BACKGROUND:We have successfully established an animal model of small ear pig in southern Yunnan province with internal tension relieving technique combined with autologous Achilles tendon for anterior cruciate ligament reconstruction,and verified the stability and reliability of the model.However,whether internal tension relieving technique can promote the ligamentalization process of autologous Achilles tendon graft has not been studied. OBJECTIVE:To investigate the differences in the process of ligamentalization between conventional reconstruction and internal reduction reconstruction of the anterior cruciate ligament by gross view,histology and electron microscopy. METHODS:Thirty adult female small ear pigs in southern Yunnan province were selected.Anterior cruciate ligament reconstruction was performed on the left knee joint with the ipsilateral knee Achilles tendon(n=30 in the normal group),and anterior cruciate ligament reconstruction was performed on the right knee joint with the ipsilateral knee Achilles tendon combined with the internal relaxation and enhancement system(n=30 in the relaxation group).The autogenous right forelimb was used as the control group;the anterior cruciate ligament was exposed but not severed or surgically treated.At 12,24,and 48 weeks after surgery,10 animals were sacrificed,respectively.The left and right knee joint specimens were taken for gross morphological observation to evaluate the graft morphology.MAS score was used to evaluate the excellent and good rate of the ligament at each time point.Hematoxylin-eosin staining was used to evaluate the degree of ligament graft vascularization.Collagen fibers and nuclear morphology were observed,and nuclear morphology was scored.Ultrastructural remodeling was evaluated by scanning electron microscopy and transmission electron microscopy. RESULTS AND CONCLUSION:(1)The ligament healing shape of the relaxation group was better at various time points after surgery,and the excellent and good rate of MAS score was higher(P<0.05).Moreover,the relaxation group could obtain higher ligament vascularization score(P<0.05).(2)The arrangement of collagen bundles and fiber bundles in the two groups gradually tended to be orderly,and the transverse fiber connections between collagen gradually increased and thickened,suggesting that the strength and shape degree of the grafts were gradually improved,but the ligament remodeling in the relaxation group was always faster than that in the normal group at various time points after surgery.(3)The diameter,distribution density,and arrangement degree of collagen fibers in the relaxation group were better than those in the normal group at all time points,especially in the comparison of collagen fiber diameter between and within the relaxation group(P<0.05).

AIM: To delineate the specific mutation responsible for retinitis pigmentosa(RP)in a Yi pedigree, and to analyze the correlation of RHO gene mutation with clinical phenotype.METHODS:A comprehensive clinical evaluation was conducted on the proband diagnosed with RP and other familial members, complemented by a thorough ophthalmic examination. Peripheral blood samples were obtained from the proband and familial members, from which genomic DNA was extracte. Subsequent whole exome sequencing(WES)was employed to identify the variant genes in the proband. The identified variant gene was validated through Sanger sequencing, then an in-depth analysis of the mutation genes was carried out using genetic databases to ascertain the pathogenic mutation sites. Furthermore, an exhaustive analysis was performed to delineate the genotype and phenotype characteristics.RESULTS:The RP pedigree encompasses 5 generations with 42 members, including 19 males and 23 females. A total of 13 cases of RP were identified, consisting of 4 males and 9 females, which conforms to the autosomal dominant inheritance pattern. The clinical features of this family include an early onset age, rapid progression, and a more severe condition. The patients were found to have night blindness around 6 years old, representing the earliest reported case of night blindness in RP families. The retina was manifested by progressive osteocytoid pigmentation of the fundus, a reduced visual field, and significantly decreased or even vanished a and b amplitudes of ERG. The combined results of WES and Sanger sequencing indicated that the proband had a heterozygous missense mutation of the RHO gene c.1040C>T:p.P347L, where the 1 040 base C of cDNA was replaced by T, causing codon 347 to encode leucine instead of proline. Interestingly, this mutation has not been reported in the Chinese population.CONCLUSION:This study confirmed that the mutant gene of RP in a Yi nationality pedigree was RHO(c.1040C>T). This variant leads to the change of codon 347 from encoding proline to encoding leucine, resulting in a severe clinical phenotype among family members. This study provides a certain molecular, clinical, and genetic basis for genetic counseling and gene diagnosis of RHO.

AIM: To delineate the specific mutation responsible for retinitis pigmentosa(RP)in a Yi pedigree, and to analyze the correlation of RHO gene mutation with clinical phenotype.METHODS:A comprehensive clinical evaluation was conducted on the proband diagnosed with RP and other familial members, complemented by a thorough ophthalmic examination. Peripheral blood samples were obtained from the proband and familial members, from which genomic DNA was extracte. Subsequent whole exome sequencing(WES)was employed to identify the variant genes in the proband. The identified variant gene was validated through Sanger sequencing, then an in-depth analysis of the mutation genes was carried out using genetic databases to ascertain the pathogenic mutation sites. Furthermore, an exhaustive analysis was performed to delineate the genotype and phenotype characteristics.RESULTS:The RP pedigree encompasses 5 generations with 42 members, including 19 males and 23 females. A total of 13 cases of RP were identified, consisting of 4 males and 9 females, which conforms to the autosomal dominant inheritance pattern. The clinical features of this family include an early onset age, rapid progression, and a more severe condition. The patients were found to have night blindness around 6 years old, representing the earliest reported case of night blindness in RP families. The retina was manifested by progressive osteocytoid pigmentation of the fundus, a reduced visual field, and significantly decreased or even vanished a and b amplitudes of ERG. The combined results of WES and Sanger sequencing indicated that the proband had a heterozygous missense mutation of the RHO gene c.1040C>T:p.P347L, where the 1 040 base C of cDNA was replaced by T, causing codon 347 to encode leucine instead of proline. Interestingly, this mutation has not been reported in the Chinese population.CONCLUSION:This study confirmed that the mutant gene of RP in a Yi nationality pedigree was RHO(c.1040C>T). This variant leads to the change of codon 347 from encoding proline to encoding leucine, resulting in a severe clinical phenotype among family members. This study provides a certain molecular, clinical, and genetic basis for genetic counseling and gene diagnosis of RHO.

Objective To analyze the status quo of cognitive frailty (CF) in community elderly patients with chronic obstructive pulmonary disease (COPD) and its correlation with sleep quality, anxiety and depression. Methods Elderly patients with COPD receiving health management in the center were selected from July 2023 to June 2024. The general data of patients were collected and Mini-Mental State Examination (MMSE), Fried Frailty Phenotype (FP), Pittsburgh Sleep Quality Index (PSQI) and Hospital Anxiety and Depression Scale (HADS) were used for investigation, and the above scores were analyzed. All patients were divided into CF group (n=129) and non-CF group (n=319) according to MMSE score and FP score. Univariate and multivariate logistic analyses were used to analyze the influencing factors of CF in elderly COPD patients. Results Pearson correlation analysis showed that MMSE score was significantly negatively correlated with PSQI score and HADS score in elderly COPD patients (P<0.05), and FP score was significantly positively correlated with PSQI score and HADS score (P<0.05). After logistic regression analysis, it was found that education level, marital status and sleep time were protective factors of CF in elderly COPD patients (P<0.05), and PSQI score and HADS score were risk factors of CF in elderly patients with COPD (P<0.05). Conclusion CF in community elderly COPD patients is related to sleep quality, sleep duration and anxiety and depression. It is necessary to take clinical measures to improve the sleep quality and psychological status, so as to avoid or slow down the occurrence of CF.

Objective:To investigate the relationship between the outcome of skin injury and urinary arsenic methylation metabolism levels in people exposed to arsenic through drinking water.Methods:Using cluster sampling method, permanent residents from drinking-water-borne endemic arsenic poisoning areas in Bayannur City, Inner Mongolia Autonomous Region were selected as survey subjects in 2004 (before water improvement). In 2017 (after water improvement), 74 survey subjects from 2004 were tracked and followed up. Urine samples were collected from survey subjects and high-performance liquid chromatography inductively coupled plasma mass spectrometry was used to detect the levels of arsenic methylation metabolites in urine. According to the "Diagnosis of Endemic Arsenic Poisoning" (WS/T 211-2015), the clinical grading (normal, suspicious, mild, moderate and severe) of skin injury of the survey subjects and the outcome of 2017 (improved, unchanged, aggravated) were assessed. A database was established and SPSS 25.0 software was used for statistical analysis.Results:The clinical grading ratios of skin injuries among survey subjects in 2004 and 2017 were compared, the differences were statistically significant (normal, suspicious, mild, moderate and severe: 38, 18, 4, 14 cases in 2004 and 27, 31, 3, 13 cases in 2017, χ 2 = 53.02, P < 0.001). Compared with 2004, in 2017, the levels of total arsenic (tAs), inorganic arsenic (iAs), monomethylarsenic (MMA), dimethylarsenic (DMA), percentage of inorganic arsenic (iAs%), and ratio of monomethylarsenic to dimethylarsenic (MMA/DMA) in the urine of survey subjects were low, and the differences were statistically significant ( Z = - 8.24, - 9.07, - 7.81, - 8.04, - 8.24, - 3.56, P < 0.001). The levels of dimethylarsenic percentage (DMA%), monomethylation rate (PMI) and dimethylation rate (SMI) were higher, and the differences were statistically significant ( Z = - 6.39, - 8.24, - 3.52, P < 0.001). In 2004, patients with different clinical grading of skin injuries had different outcomes in 2017 (χ 2 = 30.80, P < 0.001). There were statistically significant differences in tAs, iAs, MMA and DMA variation in urine among skin injury patients with different outcomes ( H = 10.62, 9.35, 8.80, 9.13, P < 0.05). Conclusions:Improving water can significantly reduce the levels of tAs, iAs, MMA, and DMA in the urine of arsenic exposed individuals. The outcome of skin injury in individuals exposed to arsenic through drinking water is related to the variation of urinary arsenic methylation metabolites tAs, iAs, MMA, and DMA.

In order to meet the market demand for the fast and accurate genetic detection of African swine fever virus(ASFV),a new method for EIS-qPCR detection was established with a rapid,high sensitivity,and pollution prevention.Primers and probes for a duplex qPCR were designed based on the conserved region of ASFV virus p72 gene and the endogenous internal standard(EIS)cytb gene sequence in pigs.An anti-contamination system was established with uracil DNA N-gly-cosylase enzyme in the reaction system.The results showed that the method can finish the rapid qPCR detection of ASFV within 30 min with a minimal detection limit of 4.12 copies/μL.Moreo-ver,the method only detected the ASFV p72 gene,and no amplifications of classical swine fever vi-rus(CSFV),pseudorabies virus(PRV),porcine parvo virus(PPV),porcine reproductive and re-spiratory syndrome virus(PRRSV)and porcine circovirus type 2(PCV2)were observed.Repeti-tive results showed a coefficient of variation below 2％.With strong anti-pollution capacity,the method can effectively eliminate false-positive amplification caused by low-dose aerosol pollution.Detection results of 146 clinical samples showed a 100％consistence with the results of the com-mercial ASFV detection kit.Compared with similar technologies,the EIS-qPCR established in this study was faster,sensitive,and suitable for the rapid diagnosis of ASFV infection in the early stage,which provided the tool for the monitoring and precise prevention and control of African swine fever.

Objective To investigate the mechanism by which lectin-like oxidized low density lipoprotein receptor-1(LOX-1)regulates hyperglycemic-induced myocardial fibroblast(CFs)fibrosis through the glycogen synthase kinase-3β(GSK-3β)/signal transducer and activator of transcription 3(STAT3)pathway.Methods CFs were isolated,cultured and identified.LOX-1 RNAi lentiviral vector was constructed and infected CFs.The experimental groups were as follows:Normal control(NC)group,High glucose(HG)group,LV-LOX-1,LV-Con group,Hypertonic(HPG)group.After LV-LOX-1 and LV-Con were infected with CFs,adding 25 mmol/L glucose to culture CFs for 24 h,they were denoted as HG+LV-LOX-1 group and HG+LV-Con group.Cells in HG+LV-LOX-1 group and HG+LV-Con group were treated with 10 μ mol/L SB216763 and 10 μ mol/L STATTIC for 24 h,respectively,and then they were recorded as HG+LV-LOX-1+SB216763 group,HG+LV-Con+SB216763 group,HG+LV-LOX-1+STATTIC group and HG+LV-Con+STATTIC group.CCK-8 was used to detect the activity of CFs,and the expression levels of mRAN and protein of LOX-1,collagen type I(COL-I),thioredoxin 5(TXNDC5),GSK-3β,STAT3,p-GSK-3β and p-STAT3 were detected by qRT-PCR and Western blot.Results CFs infected with LOX-1 RNAi lentiviral vector were obtained,which showed green under fluorescence microscopy.Compared with HG and HG+LV-Con groups,the mRNA expressions of LOX-1,COL-I and TXNDC5 were decreased in HG+LV-LOX-1 group(P<0.05).Compared with HG+LV-LOX-1 group,mRNA expressions of COL-I and TXNDC5 were decreased in HG+LV-LOX-1+SB216763 and HG+LV-LOX-1+STATTIC groups(P<0.05).Compared with HG and HG+LV-Con groups,p-GSK-3β protein expression was increased in HG+LV-LOX-1 group(P<0.05),while LOX-1,p-STAT3,COL-I,TXNDC5 protein expression was decreased in HG+LV-LOX-1 group(P<0.05).Compared with HG+LV-LOX-1 group,p-GSK-3β protein expression was increased in HG+LV-LOX-1+SB216763 group(P<0.05),while the protein expressions of p-STAT3,COL-I and TXNDC5 were decreased in HG+LV-LOX-1+SB216763 and HG+LV-LOX-1+STATTIC groups(P<0.05).Conclusion LOX-1,GSK-3β,STAT3,TXNDC5,and COL-I are involved in high glucose induced CFs fibrosis.LOX-1 promotes the expression of TXNDC5 and COL-I through GSK-3β/STAT3 pathway,and inhibition of LOX-1 can inhibit high glucose induced CFs fibrosis.

Objective To investigate the molecular mechanism of lectin-like oxidized low-density lipoprotein receptor 1(LOX-1)in the regulation of high glucose induced phagocytosis dysfunction of mouse microglia(BV2 microglia).Methods BV2 cells were cultured in vitro,lentivirus LOX-1RNAi vector(LV-LOX-1)and lentivirusempty vector(LV-Con)were constructed and divided into normal control(NC)group,HG group,LV-LOX-1 group and LV-Con group.After infecting BV2 cells with LV-LOX-1 and LV-Con,the cells were cultured with 25 mmol/L glucose for 24 h,and then divided into HG+LV-LOX-1 group and HG+LV-Con group.After treatment of HG+LV-LOX-1 and HG+LV-Con infected BV2 microglia with 15 μmol/L FH535(β-catenin inhibitor)and AEBSF(ATF6α inhibitor)for 24 h,respectively,they were denoted as HG+LV-LOX-1+FH535 group,HG+LV-Con+FH535 group,HG+LV-LOX-1+AEBSF group,and HG+LV-Con+AEBSF group.Transfection efficiency was determined by fluorescence microscopy,RT-PCR and Western blot.Cell viability was detected b CCK-8.RT-PCR and Western blot were used to detect the mRNA and protein expression of LOX-1,β-catenin,ATF6α and milk fat globular-surface growth factor Ⅷ(MFG-E8)in each group.Results After 72 h of LV-LOX-1 infection,the cells in LV-LOX-1 and LV-Con groups showed a lot of green fluorescence,but not in NC group.Compared with NC group,the mRNA and protein expression of LOX-1 and ATF6α were increased(P<0.05),while the mRNA and protein expression of MFG-E8 and β-catenin decreased in HG group(P<0.05).Compared with HG+LV-Con group,the mRNA and protein expression of LOX-1 and ATF6α were decreased(P<0.05),while the mRNA and protein expression of MFG-E8 and β-catenin increasedin HG+LV-LOX-1 group(P<0.05).Compared with HG+LV-LOX-1 group,the mRNA and protein expressions of MFG-E8 and β-catenin were decreased(P<0.05),and the mRNA and protein expressions of ATF6α and p-β-catenin and p-ATF6α were increased in HG+LV-LOX-1+FH535 group(P<0.05).Compared with HG+LV-LOX-1 group,the mRNA and protein expression were increased(P<0.05),ATF6α mRNA and protein expression and p-ATF6α protein expression were decreased MFG-E8 in HG+LV-LOX-1+AEBSF group(P<0.05).Conclusions LOX-1,MFG-E8,β-catenin and ATF6α are involved in the regulation of phagocytosis of BV2 cells.LOX-1 promotes the phagocytosis dysfunction of BV2 microglia induced by high glucose through β-catenin/ATF6α signaling pathway.

Objective:To explore the relation of depressive and anxiety symptoms to defense mechanism in transgender population.Methods:Totally 451 transgender patients in the sexual and psychological outpatient depart-ment of a hospital were selected.They were assessed with the self-Rating Depression Scale(SDS),Self-Rating Anxiety Scale(SAS)and Defense Mechanism Scale(DSQ).The SDS standard score of ≥53 was classified as having depressive symptoms,and the SAS standard score of ≥50 was classified as having anxiety symptoms.Re-sults:The detection rates of depression and anxiety were 46.8％and 28.8％respectively.Multiple linear regression analysis showed that SDS scores were positively correlated with DSQ scores of projection,conceit,complaint,with-drawal,somatization,control,isolation and identity(β=0.08-0.22),while SDS scores were negatively correlated with DSQ scores of sublimation,depression,omnipotence with incompetence and denial(0=-0.09--0.19).The SAS scores were positively correlated with the DSQ scores of projection,latent manifestation,somatization,control,isolation,identity,and consumption tendency(0=0.09-0.26),while the SAS scores were negatively cor-related with the DSQ scores of sublimation,depression,omnipotence accompanied by incompetence,and denial(β=-0.09--0.15).Conclusion:The proportion of depression and anxiety symptoms detected in the transgender group is higher,which may be related to the use of some defenses.

Objective:To establish an inductively coupled plasma-mass spectrometry(ICP-MS)method for the determination of 24 elements in polysorbate 80,and evaluate the results.Methods:The elements were determined by ICP-MS,statistical analysis was conducted using SPSS 26,and the risk assessment was carried out with reference to ICH Q3D guidelines.Results:The standard curves showed good linear relationship in the corresponding concentration range(r=0.997-0.999).The recoveries were 88.81％-134.64％(RSD＜5％,n=6).None of the elements recommended for evaluation exceeded the control threshold.Conclusion:The method is accurate,reliable and easy-to-operate,and the risk of elemental impurities in this sample is low.