Objective To optimize the method of isolating ,culturing and screening fetal mouse liver stem cells in vitro ,and to identify the potential of bi‐directional differentiation .Methods The fetal liver stem cells of mouse were isolated by the density gra‐dient centrifugation and cell difference adherence method ,the proliferation of stem cells was determined by cell plate cloning tech‐nique and MTT method;stem cells were induced for differentiation by adding DMSO and HGF .Results The isolated stem cells showed adherence within 24 h ,which were orbicular‐ovate ,closely packed ,activated within 1~2 weeks ;the positive rates of CD133 , CD49f and EPCAM were (97 .95 ± 1 .21)% ,(92 .71 ± 3 .49)% and (50 .73 ± 3 .45)% respectively ;AFP and CK19 proteins were expressed;red glycogen granules were seen by PAS after induced differentiation;ALB and HNF‐4αwere expressed .Conclusion Fe‐tal hepatic stem cells are successfully isolated by the density gradient centrifugation combined with difference adherence method ,and the isolated cells have strong stemness and proliferation ability ,as well as the ability of bi‐directional differentiation towards hepato‐cytes and bile duct epithelial cells .

[Objective] To study the factors relative to the delayed diagnosis of spontaneous dual arteriovenous fistulas (DAVF) and its influence on the prognosis.[Methods] We included 102 continuous patients diagnosed DAVF in the First Affiliated Hospital of Sun Yat-sen University,and analyzed the correlative factors and impact on outcome of diagnostic delay.Outcome was whether symptoms were non-improvement,improvement or restoration at discharge.[Results] Median delay from onset to diagnosis was 3 months (interquartile range,1 to 6).Compared with patients diagnosed carlier(diagnose time≤3 months),patients diagnosed later (diagnose time ＞ 3 months) had a lower frequency of headache (P =0.012),ptosis (P =0.035) and parenchymal lesions (P =0.001),a higher frequency of conjunctival congestion (P =0.004),tinnitus (P =0.021),visual dysfunction (P ＜ 0.001),isolated visual dysfunction (P =0.007) and delayed imaging scan (P ＜ 0.001),a higher frequency of endovascular treatment,and a lower frequency of improvement or restoration at discharge (P =0.033),in which patients with visual dysfunction had a lower frequency of improvement or restoration than those without visual dysfunction (P =0.023).Compared to those with visual dysfunction and other symptoms,patients with isolated visual dysfunction had a higher frequency of onset with paroxysmal blurring or blinding (P ＜ 0.001),two eyes involved (P ＜ 0.001) and more severe visual loss (P =0.057),a higher frequency of draining into transversesigmoid sinus (P ＜ 0.001) instead of cavernous sinus (P ＜ 0.001),and suffered intracranial hypertension all (median intracranial pressure,405 mmH2O;interquartile range,370 ～ 512 mmH2O).However,no statistically significant differences were found in the frequency of improvement or restoration at discharge between two groups (P =0.739).[Conclusion] Diagnostic delay was considerable in this cohort and was associated with outcome,especially in patients with visual dysfunction.

BACKGROUND:Long-term use of corticosteroids (hereinafter referred to as hormone) after renal transplantation could obviously lead to adverse reactions. Immunosuppressive regimen with less and no hormone has been a hot focus in the study of renal transplantation al over the world. However, reduction or withdrawal of hormones has a certain risk. At present, there is no unified scheme. Because urine protein can be immediately detected after tubular injury, to monitor urine protein can find the renal dysfunction after transplantation in recipients undergoing renal transplantation, which can gain time for clinical therapy. OBJECTIVE:To discuss the influence of hormone (prednisone) removal on the occurrence of urine protein in recipients undergoing renal transplantation. METHODS:A total of 35 recipients undergoing renal transplantation after removal of prednisone received immunosuppressive regimen of cyclosporine A or tacrolimus+mycophenolate mofetil bivalent. Initial dose of prednisone was 30 mg/d, and then gradual y reduced by 5 mg per week, and withdrawn at 1 month after renal transplantation. There were 16 cases in cyclosporine A group and 19 cases in tacrolimus group. Urine protein was measured and quantified at 3, 6, 12 and 24 months after renal transplantation and 3, 6 and 12 months after addition of prednisone in both groups. Simultaneously, serum creatinine, fasting glucose, body mass increases, the rate of acute rejection, infection, patient/graft survival at 2 years after renal transplantation and urine protein at 24 hours before and after adding hormone were recorded. RESULTS AND CONCLUSION:For the two groups, urineα1-microglobulin started to rise after 6 months of removal of prednisone. Urinary microalbumin, urinaryα1-microglobulin, and urinary transferrin ascended obviously at 12 months. Urinary protein was positive in five cases of cyclosporine A group and in three cases of tacrolimus group. At 24 months, urinary microalbumin, urinaryα1-microglobulin, urinary transferrin and urinary IgG ascended obviously. Urinary protein was positive in cyclosporine A group with 11 cases and in tacrolimus group with 10 cases. 24-hour urinary protein quantity was more than 1 g in every case. On this base, we made the patients to take more prednisone for 6 months, so urineα1-microglobulin and urinary microalbumin began to descend. Each group had one case of positive urinary protein turning to negative. Twelve months after the adjustment of the prednisone, urinary microalbumin, urinaryα1-microglobulin, and urinary transferrin descended respectively. Positive urinary protein turned into negative:in cyclosporine A group with two cases and in tacrolimus group with three cases. 24-hour urinary protein quantity was around 0.7 g. Two years after renal transplantation, serum creatinine and acute rejection rates were higher in the cyclosporine A group than in the tacrolimus group (P<0.05). No significant difference in fasting glucose, body mass increase, infections, and patient/graft survival was detectable between both groups. Results suggested that removal of prednisone greatly affected urine protein in recipients undergoing renal transplantation. In particular, at 2 years after renal transplantation, urinary microalbumin, urinaryα1-microglobulin, urinary transferrin and urinary IgG ascended obviously, and the security needs further research.

Objective To study the apoptosis of human leukemia K562 cells induced by snake venom of Agkistrodon Halys Pallas in Zhejiang Province.Methods IC_ 50 value and cytotoxity of K562 cell were detected by MTT method.Apoptotic cells were dyed by Hoechest 33258.Sub-G1 peak and cell cycle were detected by FCM.Protein expression of Bcl-2 gene was detected by FCM and western-blot method.Results The snake venom of Agkistrodon Halys Pallas in Zhejiang Province inhibited the growth of K562 cells,which appeared dose-dependent.The snake venom induced apoptosis of K562 cells.Meanwhile,protein expression of Bcl-2 was down-regulated.Conclusion Snake venom of Agkistrodon Halys Pallas in Zhejiang could induce apoptosis of human leukemia K562 cells.The mechanism may be related to down-regulation of Bcl-2 gene expression.

Epidemiological data,clinical observation and animal experiments have shown that cold wave is closely associated with the onset of stroke.When a population with stroke etiology or risk factors is under pre-stroke state,they will have stroke under the influence of various inducing factors.Cold wave is the external factor that causes the body to enter the pre-stroke state and there are many possible mechanisms.The drug intervention of stroke-prone hypertensive rats at 1 week before the cold wave can reduce the occurrence of stroke during cold wave,suggesting that it is of great significance to conduct preventive intervention to the pre-stroke population before the cold wave coming.

Abstract BACKGROUND: Calcium dobesilate (calcium dihydroxy-2, 5-benzenesulfonate) has been widely used to treat chronic venous insufficiency and diabetic retinopathy, especialy many clinical studies showed that calcium dobesilate as vasoprotective compound ameliorates renal lesions in diabetic nephropathy. However, there are few literatures reported calcium dobesilate in the treatment of chronic renal alograft dysfunction after renal transplantation. OBJECTIVE:To observe the efficacy and safety of calcium dobesilate on chronic renal dysfunction after renal transplantation. METHODS:A total of 152 patients with chronic renal alograft dysfunction after renal transplantation were enroled from the Military Institute of Organ Transplantation, Changzheng Hospital, Second Military Medical University of Chinese PLA. They were randomly divided into the treatment group (n=78) and the control group (n=74). Patients in the treatment group received 500 mg of calcium dobesilate three times daily for eight weeks. Al patients were treated with calcineurin inhibitor-based triple immunosuppressive protocols and comprehensive therapies. RESULTS AND CONCLUSION: For patients receiving calcium dobesilate, serum creatinine, blood urea nitrogen and uric acid decreased significantly at two weeks after treatment and maintained a stable level (P < 0.05). However, serum creatinine and blood urea nitrogen returned to the original level soon after drug withdrawal. No significant difference was observed in blood cel count, liver function, blood lipids, electrolytes, blood pressure and 24-hour urine output between the two groups before and after therapy (P > 0.05). Administration of calcium dobesilate did not change the general condition of patients with renal insufficiency, nor did it affect blood concentrations of the immunosuppressive agents. Calcium dobesilate may help to delay the progress of graft injury in patients with chronic renal graft dysfunction by conjugating with creatinine, ameliorating the impaired microcirculation and its antioxidant property. The decline in serum creatinine aleviates patients’ anxiety and concern arising from the elevation of creatinine. However, the negative interference with serum creatinine caused by calcium dobesilate should be cautious in order to avoid misjudgment of patients’ condition.

Objective To investigate correlation between microRNA (miR-494) and TH 17 cell differentiation in murine cervical heterotopic cardiac transplant model.Method The heterotopic cardiac transplant models of Balb/c→C57BL/6 mice were established as experimental group,and those of C57BL/6→C57BL/6 mice as control group.Real time-polymerase chain reaction(PCR) was used to detect miR-494 and interleukin(IL)-17A mRNA expression in the grafts.CD4+ T cells,CD8+ T cells and CD45+ myeloid cells were isolated from the grafts,and miR-494 and IL-17 mRNA expression was detected.In vitro,lymphocytes in the spleen from C57BL/6 mice were harvested,and CD4+ T cells were isolated with MACS and then stimulated to TH 1,TH 2,TH 17,Treg subset cells.The expression of IL-17A mRNA and miR-494 in different T subsets was examined by Reverse transcription-polymerase chain reaction(RT-PCR).Result Two grafts from each study group were harvested on the 7th day post-transplantation.In experimental group,the IL-17A mRNA expression was increased,while the expression of miR-494 was decreased as compared with control group with the difference being significant between two groups.The expression of IL-17A rnRNA in CD4+ T cells of the grafts was significantly increased,while that expression of miR-494 was decreased.In vitro,the expression of miR-494 in TH 17 cells was significantly lower than that in TH 1,TH 2 and Treg cells.Conclusion miR-494 is related closely to TH 17 cells differentiation in the transplant rejection,which may play a role in transplant rejection through regulating TH 17 cells.

Objective To investigate the efficacy and safety of conversion therapy to mizoribine (MZR) for renal transplant patients who suffered MMF or Aza adverse reaction. Methods In 56 patients with adverse reactions at different time points after renal transplantation, there were 23 cases of pulmonary infection, 14 cases of bone marrow depression, 6 cases of hepatic functional lesion and 13 cases of diarrhea. The immunosuppressive protocols of these patients were changed to CNI + MZR + Pre when the adverse reaction occurred. During the follow-up period (11 to 53 months), the effect and adverse events of conversion treatment were observed. Results After conversion treatment, 1 of 23 patients with pulmonary infection was re-infected after 26 months and finally died of heart and lung function failure. In 14 patients with bone marrow depression, blood test returned to normal in 13cases. Six patients with hepatic functional lesion were administered hepatoprotection treatment and their liver function was restored without recurrence of impaired liver function. All 13 patients with diarrhea were relieved without recurrence. The serum creatinine was 123 ± 21.3 μmol/L and 119±18. 2 μmol/L before and after the conversion therapy respectively (P＞0. 05). During the follow-up period, all patients' graft function was good. The incidence of rejection was 1.7 % (1 case). Nine patients (16. 1 %) had a higher level of uric acid after conversion. One patient had finger and toe joint pain. The symptoms were relieved after symptomatic treatment. Conclusion There were high security and good effect of conversion therapy to MZR due to MMF or Aza adverse reaction. Besides, MZR conversion therapy for renal transplantation patients provided a new option for individual immunosuppression.

Objective To compare the difference of clinical and laboratory characteristics between the patients undergoing uninephrectomy for urologic diseases and healthy living kidney donors.Methods The change of blood pressure,renal function,blood routine examination,urine protein,plasma electrolytes and blood fat in two groups preoperatively and at one week,one month,6 months and one year postoperatively were retrospectively assessed.Of the 65 living kidney donors,12 were male and 53 were female.Among 354 cases of uninephrectomy for urologic diseases,there were 291 cases of hpper urinary tract tumor,56 cases of hydronephrosis and 7 cases of severe kidney injury.Results Compared to the preoperative status,there was significant difference of hemoglobin,creatinine,urea nitrogen and urine protein in two groups.Clinical and laboratory characteristics of two groups became stable at one months postoperatively,and could reach the preoperative level at 6 months or one year postoperatively.But the creatinine was 25.71%higher than the preoperative level in healthy donors and 25.49%in patients undergoing uninephrectomy for urologic diseases.There was no significant difference between the two groups at one year postoperatively.Conclusions Two groups are safety after nephrectomy.There is no significant difference of clinical and laboratory characteristics between the 2 groups.Strict donor selection,appropriate pretransplantation evaluation,careful operation and postoperative health consulting play important roles in donors;safety and health.

Objective To discuss the diagnosis and treatment of invasive aspergillosis(IA)of lung after kidney transplantation. Methods Ten cases of IA infection of lung after kidney transplantation from January 1999 to May 2006 were reviewed.Among 10 cases.all had been examined by FOB and 3 positive.Six cases had a typical sign of IA in chest CT.Five cases had GM positive. Results Eight cases were cured including 3 cases with itraconazole,5 with amphotericin B.The other 2 died of infection. Conclusions Invasive pulmonary aspergillosis is a severe complication of renal transplantation.Early diagnosis and proper treatment can reduce the mortality.