Objective The study aims to investigate the immunological functions of the nucleocapsid (N) protein of the novel coronavirus Omicron (BA.1, BA.2) and evaluate the differences among different N proteins of mutant strains in immunogenicity. Methods By aligning sequences, the mutation sites of the Omicron (BA.1, BA.2) N protein relative to prototype strain of the novel coronavirus (Wuhan-Hu-1) were determined. The pET-28a-N-Wuhan-Hu-1 plasmid was used as template to construct pET-28a-BA.1/BA.2-N through single point mutation or homologous recombination. The three kinds of N protein were expressed in prokaryotic system, purified through Ni-NTA affinity chromatography, and then immunized into mice. The titer and reactivity of the polyclonal antibody, as well as the expression level of IL-1β and IFN-γ in mouse spleen cells, were detected using indirect ELISA and Western blot assay. Results The constructed prokaryotic expression plasmids were successfully used to express the Wuhan-Hu-1 N, BA.1 N, and BA.2 N proteins in E.coli BL21(DE3) at 37 DegreesCelsius for 4 hours. The indirect ELISA test showed that the titers of polyclonal antibody prepared by three N proteins were all 1:51 200. All three N proteins can increase the expression of IFN-γ and IL-1β cytokines, but the effect of Omicron N protein in activing two cytokines was more obvious than that of Wuhan-Hu-1 N protein. Conclusion The study obtained three new coronavirus N proteins and polyclonal antibodies, and confirmed that mutations in the amino acid sites of the N protein can affect its immunogenicity. This provides a basis for developing rapid diagnostic methods targeting N protein of different novel coronavirus variants.
Animals ; Mice ; SARS-CoV-2/genetics* ; Coronavirus Nucleocapsid Proteins/immunology* ; Nucleocapsid Proteins/isolation & purification* ; COVID-19/immunology* ; Antibodies, Viral/immunology* ; Mice, Inbred BALB C ; Interferon-gamma/metabolism* ; Interleukin-1beta/metabolism* ; Female ; Escherichia coli/metabolism* ; Mutation ; Humans

BACKGROUND:Osteoarthritis is a common degenerative joint disease,and the etiology and development of its pathogenesis are still unclear.Timely diagnosis and treatment of early osteoarthritis are crucial,and there is currently no definite and effective method.Extracellular vesicles come from a wide range of sources,including non-coding RNAs such as small RNAs,circular RNAs,and long chain non-coding RNAs.Extracellular vesicles non-coding RNAs can be directly delivered from primitive cells to neighboring or remote cells,regulating cell activity through intercellular communication and playing an important regulatory role in reshaping the bone and joint microenvironment.OBJECTIVE:To summarize the intervention effects of exosome-derived non-coding RNAs on the joint microenvironment of osteoarthritis and the progress made in clinical application,and to clarify the potential of exosome-derived non-coding RNAs in the diagnosis and treatment of osteoarthritis.METHODS:Search terms "exosomes,non-coding RNA,osteoarthritis,application,signal pathway,synovial fluid,cartilage cells,cartilage matrix,subchondral,mechanism" were used for the search on PubMed database.Finally,66 related articles were included for review analysis.RESULTS AND CONCLUSION:(1) Exosome-derived non-coding RNAs play an important regulatory role in the joint microenvironment during the pathogenesis of osteoarthritis,mainly reflected in:exosome non-coding RNAs regulating the inflammatory response in the joint,degeneration of chondrocytes and cartilage matrix,subchondral bone remodeling,and intercellular communication.(2) The non-coding RNAs in exosomes can serve as biomarkers for osteoarthritis,aiding in the early diagnosis and monitoring of disease progression and prognosis.(3) Exosome non-coding RNAs serve as therapeutic targets for osteoarthritis.Exosomes carry miRNAs to the articular chondrocytes and cartilage matrix to play a regulatory role.(4) Exosomes non-coding RNAs can improve the effect of cartilage tissue engineering by regulating gene expression and promoting intercellular communication to repair or regenerate damaged cartilage.(5) In future research,researchers should continue to explore the intervention mechanism of non-coding RNAs derived from exosomes on osteoarthritis,and apply them to clinical practice in combination with the latest research outcomes in cartilage tissue engineering,which will effectively help solve the pain of osteoarthritis patients.

BACKGROUND:Osteoarthritis is a common degenerative joint disease,and the etiology and development of its pathogenesis are still unclear.Timely diagnosis and treatment of early osteoarthritis are crucial,and there is currently no definite and effective method.Extracellular vesicles come from a wide range of sources,including non-coding RNAs such as small RNAs,circular RNAs,and long chain non-coding RNAs.Extracellular vesicles non-coding RNAs can be directly delivered from primitive cells to neighboring or remote cells,regulating cell activity through intercellular communication and playing an important regulatory role in reshaping the bone and joint microenvironment.OBJECTIVE:To summarize the intervention effects of exosome-derived non-coding RNAs on the joint microenvironment of osteoarthritis and the progress made in clinical application,and to clarify the potential of exosome-derived non-coding RNAs in the diagnosis and treatment of osteoarthritis.METHODS:Search terms "exosomes,non-coding RNA,osteoarthritis,application,signal pathway,synovial fluid,cartilage cells,cartilage matrix,subchondral,mechanism" were used for the search on PubMed database.Finally,66 related articles were included for review analysis.RESULTS AND CONCLUSION:(1) Exosome-derived non-coding RNAs play an important regulatory role in the joint microenvironment during the pathogenesis of osteoarthritis,mainly reflected in:exosome non-coding RNAs regulating the inflammatory response in the joint,degeneration of chondrocytes and cartilage matrix,subchondral bone remodeling,and intercellular communication.(2) The non-coding RNAs in exosomes can serve as biomarkers for osteoarthritis,aiding in the early diagnosis and monitoring of disease progression and prognosis.(3) Exosome non-coding RNAs serve as therapeutic targets for osteoarthritis.Exosomes carry miRNAs to the articular chondrocytes and cartilage matrix to play a regulatory role.(4) Exosomes non-coding RNAs can improve the effect of cartilage tissue engineering by regulating gene expression and promoting intercellular communication to repair or regenerate damaged cartilage.(5) In future research,researchers should continue to explore the intervention mechanism of non-coding RNAs derived from exosomes on osteoarthritis,and apply them to clinical practice in combination with the latest research outcomes in cartilage tissue engineering,which will effectively help solve the pain of osteoarthritis patients.

Objective To investigate the correlation between the expression of circulating exosome miR-485-3p and STYX with the risk of premature coronary heart disease.Methods A total of 50 inpatients with early onset coronary heart disease diagnosed by coronary angiography or CT angiography (CTA) in Af-filiated Puyang Municipal People's Hospital of Xinxiang Medical College from August to December 2023 were selected as the study group and 50 patients with excluded coronary artery disease by examination during the same period were included in the control group.The general clinical data of the two groups were collected,the plasma exosome miR-485-3p and STYX levels were detected.The degree of coronary arterial lesions in the pa-tients of the study group was evaluated by the Gensini score.The Spearman correlation analysis was used to analyze the relationship between plasma exosome miR-485-3p and STYX with LDL and Gensini score.The re-ceiver operating characteristic (ROC) curve was used to analyze the diagnostic value of plasma exosome miR-485-3p and STYX in the diagnosis of premature coronary heart disease.The multivariate logistic regression was used to determine the independent risk factors for premature coronary heart disease.Results Compared with the control group,the family history of coronary heart disease,smoking history,LDL and plasma exo-some miR-485-3p level in the study group were increased,the plasma STYX level was decreased and the differences were statistically significant (P<0.05);the Spearman correlation analysis showed that miR-485-3p was positively correlated with LDL (r=0.546) and Gensini score (r=0.485),and negatively correlated with STYX (r=-0.576).STYX was negatively correlated with LDL (r=-0.389) and Gensini score (r=-0.531).The ROC curve showed that the area under the curve of miR-485-3p,STYX and their combination in the diagnosis of premature coronary heart disease was 0.821 (95％CI:0.736-0.906),0.850 (95％CI:0.772-0.927) and 0.899 (95％CI:0.837-0.960) respectively.Conclusion The expression of circulating exosome miR-485-3p in premature coronary heart disease is up-regulated and the expression of STYX is down-regulated,the both are closely related to the degree of coronary artery lesion,which could be used as the po-tential biomarkers for the diagnosis of premature coronary heart disease.

Objective:To investigate the effects of Liuweidihuang pill on the insulin levels in sera and pancreatic islets from spontaneous mouse models of human type 2 diabetes administrated with different doses .Methods:The 6-8 week-old KK-Ay mice were randomly divided into three groups including no drug control group ,low-dose group and high-dose group,in addition C57BL/6J mice were used as a genetic control group .All the animals were given with different dose Liuweidihuang pill solutions or sterile distilled water by intragastrical administration for fifteen weeks .The fasting blood glucose ,body mass and food consumption were measured weekly .The serum insulin levels were surveyed by ELISA .And the insulin levels in the pancreas islets were detected by immunofluorescence and immunohistochemistry .Results:Decreased fasting blood glucose ,controlled body mass and food consumption ,and lower levels of insulin in the sera and pancreas islets were confirmed from the KK-Ay mice administered with Liuweidihuang pill .Furthermore,the low dose program exhibits a stronger effect .Conclusion:Liuweidihuang pill has exhibited relatively therapeutic effects in the spontaneous type 2 diabetes mice including controls of hyperglycemia and body mass and relieving insulin resistance .In addition , the low-dose regimen showed even better treatment in controlling insulin levels in the sera and pancreas islets .

OBJECTIVE: To observe the efficacy of intratympanic steroid injection as supplementary or initial treatment for sudden sensorineural hearing loss (SSNHL). METHOD: A total of 68 patients diagnosed with SSNHL were randomized into group A (45 cases including systemic steroid for 33 cases and systemic steroid + intratympanic steroid as supplementary treatment for 12 cases) and group B (23 cases, initial intratympanic steroid). Then observe the therapeutic effect in two groups. RESULT: The total effective rate was 55.6% in group A and 56.5% in group B. No statistical difference was detected between these two groups (P > 0.05). There was statistical difference after therapy of intratympanic steroid as supplementary treatment for 12 patients due to poor hearing improvement after systemic steroid in group A (P < 0.05). CONCLUSION Both systemic and intratympanic steroid injection for SSNHL are effective. The efficiency of intratympanic steroid injection as supplementary or initial treatment for SSNHL is similar to that of systemic steroid. The intratympanic steroid injection for SSNHL as initial protocol or as supplementary treatment when poor hearing improvement after systemic steroid is recommended.
Hearing Loss, Sensorineural ; drug therapy ; Hearing Loss, Sudden ; drug therapy ; Humans ; Injection, Intratympanic ; Steroids ; administration & dosage ; therapeutic use ; Treatment Outcome ; Tympanic Membrane

Objective To study the influence of donor GFR on the early renal function in recipients undergoing living donor transplantation.Methods A total of 172 living donor transplant recipients in our kidney transplantation center from 2006 to 2011 were enrolled into this study.Among them,166 were genetically related (96.5％),while 6 were genetically unrelated (spouses in 5 and other in 1).The predonation GFR was measured by isotope clearance (99mTC-DTPA with few exceptions).The range of donor GFR was 62 to 148 ml/min.The recipients were classified into two groups according to donor graft GFR level (GFR≤45 ml/min,n=76; GFR＞45 ml/min,n =96).The predonation dialysis,cold and warm ischemia time,antibody induction,immunosuppressive regimens and HLA mismatch were not significantly different between two groups.Results There were no significant differences in the incidence of postoperative acute rejection and delay graft function (DGF).The postoperative Scr of GFR＞45 ml/min group in 1 week,1 month,3 months and 1 year was lower compared with the GFR ≤45 ml/min group,and only the difference of Scr in 1 week was significantly different (P＜0.05).A repeated-measure ANOVA revealed no significant differences were found in Scr variation of two groups during the first year after transplantation.Conclusions Predonation GFR of the donor has effect on the Scr of postoperative Ⅰ week of recipients,not on the Scr within a year.Recipients with graft GFR＞45 ml/min have lower Scr levels.

Objective:To construct suitable vectors for the secretory expression of hirudin in Pichia pastoris.Methods:The α-facor-hirudin gene was amplified from pPIC9-hirudin by PCR and sub-cloned into PA0815.The multi-copy recombinant plasmid pA0815-(α-Hirudin)n was constructed.The recombinant was transformed into P.pastoris strain GS115 for induction expression and then the activity of secreted products was identified.Results:A new multi-copy vector pA0815-(αHirudin)n was successfully constructed and was capable of secreting recombinant hirudin efficiently,which was confirmed respectively by PCR and SDS-PAGE.The products possessed the activity of thrombin inhibitor.Conclusion:This result offers efficient P.pastoris stains for mass production of biological active hirudin.

0.05).CONCLUSION:There is no evidence of impaired JPS in weight-bearing positions in subjects with ACL reconstruction at a mean follow-up of 12 months after surgery compared with subjects with healthy knees.

OBJECTIVE: To evaluate the long-term toxicity of Anti-EB-virus oral liquid on beagle dogs, and to provide safety evidences for clinical experiment. METHODS: Of the total beagle dogs, three groups totaled 18 were assigned to receive Anti-EB-virus oral liquid at high (76.41 g/kg), medium (25.47 g/kg) and low dosage (8.49 g/kg), and 6 (blank control group) to receive placebo. The ratio of male to female in each group was 1∶1. The beagle dogs were administered intragastrically with drugs qd, 6 days in a week for up to 26 weeks. All the indicators were monitored and the recovery of the beagle dogs was observed. RESULTS: Dogs in high dose group vomited obviously. Gastrointestinal irritation was also noted in medium dose subgroup, but was less severe than the high dose subgroup. During the recovery stage, one dog in high dose group was strong positive in urinary protein test and one in medium dose group was strong positive in urinary occult blood test. No significant drug associated toxic reactions were noted from beagle dogs' body weight, temperature, appetite, ECG, hematology test, blood biochemical analysis, ophthalmology test, marrow test, routine urianlysis, histopathologic examination, etc. CONCLUSION: The non-toxic dose of Anti-EB-Virus Liquid on beagle dogs was 8.49 g/kg in crude drug.