Bone marrow from 20 patients suffering from aplastic anemia were cultured with fetal muscle condition medium and in three different combinations: (1) Marrow from the patients cultured alone; (2) Coculture of marrow from patient with normal marrow; and (3) Culture of normal marrow with serum from patient. It was found that: (1) Nine patients had low colony formation when culture alone; no suppression in coculture and no inhibiter factor was found in serum from patient. Stem cells might be absent or defective in these patients. (2) Five patients had low colony when cultured alone; their marrow showed marked inhibitory effect on normal marrow CFU-C in coculture. and the latter was decreased by 40-100%. Suppressor cells might have caused the aplasia in these patients. (3) The sera from five patients suppressed CFU-C of normal marrow cells, colony counts being decreased by 55-77%. The cause of aplasia in these patients might be the presence of an humoral inhibiting factor. (4) In one patient no abnormal findings were found in all three different combinations of cultures. This patient might be suffering from a defective hematopoietic environment.

Objective To evaluate the alteration of CD4+ regulatory T cells in peripheral blood from patients with ankylosing spondylitis(AS). Methods Seventy-eight AS patients and 50 healthy individuals were included in this study. The proportion of CD4+CD25+CD127lo/- T cell population in CD4+ T cells as well as that cytotoxic T lymphocytes(CTL) and NK cells in lymphocytes was evaluated by flow cytometry. Serum TGF-β and TNF-α were measured by ELISA. The inhibitory function of CD4+CD25+ regulatory T cells was measured by mixed lymphocyte culture. Results The population of CD4+CD25+CD127lo/- T cells in peripheral blood of AS patients accounted for (4.36±1.21)% of CD4+ T lymphocytes, which was significantly lower than that in healthy individuals (P<0.05). The CD4+CD25+CD127lo/- T cells population in AS patients was positively correlated with TGF-β level, but negatively with TNF-α. Compared with healthy individuals, the function of CD4+CD25+ regulatory T cells in the inhibition of alloreactive T cells was lower in AS patients, which was related to the decreased secretion of TGF-β. Conclusion The CD4+ regulatory T cells in peripheral blood of AS patients are significantly decreased and its function is defective, which leads to immune regulatory dysfunction in vivo. It may be one of immune pathogenesis mechanisms of AS.

According to the experience of teaching the science of Chinese Medical Herbs Preparation,the author gives some advice such as choosing typical drugs,providing examples and setting up comprehensive and contriving experiments etc,which is worth referring to in teaching es-pecially for those who have less time in experiments.

Abstract: To report on two patients with Coronavirus Disease 2019 (COVID-19) combined with diffuse connective tissue disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection followed for nearly 3 years, in order to understand the long-term effects on the patients' immune system. Both patients were male, aged 81-82 years, and were hospitalized with fever on January 29, 2020 and February 10, 2020, respectively. Both were diagnosed with COVID-19 after positive SARS-CoV-2 polymerase chain reaction (PCR) tests. After receiving anti-infection treatment, cough suppressants, ex‐pectorants, and symptomatic supportive treatment, their body temperature returned to normal and two consecutive PCR tests were negative for SARS-CoV-2, and they were discharged from hospital. However, due to recurring fevers and varying degrees of rheumatic disease-related symptoms, both patients were readmitted to the hospital, indicating the presence of positive auto‐ antibodies and organ involvement. One patient recovered from COVID-19 with recurrent fever, joint pain, muscle aches and subcutaneous nodules, and was subsequently diagnosed with undifferentiated connective tissue disease. The other patient developed recurrent fever, mouth ulcers and rash after recovery from COVID-19 and was subsequently diagnosed with anti neutro phil cytoplasm antibody (ANCA)-associated vasculitis (AAV). The patient was treated with glucocorticoids and immunosuppres sive drugs and the symptoms resolved rapidly and subsequent laboratory and imaging examinations showed stable condition. However, due to self-termination of medication, their symptoms quickly relapsed, and further treatment with glucocorticoids and immunosuppressive agents resulted in sustained stability of their condition. The erythrocyte sedimentation rate and hyper‐sensitive C-reactive protein remained within normal limits, and lung CT scans showed stable lesions with partial absorption.SARS-CoV-2 infection may have long-term effects on patients' immune systems, leading to abnormal immune responses and diffuse connective tissue disease. This suggests that regular follow-up observation of immune system-related diseases may be necessary for elderly patients with COVID-19.

Objective To evaluate the uncertainty of the pseudo-ginsenoside GQ (PGQ) concentration in human plasma by HPLC-MS/MS.Methods The whole process of PGQ determination by HPLC-MS/MS in human plasma was evaluated and the uncertainty caused by repeatability,weighing,standard solution preparation,biological sample preparation,extraction recovery process,recovery,instrument precision and calibration curve fitting were evaluated,respectively.The combined and expanded uncertainty values were both calculated.Results The expanded uncertainty values for low (15.16 ng·mL-1),medium (2 516.67 ng·mL-1) and high (3 902.00 ng·mL-1) levels of PGQ were 1.39,177.74 and 262.69 ng·mL-1,respectively (P =95 ％,k =2).Conclusion The uncertainty of the PGQ determination in human plasma by HPLC-MS/MS is mainly caused by recovery,repeatabihty and sample preparation at low concentration,by sample preparation and recovery at medium and high concentration.

Objective To study the effect of TCM five-tone therapy on chronic fatigue syndrome(CFS).Methods Fifty nine CFS patients were divided into the treatment group(n=30)and the control group(n=29),which received TCM five-tone therapy and common music therapy,respectively for 3 months.Both groups were assessed with fatigue Scale-14,depression status inventory and visual analogue scale.Result After treatment,the treatment group was scored lower than the control group in FS-14,DSI and VAS(all P<0.05).Conclusion TCM five-tone therapy may be more effective in decreasing the CFS patients with fatigue and depression and alleviating their pain symptoms.

Objective To compare the effects of preload with intravenous infusion of 6％ hydroxyethyl starch combined with phenylephrine or dopamine to prevent the hypotension after combined epiduralspinal anesthesia in parturient undergoing caesarean section.Methods Eighty patients with ASA class Ⅰ or Ⅱ[,were randomly divided into Dopamine group and Phenylephrine group,40 cases in each group.The 6％hydroxyethyl starch 500 ml was infused at the tate of 20 ml/(kg · h) after the intravenous catheterization was established and after the finishing of the infusion of 250 ml,the dopamine 5 mg (Dopamine group) or 200 ug phenylephrine (Phenylephrine group) were added respectively in residual liquid.After the bupivacaine was injected into the subarachnoid space,the intravenous infusion was continued at the same rate until the fetus was taken out and the blood pressure and heart rate were measured at intervals of 1 min.The blood sample of fetal cord was taken to measure ther troponin Ⅰ concentration.Results The incidence of hypotension after combined epidural-spinal anesthesia anesthesia in dopamine group (2/40) and in phenylephrine group (3/40) was with no statistical difference (P ＞ 0.05) ;The incidence of bradycardia in dopamine group (0/40) was significantly lower than that in phenylephrine group (6/40)) (P ＜0.05) ; The incidence of tachycardia in dopamine group (8/40) was significantly higher than that in phenylephrine group (1/40) (P ＜0.05) ; The troponin Ⅰ concentration of fetal cord blood in dopamine group [(0.21 ±0.07) ng/ml] and in phenylephrine group [(0.18 ±0.09)ng/ml]was with no statistical difference (P ＞0.05).Conclusion Preload with intravenous infusion of 6％ hydroxyethyl starch combined with phenylephrine or dopamine can effectively prevent the hypotension after combined epidural-spinal anesthesia in parturient undergoing caesarean section with no significant effect on the fetus and both can be chosen in terms of the heart rate of parturient before anesthesia.

AIM:To investigate the target killing effect of T lymphocytes with chimeric CD20scFv gene on Daudi cells and the activation of T lymphocytes.METHODS:Two kinds of plasmids were transfected into retrovirus-packed PA317 cell lines.The supernatant was collected from successfully transfected PA317 culture and was used to infect peripheral blood T lymphocytes.After one-week screening with G418,the cells were used to kill Daudi and K562 cells.The positive rates of AnnexinⅤ in Daudi cells were measured at different times points respectively by flow cytometry.Meanwhile,the level of IL-2 and IFN-? were determined by ELISA.RESULTS:The Annexin V positive rate was significant higher in Daudi cells compared to control K562 cell lines at 24 h.No difference of AnnexinV in Daudi cells was observed in CD20 modification T lymphocyte groups.The secretions of IL-2 and IFN-? in CD20scFv-CD80-IgGFc-CD28-? gene modified T cells co-cultured with Daudi cells were dramatically higher than that in CD20scFv-IgGFc group at 72 h.CONCLUSION:① The two kinds of genetic modified specific T cells have no significant difference in inducing early apoptosis of Daudi cells.CD28-? can't affect Daudi cell early apoptosis at the CD20scFv target killing.② The increase in the secretions of IL-2 and IFN-? is more obvious in CD20scFv-IgGFc-CD28-? group,indicating that the self-activation takes place in CD3? and CD28 modified T cells without MHC restriction and then increases the activation and killing function of T cells.

Objective To study on plasmid-mediated quinolone-resistant in Escherichia coli strains isolated from fecal samples of chicken and swine from the nine farms around our country.Methods Antimi-crobial susceptibility testing was carried out by broth microdilution testing,gyrA,gyrB,parC,qnr and aac (6')- Ⅰ b-cr were examined by PCR,and the products were sequenced.Conjugation experiment was carried out to proved that the plasmid-mediated quinolone resistance was transferable.Results In the total 818 animal isolates,qnr and aac genes were detected in 38 (4.6%) and 75 (9.2%) strains.The qnrA,qnrB,and qnrS genes were detected in 1 (0.1%),9 (1.1%) and 28 (3.4%) of the isolates.All isolates were negative for qnrC,qnrD genes.Conclusion There is a close relationship between high level quinolone resistance and plasmid-mediated quinolone resistance.The results of the current study highlight food-producing animals as a potential reservoir of antimicrobial-resistant bacteria and clinically important resistance genes.More attention should be paid to the surveillance of such strains.

Objective To characterize the roles of different quinolone resistance mechanisms in quinolone-resistant Escherichia coli isolates,including different topoisomerase point mutations,efflux pumps and outer membrane proteins.Methotis Through homologous gene recombination methods,different quinolone-resistant mechanisms of E. coli mutants were constructed and the susceptibility changes of these mutants to different antimicrobials were measured.Resuits Efflux pumps AcrAB and outer membrane protein TolC played different roles in different E. coli isolates.Compared with other mechanisms,the mutations in topoisomerases played a dominant role in quinolone resistance.Only the mutations jn parC had no effect on quinolone resistance,which further confirmed parC was the secondary target of quinolones in E.coli.Fluoroquinolone susceptible E.coli would automatically become highly resistant to quinolones after acquiring the point mutations in both gyrA(S83L,D87N)and parC(S80I,A108V),but not requiring the over-expres-sion of efflux.Conclusion The mutations in topoisomerases play a dominant role in E.coli quinolone resistance,and the mutations in both gyrA and parC are required.