To identify clinical viral diseases characterized by reproductive disorders and abortion,a quadruple qPCR method was established for simultaneous detection of PRV,PPV,PCV2 and AS-FV.Four pairs of specific primers and probes were designed according to the conserved genes of four viruses in the NCBI gene bank.The annealing temperature,primer concentration and probe concentration of the reaction were optimized,and the specificity,sensitivity and repeatability of the method were tested.The results showed that the method could not detect other pathogens except the target ones.The minimum detection limit of PRV,PPV,PCV2 and ASFV was 10 copies.Intra-group and inter-group repeatability tests showed that the coefficient of variation of C,values be-tween different batches was less than 3％,indicating that the method was highly specific,sensitive and stable.Establishment of an efficient and sensitive quadruple qPCR method provides technical reference for the clinical prevention and control of porcine pseudorabies virus disease,porcine circo-virus disease,porcine parvovirus disease and African swine fever.

To establish a TaqMan-based multiplex RT-qPCR method for the identification of Japa-nese encephalitis virus(JEV),Porcine reproductive and respiratory syndrome virus(PRRSV),and Classical swine fever virus(CSFV),this study designed and synthesized three pairs of specific primers and probes based on the conserved sequences of JEV E,PRRSV ORF6,and CSFV E2 a-vailable in the NCBI GenBank.By optimizing the reaction system and protocol,a multiplex RT-qPCR method for detecting these three viruses was developed and applied to the detection of clini-cal samples.The results showed that the established TaqMan multiplex RT-qPCR specifically am-plified the gene fragments of JEV,PRRSV,and CSFV,and did not amplify other non-target genes,indicating good specificity of the method.Intra-assay and inter-assay repeatability tests showed that the coefficient of variation(Cv)values were all below 3％,demonstrating that the method has ex-cellent repeatability.Sensitivity tests revealed that the minimum detectable amount for the recom-binant plasmids of the three viruses was 100 copies/pL.Using the established method,a total of 969 samples,including blood,aborted fetuses,semen,and deceased pigs,from 26 pig farms in Guizhou Province were tested.The detection rates were 34.3％(332/969)for JEV,28.3％(274/969)for PRRSV,and 19.8％(192/969)for CSFV.The co-infection rates were 10.1％(98/969)for JEV and PRRSV,12.1％(117/969)for JEV and CSFV,and 14.6％(141/969)for CSFV and PRRSV.Additionally,the triple co-infection rate of JEV,PRRSV,and CSFV was 7.9％(77/969).These results indicate that the TaqMan multiplex RT-qPCR method developed in this study is ef-fective for detecting these three viruses in pig farms,providing technical support for identifying vi-ral causes of reproductive disorders.

Objective To analyze the risk factors associated with intraoperative parathyroid injury during thyroid cancer surgery and develop a predictive model to evaluate its predictive efficacy.Methods A retrospective study was conducted involving 76 patients who experienced intraoperative parathyroid injury during thyroid cancer surgery between May 2021 and February 2024.These patients were included in the parathyroid injury group.For the control group,76 patients who underwent thyroid cancer surgery without intraoperative parathyroid injury during the same period were selected.Clinical data,complete blood count parameters,and other relevant variables were collected.Univariate analysis was performed to screen for potential risk factors,and Logistic regression analysis was used to identify independent risk factors.A regression predictive model was established,and the model's goodness-of-fit and predictive power were evaluated.Results Among the 76 patients,75 had temporary parathyroid function impairment and 1 had permanent parathyroid function impairment.Univariate and multivariate Logistic regression analyses showed that bilateral lymph node dissection,bilateral total lobectomy,capsule invasion,and combined hashimoto's thyroiditis were independent risk factors(P＜0.05).A predictive regression model was developed based on these factors.The Hosmer-Lemeshow test showed a x2=2.064,P=0.356,indicating good model fit.The receiver operating characteristic(ROC)curve revealed that the model had an area under the curve(AUC)of 0.713,with a 95％confidence interval(CI)of 0.639 to 0.787,suggesting good predictive efficacy.Conclusion Hashimoto's thyroiditis,capsular invasion,bilateral lymph node dissection,and bilateral total lobectomy are significant risk factors for intraoperative parathyroid injury during thyroid cancer surgery.The predictive model constructed based on these factors has good predictive efficacy.

To establish a TaqMan-based multiplex RT-qPCR method for the identification of Japa-nese encephalitis virus(JEV),Porcine reproductive and respiratory syndrome virus(PRRSV),and Classical swine fever virus(CSFV),this study designed and synthesized three pairs of specific primers and probes based on the conserved sequences of JEV E,PRRSV ORF6,and CSFV E2 a-vailable in the NCBI GenBank.By optimizing the reaction system and protocol,a multiplex RT-qPCR method for detecting these three viruses was developed and applied to the detection of clini-cal samples.The results showed that the established TaqMan multiplex RT-qPCR specifically am-plified the gene fragments of JEV,PRRSV,and CSFV,and did not amplify other non-target genes,indicating good specificity of the method.Intra-assay and inter-assay repeatability tests showed that the coefficient of variation(Cv)values were all below 3％,demonstrating that the method has ex-cellent repeatability.Sensitivity tests revealed that the minimum detectable amount for the recom-binant plasmids of the three viruses was 100 copies/pL.Using the established method,a total of 969 samples,including blood,aborted fetuses,semen,and deceased pigs,from 26 pig farms in Guizhou Province were tested.The detection rates were 34.3％(332/969)for JEV,28.3％(274/969)for PRRSV,and 19.8％(192/969)for CSFV.The co-infection rates were 10.1％(98/969)for JEV and PRRSV,12.1％(117/969)for JEV and CSFV,and 14.6％(141/969)for CSFV and PRRSV.Additionally,the triple co-infection rate of JEV,PRRSV,and CSFV was 7.9％(77/969).These results indicate that the TaqMan multiplex RT-qPCR method developed in this study is ef-fective for detecting these three viruses in pig farms,providing technical support for identifying vi-ral causes of reproductive disorders.

Objective To analyze the risk factors associated with intraoperative parathyroid injury during thyroid cancer surgery and develop a predictive model to evaluate its predictive efficacy.Methods A retrospective study was conducted involving 76 patients who experienced intraoperative parathyroid injury during thyroid cancer surgery between May 2021 and February 2024.These patients were included in the parathyroid injury group.For the control group,76 patients who underwent thyroid cancer surgery without intraoperative parathyroid injury during the same period were selected.Clinical data,complete blood count parameters,and other relevant variables were collected.Univariate analysis was performed to screen for potential risk factors,and Logistic regression analysis was used to identify independent risk factors.A regression predictive model was established,and the model's goodness-of-fit and predictive power were evaluated.Results Among the 76 patients,75 had temporary parathyroid function impairment and 1 had permanent parathyroid function impairment.Univariate and multivariate Logistic regression analyses showed that bilateral lymph node dissection,bilateral total lobectomy,capsule invasion,and combined hashimoto's thyroiditis were independent risk factors(P＜0.05).A predictive regression model was developed based on these factors.The Hosmer-Lemeshow test showed a x2=2.064,P=0.356,indicating good model fit.The receiver operating characteristic(ROC)curve revealed that the model had an area under the curve(AUC)of 0.713,with a 95％confidence interval(CI)of 0.639 to 0.787,suggesting good predictive efficacy.Conclusion Hashimoto's thyroiditis,capsular invasion,bilateral lymph node dissection,and bilateral total lobectomy are significant risk factors for intraoperative parathyroid injury during thyroid cancer surgery.The predictive model constructed based on these factors has good predictive efficacy.

To identify clinical viral diseases characterized by reproductive disorders and abortion,a quadruple qPCR method was established for simultaneous detection of PRV,PPV,PCV2 and AS-FV.Four pairs of specific primers and probes were designed according to the conserved genes of four viruses in the NCBI gene bank.The annealing temperature,primer concentration and probe concentration of the reaction were optimized,and the specificity,sensitivity and repeatability of the method were tested.The results showed that the method could not detect other pathogens except the target ones.The minimum detection limit of PRV,PPV,PCV2 and ASFV was 10 copies.Intra-group and inter-group repeatability tests showed that the coefficient of variation of C,values be-tween different batches was less than 3％,indicating that the method was highly specific,sensitive and stable.Establishment of an efficient and sensitive quadruple qPCR method provides technical reference for the clinical prevention and control of porcine pseudorabies virus disease,porcine circo-virus disease,porcine parvovirus disease and African swine fever.

This study aims to investigate the effects of Japanese encephalitis virus(JEV)on TLRs signaling pathway and its regulation of the secretion of inflammatory factors during the infection of testicular interstitial cells,In this study,the mRNA levels of TLR3,TLR7,TLR8,TRIF and MyD88 genes were detected by qPCR after 1 MOI dose of JEV was inoculated into testicular stro-mal cells at different time periods.Western blot assay was used to detect the expression levels of TLR3,TLR7,TRIF and MyD88 protein at 6 h after JEV infection,and ELISA was used to detect the expression levels of IL-1β,IL-6 and TNF-α at different time periods(6,12 and 24 h).The re-sults showed as follows:After 6 h of JEV infection,the mRNA levels of TLR3,TLR7,TRIF and MyD88 genes were significantly up-regulated(P＜0.05),and the mRNA levels of TLR8 genes were down-regulated(P＜0.05).Western blot results showed that the protein expressions of TLR3,TLR7,TRIF and MyD88 were significantly up-regulated when JEV infected testicular stromal cells for 6 h(P＜0.05),which was consistent with the corresponding mRNA transcription levels.There was no significant change in TLR8 protein expression.ELISA results showed that 6 h after JEV infection of testicular stromal cells,IL-6 was significantly increased(P＜0.01),and the expressions of IL-1β and TNF-α were not changed.TLR3,TLR7,TLR8,TRIF and MyD88 were si-lenced by siRNA,and the silenced cells were inoculated with JEV for 6 h,and IL-6 expression lev-els were detected by ELISA.The results showed that silenced TLR3,TLR7,TLR8,TRIF and MyD88 could significantly reduce the increase of IL-6 secretion induced by JEV infection(P＜0.05).These results indicated that JEV could induce the expression of inflammatory factor IL-6 by activating TLR3,TLR7 and TLR8 signaling pathway after infection of testicular stromal cells.This study provides a reference for further elucidating the mechanism of reproductive disorders caused by JEV infection.

Porcine circovirus type 2(PCV2)is the main pathogen causing porcine circovirus related diseases.PCV2 infection in pigs may lead to porcine dermatitis and nephrotic syndrome(PDNS)and weaned piglets multiple system failure syndrome(PMWS),etc.At present,the pathogenic mechanism is not fully understood.PCV2 is a single strand of negative link DNA,which can cause immune suppression in the body and lead to increased secondary susceptibility,which has a syner-gistic effect with various pig diseases and brings major economic losses to the pig industry.Al-though there are commercial vaccines,the prevention of vaccines has certain limitations and there is no effective drug treatment so far,an outbreak will threaten people's life and health and public safety,resulting in significant economic losses.In order to understand the latest progress of PCV2 escape mechanism and prevention and control,this paper summarizes the inhibition of interferon production,regulation of apoptosis,regulation of autophagy,regulation of pyroptosis and inflam-matory response,evasion of adaptive immune response,and prevention and control of PCV2,in or-der to provide new theoretical ideas for the research and prevention and control of PCV2.

Objective:To differentiate human embryonic stem cells ( hESCs ) into keratinocytes ( K-hESCs) and analyse the expression characteristics of biomarkers of K-hESCs.Methods: The hESCs of line H9 were seeded on matrigel in mTeSR1 medium.The hESCs were directly differentiated into kerati-nocytes in epithelial differentiation medium with bone morphogenetic protein 4, retinoic acid and N2 sup-plement.The karyotype of K-hESCs was analyzed, comparing the gene expression differences of K-hESCs with human gingival epithelial cells (HGECs), human immortalized oral epithelial cells (HIOECs) and HaCaT by Real-time PCR.Molecular characteristics of the cell differentiation were observed throughout the process by immunocytochemical techniques.Results:H9-hESCs were successfully differentiated into the cells that exhibited characteristics of keratinocytes in epithelial differentiation medium.The karyotype of K-hESCs was 46, XX; and the keratinocyte gene p63 expression in K-hESCs was significantly lower than that in HaCaT ( P<0.05) , but there was no significant difference of p63 expression in K-hESCs, comparing with that in HGECs and HIOECs ( P >0.05 ) .Conclusion: H9-hESCs could be directly differentiated into K-hESCs.The gene expression of K-hESCs was similar to that of epithelial cells in the early stage of monolayer cells differentiation with high proliferative activity.

To understand the mechanical properties of aortic artery of atheroselerosis (AS), the aortic artery of rat with AS was studied by mechanical test. Wistar rats were used for establishing the model. The mechanical measurements of opening angle in zero-stress state and the vessel loading test were conducted on the isolated aortic arteries of AS rats. Data on the stress-strain of aortic artery were obtained. Determination of percentage of collagen content was made with the use of electron microscope. The relationship between mechanical measurements and collagen concentration was evaluated. The opening angle in the group of AS was significantly smaller than that in control (87.74 degrees +/-9.67 degrees vs. 196.03 degrees +/- 27.76 degrees, P < 0.001). Significant decrease of material constants (alpha0, alpha1, alpha2, b0, b1, b2) in both long axis and radial axis was observed in AS group(campared with control, P < 0.05-0.001). Close relationship between the mechanical constants and the percentage of elastin and collagen content was observed (r = -0.7523 to -0.8423, P < 0.001). In conclusion, mechanical remodeling in aortic artery of AS might be related with histological remodeling.
Animals ; Aorta, Abdominal ; metabolism ; pathology ; Atherosclerosis ; metabolism ; pathology ; physiopathology ; Biomechanical Phenomena ; Collagen ; analysis ; Elastin ; analysis ; Female ; Male ; Random Allocation ; Rats ; Rats, Wistar ; Stress, Mechanical