Gut microbiota dynamics in KK-Ay mice: restoration following antibiotic treatment.

The primary aim of this study was to investigate the alterations in the microbial community of KK-Ay mice following antibiotic treatment. A comparative analysis of the gut microbiota was conducted between KK-Ay mice treated with antibiotics and those without treatment. The microbial community dynamics in antibiotic-treated KK-Ay mice were meticulously assessed over an eight-week period using 16S rDNA sequencing analysis. Simultaneously, dynamic renal function measurements were performed. The results demonstrated a marked decrease in bacterial DNA abundance following antibiotic intervention, coupled with a substantial reduction in bacterial diversity and a profound alteration in microbial composition. These observed microbiota changes persisted in the KK-Ay mice throughout the eight-week post-antibiotic treatment period. Particularly noteworthy was the reemergence of bacterial populations after two weeks or more, resulting in a microbiota composition resembling that of untreated KK-Ay mice. This transition was characterized by a significant increase in the abundance of clostridia at the class level, Lachnospirales and Oscillospirales at the order level, and Lachnospiraceae, Oscillospiraceae, and Ruminococcaceae at the family level. Concurrently, there was a notable decrease in Clostridia_UCG-014. The observed alterations in the gut microbiota of antibiotic-treated KK-Ay mice suggest a dynamic response to antibiotic intervention and subsequent restoration towards the original untreated state.

An Update on the Role and Potential Molecules in Relation to Ruminococcus gnavus in Inflammatory Bowel Disease, Obesity and Diabetes Mellitus.

Ruminococcus gnavus (R. gnavus) is a gram-positive anaerobe commonly resides in the human gut microbiota. The advent of metagenomics has linked R. gnavus with various diseases, including inflammatory bowel disease (IBD), obesity, and diabetes mellitus (DM), which has become a growing area of investigation. The initial focus of research primarily centered on assessing the abundance of R. gnavus and its potential association with disease presentation, taking into account variations in sample size, sequencing and analysis methods. However, recent investigations have shifted towards elucidating the underlying mechanistic pathways through which R. gnavus may contribute to disease manifestation. In this comprehensive review, we aim to provide an updated synthesis of the current literature on R. gnavus in the context of IBD, obesity, and DM. We critically analyze relevant studies and summarize the potential molecular mediators implicated in the association between R. gnavus and these diseases. Across numerous studies, various molecules such as methylation-controlled J (MCJ), glucopolysaccharides, ursodeoxycholic acid (UDCA), interleukin(IL)-10, IL-17, and capric acid have been proposed as potential contributors to the link between R. gnavus and IBD. Similarly, in the realm of obesity, molecules such as hydrogen peroxide, butyrate, and UDCA have been suggested as potential mediators, while glycine ursodeoxycholic acid (GUDCA) has been implicated in the connection between R. gnavus and DM. Furthermore, it is imperative to emphasize the necessity for additional studies to evaluate the potential efficacy of targeting pathways associated with R. gnavus as a viable strategy for managing these diseases. These findings have significantly expanded our understanding of the functional role of R. gnavus in the context of IBD, obesity, and DM. This review aims to offer updated insights into the role and potential mechanisms of R. gnavus, as well as potential strategies for the treatment of these diseases.

Specific Alternation of Gut Microbiota and the Role of Ruminococcus gnavus in the Development of Diabetic Nephropathy.

In this study, we aim to investigate the precise alterations in the gut microbiota during the onset and advancement of diabetic nephropathy (DN) and examine the impact of Ruminococcus gnavus (R. gnavus) on DN. Eight-week-old male KK-Ay mice were administered antibiotic cocktails for a duration of two weeks, followed by oral administration of R. gnavus for an additional eight weeks. Our study revealed significant changes in the gut microbiota during both the initiation and progression of DN. Specifically, we observed a notable increase in the abundance of Clostridia at the class level, higher levels of Lachnospirales and Oscillospirales at the order level, and a marked decrease in Clostridia_UCG-014 in DN group. Additionally, there was a significant increase in the abundance of Lachnospiraceae, Oscillospiraceae, and Ruminococcaceae at the family level. Moreover, oral administration of R. gnavus effectively aggravated kidney pathology in DN mice, accompanied by elevated levels of urea nitrogen (UN), creatinine (Cr), and urine protein. Furthermore, R. gnavus administration resulted in down-regulation of tight junction proteins such as Claudin-1, Occludin, and ZO-1, as well as increased levels of uremic toxins in urine and serum samples. Additionally, our study demonstrated that orally administered R. gnavus up-regulated the expression of inflammatory factors, including nucleotide-binding oligomerization domain-like receptor pyrin domain-containing protein 3 (NLRP3) and Interleukin (IL)-6. These changes indicated the involvement of the gut-kidney axis in DN, and R. gnavus may worsen diabetic nephropathy by affecting uremic toxin levels and promoting inflammation in DN.

Snail microbiota and snail-schistosome interactions: axenic and gnotobiotic technologies.

The microbiota in the intermediate snail hosts of human schistosomes can significantly affect host biology. For decades, researchers have developed axenic snails to manipulate the symbiotic microbiota. This review summarizes the characteristics of symbiotic microbes in intermediate snail hosts and describes their interactions with snails, affecting snail growth, development, and parasite transmission ability. We focus on advances in axenic and gnotobiotic technologies for studying snail-microbe interactions and exploring the role of microbiota in snail susceptibility to Schistosoma infection. We discuss the challenges related to axenic and gnotobiotic snails, possible solutions to address these challenges, and future research directions to deepen our understanding of snail-microbiota interactions, with the aim to develop microbiota-based strategies for controlling snail populations and reducing their competence in transmitting parasites.

Core gut microbes Cloacibacterium and Aeromonas associated with different gastropod species could be persistently transmitted across multiple generations.

BACKGROUND: Studies on the gut microbiota of animals have largely focused on vertebrates. The transmission modes of commensal intestinal bacteria in mammals have been well studied. However, in gastropods, the relationship between gut microbiota and hosts is still poorly understood. To gain a better understanding of the composition of gut microbes and their transmission routes in gastropods, a large-scale and long-term experiment on the dynamics and transmission modes of gut microbiota was conducted on freshwater snails. RESULTS: We analyzed 244 microbial samples from the digestive tracts of freshwater gastropods and identified Proteobacteria and Bacteroidetes as dominant gut microbes. Aeromonas, Cloacibacterium, and Cetobacterium were identified as core microbes in the guts, accounting for over 50% of the total sequences. Furthermore, both core bacteria Aeromonas and Cloacibacterium, were shared among 7 gastropod species and played an important role in determining the gut microbial community types of both wild and cultured gastropods. Analysis of the gut microbiota at the population level, including wild gastropods and their offspring, indicated that a proportion of gut microbes could be consistently vertically transmitted inheritance, while the majority of the gut microbes resulted from horizontal transmission. Comparing cultured snails to their wild counterparts, we observed an increasing trend in the proportion of shared microbes and a decreasing trend in the number of unique microbes among wild gastropods and their offspring reared in a cultured environment. Core gut microbes, Aeromonas and Cloacibacterium, remained persistent and dispersed from wild snails to their offspring across multiple generations. Interestingly, under cultured environments, the gut microbiota in wild gastropods could only be maintained for up to 2 generations before converging with that of cultured snails. The difference observed in gut bacterial metabolism functions was associated with this transition. Our study also demonstrated that the gut microbial compositions in gastropods are influenced by developmental stages and revealed the presence of Aeromonas and Cloacibacterium throughout the life cycle in gastropods. Based on the dynamics of core gut microbes, it may be possible to predict the health status of gastropods during their adaptation to new environments. Additionally, gut microbial metabolic functions were found to be associated with the adaptive evolution of gastropods from wild to cultured environments. CONCLUSIONS: Our findings provide novel insights into the dynamic processes of gut microbiota colonization in gastropod mollusks and unveil the modes of microbial transmission within their guts. Video Abstract.

Gut microbiota in parasite-transmitting gastropods.

BACKGROUND: Gastropoda, the largest class within the phylum Mollusca, houses diverse gut microbiota, and some gastropods serve as intermediate hosts for parasites. Studies have revealed that gut bacteria in gastropods are associated with various biological aspects, such as growth, immunity and host-parasite interactions. Here, we summarize our current knowledge of gastropod gut microbiomes and highlight future research priorities and perspectives. METHODS: A literature search was undertaken using PubMed, Web of Science and CNKI for the articles on the gut microbiota of gastropods until December 31, 2022. We retrieved a total of 166 articles and identified 73 eligible articles for inclusion in this review based on the inclusion and exclusion criteria. RESULTS: Our analysis encompassed freshwater, seawater and land snails, with a specific focus on parasite-transmitting gastropods. We found that most studies on gastropod gut microbiota have primarily utilized 16S rRNA gene sequencing to analyze microbial composition, rather than employing metagenomic, metatranscriptomic, or metabolomic approaches. This comprehensive review provided an overview of the parasites carried by snail species in the context of gut microbiota studies. We presented the gut microbial trends, a comprehensive summary of the diversity and composition, influencing factors, and potential functions of gastropod gut microbiota. Additionally, we discussed the potential applications, research gaps and future perspectives of gut microbiomes in parasite-transmitting gastropods. Furthermore, several strategies for enhancing our comprehension of gut microbiomes in snails were also discussed. CONCLUSIONS: This review comprehensively summarizes the current knowledge on the composition, potential function, influencing factors, potential applications, limitations, and challenges of gut microbiomes in gastropods, with a specific emphasis on parasite-transmitting gastropods. These findings provide important insights for future studies aiming to understand the potential role of gastropod gut microbiota in controlling snail populations and snail-borne diseases.

Jiangtang Decoction Ameliorates Diabetic Kidney Disease Through the Modulation of the Gut Microbiota.

Purpose: This study aimed to elucidate the impact of Jiangtang decoction (JTD) on diabetic kidney disease (DKD) and its association with alterations in the gut microbiota. Methods: Using a diabetic mouse model (KK-Ay mice), daily administration of JTD for eight weeks was undertaken. Weekly measurements of body weight and blood glucose were performed, while kidney function, uremic toxins, inflammation factors, and fecal microbiota composition were assessed upon sacrifice. Ultra-structural analysis of kidney tissue was conducted to observe the pathological changes. Results: The study findings demonstrated that JTD improve metabolism, kidney function, uremic toxins and inflammation, while also exerting a modulatory effect on the gut microbiota. Specifically, the genera Rikenella, Lachnoclostridium, and unclassified_c_Bacilli exhibited significantly increased abundance following JTD treatment, accompanied by reduced abundance of norank_f_Lachnospiraceae compared to the model group. Importantly, Rikenella and unclassified_c_Bacilli demonstrated negative correlations with urine protein levels. Lachnoclostridium and norank_f_Lachnospiraceae were positively associated with creatinine (Cr), indoxyl sulfate (IS) and interleukin (IL)-6. Moreover, norank_f_Lachnospiraceae exhibited positive associations with various indicators of DKD severity, including weight, blood glucose, urea nitrogen (UN), kidney injury molecule-1 (KIM-1) levels, trimethylamine-N-oxide (TMAO), p-cresyl sulfate (pCS), nucleotide-binding oligomerization domain (Nod)-like receptor family pyrin domain-containing 3 (NLRP3) and IL-17A production. Conclusion: These findings suggested that JTD possess the ability to modulate the abundance of Rikenella, Lachnoclostridium, unclassified_c_Bacilli and norank_f_Lachnospiraceae within the gut microbiota. This modulation, in turn, influenced metabolic processes, kidney function, uremic toxin accumulation, and inflammation, ultimately contributing to the amelioration of DKD.

Metagenomic Analysis Reveals Variations in Gut Microbiomes of the Schistosoma mansoni-Transmitting Snails Biomphalaria straminea and Biomphalaria glabrata.

Biomphalaria snails play a crucial role in the transmission of the human blood fluke Schistosoma mansoni. The gut microbiota of intermediate hosts is known to influence their physiological functions, but little is known about its composition and role in Biomphalaria snails. To gain insights into the biological characteristics of these freshwater intermediate hosts, we conducted metagenomic sequencing on Biomphalaria straminea and B. glabrata to investigate variations in their gut microbiota. This study revealed that the dominant members of the gut microbiota in B. glabrata belong to the phyla Bacteroidetes and Proteobacteria, which were also found to be the top two most abundant gut bacteria in B. straminea. We identified Firmicutes, Acidovorax and Bosea as distinctive gut microbes in B. straminea, while Aeromonas, Cloacibacterium and Chryseobacterium were found to be dependent features of the B. glabrata gut microbiota. We observed significant differences in the community structures and bacterial functions of the gut microbiota between the two host species. Notably, we found a distinctive richness of antibiotic resistance genes (ARGs) associated with various classes of antibiotics, including bacitracin, chloramphenicol, tetracycline, sulfonamide, penicillin, cephalosporin_ii and cephalosporin_i, fluoroquinolone, aminoglycoside, beta-lactam, multidrug and trimethoprim, in the digestive tracts of the snails. Furthermore, this study revealed the potential correlations between snail gut microbiota and the infection rate of S. mansoni using Spearman correlation analysis. Through metagenomic analysis, our study provided new insights into the gut microbiota of Biomphalaria snails and how it is influenced by host species, thereby enhancing our understanding of variant patterns of gut microbial communities in intermediate hosts. Our findings may contribute to future studies on gastropod-microbe interactions and may provide valuable knowledge for developing snail control strategies to combat schistosomiasis in the future.

Specific alterations of gut microbiota in diabetic microvascular complications: A systematic review and meta-analysis.

Background: The role of gut microbiota in diabetes mellitus (DM) and its complications has been widely accepted. However, the alternation of gut microbiota in diabetic microvascular complications (DC) remains to be determined. Methods: Publications (till August 20th, 2022) on gut microbiota in patients with DC were retrieved from PubMed, Web of Science, Embase and Cochrane. Review Manager 5.3 was performed to estimate the standardized mean difference (SMD) and 95% confidence interval (CI) and calculate alpha diversity indices and the relative abundance of gut microbiota between patients in DC v.s. DM and DC v.s. healthy controls (HC). Results: We included 13 studies assessing 329 patients with DC, 232 DM patients without DC, and 241 HC. Compared to DM, patients with DC shared a significantly lower Simpson index (SMD = -0.59, 95% CI [-0.82, -0.36], p < 0.00001), but a higher ACE index (SMD = 0.42, 95% CI[0.11, 0.74], p = 0.009). Compared to HC, DC patients held a lower ACE index (SMD = -0.61, 95% CI[-1.20, -0.02], p = 0.04). The relative abundances of phylum Proteobacteria (SMD = 0.03, 95% CI[0.01, 0.04], p = 0.003, v.s. HC) and genus Klebsiella (SMD = 0.00, 95% CI[0.00, 0.00], p < 0.00001, v.s. HC) were enriched, accompanying with depleted abundances of phylum Firmicutes (SMD = -0.06, 95% CI[-0.11, -0.01], p = 0.02, v.s. HC), genera Bifidobacterium (SMD = -0.01, 95% CI[-0.02,-0.01], p < 0.0001, v.s. DM), Faecalibacterium (SMD = -0.01, 95% CI[-0.02, -0.00], p = 0.009, v.s. DM; SMD = -0.02, 95% CI[-0.02, -0.01], p < 0.00001, v.s. HC) and Lactobacillus (SMD = 0.00, 95% CI[-0.00, -0.00], p < 0.00001, v.s. HC) in DC. Conclusions: Gut microbiota perturbations with the depletion of alpha diversity and certain short-chain fatty acids (SCFAs)-producing bacteria were associated with the pathology of DC. Therefore, gut microbiota might serve as a promising approach for the diagnosis and treatment of DC. Further investigations are required to study the mechanisms by which gut dysbiosis acts on the onset and progression of DC.

D-Ribose Induces Podocyte NLRP3 Inflammasome Activation and Glomerular Injury via AGEs/RAGE Pathway.

D-ribose levels are demonstrated to be increased in type II diabetes mellitus and increased blood D-ribose is involved in the development of diabetic complications such as diabetic encephalopathy and nephropathy. However, the mechanism mediating the pathogenic role of D-ribose in nephropathy remains poorly understood. Given that D-ribose was reported to induce advanced glycation end products (AGEs) formation, the present study tested whether D-ribose induces NLRP3 activation and associated glomerular injury via AGEs/receptor of AGEs (RAGE) signaling pathway. In vivo, C57BL/6J and Asc-/- mice were treated with D-ribose with or without AGEs inhibitor. Administration of D-ribose daily for 30 days was found to induce NLRP3 inflammasome formation in glomerular podocyte, as shown by increased co-localization of NLRP3 with apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) or caspase-1. This D-ribose-induced NLRP3 inflammasome formation was accompanied by its activation as evidenced by increased IL-1ß production, a major product of NLRP3 inflammasome. Corresponding to NLRP3 inflammasome activation, D-ribose led to significant glomerular injury in mice. All these D-ribose-induced glomerular inflammasome and associated pathological changes were markedly attenuated by deletion of Asc gene. Furthermore, the accumulation of AGEs and RAGE was found increased in glomeruli of mice receiving D-ribose. In cell studies, we also confirmed that D-ribose induced NLRP3 inflammasome formation and activation in podocytes, which was significantly blocked by caspase-1 inhibitor, YvAD. Mechanically, AGEs formation inhibition and cleavage or silencing of RAGE gene were shown to suppress D-ribose-induced NLRP3 inflammasome formation and activation, as shown by significant reduction of NLRP3 inflammasome molecular aggregation, caspase-1 activity and IL-1ß production. These results strongly suggest that relatively long term administration of D-ribose induces NLRP3 inflammasome formation and activation in podocytes via AGEs/RAGE signaling pathway, which may be one of important triggering mechanisms leading to diabetic nephropathy.

Suppression of Glucagon-Like Peptide-1 Release by Inhibition of Intestinal NLRP3 Inflammasome Activation in Asc-/- and Nlrp3-/- Mice.

The glucagon-like peptide-1 (GLP-1) is an insulinotropic hormone secreted by intestinal enteroendocrine L-cells, which plays a crucial role in glucose control, regulation, and protection from different pathological conditions such as diabetes mellitus. The present study sought to test whether GLP-1 release increases gut injury with a high-fat diet (HFD) and whether this GLP-1 release is associated with NLRP3 inflammasome activation. Our results showed that the NLRP3 inflammasome is activated in the intestinal tissue of wild-type mice on a HFD, accompanied by GLP-1 overexpression. The number of intestinal L-cells and the GLP-1 level in serum are increased in WT mice with HFD. However, in the Asc-/- and Nlrp3-/- mice, these HFD-induced intestinal and serum GLP-1 changes were suppressed. Using confocal microscopy, the colocalization of GLP-1 and FLICA that labels activated caspase-1 in intestine was decreased in the Asc-/- and Nlrp3-/- mice compared to WT mice. Mechanistically, the inhibitor of caspase-1 or HMGB1 blocker is used to demonstrate the regulatory action of NRLP3 inflammasome in GLP-1 release. It was found that the level of GLP-1 and its colocalization with IL-1ß were reduced by inhibition of the caspase-1 activity, but not altered by blockade of HMGB1 action. Our results suggest that NLRP3 inflammasome activation triggers GLP-1 production from the intestine, which is associated with IL-1ß, but not with HMGB1. These findings for the first time provide evidence that the activation of NLRP3 inflammasome in the intestine increases GLP-1 release in mice, which may serve as an adaptive response to intestinal inflammation.

Control of lysosomal TRPML1 channel activity and exosome release by acid ceramidase in mouse podocytes.

The transient receptor potential mucolipin 1 (TRPML1) channel has been reported to mediate lysosomal Ca2+ release that is involved in Ca2+-dependent lysosome trafficking and autophagic flux. However, this regulatory mechanism of lysosomal TRPML1 channel activity in podocytes remains poorly understood. In the present study, we tested whether the TRPML1 channel in podocytes mediates lysosome trafficking, which is essential for multivesicular body (MVB) degradation by lysosomes. We first demonstrated the abundant expression of TRPML1 channel in podocytes. By GCaMP3 Ca2+ imaging, we characterized the lysosomal specificity of TRPML1 channel-mediated Ca2+ release in podocytes. Given the important role of acid ceramidase (AC) in lysosome function and podocyte injury, we tested whether AC regulates this TRPML1 channel-mediated Ca2+ release and consequent lysosome-dependent MVB degradation in podocytes. Pharmacologically, it was found that TRPML1 channel activity was remarkably attenuated by the AC inhibitor carmofur. Sphingosine, as an AC product, was demonstrated to induce TRPML1-mediated Ca2+ release, which was inhibited by a TRPML1 blocker, verapamil. Using a Port-a-Patch planar patch-clamp system, we found that AC-associated sphingolipids, sphingomyelin, ceramide, and sphingosine had different effects on TRPML1 channel activity in podocytes. Functionally, the inhibition of AC or blockade of TRPML1 channels was found to suppress the interaction of lysosomes and MVBs, leading to increased exosome release from podocytes. These results suggest that AC is critical for TRPML1 channel-mediated Ca2+ release, which controls lysosome-MVB interaction and exosome release in podocytes.

Protective Effects of Kaempferol on D-Ribose-Induced Mesangial Cell Injury.

Recently, it has been found that the level of urinary D-ribose in type 2 diabetes is notably higher than that in age-matched normal control, and D-ribose is more reactive in the glycation than D-glucose and induces oxidative stress. Kaempferol is one of the main bioactive components in Astragalus membranaceus, with numerous physiological actives, such as antioxidant. The present study investigated the protective effects of kaempferol on D-ribose-treated mesangial cells. CCK-8 and LDH assay were used to test cell viability and cell toxicity. Immunofluorescence and flow cytometry were used to detect the AGE formation and ROS accumulation. GSH level was measured to reflect oxidation resistance. Cell apoptosis was evaluated by Hoechst 33258 staining, AO/EB staining, and western blot. Mitochondrial membrane integrity was detected by JC-1 staining, western blot, and RT-PCR. The change of autophagy level was tested by western blot. The results indicated that D-ribose induced not only cell damage and increased AGE formation and ROS accumulation but also GSH depletion. Further studies demonstrated that D-ribose induced mitochondrial depolarization and the activation of caspase-9/3. But kaempferol could partly block these damages. Subsequently, it was confirmed that kaempferol repaired the autophagy disturbance induced by D-ribose, and 3-MA could reverse the protective effect of kaempferol under D-ribose condition. Our study demonstrated that D-ribose induced AGE accumulation and ROS production in mesangial cell and caused mitochondrial apoptosis, but kaempferol could attenuate these changes and its protective effect might be related to the repair of autophagy.

Lysosomal regulation of extracellular vesicle excretion during d-ribose-induced NLRP3 inflammasome activation in podocytes.

The NLRP3 inflammasome is activated in the cytoplasm of cells and its products such as IL-1ß are exported through a non-classical ER-Golgi pathway. Several mechanistically distinct models including exocytosis of secretory lysosomes, microvesicles (MVs) and extracellular vehicles (EVs) have been proposed for their release. In this study, we hypothesized that the NLRP3 inflammasome product, IL-1ß in response to exogenously administrated and endogenously produced d-ribose stimulation is released via extracellular vesicles including EVs via a sphingolipid-mediated molecular mechanisms controlling lysosome and multivesicular body (MVB) interaction. First, we demonstrated that both endogenous and exogenous d-ribose induced NLRP3 inflammasome activation to produce IL-1ß, which was released via EVs in podocytes. Then, we found that colocalization of marker MVB marker VPS16 with IL-1ß within podocytes increased upon d-ribose stimulation, which was accompanied by decreased colocalization of lysosome marker Lamp-1 and VPS16, suggesting decrease in MVB inclusion of IL-1ß due to reduced lysosome and MVB interaction. All these changes were mimicked and accelerated by lysosome v-ATPase inhibitor, bafilomycin. Moreover, ceramide in podocytes was found elevated upon d-ribose stimulation, and prior treatments of podocyte with acid sphingomyelinase (Asm) inhibitor, amitriptyline, acid ceramidase (AC) inducer, genistein, or AC CRISPR/cas9 activation plasmids were found to decrease d-ribose-induced ceramide accumulation, EVs release and IL-1ß secretion due to reduced interactions of lysosome with MVBs. These results suggest that inflammasome-derived products such as IL-1ß during d-ribose stimulation are released via EVs, in which lysosomal sphingolipid-mediated regulation of lysosome function plays an important role.

D-ribose induces nephropathy through RAGE-dependent NF-κB inflammation.

Recently, aberrantly high levels of D-ribose have been discovered in type II diabetic patients. D-ribose glycates proteins more rapidly than D-glucose, resulting in the production of advanced glycation end products (AGEs). Accumulations of these products can be found in impaired renal function, but the mechanisms are poorly understood. The present study tested whether D-ribose induces renal dysfunction via the RAGE-dependent NF-κB signaling pathway. In vivo, administration of D-ribose was found to lower blood glucose and regulate insulin tolerance. Compared to controls, urine nitrogen and creatinine excretion were increased in mice receiving D-ribose and were accompanied by severe pathological renal damage. Furthermore, immunohistochemistry showed that NF-κB, AGEs, and receptor of AGEs (RAGE) increased in the kidneys of the mice with D-ribose treatment. In vitro, by western blot and immunofluorescent staining, we confirmed that D-ribose induced NF-κB activation and accumulation of AGEs and RAGE in mesangial cells. By co-immunoprecipitation, we found that the pull-down of RAGE remarkably increased the expression of NF-κB. Silencing the RAGE gene blocked the phosphorylation of NF-κB induced by D-ribose. These results strongly suggest that D-ribose induced NF-κB inflammation in a RAGE-dependent manner, which may be a triggering mechanism leading to nephropathy.

Inhibition of pannexin-1 channel activity by adiponectin in podocytes: Role of acid ceramidase activation.

The pannexin-1 (Panx1) channel has been reported to mediate the release of ATP that is involved in local tissue inflammation, obesity, and many chronic degenerative diseases. It remains unknown whether Panx1 is present in podocytes and whether this channel in podocytes mediates ATP release leading to glomerular inflammation or fibrosis. To answer these questions, we first characterized the expression of Panx channels in podocytes. Among the three known pannexins, Panx1 was the most enriched in podocytes, either cultured or native in mouse glomeruli. Using a Port-a-Patch planar patch-clamp system, we recorded a large voltage-gated outward current through podocyte membrane under the Cs+in/Na+out gradient. Substitution of gluconate or aspartate for chloride in the bath solution blocked voltage-gated outward currents and shifted the reversal potential of Panx1 currents to the right, indicating the anion permeability of this channel. Pharmacologically, the recorded voltage-gated outward currents were substantially attenuated by specific Panx1 channel inhibitors. Given the anti-inflammatory and intracellular ATP restorative effects of adiponectin, we tested whether this adipokine inhibits Panx1 channel activity to block ATP release. Adiponectin blocked Panx1 channel activity in podocytes. Mechanistically, inhibition of acid ceramidase (AC) remarkably enhanced Panx1 channel activity under control conditions and prevented the inhibition of Panx1 channel by adiponectin. Correspondingly, intracellular addition of AC products, sphingosine or sphingosine-1-phosphate (S1P), blocked Panx1 channel activity, while elevation of intracellular ceramide had no effect on Panx1 channel activity. These results suggest that adiponectin inhibits Panx1 channel activity in podocytes through activation of AC and associated elevation of intracellular S1P.

NLRP3 Inflammasome Formation and Activation in Nonalcoholic Steatohepatitis: Therapeutic Target for Antimetabolic Syndrome Remedy FTZ.

The Nod-like receptor protein 3 (NLRP3) inflammasome activation not only serves as an intracellular machinery triggering inflammation but also produces uncanonical effects beyond inflammation such as changing cell metabolism and increasing cell membrane permeability. The present study was designed to test whether this NLRP3 inflammasome activation contributes to the "two-hit" injury during nonalcoholic steatohepatitis (NASH) and whether it can be a therapeutic target for the action of Fufang Zhenzhu Tiaozhi (FTZ), a widely used herbal remedy for hyperlipidemia and metabolic syndrome in China. We first demonstrated that NLRP3 inflammasome formation and activation as well as lipid deposition occurred in the liver of mice on the high-fat diet (HFD), as shown by increased NLRP3 aggregation, enhanced production of IL-1ß and high mobility group box 1 (HMGB1), and remarkable lipid deposition in liver cells. FTZ extracts not only significantly reduced the NLRP3 inflammasome formation and activation but also attenuated the liver steatosis and fibrogenic phenotype changed. In in vitro studies, palmitic acid (PA) was found to increase colocalization of NLRP3 components and enhanced caspase-1 activity in hepatic stellate cells (HSCs), indicating enhanced formation and activation of NLRP3 inflammasomes by PA. PA also increased lipid deposition. Nlrp3 siRNA can reverse this effect by silencing the NLRP3 inflammasome and both with FTZ. In FTZ-treated cells, not only inflammasome formation and activation was substantially attenuated but also lipid deposition in HSCs was blocked. This inhibition of FTZ on lipid deposition was similar to the effects of glycyrrhizin, an HMGB1 inhibitor. Mechanistically, stimulated membrane raft redox signaling platform formation and increased O2•- production by PA to activate NLRP3 inflammasomes in HSCs was blocked by FTZ treatment. It is concluded that FTZ extracts inhibit NASH by its action on both inflammatory response and liver lipid metabolism associated with NLRP3 inflammasome formation and activation.

[Study on effect of Jiangtang decoction on AGEs-RAGE and oxidative stress in KK-Ay mice].

To elucidate the efficacy of Jiangtang decoction(JTD) on AGEs-RAGE and oxidative stress in type 2 diabetic model KK-Ay mice. Fifty KK-Ay mice were randomly divided into 5 groups as follows： model group, metformin group, low-dose, medium-dose and high-dose of JTD group, with 10 C57BL/6J as normal group. All groups are orally administrated with equal distilled water, 250 mg•kg⁻¹ metformin hydrochloride, 2, 4,8 g•kg⁻¹ JTD, equal distilled water respectively, once per day for 12 weeks. Alanine aminotransferase(ALT), creatinine(CREA), urea nitrogen(BUN),advanced glycation end products(AGEs) and receptor of glycation end products(RAGE) in blood or urine were measured during the experiments. Furthermore, on the day of the sacrifice, kidney was collected, and electronic microscopy and immunohistochemistry were performed to evaluate the protective renal effect of JTD. In addition, the levels of AGEs, RAGE, Cata-lase(CAT) and superoxide dismutase(SOD) were assessed by Western blot, Real-time PCR or ELISA to analyze the efficacy of JTD. This study demonstrated that JTD might protect kidney of KK-Ay by down-regulating the expression of AGEs, RAGE and oxidative stress.

Jiangtang decoction ameliorate diabetic nephropathy through the regulation of PI3K/Akt-mediated NF-κB pathways in KK-Ay mice.

BACKGROUND: Jiangtang decoction (JTD) is a China patented drug which contains Euphorbia humifusa Willd, Salvia miltiorrhiza Bunge, Astragalus mongholicus Bunge, Anemarrhena asphodeloides Bunge, and Coptis chinensis Franch. For decades, it has also been used clinically to treat diabetic nephropathy (DN) effectively; however, the associated mechanisms remain unknown. Thus, the present study aimed to examine the protective efficacy of JTD in DN and elucidate the underlying molecular mechanisms. METHODS: A diabetic model using KK-Ay mice received a daily administration of JTD for 12 weeks. Body weight, blood glucose, triglycerides (TGs), total cholesterol (TC), urea nitrogen (UN), creatinine (Cr), and microalbumin/urine creatinine (MA/UCREA) was measured every 4 weeks. Furthermore, on the day of the sacrifice, blood, urine, and kidneys were collected to assess renal function according to general parameters. Pathological staining was performed to evaluate the protective renal effect of JTD. In addition, the levels of inflammatory cytokines (tumor necrosis factor-α [TNF-α], interleukin [IL]-6 and intercellular adhesion molecule [ICAM]-1), insulin receptor substrate [IRS]-1, advanced glycation end products [AGEs], and receptor of glycation end products [RAGE] were assessed. Finally, the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway and involvement of nuclear factor-κB (NF-κB) was further analyzed. RESULTS: After 12 weeks of metformin and JTD administration, the mice exhibited a significant amelioration in glucose and lipid metabolism dysfunction, reduced morphological changes in the renal tissue, decreased urinary albumin excretion, and normalized creatinine clearance. JTD treatment also reduced the accumulation of AGEs and RAGE, up-regulated IRS-1, and increased the phosphorylation of both PI3K (p85) and Akt, indicating that the activation of the PI3K/Akt signaling pathway was involved. Additionally, JTD administration reduced the elevated levels of renal inflammatory mediators and decreased the phosphorylation of NF-κB p65. CONCLUSIONS: These results demonstrate that JTD might reduce inflammation in DN through the PI3K/Akt and NF-κB signaling pathways.

[Relationship between autophagy and T2DM and intervention effect of traditional Chinese medicine].

Insulin resistance and insulin secretion deficiency are main machanisms in inducing type 2 diabetes mellitus (T2DM), and mitochondria damage plays an important role in them. Research shows that autophagy is a self-protective mechanism of cells, which plays an important role in maintaining the normal structure and function of pancreatic ß cells and improving insulin resistance. Previous studies show that traditional Chinese medicine can regulate cell autophagy to influence ß cells and insulin resistance, type 2 diabetes mellitus and its complications. Thus this review will talk about the process of the relationship between autophagy and T2DM and the intervention effect of traditional Chinese medicine.