BACKGROUND:Endometrial receptivity is a key factor in embryo implantation in northwest Tibetan cashmere goats,and the expression of genes related to endometrial receptivity and their variable splicing are still unclear. OBJECTIVE:To analyze and explore genes and variable splicing events related to endometrial receptivity in northwest Tibetan cashmere goats. METHODS:On days 5 and 15 of pregnancy(representing pre receptive endometrium group and receptive endometrium group),three northwest Tibetan cashmere goats were randomly selected.Endometrial tissue was collected and stained with hematoxylin and eosin to observe tissue morphology.Immunohistochemical staining was used to detect the expression of endometrial receptive marker proteins leukemia inhibitory factor and vascular endothelial growth factor.After the total RNA was extracted and the quality test was qualified,transcriptome sequencing was performed to search differentially expressed mRNAs,lncRNAs,circRNAs,and miRNAs,perform functional prediction,and analyze alternative splicing mRNAs and lncRNAs related to endometrial receptivity. RESULTS AND CONCLUSION:(1)Compared with the pre receptive endometrium group,the expression levels of leukemia inhibitory factor and vascular endothelial growth factor proteins in the endometrial tissue of the receptive endometrium group were significantly increased.(2)The sequencing results showed that the differentially expressed genes were mostly mRNA and lncRNA genes,including 250 upregulated mRNAs,193 upregulated lncRNAs,135 downregulated mRNAs,and 123 downregulated lncRNAs,which were significantly enriched in the Wnt,Hedgehog,and Hippo signaling pathways.(3)Alternative splicing event analysis uncovered 8 differentially expressed variable splicing transcripts,which were all mRNA transcripts,including 2 downregulated and 6 upregulated,and were significantly associated with vascular endothelial growth factor receptor signaling,cell motility,and embryonic development.

This study aims to explore whether Naoxintong Capsules improve multiple cerebral infarctions and myocardial injury via promoting angiogenesis, thereby exerting a simultaneous treatment effect on both the brain and heart. Male SD rats were randomly divided into six groups: sham-operated group, model group, high-dose, medium-dose, and low-dose groups of Naoxintong Capsules(440, 220, and 110 mg·kg~(-1)), and nimodipine group(10.8 mg·kg~(-1)). Rat models of multiple cerebral infarctions were established by injecting autologous thrombus, and samples were collected and tested seven days after modeling. Evaluations included multiple cerebral infarction model assessments, neurological function scores, grip strength tests, and rotarod tests, so as to evaluate neuromotor functions. Morphological structures of brain and heart tissue were observed using hematoxylin-eosin(HE) staining, Nissl staining, and Masson staining. Network pharmacology was employed to screen the mechanisms of Naoxintong Capsules in improving multiple cerebral infarctions and myocardial injury. Neuronal and myocardial cell ultrastructures were observed using transmission electron microscopy. Apoptosis rate in brain neuronal cells was detected by TdT-mediated dUTP nick end labeling(TUNEL) staining, and reactive oxygen species(ROS) levels in myocardial cells were measured. Immunofluorescence was used to detect the expression of platelet endothelial cell adhesion molecule-1(CD31), antigen identified by monoclonal antibody Ki67(Ki67), hematopoietic progenitor cell antigen CD34(CD34), and hypoxia inducible factor-1α(HIF-1α) in brain and myocardial tissue. Western blot, and real-time quantitative polymerase chain reaction(RT-qPCR) were used to detect the expression of HIF-1α, vascular endothelial growth factor(VEGF), vascular endothelial growth factor receptor 2(VEGFR2), sarcoma(Src), basic fibroblast growth factor(bFGF), angiopoietin-1(Ang-1), and TEK receptor tyrosine kinase(Tie-2). Compared with the model group, the medium-dose group of Naoxintong Capsules showed significantly lower neurological function scores, increased grip strength, and prolonged time on the rotarod. Pathological damage in brain and heart tissue was reduced, with increased and more orderly arranged mitochondria in neurons and cardiomyocytes. Apoptosis in brain neuronal cells was decreased, and ROS levels in cardiomyocytes were reduced. The microvascular density and endothelial cells of new blood vessels in brain and heart tissue increased, with increased overlapping regions of CD31 and Ki67 expression. The relative protein and mRNA expression levels of HIF-1α, VEGF, VEGFR2, Src, Ang-1, Tie-2, and bFGF were elevated in brain tissue and myocardial tissue. Naoxintong Capsules may improve multiple cerebral infarctions and myocardial injury by mediating HIF-1α/VEGF expression to promote angiogenesis.
Animals ; Male ; Drugs, Chinese Herbal/administration & dosage* ; Rats, Sprague-Dawley ; Rats ; Cerebral Infarction/genetics* ; Hypoxia-Inducible Factor 1, alpha Subunit/genetics* ; Vascular Endothelial Growth Factor A/genetics* ; Capsules ; Signal Transduction/drug effects* ; Humans ; Brain/metabolism* ; Myocardium/metabolism* ; Apoptosis/drug effects*

OBJECTIVE: To explore the influence of oligospermia (OS) on the detection of sperm DNA fragmentation index (DFI) by fluorescence method based on artificial intelligence (AI) recognition and flow cytometry-based sperm chromatin structure assay (SCSA). METHODS: We collected semen samples from 201 males, including 50 azoospermia (AS) patients as negative controls, 90 OS patients (sperm concentration >0×10⁶/ml and <15×10⁶/ml), and 61 normal men (sperm concentration ≥15×10⁶/ml). Then we subdivided the OS patients into a mild OS (sperm concentration ≥10×10⁶/ml and <15×10⁶/ml), a moderate OS (sperm concentration ≥5×10⁶/ml and <10×10⁶/ml) and a severe/extremely severe OS group (sperm concentration >0×10⁶/ml and <5×10⁶/ml), with 30 cases in each group, and compared the results of DFI detection between the AI fluorescence method and traditional flow cytometry. RESULTS: The DFI value detected by AI fluorescence method showed statistically significant difference from that detected by flow cytometry in the AS, moderate OS and severe/extremely severe OS groups (P<0.01), the former even lower than the latter, but not in the normal control and the mild OS groups (P > 0.05). In the AS group, a dramatically lower rate of non-0 results was achieved by AI fluorescence method than by flow cytometry (8% vs 100%, P<0.01). The DFI values detected by AI fluorescence method exhibited a good linear correlation to those obtained by flow cytometry in the normal control and mild OS groups (R2 = 0.7470; R2 = 0.7180), but a poor linear correlation in the OS full-sample, moderate OS and severe/extremely severe OS groups (R2 = 0.3092; R2 = 0.3558; R2 = 0.2147). CONCLUSION The AI fluorescence method has a higher specificity and is more suitable than flow cytometry for detection of sperm DFI in OS patients. The DFI values obtained by the two methods are consistent with sperm concentration ≥10×10⁶/ml, but the accuracy of the results of detection may be affected with sperm concentration >0×10⁶/ml and <10×10⁶/ml.
Humans ; Male ; Flow Cytometry/methods* ; Oligospermia/genetics* ; Artificial Intelligence ; Spermatozoa ; Adult ; DNA Fragmentation ; Case-Control Studies ; Fluorescence

OBJECTIVE: This study investigates the efficacy and mechanisms of Qingjie Fuzheng Granules (QFG) in inhibiting colitis-associated colorectal cancer (CAC) development via RNA sequencing (RNA-seq) and 16S ribosomal RNA (rRNA) correlation analysis. METHODS: CAC was induced in BALB/c mice using azoxymethane (AOM) and dextran sulfate sodium (DSS), and QFG was administered orally to the treatment group. The effects of QFG on CAC were evaluated using disease index, histology, and serum T-cell ratios. RNA-seq and 16S rRNA analysis assessed the transcriptome and microbiome change. Key pharmacodynamic pathways were identified by integrating these data and confirmed via Western blotting and immunofluorescence. The link between microbiota and CAC-related markers was explored using linear discriminant analysis effect size and Spearman correlation analysis. RESULTS: Long-term treatment with QFG prevented AOM/DSS-induced CAC formation, reduced levels of interleukin (IL)-1β, tumor necrosis factor-alpha (TNF-α), IL-6, and interferon γ (IFN-γ), and increased CD3⁺ and CD4⁺/CD8⁺ T cells ratio, without causing hepatic or renal toxicity. A 16S rRNA analysis revealed that QFG rebalanced the Firmicutes/Bacteroidetes ratio and mitigated AOM/DSS-induced microbiota disturbances. Transcriptomics and Western blotting analysis identified the nucleotide-binding oligomerization domain-containing protein 2 (NOD2)/nuclear factor kappa-B (NF-κB) pathway as key for QFG's treatment against CAC. Furthermore, QFG decreased the abundance of Bacilli, Bacillales, Staphylococcaceae, Staphylococcus, Lactobacillales, Aerococcus, Alloprevotella, and Akkermansia, while increasing Clostridiales, Lachnospiraceae, Lachnospiraceae_NK4A136_group, Ruminococcaceae, and Muribaculaceae, which were highly correlated with CAC-related markers or NOD2/NF-κB pathway. CONCLUSION By mapping the relationships between CAC, immune responses, microbiota, and key pathways, this study clarifies the mechanism of QFG in inhibiting CAC, highlighting its potential for clinical use as preventive therapy.

Objective To investigate the risk factors for bronchopulmonary dysplasia(BPD)in twin preterm infants with a gestational age of<34 weeks,and to provide a basis for early identification of BPD in twin preterm infants in clinical practice.Methods A retrospective analysis was performed for the twin preterm infants with a gestational age of<34 weeks who were admitted to 22 hospitals nationwide from January 2018 to December 2020.According to their conditions,they were divided into group A(both twins had BPD),group B(only one twin had BPD),and group C(neither twin had BPD).The risk factors for BPD in twin preterm infants were analyzed.Further analysis was conducted on group B to investigate the postnatal risk factors for BPD within twins.Results A total of 904 pairs of twins with a gestational age of<34 weeks were included in this study.The multivariate logistic regression analysis showed that compared with group C,birth weight discordance of>25%between the twins was an independent risk factor for BPD in one of the twins(OR=3.370,95%CI:1.500-7.568,P<0.05),and high gestational age at birth was a protective factor against BPD(P<0.05).The conditional logistic regression analysis of group B showed that small-for-gestational-age(SGA)birth was an independent risk factor for BPD in individual twins(OR=5.017,95%CI:1.040-24.190,P<0.05).Conclusions The development of BPD in twin preterm infants is associated with gestational age,birth weight discordance between the twins,and SGA birth.

Traditional Chinese medicine(TCM)is a medical science about the physiological state and disease process of human body.It contains the universality of general thinking methods and the particularity of oriental cultural thinking.Starting from the necessity of the construction of clinical thinking mode of TCM and based on the philosophical features of TCM,this paper re-examines and analyzes the importance of the thinking mode for the nature of disease,and puts forward the six-dimensional differentiation system for TCM clinical practice,which covers disease differentiation,syndrome differentiation,symptom differentiation,pathogenesis differentiation,constitution differentiation and qi differentiation.Guided by the thinking mode of six-dimensional differentiation system,the TCM clinical practice exerts holistic view:disease differentiation is advocated to combine syndrome differentiation;symptom differentiation is mainly for the primary symptoms,which can clarify the transformation of pathogenesis,the progression of the disease,and the evolution of the syndromes;pathogenesis differentiation focuses on the evolution of pathogenesis,and treatment based on syndrome differentiation and modification of drugs based on symptoms are required;constitution differentiation should be made on the basis of the patient's constitution,and detailed identification of the etiology,disease location,disease nature,and progression of the disease should also be conducted;qi differentiation should be made on the basis of the comprehensive consideration of the exuberance and decline of healthy qi and pathogenic qi,thus to ensure the treatment adapting to the individuality,climate,and environment.The establishment of six-dimensional differentiation system aims to expound the internal mechanism of key points for disease differentiation,therapeutics based on syndrome differentiation,drug modification according to symptom differentiation,and differentiation of pathogenesis and illness state,which is helpful to the overall grasp of the disease,pathogenesis,syndrome and treatment in clinical practice,and will provide theoretical basis for the precise treatment of disease.

Objective To detect the levels of serum collagen type Ⅰ alpha 1 chain(COL1A1)and collagen type Ⅰ alpha 2 chain(COL1A2)in patients with intracranial aneurysm(IA),and explore their correlations with aneurysm rupture.Methods A total of 110 IA patients admitted to our hospital were regarded as the IA group and another 100 volunteers who underwent physical examination in our hospital were regarded as the control group.The expression levels of serum COL1A1 and COL1A2 were detected by ELISA.The IA patients were divided into the ruptured group(n=66)and unruptured group(n=44)according to the presence or absence of aneurysm rupture,and the clinical data and expression levels of serum COL1A1 and COL1A2 were compared between the two groups.The expression levels of serum COL1A1 and COL1A2 in patients with different Hunt-Hess grades were compared.The risk factors of aneurysm rupture in patients with IA were analyzed by multivariate Logistic regression analysis.The predictive value of serum COL1A1 and COL1A2 for aneurysm rupture in patients with IA were evaluated by receiver operating characteristic(ROC)curve.The correlation of serum COL1A1 and COL1A2 with Hunt-Hess grade for patients in rupture group was analyzed by Spearman correlation analysis.Results The expression levels of serum COL1A1 and COL1A2 for patients in the IA group were significantly higher than those in the control group(P<0.05).The number of patients with hypertension,diabetes mellitus,hyperlipidemia,aneurysm diameter>10 mm,and the expression levels of serum COL1A1 and COL1A2 in the rupture group were significantly more/higher than those in the unruptured group(P<0.05).The expression levels of serum COL1A1 and COL1A2 in patients with Hunt-Hess grades from Ⅲ to Ⅳ were significantly higher than those in patients with grades from Ⅰ to Ⅱ(P<0.05).The expression levels of serum COL1A1 and COL1A2 for patients in the rupture group were positively correlated with Hunt-Hess grade(r=0.562,0.414,P<0.05).Multivariate Logistic regression analysis showed that hypertension,diabetes mellitus,aneurysm diameter>10 mm,and increased expression levels of COL1A1 and COL1A2 were risk factors for aneurysm rupture in IA patients(P<0.05).The area under the curve(AUC)of aneurysm rupture predicted by serum COL1A1 and COL1A2 together was significantly higher than that predicted by COL1A1 alone(Z=1.905,P=0.028)and COL1A2 alone(Z=1.754,P=0.040).Conclusion The increased expression levels of serum COL1A1 and COL1A2 are risk factors for aneurysm rupture in patients with IA,and their combined prediction of aneurysm rupture in IA patients has certain clinical value.

Objective:To investigate the clinical efficacy of three-connections and four-screwings technique in the treatment of high double column acetabular fractures through a single ilioinguinal approach.Methods:A retrospective study was conducted to analyze the data of 42 patients who had been treated for high double column acetabular fractures from June 2017 to June 2020 at Trauma Ward 2, Department of Orthopedics and Traumatology, The First Hospital of Traditional Chinese Medicine of Changde. There were 19 males and 23 females with an age of (42.7±25.6) years. 29 injuries were due to a traffic accident, 12 ones to fall from a height, and one to fall. The time from injury to operation was (4.5±2.1) days. All the patients were treated by the three-connections and four-screwings technique through a single ilioinguinal approach. Briefly, the anterior column was connected and secured to the main bone using 3 routes, and the posterior column was attached and fixated to the anterior column reset using 2 or 3 of the 4 screwings. The operation time, intraoperative blood loss, fracture reduction quality, fracture healing time, hip function at the last follow-up and complications during the follow-up were recorded.Results:For this cohort, the operation time was (150.0±30.5) min, and intraoperative blood loss (300.0±50.0) mL. According to the Matta scale for postoperative acetabular fracture reduction, 34 cases were excellent, 6 cases good, and 2 cases acceptable, with an excellent and good rate of 95.2% (40/42). After operation one patient had fat liquefaction and wound exudation which responded to drainage and dressing change. The 42 patients were followed up for (15.0±3.4) months. All fractures healed after (11.0±2.0) months. By the modified Merle d'Aubigné & Postel scoring system, the hip function was evaluated at the last follow-up as excellent in 33 cases, as good in 6 cases, and as fair in 3 cases, yielding an excellent and good rate of 92.9% (39/42).Conclusions:In the treatment of high double column acetabular fractures, the three-connections and four-screwings technique through a single ilioinguinal approach can lead to fine reduction and rigid fixation by lag screw compression and neutralization plate protection. Consequently, early functional exercises can be performed to secure good therapeutic outcomes for the patients.

Aim To investigate the synergistic sensiti-zation effect of saikosaponin D(SSD)combined with temozolomide(TMZ)on glioblastoma cells(GBM)and its molecular mechanism.Methods The sensitiv-ity of RG-2,U251 and LN-428 GBM cell lines to SSD and TMZ was analyzed by CCK-8 method combined with HE staining,and the optimal compatible concen-tration was screened.The effect of HE staining com-bined with Hoechst fluorescence staining on the prolif-eration of GBM cell line was detected by clonal forma-tion experiment.The autophagosome formation of GBM cells was observed by monodansylcadaverine(MDC)staining.The expression and distribution of endoplas-mic reticulum stress-related factors and apoptosis and autophagy proteins were detected by Western blot and ICC.Results The sensitivity order of GBM cells to TMZ was RG-2＞U251＞LN-428.The results of com-bined administration showed the synergistic inhibitory effect of SSD combined with TMZ on proliferation of GBM cell lines,which was confirmed by cell cloning formation experiment.Compared with the TMZ group,Hoechst fluorescence staining showed a significant in-crease in the number of nuclear bright staining in the combined administration group.MDC fluorescence staining showed that there were more dense green parti-cles in the cytoplasm of SSD/TMZ plus group than that of TMZ group.Western blot results showed that com-pared with TMZ group,the expression of ER stress markers GRP78,CHOP,p-PERK and ATF6 signifi-cantly increased in SSD/TMZ group(P＜0.05).The expressions of apoptosis proteins caspase-12,caspase-9,caspase-3,cleaved caspase-3,Bax and autophagy proteins LC3 and Beclin-1 significantly increased(P＜0.05),which were verified by ICC test.Conclusions SSD can cooperate with TMZ to inhibit the prolifera-tion of GBM cells and induce apoptosis and autophagy,and enhance the sensitivity of GBM cells to TMZ by ac-tivating endoplasmic reticulum stress pathway.

This study aims to investigate the regulatory effects of Astragalus polysaccharide(APS) and APS combined with 5-fluorouracil(5-FU) on indoleamine-2,3-dioxygenase(IDO1) in the colon tumor microenvironment. Sixty Balb/c mice were randomized into a blank group, a model group, an APS group, an APS + 5-FU group, an APS + low-dose 5-FU group, and a 5-FU group. A tumor model was established by subcutaneous transplantation with CT-26 mouse colon cancer cells in other groups except the blank group. After successful modeling, each group was treated with corresponding drugs for 7 days. The general condition, body weight, and tumor volume of the mice were observed and measured daily during the treatment period. The mice were sacrificed at the end of treatment, and the tumor suppression rate and spleen index of the mice were calculated. Western blot and fluorescence quantitative PCR were employed to determine the protein and mRNA levels, respectively, of IDO1 in the tumor tissue of mice. High performance liquid chromatography was employed to measure the levels of tryptophan(Trp) and kynurenine(Kyn) in the tumor tissue of mice. Hematoxylin-eosin(HE) staining was performed to observe the histological changes of the tumor tissue, and immunohistochemistry to detect the changes of CD4 and CD8 expression in the tumor tissue. Compared with that in the model group, the tumor volume of mice in each treatment group significantly reduced. The body weights of mice in APS + 5-FU group and 5-FU group significantly reduced from day 4 to day 7 of treatment. In addition, the APS + 5-FU group and 5-FU group showed significantly decreased spleen index. The protein and mRNA levels of IDO1 were significantly down-regulated in the APS, APS + 5-FU, and APS + low-dose 5-FU groups. The drug interventions significantly increased the Trp content and decreased the Kyn content. The APS + 5-FU group showed significantly reduced infiltration of CD4~+ T lymphocytes and increased infiltration of CD8~+ T lymphocytes. APS inhibited the expression of IDO1 in the colon tumor microenvironment to increase CD8~+ T lymphocyte infiltration, and the combination of APS with 5-FU demonstrated better effect.
Mice ; Animals ; Tumor Microenvironment ; Colonic Neoplasms/genetics* ; Fluorouracil/pharmacology* ; Polysaccharides/pharmacology* ; CD8-Positive T-Lymphocytes/metabolism* ; RNA, Messenger/metabolism*