OBJECTIVETo study the dynamic changes of levels of hepatocyte growth factor (HGF) in tears and their association with corneal haze in rabbits early after epipolis laser in situ keratomileusis (Epi-LASIK).

METHODSTwenty-four New Zealand rabbits received Epi-LASIK with an ablation depth of 100 µm in one eye and of 150 µm in the other eye. Before and at 3, 7, 14, and 30 days after the surgery, the level of HGF in tears collected from the rabbits was measured using enzyme-linked immunosorbent assay (ELISA), and corneal haze was graded after surgery.

RESULTSIn all the rabbits, corneal epithelium healing occurred in 3 to 5 days after Epi-LASIK. Corneal haze appeared 3 days postoperatively in the rabbits accompanied by increased levels of HGF in tears. At 3, 7, 14, and 30 days after the surgery, the rabbits with an ablation depth of 150 µm showed more obvious corneal haze (P<0.05) and significantly higher levels of HGF in tears than those with an ablation depth of 100 µm (P<0.05).

CONCLUSIONIn rabbits receiving Epi-LASIK, HGF levels in tears and the grade of corneal haze show a positive correlation early after the surgery and are both related with the depth of ablation.

OBJECTIVETo prepare and characterize monoclonal antibodies (mAb) and polyclonal antibodies against nucleocapsid (N) protein of severe acute respiratory syndrome (SARS)-associated coronavirus (SARS-CoV) and to establish antibodies-based sandwich ELISA for detecting N protein of SARS-CoV, which might apply to early diagnosis of patients with SARS-CoV infection.

METHODSBALB/c mice were immunized with purified recombinant N protein of SARS-CoV for producing mAbs, and New Zealand white rabbits were immunized for producing polyclonal antibodies. The identification of antibodies was performed using indirect enzyme-linked immunosorbent assay (ELISA), indirect fluorescent-antibody assay (IFA), and Western immunoblotting. Capturing and detecting antibodies were selected by pairing the mAbs and polyclonal antibodies one by one and an antibodies-based sandwich antigen capture ELISA was used for detecting N antigen of SARS-CoV.

RESULTSNine mAbs and hyperimmune rabbit polyclonal antibodies, specifically against SARS-CoV nucleocapsid protein were obtained. Using paired ELISA assay, three mAbs N1E8, N8E1 and N10E4 were selected as capturing antibody and rabbit polyclonal antibodies as detecting antibody then triple antibodies-based sandwich ELISA was established following horseradish peroxidase (HRP)-conjugated goat anti-rabbit immunoglobulin G. The recombinant N protein was used as a standard to establish a detection sensitivity of approximated 50 pg/ml with this assay. When tested with 420 serum specimens from serologically confirmed SARS patients, the positive rates of serum N protein were 90.1%, 23% and 0%, in which sera collected from 1 to 10 days, 11 to 20 days and beyond 21 days respectively after the onset of symptoms. The specificity of the assay was 99.86% in 715 control serum specimens. There was no cross-reaction with other respiratory viruses and coronaviruses.

CONCLUSIONSpecific and high affinity mAbs and rabbit polyclonal antibodies were obtained. By paired and optimized sandwich ELISA, a sensitive and specific antigen capture ELISA was established for detecting N antigen of SARS-CoV, which might apply to early diagnosis, source tracing and epidemiological studies of SARS.

Animals ; Antibodies, Monoclonal ; biosynthesis ; Antibodies, Viral ; blood ; Enzyme-Linked Immunosorbent Assay ; Humans ; Mice ; Mice, Inbred BALB C ; Nucleocapsid ; immunology ; Rabbits ; SARS Virus ; immunology ; isolation & purification ; Sensitivity and Specificity ; Severe Acute Respiratory Syndrome ; virology

To elucidate the function of the S2 gene in equine infectious anemia virus (EIAV) and its role in the attenuation of the Chinese attenuated EIAV vaccine strains, the S2 in the EIAV vaccine strain EIAV (FDDV) was reverse-mutated and the in vitro replication character of the resultant virus was evaluated. Based on the sequence variation of the S2 gene between the EIAV virulent strains and vaccine strains, all the four vaccine-specific sites in the S2 of an EIAV(FDDV) infectious clone, pFDDV3-8, were reverse-mutated to the sequences of the virulent strain EIAV(DV115). The reverse-mutated molecular clone pFDDVS2r1-3-4-5 was used to transfect fetal donkey dermal (FDD) cells for rescuing the derived virus vpFDDVS2r1-3-4-5. The production and replication of vpFDDVS2r1-3-4-5 in FDD cells were proved by RT-PCR, immune fluorescence assay and reverse transcriptase activity assay. Typical virons of EIAV were clearly observed under the electron microscopy. The parallel analysis of the dynamic replication of the reverse-mutated viral clone vpFDDVS2r1-3-4-5 and its parental virus vpFDDV3-8 showed that the virus with four reverse mutations in the S2 replicated only slightly slower than its parental vaccine strain in FDD cells. This result implicates that the mutations in the S2 of the EIAV vaccine strains did not significantly alter the viral replication in vitro. Further studies on the in vivo replication of the reverse-mutated viral clone are required for understanding the relationship between the S2 and the attenuated pathogenesis of EIAV attenuated vaccines.
Animals ; Cell Line ; Genetic Engineering ; Haplorhini ; Infectious Anemia Virus, Equine ; genetics ; physiology ; Mutation ; Viral Proteins ; genetics ; metabolism ; Viral Vaccines ; genetics ; Virus Replication

Objective At present no deep investigation has been done on the relationship between handgrip strength and appendicular lean mass (ALM) in patients with chronic obstructive pulmonary disease (COPD). The study aimed to explore the present situation of handgrip strength in patients with stable COPD and relationship between handgrip strength and ALM.Methods A total of 93 patients with stable COPD who hospitalized in our department from August 2016 to July 2017 were selected for the study. All the patients underwent handgrip strength test, body composition analysis, as well as the analysis of the relationship between handgrip strength and ALM.Results Multivariate linear regression analysis showed age(X1), education(X2), smoking(X3), course of disease(X4) and lower limb lean mass(X5) could be taken as predictive factors for the variation degree of handgrip strength (R2=50.5%), and multiple linear regression equation was Y=9.959-4.315X1+1.397X2+2.679X3-1.526X4+1.538X5. The variation degree decreased to 48.1% when the course of disease was removed from the model, 28.3% when the limb lean mass was removed, 26.5% when two variables were removed. The correlation coefficients of ALM, upper limb, lower limb and torso lean mass (\[22.32±3.25\]kg, \[6.48±1.05\]kg, \[15.83±2.26\]kg, \[22.27±3.22\]kg) with handgrip strength (\[32.27±7.27\]kgf) were respectively 0.484, 0.436, 0.496 and 0.496 (P＜0.01).Conclusion The handgrip strength in patients with stable COPD is closely associated with ALM, and the course of disease and the lower limb lean mass greatly affect the handgrip strength. Clinical workers should pay extra attention to the relationship in order to give timely clinical intervention.

BACKGROUND:The angiogenesis may be related to the proliferation of neural stem ceils,but there is still no unified view.OBJECTIVE:To observe the influence of angiogenesis on neural stem cell proliferation in the subventricular zone of rats after focal cerebral ischemia/reperfusion.METHODS:Male Sprague-Dawley rats were randomly divided into normal group,sham group,vascular endothelial growth factor (VEGF)+cerebral ischemia/reperfusion group,normal saline (NS)+cerebral ischemia/reperfusion group.The injection was done via the lateral cerebral ventricle.Then,each group was subdivided into four groups (1,2,7,14 days after ischemia/reperfusion).Focal cerebral ischemia/reperfusion models were made by the thread method.After modeling,the corresponding intervention was given in each group.The expression changes of Nestin and vWF mRNA in the subventricular zone were detected in all groups by immunohistochemical staining and real-time PCR,respectively.RESULTS AND CONCLUSION:There was a certain increase in vWF and Nestin positive expression in the subventricular zone after cerebral ischemia/reperfusion.At 7 days after ischemia,the expression of vWF mRNA and Nestin reached the peak,indicating the proliferation of neural stem cells in the subventricular zone after cerebral ischemia/reperfusion is associated with the time of angiogenesis.In addition,the expression of vWF mRNA and Nestin was significantly higher in the VEGF+cerebral ischemia/reperfusion group than the other two groups,indicating angiogenesis could promote the proliferation of neural stem ceils in the subventricular zone of rats after cerebral ischemia/reperfusion.

OBJECTIVETo perform theory construction and empirical study of the competency model of rural general practitioners.

METHODSThrough literature study, job analysis, interviews, and expert team discussion, the questionnaire of rural general practitioners competency was constructed. A total of 1458 rural general practitioners were surveyed by the questionnaire in 6 central provinces. The common factors were constructed using the principal component method of exploratory factor analysis and confirmatory factor analysis. The influence of the competency characteristics on the working performance was analyzed using regression equation analysis.

RESULTSThe Cronbach 's alpha coefficient of the questionnaire was 0.974. The model consisted of 9 dimensions and 59 items. The 9 competency dimensions included basic public health service ability, basic clinical skills, system analysis capability, information management capability, communication and cooperation ability, occupational moral ability, non-medical professional knowledge, personal traits and psychological adaptability. The rate of explained cumulative total variance was 76.855%. The model fitting index were Χ(2)/df 1.88, GFI=0.94, NFI=0.96, NNFI=0.98, PNFI=0.91, RMSEA=0.068, CFI=0.97, IFI=0.97, RFI=0.96, suggesting good model fitting. Regression analysis showed that the competency characteristics had a significant effect on job performance.

CONCLUSIONSThe rural general practitioners competency model provides reference for rural doctor training, rural order directional cultivation of medical students, and competency performance management of the rural general practitioners.

Clinical Competence ; General Practitioners ; Humans ; Models, Theoretical ; Professional Competence ; Rural Health Services ; organization & administration ; Rural Population ; Surveys and Questionnaires

The study was aimed to investigate the factor V coagulation activity (FV:C), and to evaluate FVgene polymorphisms and activated protein C resistance (APCR) in the patients with venous thromboembolism (VTE). 95 patients with VTE and 95 normal controls were investigated for FV gene polymorphisms. FV Leiden, FVCambridge, and FVHong Kong were detected by PCR, MnlI and BstNI digestion respectively. FVAsp79His and FVI359T were detected by MassARRAY. FV:C and APCR in 65 patients with VTE and 60 normal controls were determined by a one-stage clotting method and the APTT-based assays respectively. The results showed that the mean levels of plasma FV:C were significantly higher in VTE group than that in controls (108.03% +/- 28.29% vs 95.17% +/- 29.75%) (P = 0.008), the incidence of APCR were 20.0% (13 of 65 cases) in patients with VTE and 5.0% (3 of 60 cases) in normal controls (P = 0.012). FV Leiden, FVCambridge, FVHong Kong, FVAsp79His and FVI359T mutations were not found in two groups. It is concluded that the increased plasma level of FV:C is a risk factor for VTE. There is APCR in both groups, APCR is also a risk factor to VTE. APCR may not be associated with mutations of FV Leiden, FVCambridge, FVHong Kong, FVAsp79His and FV I359T polymorphisms, other factors need to study further in APCR.
Activated Protein C Resistance ; complications ; genetics ; Adolescent ; Adult ; Aged ; Aged, 80 and over ; Factor V ; genetics ; metabolism ; Female ; Humans ; Male ; Middle Aged ; Polymorphism, Genetic ; Protein C ; metabolism ; Venous Thromboembolism ; blood ; complications ; genetics ; Young Adult

This study was purposed to explore the anti-leukemic mechanism of quercetin (Que) in vivo and it enhancing chemotherapeutic effect of adriamycin (ADR) by establishing the quercetin-treated P388 transplanted nude mouse model. The P388 leukemic cells in logarithmic growth phase were taken and injected subcutaneously into BALB/c nude mice so as to establish the leukemia-transplanted nude mouse model. The model mice were treated by quercetin, ADR and their combination, and the survival changes of model mice were observed, the hemogram and peripheral blood cell count examination were performed regularly; the cell cycle was detected and the influence of quercetin on cell proliferation was analyzed by flow cytometry; the caspase-3 protein expression level was detected by ELISA; the mRNA and protein changes of NF-κB, BCL-2, BAX were measured by real-time quantitative flourascence PCR and Western blot respectively. The results indicated that the quercetin and adriamycin could significantly prolong the survival of P388 leukemia nude mice, and their combination displayed significantly prolonged effect. Quercetin and adriamycin alone or in their combination could reduce the ratio of G0/G1 phase in mice, the cell ratio in S phase and G2/M phase increased, and the effects of the combination group were more significant than that of the single agent groups. Quercetin could activate caspase-3 and promote leukemic cell apoptosis. Meanwhile, quercetin could down-regulate the expression of BCL-2 and NF-κB gene, and up-regulate the expression of BAX gene. It is concluded that through modulating the expression of apoptosis-related genes, the quercetin can inhibit leukemia cell proliferation, promote apoptosis, and enhance the chemotherapeutic effects of adriamycin. These results provide some valuable data for further research and development of quercetin as a new and effective anti-leukemic drug.
Animals ; Apoptosis ; Caspase 3 ; Cell Cycle ; Cell Line, Tumor ; Cell Proliferation ; Doxorubicin ; pharmacology ; Drug Synergism ; Leukemia ; drug therapy ; pathology ; Mice ; NF-kappa B ; Quercetin ; pharmacology

OBJECTIVE: To analyze the clinical efficacy of haploidentical hematopoietic stem cell transplantation (haplo-HSCT) by using parental donors on thalassemia patients. METHODS: The 13 thalassemia patients treated by haplo-HSCT using parental donors in our hospital from July 1, 2016, to July 1, 2020 were retrospectively reviewed. Hematopoiesis reconstitution, the incidence of GVHD, infections and the long-term survival of the patients were analyzed. RESULTS: Twelve of the 13 patients were successfully implanted, the success rate of implantation was 92.3%. The median time of neutrophil and platelet engraftment was 12.5 days (range, 9-22 days) and 21 days (range,12-34 days), respectively. One patient achieved primary graft failure. Three (25%) patients developed to acute GVHD (aGVHD) and achieved complete remission after treatment. Chronic GVHD developed in three (25%) patients, one of them was extensive and under treatment, while one patient developed to severe bacterial infection (7.7%). CMV viremia was diagnosed in two patients (15.4%). There were no patients developed to CMV disease. Three (23.1%) patients achieved EB viremia after transplantation, one of them developed to EBV-related lymphocytic proliferative disease, while there were no patients showed invasive fungal infection. At the last follow-up, all patients survived, twelve of them were free from transfusion dependency. There were no transplant-related deaths. Projected overall and thalassemia-free survival at three years was 100% and 92.3%, respectively. CONCLUSION The transplant protocol of haplo-HSCT by using parental donors in patients with thalassemia has reliable source of donors, high incidence of successful implantation and low incidence of GVHD, which can be used as an effective way to increase the source of donors in children with thalassemia.
Child ; Cytomegalovirus Infections ; Graft vs Host Disease ; Hematopoietic Stem Cell Transplantation ; Humans ; Parents ; Retrospective Studies ; Thalassemia/therapy* ; Transplantation Conditioning/methods* ; Treatment Outcome ; Viremia

OBJECTIVE: To investigate the expression of CD56 in multiple myeloma (MM) cells and its relationship between extramedullary disease and extramedullary relapse. METHODS: Clinical data of 99 patients with MM treated in our hospital from January 2015 to December 2019 was retrospectively analyzed. The patients were divided into positive group and negative group according to the expression of CD56. The relationship between CD56 and multiple myeloma extramedullary disease, extramedullary relapse was analyzed. RESULTS: Among 99 newly diagnosed patients with MM, the positive rate of CD56 was 65%, and the incidence of extramedullary disease of patients in the CD56 positive group was lower than that in the CD56 negative group (17.19% vs 48.57%) (P<0.01). Meanwhile, the incidence of extramedullary relapse of patients in the CD56 positive group was lower than that in the CD56 negative group (1.56% vs 34.29%) (P<0.01). CONCLUSION CD56 is highly expressed in MM, and its low expression is associated with the occurrence of extramedullary disease and extramedullary relapse, which suggests that CD56 may be an important indicator for predicting the occurrence of extramedullary disease and extramedullary relapse.
CD56 Antigen ; Humans ; Multiple Myeloma ; Neoplasm Recurrence, Local ; Retrospective Studies