of phloretin on endothelial injury and endothelial-mesenchymal transformation (EndMT). Our data reveal a new role of phloretin in protection of diabetic endothelial damage via AMPK-dependent anti-EndMT activation, and also provide a potential therapeutic way for diabetic endothelial damage and its subsequent cardiovascular complications.

A sensitive method for the quantification of short-chain fatty acids by benzyl chloroformate derivatization combined with GC-MS.

Short-chain fatty acids (SCFAs) were identified as critical markers in the diagnosis of chronic and metabolic diseases, but a sensitive and stable method to determine SCFAs in feces is a challenge for analysts due to the high volatility. Herein, a sensitive and accurate method to determine SCFAs adopting precolumn derivatization coupled with gas chromatography-mass spectrometry (GC-MS) has been developed. Benzyl chloroformate (BCF) was chosen as the reaction reagent and emulsified derivatization was applied to homogenize the reaction system. Higher sensitivity, wider application and satisfactory derivatization efficiency were obtained using the developed method. An excellent method validation showed a good linearity ranging from 0.9947 to 0.9998. At the same time, the intra-day and inter-day precision were achieved in the range of 0.56% to 13.07%. The lower limits of detection of all target analytes varied from 0.1 to 5 pg. The recovery ranged from 80.87% to 119.03%, and storage stability under three different conditions was also determined. This method was also successfully applied to the analysis of SCFAs in mice fecal samples to illustrate the significant differences between normal and type 2 diabetes mellitus mice.

Separation and determination of the enantiomers of lactic acid and 2-hydroxyglutaric acid by chiral derivatization combined with gas chromatography and mass spectrometry.

Lactic acid and 2-hydroxyglutaric acid are chiral metabolites that have two distinct d- and l-enantiomers with distinct biochemical properties. Perturbations of a single enantiomeric form have been found to be closely related to certain diseases. Therefore, the ability to differentiate the d and l enantiomers is important for these disease studies. Herein, we describe a method for the separation and determination of lactic acid and 2-hydroxyglutaric acid enantiomers by chiral derivatization (with l-menthol and acetyl chloride) combined with gas chromatography and mass spectrometry. The two pairs of above-mentioned enantiomers exhibited linear calibration curves with a correlation coefficient (R2 ) exceeding 0.99. The measured data were accurate in the acceptable recovery range of 88.17-102.30% with inter- and intraday precisions (relative standard deviations) in the range of 4.23-17.26%. The limits of detection for d-lactic acid, l-lactic acid, d-2-hydroxyglutaric acid, and l-2-hydroxyglutaric acid were 0.13, 0.11, 1.12, and 1.16 µM, respectively. This method was successfully applied to analyze mouse plasma. The d-lactic acid levels in type 2 diabetes mellitus mouse plasma were observed to be significantly higher (P < 0.05, t-test) than those of normal mice, suggesting that d-lactic acid may serve as an indicator for type 2 diabetes mellitus.

Screening of lipase inhibitors in Folium Mori with lipase-linked magnetic microspheres by high-performance liquid chromatography and evaluation in diabetic mice.

A new method for the screening of compounds with hypoglycemic effect from traditional Chinese medicines employing high-performance liquid chromatography and lipase-linked magnetic microspheres has been proposed. We hypothesized that the interaction of traditional Chinese medicine extracts with lipase-linked magnetic microspheres should decrease the concentration of compounds with hypoglycemic effect. Using this approach, the potential lipase inhibitors in Folium Mori extract were investigated. First, lipase was immobilized on magnetic microspheres by a chemical method. Then, by comparing the chromatograms of samples before and after the interaction with lipase-linked magnetic microspheres, seven compounds of Folium Mori extract were identified. It was found that protocatechuic acid, chlorogenic acid, protocatechualdehyde, rutin, isoquercitrin, astragalin, and dicaffeoylquinic acid B had evident combination with lipase-linked magnetic microspheres. Their hypoglycemic effects were verified in streptozocin-induced diabetic mice. In the present study, astragalin was verified to improve the glucose tolerance and lower the level of glucose in streptozocin-induced diabetic mice, which indicated that astragalin might be a new highly efficient lipase inhibitor. Based on these significant results, this method could be a convenient approach to screen potential lipase inhibitors from traditional Chinese medicines. Meanwhile, it also could be expanded to screen other active compounds in traditional Chinese medicines.

Dominant negative FADD dissipates the proapoptotic signalosome of the unfolded protein response in diabetic embryopathy.

Endoplasmic reticulum (ER) stress and caspase 8-dependent apoptosis are two interlinked causal events in maternal diabetes-induced neural tube defects (NTDs). The inositol-requiring enzyme 1α (IRE1α) signalosome mediates the proapoptotic effect of ER stress. Diabetes increases tumor necrosis factor receptor type 1R-associated death domain (TRADD) expression. Here, we revealed two new unfolded protein response (UPR) regulators, TRADD and Fas-associated protein with death domain (FADD). TRADD interacted with both the IRE1α-TRAF2-ASK1 complex and FADD. In vivo overexpression of a FADD dominant negative (FADD-DN) mutant lacking the death effector domain disrupted diabetes-induced IRE1α signalosome and suppressed ER stress and caspase 8-dependent apoptosis, leading to NTD prevention. FADD-DN abrogated ER stress markers and blocked the JNK1/2-ASK1 pathway. Diabetes-induced mitochondrial translocation of proapoptotic Bcl-2 members mitochondrial dysfunction and caspase cleavage were also alleviated by FADD-DN. In vitro TRADD overexpression triggered UPR and ER stress before manifestation of caspase 3 and caspase 8 cleavage and apoptosis. FADD-DN overexpression repressed high glucose- or TRADD overexpression-induced IRE1α phosphorylation, its downstream proapoptotic kinase activation and endonuclease activities, and apoptosis. FADD-DN also attenuated tunicamycin-induced UPR and ER stress. These findings suggest that TRADD participates in the IRE1α signalosome and induces UPR and ER stress and that the association between TRADD and FADD is essential for diabetes- or high glucose-induced UPR and ER stress.

Dapagliflozin ameliorates hepatic steatosis via suppressing LXRα-mediated synthesis of lipids and bile acids.

Nonalcoholic fatty liver disease (NAFLD) prevalence is rising globally with no pharmacotherapies approved. Hepatic steatosis is closely associated with progression and prognosis of NAFLD. Dapagliflozin, kind of sodium-glucose cotransporter 2 (SGLT2) inhibitor, was found to improve NAFLD in clinical trials, while the underlying mechanism remains poorly elucidated. Here, we reported that dapagliflozin effectively mitigated liver injury and relieved lipid metabolism disorders in vivo. Further investigation showed that dapagliflozin markedly suppressed Liver X Receptor α (LXRα)-mediated synthesis of de novo lipids and bile acids (BAs). In AML12 cells, our results proved dapagliflozin decreased lipid contents via inhibiting the expression of LXRα and downstream liposynthesis genes. Proteosome inhibitor MG132 eliminated the effect of dapagliflozin on LXRα-mediated signaling pathway, which suggested that dapagliflozin downregulated LXRα expression through increasing LXRα degradation. Knockdown of LXRα with siRNA abolished the reduction of lipogenesis from dapagliflozin treatment, indicating that LXRα might be the pivotal target for dapagliflozin to exhibit the aforementioned benefits. Furthermore, the data showed that dapagliflozin reversed gut dysbiosis induced by BAs disruption and altered gut microbiota profile to reduce intestinal lipids absorption. Together, our study deciphered a novel mechanism by which dapagliflozin relieved hepatic steatosis and highlighted the potential benefit of dapagliflozin in treating NAFLD.

Trehalose prevents neural tube defects by correcting maternal diabetes-suppressed autophagy and neurogenesis.

Preexisting maternal diabetes increases the risk of neural tube defects (NTDs). The mechanism underlying maternal diabetes-induced NTDs is not totally defined, and its prevention remains a challenge. Autophagy, an intracellular process to degrade dysfunction protein and damaged cellular organelles, regulates cell proliferation, differentiation, and apoptosis. Because autophagy impairment causes NTDs reminiscent of those observed in diabetic pregnancies, we hypothesize that maternal diabetes-induced autophagy impairment causes NTD formation by disrupting cellular homeostasis, leading to endoplasmic reticulum (ER) stress and apoptosis, and that restoration of autophagy by trehalose, a natural disaccharide, prevents diabetes-induced NTDs. Embryos from nondiabetic and type 1 diabetic mice fed with or without 2 or 5% trehalose water were used to assess markers of autophagy, ER stress, and neurogenesis, numbers of autophagosomes, gene expression that regulates autophagy, NTD rates, indices of mitochondrial dysfunction, and neuroepithelial cell apoptosis. Maternal diabetes suppressed autophagy by significantly reducing LC3-II expression, autophagosome numbers, and GFP-LC3 punctate foci in neuroepithelial cells and by altering autophagy-related gene expression. Maternal diabetes delayed neurogenesis by blocking Sox1 neural progenitor differentiation. Trehalose treatment reversed autophagy impairment and prevented NTDs in diabetic pregnancies. Trehalose resolved homeostatic imbalance by correcting mitochondrial defects, dysfunctional proteins, ER stress, apoptosis, and delayed neurogenesis in the neural tubes exposed to hyperglycemia. Our study demonstrates for the first time that maternal diabetes suppresses autophagy in neuroepithelial cells of the developing neural tube, leading to NTD formation, and provides evidence for the potential efficacy of trehalose as an intervention against hyperglycemia-induced NTDs.

In vitro nephrotoxicity and quantitative UPLC-MS analysis of three aristololactams in Houttuynia cordata.

Three analogues of aristololactam Ⅰ (AL Ⅰ), AL AⅡ, AL FⅠ and AL BⅡ, had been isolated from Houttuynia cordata, a commonly used medicinal and edible plant with heat-removing and toxin-removing functions. Considering the significant nephrotoxicity of AL Ⅰ, this study evaluated the toxicity of these three aristololactams (ALs) on human proximal tubular epithelial cells (HK-2) by MTT assay, ROS assay, ELISA tests and cytologic morphology observation. Furthermore, the distribution of the three ALs in H. cordata were investigated by UPLC-MSn recognition and quantitation in SIM mode, so as to estimate primarily the safety of the plant. The results showed that all the three ALs in H. cordata had comparative cytotoxicity as AL I with the IC50 values from 3.88 µM to 20.63 µM, caused high levels of cellular levels of reactive oxygen species (ROS) in HK-2 cells, exhibited the potential to cause renal fibrosis by remarkably increasing the levels of transforming growth factor-ß (TGF-ß1) and fibronectin (FN), and induced fibrous changes in morphology of HK-2 cells. The contents of the three ALs varied significantly in 30 batches of H. cordata from different regions and parts. Overall, the aerial part contained much more ALs (3.20 - 108.19 µg/g) than the underground part (0.95 - 11.66 µg/g), and flowers had the highest contents. Besides, no ALs were detected in the water extract of any part of H. cordata. This work revealed that the aristololactams in H. cordata had similar in vitro nephrotoxicity as AL Ⅰ and were mainly distributed in the aerial part of the plant.

Dapagliflozin ameliorates diabetic renal injury through suppressing the self-perpetuating cycle of inflammation mediated by HMGB1 feedback signaling in the kidney.

Dapagliflozin, the Sodium-glucose cotransporter 2 (SGLT2) inhibitor class of glucose-lowering agents, has shown the significantly nephroprotective effects to reduce the risk of kidney failure in diabetes. However, the underlying mechanisms are incompletely understood to explain the beneficial effects of dapagliflozin on kidney function. Here, we demonstrated that the administered of dapagliflozin for 12 weeks improved the proteinuria, histomorphology damage, oxidative stress, and macrophage infiltrations in the kidney of streptozotocin (STZ)-induced diabetic mice. Meanwhile, dapagliflozin attenuated the renal inflammation and fibrosis by reducing the pro-inflammatory factors interleukin-6 (IL-6), IL-1ß, and tumor necrosis factor α (TNF-α) and anti-fiber factor fibronectin (FN) and elevating the anti-inflammatory factor IL-10. Our data revealed that dapagliflozin exerted anti-inflammatory effects by inhibiting the activation of high mobility group box 1 (HMGB1)/TLR2/4/NF-κB signaling pathway. Consistently, we found that dapagliflozin suppressed the expression of HMGB1 and downstream TLR2/4/NF-κB signaling proteins in the human proximal tubular (HK-2) stimulated by high glucose and lipids or HMGB1 and RAW264.7 cells stimulated by IL-1ß, respectively. Further experiments were performed in the indirect co-culture model of RAW264.7 and HK-2 cells induced by high glucose and lipids. The results again confirmed the effects of dapagliflozin on alleviating inflammatory response and regulating the proportions of M1/M2 macrophage. It is indicated that the feedback signaling of HMGB1 between the tubules and macrophage involves in the persistence of the inflammation. These data demonstrate that dapagliflozin suppress the self-perpetuating inflammation by blocking the feedback loop of HMGB1 in the kidney, which contribute to ameliorate the renal injury in diabetes.

Quantitative profiling of carboxylic compounds by gas chromatography-mass spectrometry for revealing biomarkers of diabetic kidney disease.

Diabetic kidney disease (DKD), a common microvascular complication of diabetes, currently lacks specific diagnostic indicators and therapeutic targets, resulting in miss of early intervention. To profile metabolic conditions in complex and precious biological samples and screen potential biomarkers for DKD diagnosis and prognosis, a rapid, convenient and reliable quantification method for carboxyl compounds by gas chromatography-mass spectrometry (GC-MS) was established with isobutyl chloroformate derivatization. The derivatives were extracted with hexane, injected into GC-MS and quantified with selected ion monitoring mode. This method showed excellent linearity(R2 > 0.99), good recoveries (81.1%-115.5%), good repeatability (RSD < 20%) and sensitivity (LODs: 0.20-499.90 pg, LOQs: 2.00-1007.00 pg). Among the 37 carboxyl compounds analyzed, 12 metabolites in short-chain fatty acids (SCFAs) metabolism pathway and amino acid metabolism pathway were linked with DKD development and among them, 6 metabolites were associated with both development and prognosis of DKD in mice. In conclusion, a reliable, convenient and sensitive method based on isobutyl chloroformate derivatization and GC-MS analysis is established and successfully applied to quantify 37 carboxyl compounds in biological samples of mice and 12 potential biomarkers for DKD development and prognosis are screened.

Effects of sodium-glucose co-transporter 2 inhibitors on liver fibrosis in non-alcoholic fatty liver disease patients with type 2 diabetes mellitus: An updated meta-analysis of randomized controlled trials.

BACKGROUND AND AIM: Sodium-glucose co-transporter 2 inhibitors (SGLT2i) has been verified to improve Non-alcoholic fatty liver disease (NAFLD) in previous clinical practice. We mainly aim to investigate the effects of SGLT2i on liver fibrosis in NAFLD patients with type 2 diabetes mellitus (T2DM). METHODS: We conducted a comprehensive literature search utilizing the databases PubMed, Embase, Web of Science, and Cochrane Library, and extracted continuous data in the form of mean and standard deviation of the difference before and after treatment. RevMan 5.3 software was used to chart the pooled forest plot and perform heterogeneity, sensitivity and subgroup analysis. This study is conducted under the protocol registered with the Platform of Registered Systematic Review and Meta-analysis Protocols (INPLASY protocol 4946, INPLASY202360058). RESULTS: A total of 16 articles involving 699 patients were included. Indicators of liver fibrosis, containing Liver Stiffness Measurement (LSM), Controlled Attenuation Parameter (CAP), Serum ferritin, Serum type 4 collagen 7s, and FIB-4 index, were found to be considerably reduced by SGLT2i medication and subgroup analysis manifested pronounced dose-dependence. Additionally, SGLT2i therapy decreased BMI, lipid buildup and insulin resistance. CONCLUSIONS: SGLT2 inhibitors significantly ameliorated liver fibrosis and liver fat content, improved body conditions and insulin resistance, demonstrating that SGLT2i might reduce the risk of the progression of liver fibrosis and have a positive effect on NAFLD patients with T2DM.

FTY-720 alleviates diabetes-induced liver injury by inhibiting oxidative stress and inflammation.

We aimed to investigate the protective effect of FTY-720 on liver injury and explore its potential mechanism in diabetic mice. The diabetic mouse model was induced with streptozotocin and FTY-720 was administered for 12 weeks. We assayed biocharacters and liver function and used histopathology staining to evaluate the protective effects of FTY-720 against diabetic liver injury. Levels of oxidative stress and inflammation in the liver were observed. mRNA and protein levels of essential enzymes for glucose metabolism were quantified in the liver and the protein expression of TLR4, HIF1α and NF-κB was determined. In vivo results revealed that FTY-720 significantly lowered blood glucose and lipids and improved liver function and alleviated liver fibrosis in diabetic mice. FTY-720 reduced oxidative stress and inflammation, with the increased catalase activity and reduced levels of malondialdehyde, myeloperoxidase, IL-1ß, IL-6, TNF-α, TGF-ß, and MCP1. Furthermore, FTY-720 modulated glucose metabolism in liver and elevated the ATP production, showing the promotion of glycogenesis and glycolysis and inhibition of gluconeogenesis. Moreover, FTY-720 inhibited the expression of TLR4 and HIF1α, contributing to restoration of liver function. In conclusion, FTY-720 ameliorates diabetes-induced liver injury and improves glucose homeostasis by inhibiting oxidative stress and inflammation and may be a promise drug for treatment of liver disease.

SOD1 suppresses maternal hyperglycemia-increased iNOS expression and consequent nitrosative stress in diabetic embryopathy.

OBJECTIVE: Hyperglycemia induces oxidative stress and increases inducible nitric oxide synthase (iNOS) expression. We hypothesized that oxidative stress is responsible for hyperglycemia-induced iNOS expression. STUDY DESIGN: iNOS-luciferase activities, nitrosylated protein, and lipid peroxidation markers 4-hydroxynonenal and malondialdehyde were determined in parietal yolk sac-2 cells exposed to 5 mmol/L glucose or high glucose (25 mmol/L) with or without copper zinc superoxide dismutase 1 (SOD1) treatment. Levels of iNOS protein and messenger RNA, nitrosylated protein, and cleaved caspase-3 and -8 were assessed in wild-type embryos and SOD1-overexpressing embryos from nondiabetic and diabetic dams. RESULTS: SOD1 treatment diminished high glucose-induced oxidative stress, as evidenced by 4-hydroxynonenal and malondialdehyde reductions, and it blocked high glucose-increased iNOS expression, iNOS-luciferase activities, and nitrosylated protein. In vivo SOD1 overexpression suppressed hyperglycemia-increased iNOS expression and nitrosylated protein, and it blocked caspase-3 and -8 cleavage. CONCLUSION: We conclude that oxidative stress induces iNOS expression, nitrosative stress, and apoptosis in diabetic embryopathy.

Aggravated renal fibrosis is positively associated with the activation of HMGB1-TLR2/4 signaling in STZ-induced diabetic mice.

Diabetic kidney dysfunction is closely associated with renal fibrosis. Although the suppression of fibrosis is crucial to attenuate kidney damage, the underlying mechanisms remain poorly understood. In this study, renal injury in diabetic mice was induced by the intraperitoneal injection of streptozotocin (100 or 150 mg/kg) for 2 consecutive days. In the model mice, remarkable renal injury was observed, manifested by albuminuria, swelling of kidneys, and histopathological characteristics. The renal fibrosis was obviously displayed with high-intensity staining of fibrin, type IV collagen (Col IV), and fibronectin. The levels of Col IV and transforming growth factor-ß1 were significantly increased in diabetic mice kidneys. The aggravated fibrotic process was associated with the overexpression of HMGB1, TLR2/4, and p-NF-κB. Furthermore, a high expression of F4/80 and CD14 indicated that macrophage infiltration was involved in perpetuating inflammation and subsequent fibrosis in the kidneys of diabetic mice. The results demonstrate that the severity of renal fibrosis is positively associated with the activation of HMGB1/TLR2/4 signaling in diabetes.

Oral subacute nephrotoxicity of aristololactam I in rats.

Aristolactams (ALs) have been recognized as one kind of metabolites of aristolochic acids (AAs), the nephrotoxic components of Aristolochiaceae plants, and are more widely distributed than AAs in herbal medicines. This study evaluated the oral subacute nephrotoxicity of aristolactam I (AL I), a representative compound of ALs. AL I was intragastrically administered to rats at 20 mg·kg-1·d-1 for 10 or 20 days, with aristolochic acid I (AA I) used as positive control at the same dose. After 10-day treatment, AL I led to a significant increase in early renal injury-related indices in urine and obvious histopathological lesions in kidneys, including degeneration of tubular epithelial cells, inflammatory cell infiltration and fibrosis. The lesions induced by AL I were significantly aggravated after 20-day exposure. However, AL I induced less histopathological damage in kidneys than AA I in both 10- and 20-day groups. Our results indicated that oral AL I caused nephrotoxicity by inducing oxidative stress, inflammation, and overactivation of the complement system as AA I did. Three detected apoptosis-associated indicators were not affected by AL I but remarkably increased by AA I. In summary, oral AL I induced evident renal damage in rats after only 10 days of treatment, and the damage was aggravated after 20 days. However, AL I was obviously less nephrotoxic than AA I via oral gavage.

SOD1 overexpression in vivo blocks hyperglycemia-induced specific PKC isoforms: substrate activation and consequent lipid peroxidation in diabetic embryopathy.

OBJECTIVE: Oxidative stress plays a causative role in diabetic embryopathy. We tested whether mitigating oxidative stress, using superoxide dismutase 1 (SOD1) transgenic (Tg) mice, would block hyperglycemia-induced specific protein kinase C (PKC) isoform activation and its downstream cascade. STUDY DESIGN: Day 8.5 embryos from nondiabetic wild-type control (NC), diabetic mellitus wild-type (DM), and diabetic SOD1-Tg mice (DM-SOD1-Tg) were used for detection of phosphorylated (p-) PKCα/ßII and p-PKCδ, and levels of 2 prominent PKC substrates, phosphorylated myristoylated alanine-rich protein kinase C substrate (MARCKS) and receptor for activated C kinase 1 (RACK1), and lipid peroxidation markers, 4-hydroxynonenal (4-HNE) and malondialdehyde (MDA). RESULTS: Levels of p-PKCα/ßII, p-PKCδ, p-MARCKS, 4-HNE, and MDA were significantly elevated in the DM group compared with those in the NC group and the DM-SOD1-Tg group. The NC and DM-SOD1-Tg groups had comparable levels of these protein and lipid peroxidation markers. RACK1 levels did not differ among the 3 groups. CONCLUSION: Mitigating oxidative stress by SOD1 overexpression blocks maternal hyperglycemia-induced activation of specific PKC isoforms and downstream cascades.

Total glucosides of Paeony restores intestinal barrier function through inhibiting Lyn/Snail signaling pathway in colitis mice.

BACKGROUND: Inflammatory bowel disease (IBD) is an autoimmune disease. The pathogenesis of IBD is complicated and intestinal mucosal barrier damage is considered as the trigger factor for the initiation and recurrence of IBD. Total Glucosides of Paeony (TGP) has shown good inhibitory effects on immune-inflammation in clinic studies. However, its effect and mechanism on IBD are largely unknown. PURPOSE: The purpose of this study is to evaluate the effect and mechanism of TGP on IBD. STUDY DESIGN: DSS-induced colitis mouse model was used. TGP was given by gavage. Caco-2 cells were stimulated by outer membrane vesicles (OMV) to establish an in vitro model. METHODS: C57BL/6 mice were divided into normal control group, model group, mesalazine group, paeoniflorin (PA) group, high-dose group of TGP, and low-dose group of TGP. The model was induced with 2.5% DSS for 7 days, and TGP was intragastrically administered for 10 days. The therapeutic effect of TGP was evaluated by symptoms, histochemical analysis, RT-qPCR and ELISA. The mechanism was explored by intestinal permeability, Western blot and immunofluorescence in vivo and in vitro. RESULTS: Our results showed that TGP could significantly improve the symptoms and pathological changes, with reduced levels of TNF-α, IL-17A, IL-23 and IFN-γ in the colon tissues and serum under a dose-dependent manner. TGP also reduced the intestinal permeability and restored the protein expression of tight junction and adherens junction proteins of intestinal epithelial cells in vivo and in vitro. Furthermore, TGP could inhibit the expression of p-Lyn and Snail and prevent Snail nuclear localization, thereby maintaining tight and adherens junctions. CONCLUSION: TGP effectively improves the symptoms of DSS-induced colitis in mice, protects the intestinal epithelial barrier by inhibiting the Lyn/Snail signaling pathway, and maybe a promise therapeutic agent for IBD treatment.

Modulating the gut microbiota and inflammation is involved in the effect of Bupleurum polysaccharides against diabetic nephropathy in mice.

Dysbiosis of gut microbiota and low grade inflammation has gradually become a highly potential therapeutic agent for diabetic nephropathy (DN). It has been reported that a large number of polysaccharides have positive effects on DN, including Bupleurum polysaccharides. However, the mechanism remained unclear. This study selected two Bupleurum polysaccharides from different origins to investigate the potential relationship between kidney and gut. Diabetic mice model was established by streptozotocin (STZ, 100 mg/kg) and the treatment groups were treated with two Bupleurum polysaccharides (60 mg/kg) for 6 weeks, respectively. The results showed that the administration of Bupleurum polysaccharides ameliorated diabetic nephropathy induced by STZ. Blood glucose, blood creatinine and urine albumin were decreased after the oral administration of Bupleurum polysaccharides. And the dysbiosis of gut microbiota was modulated with higher diversity and gut protective microbiota. The gut barrier was also improved and the expression of inflammatory response both in kidney and colon was reduced. These results provided the evidence that modulating the gut microbiota and inflammation was involved in the effect of Bupleurum polysaccharides against diabetic nephropathy in mice and laid the foundation for the deeper, specific mechanism research on the interaction between kidney and gut.

Houttuynia cordata polysaccharide alleviated intestinal injury and modulated intestinal microbiota in H1N1 virus infected mice.

Houttuynia cordata polysaccharide (HCP) is extracted from Houttuynia cordata, a key traditional Chinese medicine. The study was to investigate the effects of HCP on intestinal barrier and microbiota in H1N1 virus infected mice. Mice were infected with H1N1 virus and orally administrated HCP at a dosage of 40 mg(kg-1(d-1. H1N1 infection caused pulmonary and intestinal injury and gut microbiota imbalance. HCP significantly suppressed the expression of hypoxia inducible factor-1α and decreased mucosubstances in goblet cells, but restored the level of zonula occludens-1 in intestine. HCP also reversed the composition change of intestinal microbiota caused by H1N1 infection, with significantly reduced relative abundances of Vibrio and Bacillus, the pathogenic bacterial genera. Furthermore, HCP rebalanced the gut microbiota and restored the intestinal homeostasis to some degree. The inhibition of inflammation was associated with the reduced level of Toll-like receptors and interleukin-1ß in intestine, as well as the increased production of interleukin-10. Oral administration of HCP alleviated lung injury and intestinal dysfunction caused by H1N1 infection. HCP may gain systemic treatment by local acting on intestine and microbiota. This study proved the high-value application of HCP.