Enhanced trimethylamine metabolism and gut dysbiosis in type 2 diabetes mellitus with microalbumin.

Background: Abnormal gut microbiota and blood trimethylamine-N-oxide (TMAO) metabolome have been reported in patients with type 2 diabetes mellitus (T2DM) and advanced diabetic nephropathy. This study aimed to investigate the gut microbiota profiles and a group of targeted urine metabolic characteristics in T2DM patients with or without microalbuminuria, to determine the correlation between the gut microbiota composition, trimethylamine (TMA) metabolism, and the clinical features during progression of diabetic kidney disease (DKD). Methods: This study included 26 T2DM patients with microalbuminuria (Micro), 26 T2DM patients with normoalbuminuria (Normo), and 15 healthy controls (HC). Urine and Fecal samples were detected using ultra performance liquid chromatography tandem mass spectrometry and 16S ribosomal DNA gene sequencing, respectively. Results: The TMAO/TMA ratio decreased gradually during the HC-Normo-Micro transition. The levels of TMA, choline and betaine were significantly different between the HC group and the T2DM patients belonging to both Normo and Micro groups. At the operational taxonomic unit (OTU) level, the gut microflora diversity was significantly reduced in the Micro groups compared to the HC groups and the Normo groups. Taxonomic analyses revealed significant consumption in the relative abundances of eight bacterial genera and significant enrichment of two bacterial genera during the HC-Normo-Micro transition. Furthermore, the relative abundances of six bacterial genera, namely, Ruminococcus_1, [Eubacterium]_ruminantium_group, Roseburia, Faecalibacterium, Fusicatenibacter and Coprococcus_3 exhibited significant differences, and were associated with elevated urinary albumin creatinine ratio (UACR), TMAO/TMA, TMA and its precursors in the Micro group compared with the other groups. Conclusion: The imbalance of gut microbiota has occurred in patients with early-stage DKD, and the consumption of short-chain fatty acid-producing bacteria were associated with the accumulation of TMA and UACR.

Butyrate reverses ferroptosis resistance in colorectal cancer by inducing c-Fos-dependent xCT suppression.

Ferroptosis has emerged to be a promising approach in cancer therapies; however, colorectal cancer (CRC) is relatively insensitive to ferroptosis. Exactly how the gut microenvironment impacts the ferroptotic sensitivity of CRC remains unknown. Herein, by performing metabolomics, we discovered that butyrate concentrations were significantly decreased in CRC patients. Butyrate supplementation sensitized CRC mice to ferroptosis induction, showing great in vivo translatability. Particularly, butyrate treatment reduced ferroptotic resistance of cancer stem cells. Mechanistically, butyrate inhibited xCT expression and xCT-dependent glutathione synthesis. Moreover, we identified c-Fos as a novel xCT suppressor, and further elucidated that butyrate induced c-Fos expression via disrupting class I HDAC activity. In CRC patients, butyrate negatively correlated with tumor xCT expression and positively correlated with c-Fos expression. Finally, butyrate was found to boost the pro-ferroptotic function of oxaliplatin (OXA). Immunohistochemistry data showed that OXA non-responders exhibited higher xCT expression compared to OXA responders. Hence, butyrate supplementation is a promising approach to break the ferroptosis resistance in CRC.

RP11-296E3.2 acts as an important molecular chaperone for YBX1 and promotes colorectal cancer proliferation and metastasis by activating STAT3.

BACKGROUND: RP11-296E3.2 is a novel long noncoding RNA (lncRNA) associated with colorectal cancer (CRC) metastasis, that was reported in our previous clinical studies. However, the mechanisms of RP11-296E3.2 in colorectal tumorigenesis remain elusive. METHODS: RNA sequencing (RNA-seq), Fluorescence in situ hybridization (FISH), Transwell assays and others, were performed to evaluate the function of RP11-296E3.2 for proliferation and metastasis in vitro. In situ and metastatic tumor models were performed to evaluate the function of RP11-296E3.2 for proliferation and metastasis in vivo. RNA-pulldown, RNA-interacting protein immunoprecipitation (RIP), tissue microarray (TMA) assay, a luciferase reporter assay, chromatin immunoprecipitation (ChIP) and others were performed to explore the mechanisms by which RP11-296E3.2 regulates CRC tumorigenesis. RESULTS: RP11-296E3.2 was confirmed to be associated with CRC cell proliferation and metastasis in vitro and in vivo. Mechanistically, RP11-296E3.2 directly bound to recombinant Y-Box Binding Protein 1 (YBX1) and enhanced signal transducer and activator of transcription 3 (STAT3) transcription and phosphorylation. YBX1 promoted the CRC cell proliferation and migration, while knockdown of RP11-296E3.2 attenuated the effects of YBX1 on CRC cell proliferation, and metastasis and the expression of several related downstream genes. We are the first to discover and confirm the existence of the YBX1/STAT3 pathway, a pathway dependent on RP11-296E3.2. CONCLUSION: Together, these novel findings show that the RP11-296E3.2/YBX1 pathway promotes colorectal tumorigenesis and progression by activating STAT3 transcription and phosphorylation, and suggest that RP11-296E3.2 is a potential diagnostic biomarker and therapeutic target in CRC.

Bioinformatics Profiling and Experimental Validation of 4 Differentially-Expressed LIM Genes in the Course of Colorectal-Adenoma-Carcinoma.

BACKGROUND LIM domain proteins play crucial roles in tumors by interacting with diverse proteins. However, their roles in the course of colorectal mucosa-adenoma-carcinoma remain unclear. This study aimed to depict their dynamic expression profiles and elucidate their potential functions in this transition course. MATERIAL AND METHODS Differentially-expressed LIM proteins (DELGs) in paired adenomas, carcinomas, and mucosae were identified using the GEO dataset (GSE 117606) and validated by immunohistochemistry using our tissue microarray. Kaplan-Meier survival analysis, WGCNA, module-trait analysis, and KEGG enrichment were conducted. The correlation of DELGs expression levels with immune infiltration was assessed using the ESTIMATE package and TISCH database. The role of DELGs of interest was validated using cell proliferation, migration, and invasion assays. RESULTS Four DELGs were identified - LMO3, FHL1, NEBL, and TGFB1I1 - all of which were of significance in prognosis. Module-trait correlation and KEGG enrichment revealed their involvement in cancer-related signaling. Immunohistochemistry showed gradual downregulation of LMO3 but upregulation of NEBL in the mucosa-adenoma-carcinoma sequence. The opposite expression patterns were observed for FHL1 and TGFB1I1 in tumor epithelium and mesenchyme. High expression levels of the DELGs were correlated with increased infiltration of NK, NKT, and macrophages, except for NEBL. Importantly, LMO3 inhibited proliferation, migration, and invasion of colon epithelial cells. CONCLUSIONS This study identified 4 differentially-expressed LIM genes - LMO3, FHL1, TGFB1I1, and NEBL - and revealed they were involved in the mucosa-adenoma-carcinoma sequence via regulating cancer-related pathways, influencing epigenetic field, or affecting immune infiltration. Our findings provide new insights into the roles of LIM proteins in the course of mucosa-adenoma-carcinoma.

Graphene Nanosheets as Lubricant Additives: Effects of Nature and Size on Lubricating Performance.

Graphene has been widely investigated as an additive in lubricating oils to enhance their tribological performance. Here, the effects of the nature and size of the graphene nanosheets on the tribological performance were investigated with the hydrogenated hydroxyl-terminated polybutadiene dioctoate (O-HHTPB-O) as a model base oil after alkylation of the graphene oxide (GO) of different sizes with 1-dodecylamine (DA) and reduction. The 1-dodecylamine-modified graphene oxide (DA-GO) showed better dispersibility in the O-HHTPB-O base oil and subsequently better tribological performance than the reduced one (DA-rGO) for both the larger graphene oxide nanosheets (GOL) and the smaller graphene oxide nanosheets (GOS). The DA-GOS exhibited better wear-reduction performance than the DA-GOL, owing to its smaller size and higher polarity. Although the DA-GOL could be ground during the friction, the friction and wear in the original period affected the complete period lubricating performance.

Triptolide Alleviates Podocyte Epithelial-Mesenchymal Transition via Kindlin-2 and EMT-Related TGF-ß/Smad Signaling Pathway in Diabetic Kidney Disease.

Diabetes-induced chronic kidney diseases are widespread and decrease the quality of life for millions of affected individuals in China. To date, no therapies effectively alleviate these conditions. Triptolide, a traditionally used Chinese medicine, has shown promise in treating renal diseases. Here, the study aimed to decipher the exact mechanism by which it functions. It was hypothesized that triptolide might prevent the epithelial-mesenchymal transition (EMT) of podocytes by activating the kindlin-2 and TGF-ß/Smad pathways. Triptolide or telmisartan was intragastrically administered to 9-week-old db/db and dm/dm mice with diabetic nephropathy (DN) for 12 weeks. In addition, biochemical parameters and body weight were detected. WT-1, nephrin, podocin, E-cadherin, and α-SMA were determined by immunohistochemistry in the renal tissues of treated mice. Protein and mRNA expression of podocyte EMT markers, kindlin-2 and TGF-ß/Smad, were analyzed to elucidate the underlying mechanism. It was observed that triptolide treatment relieved structural injuries and functional variations in diabetic mice. It also increased the protein and mRNA levels of nephrin, podocin, and E-cadherin and decreased the expression of α-SMA in diabetic mice. The protein and mRNA expressions of TGF-ß1, p-SMAD3, and kindlin-2 decreased in diabetic kidneys following triptolide treatment. The findings demonstrated that triptolide might protect podocytes during DN by inhibiting podocyte EMT through inactivation of kindlin-2, combined with the downregulation of P-SMAD3 in the TGF-ß/Smad signaling pathway.

Study on the protective effect of berberine treatment on sepsis based on gut microbiota and metabolomic analysis.

Introduction: Sepsis, an infection with multiorgan dysfunction, is a serious burden on human health. Berberine (BBR), a bioactive component, has a protective effect on sepsis and the effect may be related to gut microbiota. However, studies on the role of BBR with gut microbiota in sepsis are lacking. Therefore, this study investigated the ameliorative effects and the underlying mechanisms of BBR on cecal ligature and puncture (CLP) rats. Methods: This study has observed the effect of BBR on pathological injury, Inflammation, intestinal barrier function, gut microbiota, and metabolite change in CLP rats by Hematoxylin-eosin staining, enzyme-linked immunosorbent assays, flow cytometry, 16S rDNA, and metabolomics analyses. Results: The inhibition effects of BBR treatment on the histological damage of the lung, kidney, and ileum, the interleukin (IL)-1b, IL-6, IL-17A, and monocyte chemokine-1 levels in serum in CLP rats were proved. Also, the BBR inhibited the diamine-oxidase and fluorescein isothiocyanate-dextran 40 levels, suggesting it can improve intestinal barrier function disorders. The cluster of differentiation (CD) 4+, CD8+, and CD25+ Forkhead box protein P3 (Foxp3) + T lymphocytes in splenocytes were up-regulated by BBR, while the IL-17A+CD4+ cell level was decreased. The abundance of gut microbiota in CLP rats was significantly different from that of the sham and BBR treatment rats. The significantly changed metabolites in the serum mainly included carbohydrates, phenols, benzoic acids, alcohols, vitamins et al. Additionally, this study predicted that the biological mechanism of BBR to ameliorate sepsis involves glycolysis-, nucleotide-, and amino acid-related metabolic pathways. Discussion: This study proved the strong correlation between the improvement effect of BBR on sepsis and gut microbiota and analyzed by metabolomics that gut microbiota may improve CLP rats through metabolites, providing a scientific basis for BBR to improve sepsis and a new direction for the study of the biological mechanism.

Elevated expansion of follicular helper T cells in peripheral blood from children with acute measles infection.

BACKGROUND: Measles outbreaks have threatened the global elimination and eradication of measles in recent years. Measles virus (MeV)-specific antibodies are successful in clearing MeV infection. Follicular helper T (Tfh) cells play a crucial role in promoting antibody production. This study investigated the potential role of Tfh cells in peripheral blood mononuclear cells (PBMCs) from children with acute MeV infection. RESULTS: The frequencies of CXCR5+CD4+ Tfh, ICOShigh Tfh, and PD-1high Tfh cells in PBMCs and levels of IL-6 and IL-21 in plasma were significantly elevated in patients with acute MeV infection. Moreover, a positive correlation was discovered among the frequency of ICOShigh Tfh cells, plasma levels of IL-21 and optical density (OD) values of MeV-specific IgM antibodies in the patients with acute MeV infection. However, elevated plasma MeV-specific NAb titres were not associated with the frequency of Tfh, ICOShigh Tfh, or PD-1high Tfh cells in the patients with acute MeV infection. CONCLUSION: These results suggest that an elevated Tfh cell frequency and associated molecules possibly play a key role in children with acute MeV infection, which contributes to the prevention and treatment of MeV infection in children.

Clinical characteristics of diabetic nephropathy in patients with type 2 diabetic mellitus manifesting heavy proteinuria: A retrospective analysis of 220 cases.

AIMS: To determine the predictability of diagnosing diabetic nephropathy (DN) versus non-diabetic renal disease (NDRD) from clinical and laboratory data in Chinese patients with type 2 diabetes mellitus (T2DM) manifesting heavy proteinuria. METHODS: We retrospectively analyzed the clinical and laboratory data of patients with T2DM manifesting heavy proteinuria who underwent renal biopsy from January 2014 to December 2017. RESULTS: According to renal biopsy, 220 patients were finally enrolled, including 109 cases diagnosed with DN alone (49.55%), 94 with NDRD alone (42.73%) and 17 with DN plus superimposed NDRD (7.73%). Multivariate analysis showed the significant risk factors for DN alone were age, duration of diabetes, presence of retinopathy, 24-h proteinuria, serum albumin and SBP. Presence of retinopathy achieved the highest overall diagnostic efficiency with the area under the curve of 0.852, sensitivity of 78.9% and specificity of 91.5%. The combined diagnosis with four indicators (duration of diabetes, retinopathy, SBP, and serum albumin) showed the area under the curve of 0.938, sensitivity of 88.1% and specificity of 87.2%. CONCLUSIONS: The prevalence of DN is high in patients with T2DM manifesting heavy proteinuria. Renal biopsy should be performed in diabetics in the atypical clinical scenario.

[Identification on host animals for plague by DNA barcoding technology in Shaanxi province].

OBJECTIVE: To apply the DNA barcoding technology for identification on host animal and to establish the host animal DNA bar code database on natural foci of plague in Shaanxi. METHODS: 139 host animals belonging to 3 orders, 6 families and 12 genera and 62 residues belonging to 7 species from 8 different parts of the province, were detected. DNA barcoding technology was used to analyze the DNA CO I gene sequence on the natural foci of plague in Dingbian county. RESULTS: The intra-specific genetic distance was less than 2% while the inter-specific distance ranged from 8.9% to 15.1%. Fourteen major clusters were apparently showed on a Neighbor-Joining tree. Residue samples could be detected regarding the objective gene. Alashan ground squirrel was previously noticed to carry 14 major clusters, which were previously mistakenly named as Citellus dauricus in Dingbian county. CONCLUSION: DNA barcoding technology could overcome the shortcomings caused by the morphological identification so could be used to identify the host animal and residues in the natural focus of plague.

Simultaneous determination of puerarin, daidzin, daidzein, paeoniflorin, albiflorin, liquiritin and liquiritigenin in rat plasma and its application to a pharmacokinetic study of Ge-Gen Decoction by a liquid chromatography-electrospray ionization-tandem mass spectrometry.

A liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) method was developed and validated for simultaneous determination of seven constituents including puerarin, daidzin, daidzein, paeoniflorin, albiflorin, liquiritin and liquiritigenin in rat plasma using schisandrin as the internal standard (IS). The plasma samples were pretreated by a one-step direct protein precipitation with acetonitrile. The chromatographic separation was carried out on a C18 column with a gradient mobile phase consisting of acetonitrile and water (containing 0.1% formic acid and 5mM ammonium acetate). All analytes and IS were quantitated through electrospray ionization in positive ion multiple reaction monitoring (MRM) mode. The mass transitions were as follows: m/z 417.5→297.2 for puerarin, m/z 417.1→255.2 for daidzin, m/z 255.2→152.4 for daidzein, m/z 498.1→179.3 for paeoniflorin, m/z 481.1→197.3 for albiflorin, m/z 436.2→257.3 for liquiritin, m/z 257.2→137.3 for liquiritigenin and m/z 415.0→384.2 for IS, respectively. All calibration curves exhibited good linearity (r>0.9979) over a wide concentration range for all components. The intra-day and inter-day precisions (RSD) at three different levels were both less than 14.3% and the accuracies (RE) ranged from -13.2% to 14.8%. The extraction recoveries of the seven compounds ranged from 72.9% to 117.4%. The validated method was successfully applied to pharmacokinetic study of the seven components in female rat plasma after oral administration of Ge-Gen Decoction aqueous extract.