ObjectiveTo investigate the mechanism of Zuogui Wan （ZGW） in improving ovarian inflammatory microenvironment and stemness of ovarian germline stem cells （OSCs） for treating ovarian aging via the cyclic guanosine monophosphate/adenosine monophosphate synthase （cGAS）/stimulator of interferon genes （STING） signaling pathway. MethodsForty C57BL/6 female mice were randomly divided into a blank group， a model group， a low-dose ZGW group （2.7 g·kg^-1）， a high-dose ZGW group （5.4 g·kg^-1）， and an estradiol valerate group （0.15 mg·kg^-1）， with 8 mice in each group. Except the blank group， all other groups received a single intraperitoneal injection of cyclophosphamide at 120 mg·kg^-1 to establish an ovarian aging mouse model. After successful modeling， each group was continuously administered for 4 weeks， once daily. The physiological status of the mice was observed， and the ovarian index was calculated. The estrus cycle of the mice was monitored. Hematoxylin-eosin （HE） staining was used to observe pathological changes in ovarian tissue. Enzyme-linked immunosorbent assay （ELISA） was used to detect serum sex hormone levels. Serum inflammatory factors interleukin-1β （IL-1β）， tumor necrosis factor-α （TNF-α）， and mouse interleukin-6 （IL-6） levels were detected using kits. Western blot was used to detect the protein expression of ovarian cGAS， STING， p-STING， TANK-binding kinase 1 （TBK1）， p-TBK1， interferon-induced transmembrane protein 3 （Fragilis）， and Vasa homolog protein （MVH）. Quantitative real-time polymerase chain reaction （Real-time PCR） was used to detect the mRNA expression of inflammatory factors in ovarian tissue. Immunofluorescence double labeling was performed to locate OSCs in ovarian tissues， and fluorescence intensities of OSCs markers MVH and octamer binding transcription factor 4 （Oct4） were calculated. ResultsCompared with the blank group， the model group showed reduced body weight， ovarian wet weight， and ovarian index （P<0.01） and a disordered estrus cycle （P<0.01）. In addition， the levels of serum follicle-stimulating hormone （FSH）， TNF-α， IL-6， and IL-1β were increased （P<0.01）， while anti-Müllerian hormone （AMH） and estradiol （E₂） levels were decreased （P<0.01）. The protein expression of cGAS， p-STING/STING， and p-TBK1/TBK1 in ovarian tissue was increased （P<0.05， P<0.01）， while that of OSCs stemness factors MVH and Fragilis was reduced （P<0.01）. Immunofluorescence indicated a reduction in MVH and Oct4 expression in OSCs （P<0.01）. The mRNA expression of inflammatory factors TNF-α， IL-6， and IL-1β in ovarian tissue was increased （P<0.05， P<0.01）. Compared with the model group， the treatment groups exhibited improved body weight， ovarian wet weight， and ovarian index （P<0.05） and a reduced rate of estrus cycle disorder （P<0.05， P<0.01）. The levels of serum FSH， TNF-α， IL-6， and IL-1β were decreased （P<0.05， P<0.01）， while AMH and E₂ levels were increased （P<0.01）. The protein expression levels of cGAS， p-STING/STING， and p-TBK1/TBK1 in ovarian tissue were decreased （P<0.05）， while the protein expression of MVH and Fragilis was increased （P<0.05）， and the fluorescence intensities of MVH and Oct4 were increased （P<0.05， P<0.01）. The mRNA expression of inflammatory factors in ovarian tissue was decreased （P<0.05）. ConclusionZGW alleviate ovarian inflammatory response， regulate ovarian microenvironment homeostasis， and maintain stemness of OSCs in ovarian aging mice probably by modulating the cGAS-STING signaling pathway， thereby improving ovarian function and delaying ovarian aging.

OBJECTIVES: The gut microbiota plays a crucial role in the pathophysiology of various types of diabetes. However, the causal relationship between them has yet to be systematically elucidated. This study aims to explore the potential causal associations between gut microbiota and diabetes using a two-sample Mendelian randomization (MR) analysis, based on multiple taxonomic levels. METHODS: Eligible instrumental variables were extracted from the selected genome-wide association study (GWAS) data on gut microbiota. These were combined with GWAS datasets on type 1 diabetes (T1D), type 2 diabetes (T2D), and gestational diabetes mellitus (GDM) to conduct forward MR analysis, sensitivity analysis, reverse MR analysis, and validation of significant estimates. Microbial taxa with causal effects on T1D, T2D, and GDM were identified based on a comprehensive assessment of all analytical stages. RESULTS: A total of 2 179, 2 176, and 2 166 single nucleotide polymorphisms (SNP) were included in the MR analyses for gut microbiota with T1D, T2D, and GDM, respectively. MR results indicated causal associations between: Six microbial taxa (Eggerthella, Lachnospira, Bacillales, Desulfovibrionales, Parasutterella, and Turicibacter) and T1D; 9 microbial taxa (Verrucomicrobia, Deltaproteobacteria, Actinomycetales, Desulfovibrionale, Actinomycetaceae, Desulfovibrionaceae, Actinomyces, Alcaligenaceae, and Lachnospiraceae NC2004 group) and T2D; 10 microbial taxa (Betaproteobacteria, Coprobacter, Ruminococcus2, Tenericutes, Clostridia, Methanobacteria, Mollicutes, Methanobacteriales, Methanobacteriaceae, and Methanobrevibacter) and GDM. CONCLUSIONS This study identified specific gut microbial taxa that may significantly increase or decrease the risk of developing diabetes. Some findings were fully replicated in independent validation datasets. However, the underlying biological mechanisms of these causal relationships warrant further investigation through mechanistic studies and population-based research.
Gastrointestinal Microbiome/genetics* ; Humans ; Mendelian Randomization Analysis ; Genome-Wide Association Study ; Diabetes Mellitus, Type 2/genetics* ; Diabetes Mellitus, Type 1/genetics* ; Female ; Polymorphism, Single Nucleotide ; Diabetes, Gestational/genetics* ; Pregnancy

OBJECTIVES: To investigate the therapeutic mechanism of 2,6-dimethoxy-1,4-benzoquinone (DMQ) for alleviating dextran sulfate sodium (DSS)-induced ulcerative colitis (UC) in mice. METHODS: Eighteen male C57BL/6J mice were equally randomized into control group, DSS group and DMQ treatment group. In DSS and DMQ groups, the mice were treated with DSS in drinking water to induce UC, and received intraperitoneal injections of sterile PBS or DMQ (20 mg/kg) during modeling. The changes in body weight, disease activity index (DAI), colon length, spleen weight, and colon histological scores of the mice were examined, and the percentages of Th17 and IFN-γ⁺ CD8⁺ T cells in the mesenteric lymph nodes and spleen were analyzed using flow cytometry. The expressions of tight junction proteins (Occludin and ZO-1), proteins associated with inflammasome activation (caspase-1 and p20), IL-1β and TNF-α in the colon tissues were detected using Western blotting or ELISA. In the cell experiment, mouse bone marrow-derived macrophages (BMDMs) primed with lipopolysaccharide (LPS) were treated with DMQ, followed by stmulation with nigericin to activate the classical NLRP3 inflammasome pathway. In cultured human peripheral blood mononuclear cells (PBMCs) treated with either LPS alone or LPS plus nigericin, the effects of DMQ on inflammasome activation, pyroptosis, and cytokine release were evaluated via Western blotting, ELISA, and flow cytometry. RESULTS: In DSS-treated mice, DMQ treatment significantly alleviated DSS-induced body weight loss, colon shortening, spleen enlargement, and colon inflammation. The DMQ-treated mice showed significantly reduced percentages of Th17 cells and IFN-γ⁺ CD8⁺ T cells in the mesenteric lymph nodes and spleen, with increased occludin and ZO-1 expressions and decreased caspase-1 expression in the colon tissue. DMQ obviously inhibited classical NLRP3 inflammasome activation in mouse BMDMs and both the classical and alternative pathways of NLRP3 activation in human PBMCs, causing also suppression of caspase-1-dependent pyroptosis. CONCLUSIONS DMQ ameliorates DSS-induced UC in mice by inhibiting NLRP3 inflammasome activation.
Animals ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Mice, Inbred C57BL ; Colitis, Ulcerative/metabolism* ; Dextran Sulfate/adverse effects* ; Male ; Inflammasomes/metabolism* ; Mice ; Benzoquinones/therapeutic use* ; Th17 Cells ; Caspase 1/metabolism*

Objective This study investigated the regulatory effect of high mobility group protein B1 (HMGB1) in the peripheral blood of patients with primary immune thrombocytopenia (ITP) on myeloid dendritic cells (mDC) and Th17/regulatory T cells (Treg) balance. Methods The study enrolled 30 newly diagnosed ITP patients and 30 healthy controls.Flow cytometry was used to measure the proportion of mDC, Th17, and Treg cells in the peripheral blood of ITP patients and healthy controls. ELISA was conducted to quantify the serum levels of HMGB1, interleukin 6 (IL-6), IL-23, IL-17, and transforming growth factor β(TGF-β). The mRNA levels of retinoic acid-related orphan receptor γt(RORγt) and forehead box P3(FOXP3) were detected by real-time PCR. The correlation between the abovementioned cells, cytokines, and platelet count was assessed using Pearson linear correlation analysis. Results The proportion of Th17 cells and the expression levels of HMGB1, IL-6, IL-23, IL-17 and the level of RORγt mRNA in the peripheral blood of ITP patients were higher than those in healthy controls. However, the Treg cell proportion and TGF-β level were lower in ITP patients than those in healthy controls. In patients with ITP, the proportion of mDC and the level of FOXP3 mRNA did not show significant changes. The proportion of mDC cells was significantly correlated with the expression of IL-6 and IL-23. Moreover, the expression of HMGB1 showed a significant correlation with the expression of mDC, IL-6, IL-23, RORγt mRNA, and IL-17. Notably, both the proportion of mDC cells and the expression of HMGB1 were negatively correlated with platelet count. Conclusion The high expression of HMGB1 in peripheral blood of ITP patients may induce Th17/Treg imbalance by promoting the maturation of mDC and affecting the secretion of cytokines, thereby potentially playing a role in the immunological mechanism of ITP.
Humans ; Th17 Cells/cytology* ; HMGB1 Protein/genetics* ; T-Lymphocytes, Regulatory/cytology* ; Female ; Male ; Dendritic Cells/metabolism* ; Adult ; Middle Aged ; Purpura, Thrombocytopenic, Idiopathic/genetics* ; Nuclear Receptor Subfamily 1, Group F, Member 3/genetics* ; Young Adult ; Interleukin-23/blood* ; Interleukin-17/blood* ; Interleukin-6/blood* ; Forkhead Transcription Factors/genetics* ; Myeloid Cells/cytology* ; Aged

OBJECTIVE: To explore the clinical features and management of a child with Chitayat syndrome. METHODS: A child presented at the Fengqing People's Hospital on August 8 2019 was selected as the study subject. Clinical data of the child were retrospectively analyzed. Peripheral blood samples were collected from the child and his father and sister. Whole-exome sequencing (WES) was carried out. Candidate variant was verified by Sanger sequencing. Genome Browser, AlphaFold, and PolyPhen-2 were employed for protein structure simulation and amino acid sequence conservation analysis. Pathogenicity of the variant was rated based on guidelines from the American College of Medical Genetics and Genomics (ACMG). Literature was retrieved from databases including CNKI, Wanfang, and PubMed using the keyword "Chitayat syndrome". The clinical characteristics and prognosis of patients with Chitayat syndrome were reviewed and analyzed. This study was approved by the Ethics Committee the Seventh Affiliated Hospital of Sun Yat-sen University (Ethics No.: KY-2024-086-01). RESULTS: The child was born at full term and had special facial features, skeletal abnormalities, recurrent respiratory tract infections and global developmental delay. WES and Sanger sequencing revealed that he has harbored a heterozygous c.266A>G p.(Tyr89Cys) variant of the ERF gene. Protein structure modeling suggested that the mutant protein has increased spatial distance between the side chain group and DNA, which may reduce its binding affinity to DNA. Amino acid sequence analysis indicated that the p.Tyr89 residue is highly conserved across multiple species. The variant was therefore classified as pathogenic (PM1+PM2_Supporting+PM6+PS1+PP3). The patient was diagnosed with "Chitayat syndrome". Nutritional support and rehabilitation training were recommended, though the child had died of severe pneumonia at 13 months old. Literature retrieval has collected 7 relevant articles, which involved 14 cases of Chitayat syndrome confirmed by genetic testing. Together with our case, all patients had facial dysmorphisms and skeletal deformities. Fourteen patients (93.3%) had respiratory distress. Seven of them (46.7%) had recurrent respiratory infections and 7 (46.7%) were confirmed with respiratory tract malacia. Eight (53.3%) patients had neuropsychological retardation, while 8 (53.3%) had growth delay. The main interventions for Chitayat syndrome include respiratory and nutritional support, and rehabilitation training for developmental delays. CONCLUSION Chitayat syndrome is rarely seen and its clinical manifestations may vary. Airway management and early intervention of developmental delay are important for improving the prognosis.
Humans ; Male ; Exome Sequencing ; Female ; Mutation ; Child, Preschool ; Infant ; Developmental Disabilities/genetics*

Objective:To explore the clinical features and management of a child with Chitayat syndrome.Methods:A child presented at the Fengqing People′s Hospital on August 8 2019 was selected as the study subject. Clinical data of the child were retrospectively analyzed. Peripheral blood samples were collected from the child, father and sister. Whole-exome sequencing (WES) was carried out. Candidate variant was verified by Sanger sequencing. Genome Browser, AlphaFold, and PolyPhen-2 were employed for protein structure simulation and amino acid sequence conservation analysis. Pathogenicity of the variant was rated based on guidelines from the American College of Medical Genetics and Genomics (ACMG). Literature was retrieved from databases including CNKI, Wanfang, and PubMed using the keyword "Chitayat syndrome". The clinical characteristics and prognosis of patients with Chitayat syndrome were reviewed and analyzed. This study was approved by the Ethics Committee the Seventh Affiliated Hospital of Sun Yat-sen University (Ethics No.: KY-2024-086-01).Results:The child was born at full term and had special facial features, skeletal abnormalities, recurrent respiratory tract infections and global developmental delay. WES and Sanger sequencing revealed that she has harbored a heterozygous c. 266A>G p. (Tyr89Cys) variant of the ERF gene. Protein structure modeling suggested that the mutant protein has increased spatial distance between the side chain group and DNA, which may reduce its binding affinity to DNA. Amino acid sequence analysis indicated that the p. Tyr89 residue is highly conserved across multiple species. The variant was therefore classified as pathogenic (PM1+ PM2_Supporting+ PM6+ PS1+ PP3). The patient was diagnosed with "Chitayat syndrome". Nutritional support and rehabilitation training were recommended, though the child had died of severe pneumonia at 13 months old. Literature retrieval has collected 7 relevant articles, which involved 14 cases of Chitayat syndrome confirmed by genetic testing. Together with our case, all patients had facial dysmorphisms and skeletal deformities. Fourteen patients (93.3%) had respiratory distress. Seven of them (46.7%) had recurrent respiratory infections and 7 (46.7%) were confirmed with respiratory tract malacia. Eight (53.3%) patients had neuropsychological retardation, while 8 (53.3%) had growth delay. The main interventions for Chitayat syndrome include respiratory and nutritional support, and rehabilitation training for developmental delays. Conclusion:Chitayat syndrome is rarely seen and its clinical manifestations may vary. Airway management and early intervention of developmental delay are important for improving the prognosis.

Objective:To determine if preoperative 68Ga-fibroblast activation protein inhibitor (FAPI)-04 PET/MR could contribute to predicting pathological complete response (pCR) in patients with gastrointestinal cancer undergoing neoadjuvant immunotherapy. Methods:In this retrospective study, 35 patients (23 males, 12 females, age (59.1±7.9) years) with gastrointestinal cancer who underwent 68Ga-FAPI-04 PET/MR after receiving neoadjuvant immunotherapy between February 2021 and January 2024 were enrolled. Clinical data, PET imaging parameters including SUV, peak of SUV normalized by lean body mass (SUL peak), FAPI-positive tumor volume (FTV), and total FAPI-positive lesion burden (TLF), and pathological data were collected and analyzed. Patients were divided into pCR group and non-pCR group, and the independent-sample t test or Mann-Whitney U test was performed to compare those parameters between the 2 groups. ROC curve analysis (Delong test) was performed to evaluate the diagnostic efficiency of each parameter to predict pCR. Results:The overall pCR rate of the neoadjuvant therapy was 40.0%(14/35). In the visual evaluation, 68Ga-FAPI-04 PET was limited in predicting pCR, showing false positivity in 12 patients and false negative in 1 patent. While SUV max( t=2.50, P=0.018), SUL peak( t=3.11, P=0.004), FTV( U=3.00, P=0.030) and TLF( U=2.96, P=0.042) in non-pCR group were all higher than those in pCR group. The predictive efficiency of FTV <1.925cm 3 for pCR was better than the efficiency of PET visual evaluation ( Z=3.61, P<0.001), with the prediction accuracy of 82.86%(29/35). Conclusions:68Ga-FAPI-04 PET/MR may provide an effective clinical tool for guiding further treatment of patients with gastrointestinal cancer undergoing neoadjuvant immunotherapy. The quantitative features derived from 68Ga-FAPI-04 PET appear promising in predicting pCR, which are expected to provide a reference for avoiding surgery.

Objective:To characterize the vaginal flora of pregnant women during the second trimester using full-length 16S rRNA sequencing.Methods:A total of 142 pregnant women were systematically sampled from a pregnancy cohort. Vaginal swabs were collected for full-length 16S rRNA gene sequencing，and bioinformatics analysis was performed to characterize the vaginal microbiota and identify associated influencing factors.Results:Among the 142 pregnant women，the most frequently detected species were Lactobacillus iners（83.10%，118/142）and Lactobacillus crispatus（49.30%，70/142）. The majority of samples（90.85%，129/142）were classified as Lactobacillus-dominant vagitypes，with the Lactobacillus iners vagitype accounting for 48.59%（69/142）and the Lactobacillus crispatus vagitype accounting for 38.73%（55/142）. The vaginal microbiota was clustered into five community state types（CSTs）：Ⅰa，Ⅰb，Ⅲa，Ⅲb，and Ⅳ. The most prevalent CSTs were Lactobacillus iners-dominated CST-Ⅲ（51.41%，73/142）and Lactobacillus crispatus-dominated CST-Ⅰ（24.65%，35/142）. No samples were classified as CST-Ⅱ or CST-Ⅴ. A significant negative correlation was observed between Lactobacill and vaginosis-associated bacteria. Age，alcohol consumption，smoking，and vaginal treatments showed significant associations or trends toward significance with various Alpha diversity indices. Vaginal douching was associated with CST clustering，while obstetric history（primiparity，previous miscarriage history）was associated with vagitype classification. However，no significant associations were identified between maternal baseline characteristics and Beta diversity indices. Conclusions:Full-length 16S rRNA gene sequencing reveals that the vaginal microbiota of pregnant women is dominated by Lactobacillus iners and Lactobacillus crispatus. Maternal age，lifestyle factors such as smoking and alcohol consumption，and obstetric history are significantly associated with variations in vaginal microbiota composition.

Objective:To develop a camelid single-domain antibody (SdAb) recognizing linear B-cell epitopes in glutamate dehydrogenase of Clostridium difficile(CD-GDH), and to apply it in Western blot and ELISA. Methods:Purified recombinant CD-GDH was used as bait to screen phage-displayed camelid SdAb library and obtain positive clones. Then those clones were confirmed by Western blot, and their variable domain of heavy chain of heavy chain antibody(VHH) nucleotide sequence were determined. The VHH sequence was synthesized after codon optimization and cloned into the expression vector pET28a. The SdAb was then expressed and purified, and its ability to detect CD-GDH protein in multiple assays was further explored.Results:Six positive clones were obtained, among which clone GA4 was chosen for recombinant expression in Escherichia coli and further purification. The purified GA4 binded well with CD-GDH with a Kd value of 3 nmol/L. In Western blot and ELISA, GA4 was proven to be able to selectively detect both recombinant and endogenous CD-GDH. Conclusions:A camelid SdAb targeting a linear B-cell epitope in CD-GDH is successfully developed, which provides a very useful tool for detecting CD-GDH.

Objective:To explore the impact of noise and dust exposure in the mechanical manufacturing industry on the risk of hypertension and abnormal electrocardiogram in workers, and their combined effects, in order to provide support for the prevention and treatment of occupational related diseases among workers.Methods:In January 2024, A convenience sampling method was used to study 2802 on-the-job workers who underwent occupational health check-ups from January 2023 to December 2023 at a machinery manufacturing enterprise in Baotou, Inner Mongolia. Blood pressure and electrocardiogram results were analyzed in the noise group, dust group, dust noise group and control group according to the exposure factors. For count data, the chi-square test was employed to analyze differences among groups. Additionally, a binary logistic regression model was utilized to assess the impact of dust and noise exposure on the prevalence of hypertension and electrocardiogram abnormalities.Results:The stratified analysis results showed that the differences in hypertension prevalence among the four groups were statistically significant ( P<0.001) in males, age groups >30-40 years, >40-50 years, >50 years, different exposure durations, and different enterprise sizes. For ECG abnormalities, significant differences were observed among the four groups ( P<0.001) in males, the age group >30-40 years, different enterprise sizes, and those with exposure durations ≤15 years. The trend test for hypertension prevalence across different age groups revealed that as age increased, the prevalence of hypertension showed an upward trend in the noise-exposed group, dust-exposed group, and combined dust-noise-exposed group ( χ2=10.76, 4.25, 6.60, P<0.001, 0.039, 0.010) . Binary regression model analyses revealed that the the risk of hypertension in the noise group, dust group and dust noise group was 2.63 times ( OR=2.63, 95% CI: 1.89~3.67, P<0.001) , 2.36 times ( OR=2.36, 95% CI: 1.76~3.16, P<0.001) and 2.69 times ( OR=2.69, 95% CI: 2.14~3.38, P<0.001) . Using ECG abnormalities as the dependent variable and incorporating the statistically significant variables from Table 1 as independent variables into the binary logistic regression model, the results showed no statistically significant differences in the risk of ECG abnormalities in the dust-exposed, noise-exposed, and combined dust-noise-exposed groups compared to the control group ( P> 0.05) . Conclusion:Combined exposure to dust and noise increases the risk of hypertension in workers and has a combined effect.