Objective To generate HLA - A * 0201 and A * 2403 restricted HBcAg - specific cytotoxic T lymphocyte (CTL) clones. Methods Peripheral blood mononuclear cells (PBMCs) were derived from a HLA - A * 0201/2403 - positive patient with chron- ic hepatitis B . PBMCs were stimulated respectively using two synthetic peptides( HBc18 ～ 27 and HBc 117 ～ 125) , and epitope -specific CTL clones were generated by limiting dilution technique with PHA alone or combined with synthetic peptides. The cell clones were then characterized by IF staining,FCM and LDH release. Results After 2 weeks of in vitro stimulating PBMCs, specific CTL lines were established. 29 T clones were generated from those CTL lines using HBc18 ～27 stimulating. Among the 29 clones ,28 clones belonged to CD8~+ T cells and all displayed cytolytic activity. 12 CD8 +T clones were generated from those CTL lines using HBc117 ～ 125 stimulating. Specific cytotoxic activity was observed in 9 of those clones. The other three displayed less cytolytic activity. In the process of cloning, PHA was used alone or combined with synthetic peptides,and the achievement showed a rate of 15.62% and 14.58% . Conclusion HBc18 ～ 27 and HBc117 ～ 125 are capable of activating CD8~+ T cells in PBMCs of HLA - A * 0201 /2403 patient with chronic hepatitis B. In the process of CD8~+T cloning, synthetic peptides could not increase the rate of successful cloning.

Objective To investigate cytokine levels in serum and culture medium of peripheral blood mononuclear cells(PBMC) from patients with hepatitis B virus(HBV)infection.Methods PBMCs isolated from fresh heparinized blood were cultured and stimulated with rHBcAg.After 72h at 37℃ 5% CO2 in air,the culture supernatant was collected.Levels of interferon-gamma(IFN-?) and interleukin(IL)-4 in blood serum in spontaneous and supernatant were measured by enzyme-linked immunosorbent assay(ELISA).Results Serum IFN-? levels in patients with acute self-limited hepatitis B(AH) and chronic hapetiti B(CHB) were significantly higher than those in normal control(NC)(P

Objective To establish immortalized B lympho-blastcell line(LCL) from patients with hepatitis B virus(HBV) infection in vitro.Methods Immortalized B-lymphoblastoid cell lines were established by EB virus transformation of the peripheral blood B lymphocytes and Cyclosporin A(Cys A) restraining T cells.HLA-A gene were measured in blood mononuclear cells by PCR-SSP.Results Altogether 16 immortalized lymphoblastoid cell lines were established successfully in vitro.HLA-A alleles were detected,including 1101,0201,0101,2403,et al.Conclusion The LCLs by EB virus transformation provides a resource of target cell for further research on the cellular immunity of patients with HBV infection.

Objective Cancer, a disease induced by abnormally regulated cell growth and apoptosis, is imposing a global threat to human health.This study was to explore the effects of Chinese herbal extracts ( CHE) in inducing the apoptosis and inhibiting the proliferation of human lung cancer cells. Methods Human lung cancer A549 cells were divided into a negative control, a high-dose CHE (680 ng/mL), a medium-dose CHE (340 ng/mL), and a low-dose CHE (170 ng/mL) group.The inhibitory effect of CHE on the proliferation of the lung cancer cells was detected by CCK8 and LDH assays, the apoptosis of the cells was assessed by fluorescence microscopy and flow cytometry, and the expressions of hTERT mRNA, cleaved caspase-3 and cleaved PARP were deter-mined by RT-PCR and Western blot. Results CHE inhibited the proliferation of the A549 cells with an IC50 value of 510 ng/mL. Treatment with high-dose CHE for 48 hours significantly suppressed the proliferation of the cells, induced the release of LDH, and promo-ted the apoptosis of the cells by 72.3%.RT-PCR and Western blot showed that 24-hour treatment with medium-dose CHE reduced the expression of hTERT mRNA by 4 times that of the negative control and up-regulated the expressions of cleaved caspase-3 and cleaved
PARP. Conclusion Chinese herbal extracts can induce cell apoptosis by decreasing the expression of hTERT mRNA and increasing those of the cleaved caspase 3 and cleaved PARP proteins.

The change of PNA receptors on the surface of thymocytes in protein deficient mice was investigated. After the mice were fed with protein deficient diet, the body weight continuously declined and the thymus gradually a-trophied. Qualitative test of FITC-PNA indicated the number of PNA positive cells was reduced and the fluorescent antibody reaction on the surface of PNA positive cells weakened. Quantitative test of FITC-PNA showed the amount of FITC-PNA coupled on the surface of PNA positive cells was mark- edly decreased as compared with the cotrol (P

We have observed and studied the immune response, ultrastructure and phagocytosis of peritoneal macrophages (M?) of mice in protein deficiency by means of indirect fluorescent antibody test (IFAT), immunoenzymatic staining technique (IEST),fluorescence isothiocyanate antibody (FITC-Ab) quantitative assay, M? phagocytosis test, scanning electron microscopy (SEM) and im-munoelectron microscopy (IEM).The results showed that the body weight of mice was continuously declined after fed protein deficient diet. In the same time fluorescence reaction and enzyme stain on the M? surface was retarded. The amount of FITC-Ab on the M? membrane was decreased. The villi on the M? surface were shortened, the positive rates and positive degree of cells were lowered,the reaction of cell membrane and nuclear membrane was retarded in SEM and IEM.The phagocytic function of M? was inhibited.The results showed that in protein deficiency, the immune reaction, structure and function of peritoneal M? of mice were markedly affected.

Objective To study the effects of social isolation (SI)on the spatial and nonspatial cognitive ability in mice.Methods The postnatal 21 day kunming mice were divided into control group,SI 2 weeks group,SI 4 weeks group,SI 8 weeks group and SI 2 weeks gregarious group according to randomized block design,with ten animals each.SI 2 weeks group,SI 4 weeks group and SI 8 weeks group were isolated for 2,4 and 8 weeks respectively,SI 2 weeks gregarious group would be housed under normal grouped housing condition after 2 weeks isolation until adult,the relative control groups were the same age as the relative SI and SI gregarious group.All animals were measured the spatial and nonspatial cognitive ability by carrying the object recognition test(ORT) and object location test (OLT) after the treatment.Results In the ORT,compared to the relative control group,the discrimination index in the SI 2 weeks group,SI 4 weeks group and SI 8 weeks group ( ( - 0.03 ± 0.003 ),( - 0.11 ±0.02) and( - 0.21 ± 0.02 ) respectively) were strikingly lower than the relative control group ( ( 0.29 ± 0.03 ),(0.13±0.07) and (0.09 ±0.03) respectively) (P＜0.05).In the OLT,compared to the relative control group,the discrimination index in the SI 2 weeks group,SI 4 weeks group and SI 8 weeks group( ( -0.15 ±0.02),( -0.30± 0.02),( - 0.32 ± 0.02 ) respectively ) were strikingly lower than the relative control group ( (0.33 ± 0.02 ),(0.41 ± 0.03 ),(0.27 ± 0.04)respectively)(P＜ 0.05 ),while the SI 2 weeks gregarious group with the resocialization to the normal housing condition showed no change.Conclusions 2 weeks,4 weeks and 8 weeks isolation on mice lead to the spatial and nonspatial cognition deficits,while the resocialization to the normal housing condition could recover the damage.

Objective:To analyze the risk factors of bronchopulmonary dysplasia(BPD)in very preterm infants(VPI), and to provide scientific basis for the prevention and treatment of BPD in VPI.Methods:A prospective multicenter study was designed to collect the clinical data of VPI in department of neonatology of 28 hospitals in 7 regions from September 2019 to December 2020.According to the continuous oxygen dependence at 28 days after birth, VPI were divided into non BPD group and BPD group, and the risk factors of BPD in VPI were analyzed.Results:A total of 2 514 cases of VPI including 1 364 cases without BPD and 1 150 cases with BPD were enrolled.The incidence of BPD was 45.7%.The smaller the gestational age and weight, the higher the incidence of BPD( P<0.001). Compared with non BPD group, the average birth age, weight and cesarean section rate in BPD group were lower, and the incidence of male infants, small for gestational age and 5-minute apgar score≤7 were higher( P<0.01). In BPD group, the incidences of neonatal respiratory distress syndrome(NRDS), hemodynamically significant patent ductus arteriosus, retinopathy of prematurity, feeding intolerance, extrauterine growth restriction, grade Ⅲ~Ⅳ intracranial hemorrhage, anemia, early-onset and late-onset sepsis, nosocomial infection, parenteral nutrition-associated cholestasis were higher( P<0.05), the use of pulmonary surfactant(PS), postnatal hormone exposure, anemia and blood transfusion were also higher, and the time of invasive and non-invasive mechanical ventilation, oxygen use and total hospital stay were longer( P<0.001). The time of starting enteral nutrition, cumulative fasting days, days of reaching total enteral nutrition, days of continuous parenteral nutrition, days of reaching 110 kcal/(kg·d) total calorie, days of reaching 110 kcal/(kg·d) oral calorie were longer and the breastfeeding rate was lower in BPD group than those in non BPD group( P<0.001). The cumulative doses of amino acid and fat emulsion during the first week of hospitalization were higher in BPD group( P<0.001). Multivariate Logistic regression analysis showed that NRDS, invasive mechanical ventilation, age of reaching total enteral nutrition, anemia and blood transfusion were the independent risk factors for BPD in VPI, and older gestational age was the protective factor for BPD. Conclusion:Strengthening perinatal management, avoiding premature delivery and severe NRDS, shortening the time of invasive mechanical ventilation, paying attention to enteral nutrition management, reaching whole intestinal feeding as soon as possible, and strictly mastering the indications of blood transfusion are very important to reduce the incidence of BPD in VPI.