Objective:To explore if there is premature senescence of chondrocytes in patients with Kashin-Beck disease (KBD) and osteoarthritis.Methods:Five knee cartilage samples of KBD, osteoarthritis and control groups were collected, respectively, from the Second Affiliated Hospital of Xi'an Jiaotong University. DNA was then extracted from cartilage samples and DNA methylation was analyzed by Illumina Infinium HumanMethylation450 BeadChip. At the same time, based on genome-wide methylation data, the online DNA methylation aging clock calculator (https://dnamage.genetics.ucla.edu/home) was used to calculate the DNA methylation age (DNAm age) of samples, and the results were compared with their actual ages.Results:In the comparison between KBD group and control group, 1 212 differentially methylated CpG sites were found, including 497 hypermethylated CpG sites and 715 hypomethylated CpG sites, corresponding to 264 hypermethylated genes and 368 hypomethylated genes, respectively. In the comparison between osteoarthritis group and control group, 656 differentially methylated CpG sites were found, including 343 hypermethylated CpG sites and 313 hypomethylated CpG sites, corresponding to 177 hypermethylated genes and 174 hypomethylated genes, respectively. In the above comparison, 367 overlapped CpG sites (corresponding to 182 genes) were found, which were differentially methylated in both KBD and control groups and osteoarthritis and control groups. The results of DNA methylation aging clock showed that the average age acceleration differences between DNAm age and actual age of KBD, osteoarthritis and control groups were 2.549, 0.017, and - 5.364 years, respectively, the DNAm ages of KBD and osteoarthritis groups were greater than the actual ages.Conclusion:The chondrocytes show premature senescence in both KBD and osteoarthritis.

Schizophrenia is a mental illness that can seriously affect a patient's cognitive and social communication. Because schizophrenia is affected by multiple factors, the cause of schizoophrenia is still unclear. Gene transcriptome sequencing (RNA-seq) is a new research method to study the morphological structure, and gene differential expression and function; RNA-Seq can sequence hundreds of thousands to millions of DNA molecules at a time, providing more effective data for biological transcription information. This paper reviews the principles and technical characteristics of RNA-Seq, and application of RNA-Seq in schizophrenia, which provide new ideas for schizophrenia research.

Objective:To investigate the gene transcription level changes in the amygdala of social isolation rearing models of schizophrenia to determine the pathogenic genes and their related pathways of schizophrenia.Methods:A total of 29 3-week-old SPF C57BL/6J male mice were randomly divided into control group ( n=16) and model group ( n=13); 4 mice were raised in each transparent mouse cage in the control group, and 1 mouse was raised in each transparent mouse cage in the model group; mice in each cage could see their surrounding mice but could not touch each other. Mice in both groups were fed for 4 weeks and then subjected to open field experiment, pre-pulse inhibition experiment and new object recognition experiment within one week. After the experiment, mice were sacrificed by spinal dislocation, and the amygdala was taken for transcriptome sequencing. The topGO software was used for gene ontology (GO) functional enrichment analysis of differentially expressed genes (DEGs), and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis was performed using KEGG database. Results:(1) Animal experiment: compared with the control group, the model group had significantly increased movement distance in the open field experiment ([1 239.20±106.35] m vs. [1 845.53±143.65] m, t=3.464, P=0.002), significantly decreased activity time in the central region 5 min before experiment ([13.15±1.41] s vs. [8.47±1.19]) s, t=2.464, P=0.020). Compared with the control group, the model group had significantly lower percentage of deficient prepulse inhibition (PPI) of 78 dB ([22.28±1.53] % vs. [14.59±2.75] %, t=2.629, P=0.013), and deficient PPI of 88 dB ([32.83±3.39] % vs. [18.44±3.07] %, t=3.081, P=0.005). Compared with the control group, the model group had significantly decreased ratio of time exploring new objects/time exploring former objects ([80.5±2.2]% vs. [71.0±3.6]%, t=2.356, P=0.026). (2) Bioinformatics analysis: a total of 96 DEGs were found, of which 42 were with up-regulated expressions ( Th, Crlf1, etc.), and 54 were with down-regulated expressions ( Prkcd, etc.). Th and Crlf1 were positively correlated ( r=0.940, P=0.018). GO enrichment results suggested that DEGs were enriched in projection function of plasma membrane boundary cells, neuronal differentiation, and cell apoptosis. KEGG enrichment results suggested that DEGs were enriched in WNT signaling pathway, apoptosis pathway and tyrosine metabolism pathway. Protein network interaction analysis suggested that Wnt6, Tcf712, Pitx2, Tcf7 and Cd4 were key proteins. Conclusion:DEGs such as Th, Prkcd, Lrrc74b, Fadd, Wnt6, Ror2, Notum, and Tcf7l2, and their related signaling pathways may be related to schizophrenia in the amygdala of social isolation rearing mice.

Objective To detect the metabolites of Fusarium poae strain isolated from self-produced grain in a new-onset patient with Kaschin-Beck disease,and to provide a scientific basis for studying the relationship between mycotoxins and Kaschin-Beck disease.Methods The strain of Fusarium poae was isolated from the self-produced spike wheat in a family of patient with new-onset Kaschin-Beck disease in Guide County,Qinghai Province in 2016.The isolated Fusarium poae strain was inoculated into potato dextrose agar (PDA) medium for enrichment culture.The metabolites of the fermentation were separated and purified,and the mycotoxins in the metabolite were detected by liquid chromatography-mass spectrometry (LC-MS).The full-spectrum identification was carried out by using the network database Metlin.Results The metabolites of this strain were identified by LC-MS and Metlin full spectrum,and mycotoxins such as trichothecenes,Fumonisin and Zearalenone (ZEA) were detected.A total of 1 601 material characteristic peaks were detected in positive ion mode,the most abundant mycotoxins were Fumonisin FP3,A2,FP1;and a total of 1 000 material characteristic peaks were detected in the negative ion mode,the most abundant were ZEA and monoacetoxyscirpenol.Conclusion The isolated metabolites of Fusarium poae strain mainly contain the three major classes of mycotoxins including trichothecenes,Fumonisins and ZEA.

Hepatocellular carcinoma is still a severe threat to people′s health of China. Most patients have advanced disease at the time of first diagnosis and lose the opportunity of radical surgery. In the past, the traditional medical drug treatment and radiotherapy are ineffective, which make the treatment of hepatocellular carcinoma into a bottleneck. With the emergence of target therapy represented as tyrosine kinase inhibitors and immunotherapy represented by programmed death-1 antibody and programmed death-ligand 1 antibody, the treatment of hepatocellular carcinoma has entered a new era and patients with advanced hepatocellular carcinoma have seen a new hope. The systemic therapy represented by target therapy and immunotherapy has not only greatly improved the survival of patients with advanced hepatocellular carcinoma, but also changed the treatment concept of hepatocellular carcinoma from single-drug therapy to combined therapy with multiple means. The treatment of hepatocellular carcinoma has changed from the era of surgery as the king to the era of surgery as the mainstay of whole-process management and comprehensive treatment. The authors review previous studies and their own experience to elaborate on the comprehensive treat-ment strategy for hepatocellular carcinoma based on surgical treatment in the era of targeted therapy and immunotherapy.

Objective:To explore the risk factors associated with the clinical progression of COVID-19 infection in kidney transplant(KT)recipients during the spread of Omicron variant and evaluate the effectiveness of anti-RNA virus agents in blocking the clinical progression of COVID-19 in these recipients.Methods:Retrospective analysis was conducted for the clinical data on COVID-19 infection in 232 KT recipients followed up from December 4, 2022 to January 31, 2023 at Department of Renal Transplantation, Organ Transplantation Center, Beijing Tsinghua Changgung Hospital.Inclusion criteria were age ≥18 years and stable kidney function without renal replacement therapy.The follow-up time was 30 days after COVID-19 infection.Based upon whether or not there was an infection of COVID-19, KT recipients were divided into two groups of infection(181 cases)and non-infection(51 cases). In infection group, recipients were further assigned into two sub-groups of disease progression(n=23)and stable(158 cases)according to whether or not there was a progression to severe disease.Various factors such as gender, age, body mass index(BMI), time after transplantation, underlying diseases(history of hypertension, diabetes mellitus, coronary heart disease & chronic lung disease), smoking history and dosing of anti-RNA virus agents were collected.Pearson χ2 test or Fisher's exact probability method was utilized for examining enumeration data while Mann-Whitney U test for measurement data.Univariate Logistic regression analysis was conducted and variables with P<0.05 were included into multifactorial Logistic regression analysis to identify independent risk factors for clinical progression of COVID-19 infection in KT recipients. Results:Among 232 KT recipients, infection rate of COVID-19 was 78.0%(181/232). The clinical classification was mild(112 cases), moderate(46 cases), severe(21 cases)and critical(n=2 cases). The severe rate was 12.7%(23/181). After infection with COVID-19, the proportion of KT recipients aged ≥65 years progressing from mild/moderate to severe was higher than those aged<65 years[38.5%(5/13)vs 10.7%(18/168)]. The difference was statistically significant( P=0.014); The proportion of diabetic KT recipients progressing from mild/moderate to severe was higher than those without diabetes[19.1%(13/68)vs 8.8%(10/113)]. The difference was statistically significant( P=0.045). Univariate Logistic analysis showed similar results.Age≥65 years( OR=5.21, 95% CI: 1.54-17.64, P=0.008)or diabetes mellitus( OR=2.44, 95% CI: 1.003-5.911, P=0.049)were the risk factors for COVID-19 infection recipients progressing from mild/moderate to severe disease.Multivariate Logistic analysis revealed that age ≥65 years( OR=4.03, 95% CI: 1.14-14.34, P=0.031)was an independent risk factor for COVID-19 infection recipients progressing from mild/moderate to severe.Among 181 cases of COVID-19 mild/medium infected patients, 18 cases received nimativir/ritonavir and 10 cases had azvudine for anti-RNA virus treatment.However, none of them progressed to severe; 153 cases did not use anti-RNA virus drugs and 23 cases(15.0%)progressed to severe disease and the difference was statistically significant( P=0.028). Among 23 severe cases, 14 cases received nirmatrelvir/ritonavir and 2 cases had azivudine for anti-RNA virus treatment.The former did not progress to critical disease while 1 case in the latter progressed to critical illness and death; 1/7 recipients not using anti-RNA virus agents progressed to critical illness and died while another 6 cases did not progress to critical illness. Conclusions:KT recipients aged ≥65 years or diabetes mellitus have a greater risk of progression from mild/moderate to severe disease after COVID-19 infection.Among them, age ≥65 years is an independent risk factor for patients with COVID-19 infection to progress from mild/moderate to severe.Antiviral treatment with nirmatrelvir/ritonavir or azivudine in KT recipients during mild/moderate stage of COVID-19 infection can significantly reduce the rate of severe disease.Treatment with Nirmatrelvir/Ritonavir is still effective in the severe stage.

Objective:To investigate the role of a disintegrin and metalloprotease 12 (ADAM12) gene in chondrocyte injury in patients with Kashin-Beck disease (KBD) and its impact on genes related to insulin-like growth factor binding protein (IGFBP).Methods:Articular cartilage samples were obtained from 5 patients with KBD and 5 control subjects admitted to Honghui Hospital Affiliated to Xi'an Jiaotong University. Chondrocytes were extracted and cultured in vitro. Quantitative real-time PCR (qRT-PCR) and Western blotting were used to detect the expression levels of ADAM12 mRNA and protein in chondrocytes of patients with KBD and control subjects, respectively. Subsequently, ADAM12 gene overexpression was performed using lentivirus in chondrocytes of patients with KBD. MTT assay was used to detect changes in cell viability after ADAM12 gene overexpression, and qRT-PCR was used to detect the mRNA expression levels of chondrocyte differentiation related genes SRY-box transcription factor 9 (SOX9) and type Ⅱ collagen (COLⅡ), apoptosis-related gene B-cell lymphoma/leukaemia-2-associated X protein (BAX), and anabolic related genes IGFBP3 and IGFBP5. Results:The expression levels of ADAM12 mRNA and protein in chondrocytes of patients with KBD (0.57 ± 0.05, 0.81 ± 0.07) were significantly lower than those of control subjects (1.00 ± 0.00, 1.00 ± 0.00), and the differences were statistically significant ( t = - 24.50, - 3.61, P < 0.05). The results of MTT assay showed that the cell viability of chondrocytes in ADAM12 overexpression group (1.09 ± 0.05) was higher than that in empty vector control group (1.00 ± 0.08), and the difference was statistically significant ( t = 4.12, P = 0.031). The results of qRT-PCR showed that compared with empty vector control group, the mRNA expression levels of IGFBP3 (2.35 ± 0.79 vs 0.96 ± 0.25), IGFBP5 (2.13 ± 0.30 vs 0.98 ± 0.34), SOX9 (2.92 ± 0.51 vs 0.94 ± 0.36) and COLⅡ (6.45 ± 2.81 vs 0.87 ± 0.19) in ADAM12 overexpression group were significantly increased, and the differences were statistically significant ( t = 3.19, 5.16, 6.27, 4.10, P < 0.05); while the expression level of BAX mRNA (0.31 ± 0.06 vs 1.02 ± 0.22) was significantly decreased, and the difference was statistically significant ( t = - 11.16, P < 0.001). Conclusion:The ADAM12 gene may have a role in inhibiting apoptosis and promoting differentiation in chondrocyte injury in patients with KBD, and its overexpression can increase expression of IGFBP3 and IGFBP5.

Objective:To investigate the impact of matrix metalloproteinase 13 (MMP13) and low-density lipoprotein receptor-related protein 1 (LRP1) on autophagy of articular chondrocytes in patients with Kashin-Beck disease (KBD).Methods:Human articular cartilage samples obtained from 4 KBD patients and 4 control subjects were collected from Shaanxi Institute for Endemic Disease Prevention and Control, and the expression levels of MMP13 and LRP1 in cartilage tissue were determined using immunohistochemistry (IHC). Chondrocytes were extracted and cultured in vitro, the mRNA and protein expression levels of LRP1 and the autophagy related genes [Beclin 1 (BECN1), microtubule associated protein 1 light chain 3 (LC3)], cartilage injury related genes [MMP13, caspase-3 (CASP3)], chondrocyte differentiation related genes [collagen type Ⅱ alpha 1 chain (COL2A1), and SRY-box transcription factor 9 (SOX9)] were detected by real-time fluorescence quantitative PCR (qRT-PCR) and Western blot (WB), respectively. Chondrocytes from 3 KBD patients were extracted, and MMP13 gene silencing experiment was performed by RNA interference (RNAi) technology, the mRNA and protein expression levels of the above genes were detected by qRT-PCR and WB, respectively. In addition, the antagonist receptor associated protein (RAP) of LRP1 was used to block the LRP1 of human normal chondrocytes (C28/I2 cells), and qRT-PCR and WB were used to detect the mRNA and protein expression levels of LRP1, chondrocyte autophagy, differentiation and cartilage injury related genes, respectively. Results:The IHC results showed that the expression levels of MMP13 (1.67 ± 0.21, 0.59 ± 0.15, 0.51 ± 0.12) in the surface, middle, and deep layers of cartilage tissue of KBD patients were significantly higher than those of control subjects (0.25 ± 0.03, 0.26 ± 0.04, 0.06 ± 0.01), and the differences were statistically significant ( t = - 11.38, P < 0.001; t = - 3.82, - 6.26, P = 0.019, 0.003). The expression levels of LRP1 (0.10 ± 0.02, 0.03 ± 0.01, 0.17 ± 0.03) were significantly lower than those of control subjects (1.63 ± 0.40, 0.44 ± 0.12, 0.34 ± 0.08), and the differences were statistically significant ( t = 6.61, 5.61, 3.64, P = 0.003, 0.005, 0.022). The mRNA and protein expression levels of MMP13, CASP3, SOX9 in chondrocytes of KBD patients were significantly higher than those of control subjects, and the differences were statistically significant ( P < 0.05). The mRNA expression levels of LRP1, LC3, COL2A1 were significantly lower than those of control subjects, and the differences were statistically significant ( P < 0.05). After silencing the MMP13 gene in chondrocytes of KBD patients, there were no significant differences in the mRNA and protein expression levels of LRP1, BECN1, LC3, CASP3, COL2A1, and SOX9 ( P > 0.05). After blocking LRP1 with RAP, the protein expression levels of LRP1, BECN1, LC3, MMP13, COL2A1 and SOX9 in chondrocytes were significantly lower than those in control group ( P < 0.05). Conclusions:There is no direct correlation between MMP13 and abnormal autophagy of articular chondrocytes in KBD patients. After blocking LRP1, the expression of the autophagy related genes BECN1 and LC3 in chondrocytes is decreased.

Objective Aiming at solving the problems of pain on the anteromedial tibia, tibial component loosening and osteoarthritis progression after unicompartmental knee arthroplasty (UKA), the influence of different geometric shapes of tibial component pegs on stress distributions in tibia was analyzed by finite element method. Methods The finite element models with UKA were established and validated. Geometric shapes of tibial component were designed. Under the same loading condition, the tibial components with double-peg, single-keel, double-keel and cross-star were studied for finite element analysis and compared with intact model, so as to evaluate the influence of tibial component with different shapes on stresses of cortical bone in anteromedial tibia, cancellous bone under tibial component and cartilage in contralateral tibia. Results Compared with the intact model, the peak stress of cortical bone in anteromedial tibia with double-peg, single-keel, double-keel and cross-star tibial components increased by 56.1%, 55.9%, 54.5% and 68-4%, respectively. The peak stress of cancellous bone under tibial component with single-keel and double-keels decreased by 8.1% and 15.6% respectively, while the peak stress of cancellous bone under tibial component with double-peg and cross-star increased by 67-9% and 121-5%, which were higher than the fatigue yield stress of cancellous bone. The peak stress of cartilage in contralateral tibia with double-peg, single-keel, double-keel and cross-star tibial components decreased by 42.1%, 26.6%, 24.2% and 28.5%, respectively. ConclusionsThe load distribution of the medial and lateral tibia changed after UKA operation, and a greater load was observed on the replacement side. Single-keel and double-keel tibial components were more effective in reducing stresses on cortical bone in anteromedial tibia and cancellous bone, while the stress distribution in tibia with single-keel tibial component was closer to that of the intact tibia. The research findings can provide theoretical references for designing single-keel tibial component of unicompartmental knee prosthesis which conforms to better mechanical properties of the knee joint.