ObjectiveTo investigate the effect of Tougu Xiaotong capsules （TGXTC） on the regulation of chondrocyte PANoptosis， delay of chondrocyte degeneration， and improvement of the symptoms in knee osteoarthritis （KOA）. MethodsIn vivo experiments： 50 male C57BL/6 mice were randomly assigned into five groups （n=10 per group）： sham operation group， model group， low-dose TGXTC group （7.2 g·kg^-1）， high-dose TGXTC group （14.4 g·kg^-1）， and diclofenac sodium group （0.05 g·kg^-1）. Except for the sham group， KOA models were established in all other groups using the modified Hulth method. Following successful model induction， the TGXTC groups received daily oral gavage of 7.2 or 14.4 g·kg^-1 for 6 weeks， while the diclofenac sodium group received 0.05 g·kg^-1 solution daily over the same duration. Model evaluation was performed using Lequesne MG score； micro-computed tomography （micro-CT） was used to scan the knee， hematoxylin-eosin （HE） staining and safranin O-fast green staining were used to observe the morphology of cartilage， transmission electron microscopy （TEM） was used to determine ultrastructural changes of PANoptosis. Multiple immunofluorescence （IF） co-localization assays was performed to detect the co-localization of cleaved Caspase-3， receptor-interacting protein 3 （RlPK3）， and the N-terminal domain of gasdermin D （GSDMD-N） in cartilage tissue， while western blot was employed to detect the expression levels of cleaved Caspase-3， RIPK3， and GSDMD-N. In vitro experiments： The knee cartilages of 4-week-old SD rats were isolated， and a chondrocyte in vitro culture system was established through mechanical digestion with 0.2% type Ⅱ collagenase. Second-generation chondrocytes were divided into three groups： the control group， the model group （pretreated with 10 mg·L^-1 lipopolysaccharide （LPS） for 24 h followed by treatment with 1 μmol·L^-1 nigericin for 4 h）， and the TGXTC treatment group （pretreated with 10 mg·L^-1 LPS for 24 h， followed by exposure to 1 μmol·L^-1 nigericin for 4 h and subsequently treated with 100 mg·L^-1 TGXTC for an additional 24 h）. The levels of reactive oxygen species （ROS）， apoptosis， necroptosis， and pyroptosis of chondrocytes were evaluated via fluorescence microscopy following staining with ROS detection， AO/EB and YO-PRO-1/PI staining kits. Transmission electron microscopy was utilized to investigate the ultrastructural changes associated with PANoptosis in cartilage tissue of KOA mice. Inflammatory cytokine levels （IL-1β and IL-18） were measured using ELISA. Western blot was conducted to assess protein expressions related to PANoptosis， including cleaved Caspase-3， cleaved Caspase-8， RIPK3， ZBP1， GSDMD-N， and NLRP3. ResultsCompared with the sham group， the Lequesne MG scores were significantly up-regulated（P<0.01） in the model group， and the pathological changes of cartilage were significantly， with joint spaces narrower， osteophyte formation increased， secere abrasion of cartilage surface. Ultrastructural analysis revealed pronounced chondrocyte apoptosis， necroptosis， and pyroptosis， along with markedly elevated expression of cleaved Caspase-3， RlPK3， and GSDMD-N in cartilage tissue （P<0.01）. In addition， The mean fluorescence intensities of ROS， orange-red fluorescence in AO/EB staining， green fluorescence and red fluorescence in YO-PRO-1/PI staining were increased of chondrocyte in the model group （P<0.01） . The levels of inflammatory factors IL-1β and IL-18 in the supernatant were increased （P<0.01）. The expression of PANoptosis related proteins （cleaved Caspase-3， cleaved Caspase-8， RIPK3， ZBP1， GSDMD-N， and NLRP3） were also significantly upregulated（P<0.05）. Compared to the model group， the TGXTC group demonstrated a significant improvement in various parameters of mice. These included a reduction in the Lequesne MG score， an increase in joint space， a decrease in osteophyte formation， diminished cartilage damage， reduced release of ROS， and alleviation of apoptotic， necroptotic， and pyroptotic processes in chondrocytes. Additionally， mitochondrial swelling and endoplasmic reticulum dilation were also mitigated. The levels of ROS as well as IL-1β and IL-18 were significantly decreased （P<0.05）. Furthermore， the expression levels of proteins associated with PANoptosis in cartilage tissue showed marked reductions （P<0.05）. Similar results were observed in chondrocytes： cleaved Caspase-3， cleaved Caspase-8， RIPK3， ZBP1， GSDMD-N， and NLRP3 exhibited significant decreases as well （P<0.05）. ConclusionTGXTC may mitigate chondrocytes degeneration and alleviate KOA symptoms by reducing oxidative stress and suppressing the activation of PANoptosis pathways.

ObjectiveTo investigate the effect of Tougu Xiaotong capsules （TGXTC） on the regulation of chondrocyte PANoptosis， delay of chondrocyte degeneration， and improvement of the symptoms in knee osteoarthritis （KOA）. MethodsIn vivo experiments： 50 male C57BL/6 mice were randomly assigned into five groups （n=10 per group）： sham operation group， model group， low-dose TGXTC group （7.2 g·kg^-1）， high-dose TGXTC group （14.4 g·kg^-1）， and diclofenac sodium group （0.05 g·kg^-1）. Except for the sham group， KOA models were established in all other groups using the modified Hulth method. Following successful model induction， the TGXTC groups received daily oral gavage of 7.2 or 14.4 g·kg^-1 for 6 weeks， while the diclofenac sodium group received 0.05 g·kg^-1 solution daily over the same duration. Model evaluation was performed using Lequesne MG score； micro-computed tomography （micro-CT） was used to scan the knee， hematoxylin-eosin （HE） staining and safranin O-fast green staining were used to observe the morphology of cartilage， transmission electron microscopy （TEM） was used to determine ultrastructural changes of PANoptosis. Multiple immunofluorescence （IF） co-localization assays was performed to detect the co-localization of cleaved Caspase-3， receptor-interacting protein 3 （RlPK3）， and the N-terminal domain of gasdermin D （GSDMD-N） in cartilage tissue， while western blot was employed to detect the expression levels of cleaved Caspase-3， RIPK3， and GSDMD-N. In vitro experiments： The knee cartilages of 4-week-old SD rats were isolated， and a chondrocyte in vitro culture system was established through mechanical digestion with 0.2% type Ⅱ collagenase. Second-generation chondrocytes were divided into three groups： the control group， the model group （pretreated with 10 mg·L^-1 lipopolysaccharide （LPS） for 24 h followed by treatment with 1 μmol·L^-1 nigericin for 4 h）， and the TGXTC treatment group （pretreated with 10 mg·L^-1 LPS for 24 h， followed by exposure to 1 μmol·L^-1 nigericin for 4 h and subsequently treated with 100 mg·L^-1 TGXTC for an additional 24 h）. The levels of reactive oxygen species （ROS）， apoptosis， necroptosis， and pyroptosis of chondrocytes were evaluated via fluorescence microscopy following staining with ROS detection， AO/EB and YO-PRO-1/PI staining kits. Transmission electron microscopy was utilized to investigate the ultrastructural changes associated with PANoptosis in cartilage tissue of KOA mice. Inflammatory cytokine levels （IL-1β and IL-18） were measured using ELISA. Western blot was conducted to assess protein expressions related to PANoptosis， including cleaved Caspase-3， cleaved Caspase-8， RIPK3， ZBP1， GSDMD-N， and NLRP3. ResultsCompared with the sham group， the Lequesne MG scores were significantly up-regulated（P<0.01） in the model group， and the pathological changes of cartilage were significantly， with joint spaces narrower， osteophyte formation increased， secere abrasion of cartilage surface. Ultrastructural analysis revealed pronounced chondrocyte apoptosis， necroptosis， and pyroptosis， along with markedly elevated expression of cleaved Caspase-3， RlPK3， and GSDMD-N in cartilage tissue （P<0.01）. In addition， The mean fluorescence intensities of ROS， orange-red fluorescence in AO/EB staining， green fluorescence and red fluorescence in YO-PRO-1/PI staining were increased of chondrocyte in the model group （P<0.01） . The levels of inflammatory factors IL-1β and IL-18 in the supernatant were increased （P<0.01）. The expression of PANoptosis related proteins （cleaved Caspase-3， cleaved Caspase-8， RIPK3， ZBP1， GSDMD-N， and NLRP3） were also significantly upregulated（P<0.05）. Compared to the model group， the TGXTC group demonstrated a significant improvement in various parameters of mice. These included a reduction in the Lequesne MG score， an increase in joint space， a decrease in osteophyte formation， diminished cartilage damage， reduced release of ROS， and alleviation of apoptotic， necroptotic， and pyroptotic processes in chondrocytes. Additionally， mitochondrial swelling and endoplasmic reticulum dilation were also mitigated. The levels of ROS as well as IL-1β and IL-18 were significantly decreased （P<0.05）. Furthermore， the expression levels of proteins associated with PANoptosis in cartilage tissue showed marked reductions （P<0.05）. Similar results were observed in chondrocytes： cleaved Caspase-3， cleaved Caspase-8， RIPK3， ZBP1， GSDMD-N， and NLRP3 exhibited significant decreases as well （P<0.05）. ConclusionTGXTC may mitigate chondrocytes degeneration and alleviate KOA symptoms by reducing oxidative stress and suppressing the activation of PANoptosis pathways.

Objective: To investigate the mechanism of action of Prim-O-glucosylcimifugin (POG) to improve cholesterol metabolism in osteoarthritic (OA) chondrocytes based on the long noncoding RNA nuclear-enriched transcript 1 (lncRNA NEAT1)/microRNA-128-3p (miR-128-3p) pathway. Methods: For in vivo experiments, 60 mice were divided into the normal, sham operation, model, and POG groups using the random number table method, with 15 mice per group. The osteoarthritis mouse model was constructed using the modified Hulth method in the model and POG groups. Mice in the POG group were administered 30 mg/(kg·d)POG by gavage. The other groups were administered an equal amount of normal saline for 8 weeks. The cartilage tissue structure of mice in each group was observed using hematoxylin and eosin staining. Real-time PCR was used to detect changes in the lncRNA NEAT1 and miR-128-3p mRNA expression levels in the cartilage tissues of mice. Western blotting was used to detect the protein expressions of ATP-binding cassette transporter A1 (ABCA1), liver X receptor β (LXRβ), matrix metalloprotein-3 (MMP-3), and B-lymphoblastoma-2-associated X protein (Bax) in articular cartilage of mice. An enzyme-linked immunosorbent assay was used to measure the tumor necrosis factor-α (TNF-α) content in the synovial fluid of mice. A biochemical microplate assay was used to measure the total cholesterol level in the synovial fluid of mice. The in vitro experiments were divided into the negative control, interleukin-1β(IL-1β), IL-1β+ POG, IL-1β+ oe-lncRNA NEAT1, IL-1β+ oe-lncRNA NEAT1 + POG, IL-1β + miR-128-3p inhibition, and IL-1β+ miR-128-3p inhibition+ POG groups. An OA model was established by inducing chondrocytes with IL-1β for 24 h, and 90 mg/L of POG and miR-128-3p inhibitor(50 nmol/L) were administered for 48 h as an intervention. lncRNA NEAT1 expression in chondrocytes was detected using fluorescence in situ hybridization. A dual luciferase assay was used to detect the targeting relationship between lncRNA NEAT1 and miR-128-3p. Lentiviral plasmids overexpressing lncRNA NEAT1 were used to transfect mouse chondrocytes. Real-time PCR was used to detect the effect of lncRNA NEAT1 overexpression on the mRNA level of miR-128-3p in chondrocytes. Western blotting was used to detect ABCA1, LXRβ, MMP-3, and Bax protein expression in chondrocytes after lncRNA NEAT1 overexpression and miR-128-3p inhibition. Results: POG significantly reduced OA cartilage tissue damage. Compared with the model group, the lncRNA NEAT1 mRNA level decreased, whereas the miR-128-3p mRNA level increased in the cartilage tissue of the POG group (P<0.05). Compared with the model group, ABCA1 and LXRβ protein expression increased in the POG group, whereas MMP-3 and Bax protein expression decreased (P<0.05). The TNF-α levels decreased in the POG group compared to the model group (P<0.05). Compared with the model group, the total cholesterol level in the synovial fluid of the joint of mice in the POG group decreased (P<0.05). The mean fluorescence intensity of lncRNA NEAT1 in the IL-1β+ POG group decreased compared with the IL-1β group (P<0.05). The relative luciferase activity in the miR-128-3p mimics group bound to the lncRNA NEAT1-WT plasmid decreased compared with the miR-128-3p negative control group (P<0.05). The lncRNA NEAT1 mRNA levels decreased, whereas the miR-128-3p mRNA levels increased in the IL-1β+ oe-lncRNA NEAT1 + POG group compared with the IL-1β+ oe-lncRNA NEAT1 group (P<0.05). Compared with the IL-1β+ POG group, ABCA1 and LXRβ protein expression decreased, whereas MMP-3 and Bax protein expression increased (P<0.05). Conclusion POG mediates lncRNA NEAT1/miR-128-3p to improve cholesterol metabolism in OA chondrocytes.

Objective To investigate the levels of vascular endothelial growth factor(VEGF)and vas-cular endothelial growth factor receptor 2(VEGFR2)in serum and urine of the children patients with primary nephrotic syndrome(PNS),and to conduct the correlation analysis on the experimental indexes of 24 h urine protein quantitation,blood albumin(ALB),total cholesterol(TC),blood urea nitrogen(BUN)and creatinine(Scr).Methods Thirty children patients with PNS without hormone treatment admitted and treated in this hospital from January 2020 to December 2023 were selected as the study subjects and divided into the hormone sensitive nephrotic syndrome(SSNS)group(n=12)and steroid dependent nephrotic syndrome(SDNS)group(n=18)according to the cortical hormone treatment effect.Twenty healthy children with matched age and sex visiting in the pediatric department during the same period served as the control group.The differ-ences of VEGF and VEGFR2 levels in blood and urine were compared among 3 groups and their correlation with the experimental indexes was analyzed.Results Compared with the control group,the blood VEGF lev-el,urine VEGF and BUN levels in the SDNS group were increased,blood VEGFR2 and urine VEGFR2,24 h urine protein quantitation and TC level in the SSNS group and SDNS group were increased,the ALB level was decreased,and the differences were statistically significant(P<0.05).The Pearson correlation analysis re-sults showed that blood VEGF level was positively correlated with the 24 h urine protein quantitation and TC level,and negatively correlated with the ALB level(P<0.05);the urine VEGF level was positively correlated with 24 urine protein quantitation and TC level,and negatively correlated with the Scr level(P<0.05);blood VEGF2 level was positively correlated with the 24 h urine protein quantitation and the TC level,and negative-ly correlated with the ALB level(P<0.05);the urine VEGF2 level was positively correlated with the 24 h u-rine protein quantitation and TC level,and negatively correlated with the ALB level(P<0.05).Conclusion The blood and urine VEGF and VEGFR2 may serve as the novel indicators for the early diagnosis,prediction and hormone sensitivity evaluation in the children patients with PNS.

Objective To investigate the role and underlying mechanism of atorvastatin on hyper-glycemia induced hemorrhagic transformation(HT)in a mouse model of cerebral ischemia.Meth-ods A total of 36 SPF-grade male C57BL/6 mice were randomly divided into sham operation group,HT model group and atorvastatin group,with 12 mice in each group.HE staining was used to observe cerebral hemorrhage,immunofluorescent staining was employed to detect the integrity of blood-brain barrier,and Western blotting was applied to measure the protein expression of IgG,ZO-1,occludin,claduin5,MMP-2 and-9 in ischemic penumbra brain tissues.Results Com-pared with sham operation group,the neurological deficit score,mortality rate,HT incidence,HT grading score,IgG fluorescence intensity,and protein levels of IgG,MMP-2 and-9 were signifi-cantly increased,while the protein levels of ZO-1,occludin and claudin5 were obviously decreased in the HT model group(P＜0.01).Atorvastatin treatment resulted in significantly lower neuro-logical deficit score(2.73±1.19 vs 3.91±0.94),mortality rate(16.7％vs 41.6％),HT incidence(58.3％vs 91.6％),HT grading score(1.00±1.04 vs 2.58±1.13),IgG fluorescence intensity(504.30±105.52 a.u vs 859.91±153.28 a.u),and protein levels of IgG(4.55±1.40 vs 12.06± 3.73),MMP-2(1.87±0.41 vs 2.95±0.68)and-9(1.47±0.24 vs 2.12±0.23)(P＜0.05,P＜0.01),and increased protein levels of ZO-1(1.55±0.20 vs 0.53±0.10),occludin(0.92±0.11 vs 0.35±0.07)and claudin5(0.58±0.04 vs 0.30±0.05)(P＜0.01)when compared with the HT model group.Conclusion Atorvastatin can reduce the permeability of blood-brain barrier by in-hibiting the activation of MMP-2 and MMP-9 and up-regulating the protein levels of ZO-1,occlu-din and claudin5,and thus attenuate hyperglycemia-induced HT.

Objective To establish a genetic monitoring method for laboratory quails.Methods Quail microsatellite loci were searched in the literature,and microsatellite DNA loci suitable for quails were screened by an interspecific transfer method in closely related species,namely chickens and ducks.Quail liver DNA was extracted as a template,and the corresponding loci were screened by PCR amplification and agarose gel electrophoresis.On the basis of amplification of the selected microsatellite loci,the number of alleles,polymorphisms,and microsatellite loci combinations for quail genetic quality detection were selected and detection method were developed.Results We preliminary determined 23 microsatellite loci for genetic monitoring of closed-colony laboratory quails.Conclusions A genetic monitoring method for laboratory quails was preliminary developed.

Objective To investigate the mechanism by which Tougu Xiaotong Capsule(TGXTC)alleviates chondrocyte degeneration in knee osteoarthritis(KOA).Methods Thirty 2-month-old C57BL/6 mouse models of KOA established using the Hulth method were randomized into model group,TGXTC group,and diclofenac sodium group and received treatment with saline,TGXTC(368 mg/kg),and diclofenac sodium(10 mg/kg)by gavage,respectively,with another 10 untreated mice as the blank control group.All interventions were administered 6 times a week for 4 weeks.After the treatments,structural changes in the cartilage tissue were observed with morphological staining,and Nav1.7 mRNA expression and the protein expression levels of Nav1.7,MMP-3,ADAMTS-5,and COX-2 were detected using RT-qPCR and Western blotting.Fluorescence in situ hybridization(FISH)was used to detect Nav1.7 expression in the chondrocytes.In cultured KOA chondrocytes,the effect of TGXTC and lentivirus-mediated Nav1.7 knockdown on MMP-3,MMP-13,ADAMTS-4,ADAMTS-5,and COX-2 protein expressions were assessed with Western blotting.Results In KOA mice treatments with TGXTC and diclofenac sodium both significantly alleviated structural damage of the cartilage layer,reduced Nav1.7 protein and mRNA expressions and lowered the expressions of MMP-3,ADAMTS-5,and COX-2 proteins in the cartilage tissues.FISH results indicated that TGXTC treatment significantly reduced IL-1β-induced Nav1.7 expression in the chondrocytes.In Nav1.7 knockdown experiment,Nav1.7 levels were significantly lower in IL-1β+sh-Nav1.7 group than in IL-1β group,and also lower in IL-1β+TGXTC group than in IL-1β+sh-Nav1.7+TGXTC group.TGXTC treatment significantly inhibited IL-1β-induced elevation of MMP-3,MMP-13,ADAMTS-4,ADAMTS-5 and COX-2 protein expressions in the chondrocytes,but its effects were strongly weakened by Nav1.7 knockdown.Conclusion TGXTC alleviates extracellular matrix metabolic disorder in KOA chondrocytes by regulating Nav1.7,thereby mitigating chondrocyte degeneration in KOA mice.

Diabetic foot(DF),a primary chronic complication of diabetes mellitus,contributes to a major disability and mortality in diabetic patients.DF is diagnosed mainly depending not only on clinical manifestations,signs,and related inspection,but also on recently emerging diagnostic means:biological markers.Inflammatory biomarkers are preferably used for its superiority in DF early diagnosis.In recently years,thanks to advancements of biological technologies,biomarkers such as procalcitonin(PCT),C-reactive protein(CRP),interleukins(ILs),and tumor necrosis factor-alpha(TNF-α)have been comprehensively used in DF diagnosis.Moreover,biomarkers of genomics,proteomics,metabolomics,and metagenomics have been employed as well.In this review,we aim to com-prehensively review the role of serum biomarkers in DF diagnosis and risk stratification,elaborating on the current research status in applying serum biomarkers for DF prevention,diagnosis,and prognosis assessment.

Objective To investigate the mechanism by which Tougu Xiaotong Capsule(TGXTC)alleviates chondrocyte degeneration in knee osteoarthritis(KOA).Methods Thirty 2-month-old C57BL/6 mouse models of KOA established using the Hulth method were randomized into model group,TGXTC group,and diclofenac sodium group and received treatment with saline,TGXTC(368 mg/kg),and diclofenac sodium(10 mg/kg)by gavage,respectively,with another 10 untreated mice as the blank control group.All interventions were administered 6 times a week for 4 weeks.After the treatments,structural changes in the cartilage tissue were observed with morphological staining,and Nav1.7 mRNA expression and the protein expression levels of Nav1.7,MMP-3,ADAMTS-5,and COX-2 were detected using RT-qPCR and Western blotting.Fluorescence in situ hybridization(FISH)was used to detect Nav1.7 expression in the chondrocytes.In cultured KOA chondrocytes,the effect of TGXTC and lentivirus-mediated Nav1.7 knockdown on MMP-3,MMP-13,ADAMTS-4,ADAMTS-5,and COX-2 protein expressions were assessed with Western blotting.Results In KOA mice treatments with TGXTC and diclofenac sodium both significantly alleviated structural damage of the cartilage layer,reduced Nav1.7 protein and mRNA expressions and lowered the expressions of MMP-3,ADAMTS-5,and COX-2 proteins in the cartilage tissues.FISH results indicated that TGXTC treatment significantly reduced IL-1β-induced Nav1.7 expression in the chondrocytes.In Nav1.7 knockdown experiment,Nav1.7 levels were significantly lower in IL-1β+sh-Nav1.7 group than in IL-1β group,and also lower in IL-1β+TGXTC group than in IL-1β+sh-Nav1.7+TGXTC group.TGXTC treatment significantly inhibited IL-1β-induced elevation of MMP-3,MMP-13,ADAMTS-4,ADAMTS-5 and COX-2 protein expressions in the chondrocytes,but its effects were strongly weakened by Nav1.7 knockdown.Conclusion TGXTC alleviates extracellular matrix metabolic disorder in KOA chondrocytes by regulating Nav1.7,thereby mitigating chondrocyte degeneration in KOA mice.

Hazardous environmental factors as well as occupational factors can lead to elevated incidence of diseases including tumors, and specific molecular biomarkers are needed to guide the diagnosis and treatment of diseases. In recent years, ubiquitin-specific protease 14 (USP14) has gradually attracted the attention of researchers. USP14 is widely expressed in various organs of human body and regulates the stability and degradation of important proteins in various signaling pathways. Studies have shown that its abnormal expression is highly correlated with tumors, neurodegenerative diseases, autophagy, immune response, and viral infections, and is involved in the regulation of various classic signaling pathways. It has been shown to play a key role in the development of various human diseases and can be used as a diagnostic and prognostic molecular biomarker and therapeutic target in the development of tumors. This paper reviewed the current status of research on the structure and regulation of USP14 and its function in physiological and pathological processes, with the aim of providing a reference for research on diseases or injuries caused by environmental and occupational factors.