Objective To assess the effects ofberberine on autophagy in C3H10T1/2 adipocytes.Methods C3H10T1/2 cells,a pluripotent stem-cell line of mesodermal origin,were induced to differentiate into mature adipocytes,and then were treated with berberine,The expression of autophagy marker protein LC3 Ⅱ/Ⅰ and autophagy substrate P62 was determined by Western blot ; after treating C3 H 10T1/2 cells with berberine and lysosomal inhibitor,chloroquine,autophagy flux was assessed by Western blot.Autophagosome was observed by transmission electron microscopy after berberine treatment.Results (1) After berberine treatment,the expression of LC3 Ⅱ / Ⅰin C3H10T1/2 adipocytes was reduced and P62 was increased in a time-and dose-dependent manner(P＜0.05 or P＜0.01) ; (2) Following treatment with chloroquine and berberine,the protein level of LC3 Ⅱ/Ⅰ was decreased (P＜0.05); (3) The number of autophagosome was decreased apparently after berberine treatment.Conclusions Berberine inhibits autophagy in C3H10T1/2 adipocytes.

Objective To investigate the clinical,endoscopic and pathologic features in the differential diagnosis between Crohn' s disease (CD) and intestinal tuberculosis (ITB).Methods The complete clinical data of 107 patients with CD and 69 patients with ITB in our hospital from January 2011 to January 2012 were retrospectively analyzed.The diagnostic value of the clinical and endoscopic scoring system was evaluated.Results CD occurred mainly in male.The salient features of CD included long duration of disease high incidence of colectomy.Comparing with patients with ITB,patients with CD have more cases of diarrhea,hematochezia,abdominal mass,intestinal obstruction,intestinal hemorrhage,perianal lesions,and extraintestinal manifestations (all P ＜ 0.05).It's more frequent to have positive results of anti-Saccharomyces cerevisiae antibody (ASCA),perinuclear antineutrophil cytoplasmic antibody (pANCA) and fecal occult blood in CD patients,as well as low albumin,high C-reactive protein (CRP),elevated platelet count and hematocrit (P ＜ 0.05 or P ＜ 0.01).The salient features of ITB included low fever,night sweats,active parenteral tuberculosis,increased erythrocyte sedimentation rate (ESR),chest X-ray abnormalities,the positive PPD (purified protein derivatives tuberculin) and T-SPOT (P ＜ 0.05 or P ＜0.01).Based on the imaging,CD often involved the small intestine,such as the intestinal stricture and abdominal abscess (P ＜ 0.05),while mesenteric lymphadenopathy was more common in ITB (P ＜ 0.05).The endoscopic examination showed that some patterns of disease involvement such as fissure-shape ulcer [41.12％ (44/107) vs 5.80％ (4/69)],cobblestone sign [15.89％ (17/107) vs 4.35％ (3/69)],lesions over four segment [24.30％ (26/107) vs 7.25％ (5/69)],rectum involvement [17.76％ (19/107) vs 5.80％ (4/69)],ileocecal valve stenosis [21.50％ (23/107) vs 8.70％ (6/69)] and mucosal bridge [5.61％ (6/107) vs 0(0/69)] were more frequent in CD patients than those in ITB patients(P ＜ 0.01 or P ＜0.05).However circular ulcers[37.68％ (26/69) vs 9.35％ (10/107)],rat-bite-like ulcers[24.64％ (17/69) vs 12.15％ (13/107)],persistent open ileocecal valves [39.13％ (27/69) vs 19.63％ (21/107)],tuberous and polypoid lesions [36.23％ (25/69) vs 20.56％ (22/107),37.68％ (26/69) vs 22.43％ (24/107)] were more common in ITB (P ＜ 0.01 or P ＜ 0.05).In terms of pathological findings,certain characteristic features such as transmural inflammation [5.61％ (6/107) vs 0 (0/69)],fissure-liked ulcers [14.02％ (15/107) vs 4.35％ (3/69)],non-caseous granulomas [5.61％ (6/107) vs 0(0/69)],lymphoid hyperplasia [16.82％ (18/107) vs 5.80％ (4/69)] and crypt abscess [9.35％ (10/107) vs 1.45％ (1/69)] were more common in CD than those in ITB(P ＜ 0.05).According to the clinical and endoscopic scoring system,the positive diagnostic rate of CD was 50.47 ％ (54/107)and of ITB was 66.67 ％ (46/69) (P ＜ 0.05).Conclusions The differential diagnosis between CD and ITB should be considered carefully based on clinical,endoscopic,pathological characteristics.The clinical and endoscopic scoring system may contribute to distinguish CD and ITB.

Objective To compare the expressions of Beclin 1 and its 11 interacting proteins in subcutaneous fat and visceral fat,and to observe the expressions of these genes in 3T3-L1 adipocytes differentiation process.Methods In 3T3-L1 adipocytes,cells were harvested at day 0,2,4,6,8 in differentiation process,and then proteins or mRNAs were obtained and followed with Western blot and realtime PCR.Mice tissue proteins or mRNAs were detected with Western blot and realtime PCR.Results In 3T3-L1 adipocytes differentiation process,expressions of LC3,Beclin 1,and TNFAIP3 continued to rise,expression of BIRC5 was high to low,FEZ1 continued at a low level.In mice,expressions of LC3,Beclin 1,YWHAQ,FEZI,BAD,WAC,TNFAIP3,and GOPC in visceral fat were significantly higher than those in subcutaneous fat,while expression of SLAMF1was higher in subcutaneous fat.Conclusion Autophagy plays an important role in 3T3-L1 differentiation process and may be tissue-specific in visceral fat and subcutaneous fat.

Objective To characterize the molecular characteristics of hemagglutinin (HA) and neuraminidase (NA) of influenza B viruses isolated in Jiangsu province,2011.Methods Thirteen strains of influenza B virus in different areas and epidemic period in Jiangsu province,2011 were selected for whole-genome sequencing,and analysis of molecule epidemic characteristics for HA and NA was carried out by bioinformatics method.Results Of the 13 randomly selected influeuza B strains,10 strains were assorted to Victoria lineage strains with NA genes from Yamagata lineage,and 3 strains belong to Yamagata lineage.Compared nucleotide and amino acid sequences of HA and NA genes with their vaccine strains respectiuely,196/197 glycosylation site appeared on HA1 gene in Yamagata/Victoria isolates virus.Conclusion Both B/Victoria and B/Yamagata lineage viruses co-circulated in Jiangsu province,and reassortant virus of Victoria lineage were predominant virus.

0bjective To investigate the pattern of disease relapse and prognostic risk factor of patients with cervical carcinoma and pelvic lymph node metastasis.Methotis A total of 124 cases of International Federation of Gynecology and Obstetricfi(FIG0)I bl-Ⅱ a cervical carcinoma with pelvic node metastasis who were treated at the Cancer Center of Sun Yat-sen University during January 1994 to December 2001 were selected for this study.Prognosis and recurrence were retrospectively analyzed using the clinico.pathological data.Results The overall 5 year flun,ival and 5 year disease-free survival were 63.3% and 61.4%.respectively.Overall recurrence rate was 39.5%(49/124),among which intra-pelvic relapse (61.0%,25/41)was significantly more common than extra-pelvic relapse(31.7%,13/41;P=0.008).Multivariate analysis identified involvement of common iliac node as an independent prognostic factor(P=O.035).According to this factor,node-positive patients could be divided into low risk group(without common iliac node involvement,104 cases)and hiSh risk group(with common iliac node involvement,20 cases).The 5 year disease-free survival were 69.4%and 24.5%respectively,with a significant difference(P=0.003).Intra.pelvic relapse was observed in 22.1%(23/104)of low risk and 25.0% (5/20)of high risk group respectively,with no significant difference(P>0.05).However extra-pelvicrelapse wag seen in 7.7%(8/104)of low risk and 40.0%(8/20)of hish risk group,with a significant difference(P<0.01).Conclusions Common iliac node involvement is a significant factor influencing the prognosis of patients with cervical carcinoma and pelvic lymph node metastasis.Patients with positive common iliae nodes have significantly decreased 5 year disease.free survival and hishcr extra-pelvic disease recurrence rates compared with those whose common iliac nodes are negative.These findings provide impo.rtant data for design of individualized treatment mode.

Objective:To explore the possibility of using peritoneal alternatively activated M2 macrophages to prevent rejection after islet allotransplantation in a murine model.Methods:Peritoneal monocytes from C57BL/6 mice were induced and modulated to M2 and M0 macrophages in vitro,then the phenotype of macrophage was assessed by flow cytometry.C57BL/6 mice were induced diabetic by streptozotocin (STZ) injection and transplanted with islets isolated from BALB/c mice under the left kidney capsule.The recipients were randomly divided to 3 groups (n=8).A total of 2.5× 106 M2 macrophages were injected intravenously at 0,3,7 d after transplantation in islet+M2 group;2.5×106 M0 macrophages were injected intravenously at 0,3,7 d after transplantation in islet+M0 group;the mice in islet+PBS group were injected with PBS.Blood glucose was monitored after transplantation.On day 10 after transplantation,2 recipients in each group were randomly selected and sacrificed,and the left kidneys were resected for pathological examination.Results:Achievement of euglycemia was significantly prolonged after islet transplantation in the islet+M2 group than that in the other two groups (P＜0.01).The median survival time of islet allografts in the islet+PBS group,the islet+M0 group,and the islet+M2 group were 6.5 (4-10),7.5 (4-10),and 24(＞ 15) d,respectively.Pathological examination also showed that the grafts in islet+M2 group remained an intact structure with positive insulin stain and no apparent lymphocytes infiltration,while the graft was rejected in other 2 groups with negative insulin stain and massive lymphocytes infiltration.Conclusion:Peritoneal alternatively activated M2 macrophages can prevent rejection after islet allotransplantation,prolong the survival time of islet allografts and enhance the tolerance of the recipient to blood glucose in mice.

Objective: To observe the relieving cough and reducing sputum effects of total alkaloid in Atalantia buxifolia and evaluate the safety preliminarily.Methods: The relieving cough and reducing sputum effects of total alkaloid in Atalantia buxifolia were studied by the cough model caused by the irritation of ammonia water and the phenol red output of trachea in mice.The acute toxicity test and maximum tolerance test were carried out to evaluate the safety.Results: The total alkaloid in Atalantia buxifolia at low dose could obviously prolong cough incubation period and decrease cough times in mice, and that at high dose could significantly increase the secretion of phenol red in respiratory tract, and compared with those in the blank group, the differences were statistically significant (P<0.05).In the acute toxicity test, no death showed after the administration with maximum tolerance dosage, and the rate of weight growth had no difference between the blank group and the model group (P>0.05).Conclusion: The relieving cough and reducing sputum effects of total alkaloid in Atalantia buxifolia are notable in the cough model caused by the irritation of ammonia water and the phenol red output of trachea in mice.The maximum tolerable dose test shows the total alkaloid in Atalantia buxifolia is safe.

Objective To investigate the effect of adenovirus-bone morphogenic protein 9 ( Ad-BMP9 ) on osteogenic differentiation of immortalized calvarial mesenchymal progenitor cells ( iCALs ) .Methods iCALs were infected with adenoviral vectors encoding BMP-9 or green fluorescent protein ( GFP) and the early osteogenic differentiation was assessed by detecting alkaline phosphatase (ALP) activity after being cultured for 3, 5 and 7 days.14 days after infection, alizarin red S staining was performed to study the formation of osteogenic calcium nodules .The expression of osteogenic marker genes Runx2 and OCN was assessed by quantitative real-time ( RT )-PCR and Western blotting .Results Significant increases in ALP activity and in the expressions of Runx 2 and OCN were detected in BMP-9 treated iCALs compared with GFP-treated cells(P<0.05).Meanwhile, alizarin red S staining showed that more mineralized nodules were found in the BMP-9 induced group .Conclusion BMP-9 can promote the osteogenic differentiation of iCALs .

Aim To investigate the neuroprotective effect of verbascoside,one of the phenylethanoids isolated from the stems of Cistanche salsa,on MPP+-induced injury in SHSY5Y cells.Methods SHSY5Y cells were exposed to various doses of verbascoside for 12 h,and then treated with 200 ?mol?L-1 MPP+ for 24 h.The cell viability was observed with MTT assay;reactive oxygen species,the mitochondrial membrane potential and the percentage of apoptosis were measured by flowcytometer;the activation of caspase-3 was measured with the caspase-3 activity assay kit;the expression of Bcl-2 was measured with Western blot.Results Following treatment with MPP+ for 24 h,MPP+ induced a significant decrease of cell viability;apoptosis percentage were 38.9%;accumulation of intracellular ROS,increase of caspase-3 activity and the decreases in mitochondrial membrane potential were detected.However,pretreatment with verbascoside (0.1,1 or 10 mg?L-1) for 12 h exhibited cytoprotective effects in a dose-dependent manner.Verbascoside obviously enhanced cell viability,and significantly reduced the number of cells labeled with Annexin-V.The percentage of apoptosis neurons was significantly decreased to 29.5%,15.3% and 8.6% respectively.Flowcytometer showed the verbascoside attenuated the accumulation of ROS and the MPP+-induced collapse of mitochondrial membrane potential in SHSY5Y cells.And significant decreases were detected in caspase-3 activity compared with the MPP+-treated cells at the same time point.Moreover,pretreatment with verbascoside promoted the expression of Bcl-2.Conclusions verbascoside had the neuroprotective capacity to antagonize MPP+-induced apoptosis in SHSY5Y cells,and might be useful in treating Parkinson disease.

AIM: To investigate the neuroprotective effect of tubuloside B,one of the phenylethanoids isolated from the stems of Cistanche salsa,on H2O2-induced injury in PC12 cells.METHODS: PC12 cells were exposed to various doses of tubuloside B for 12 h,then treated with H2O2 at concentration of 100 ?mol/L for 24 h.The cell viability was observed with MTT assay.Reactive oxygen species and the mitochondrial membrane potential were measured with laser scanning confocal microscopy(LSCM).The DNA content and percentage of apoptosis were assayed by DNA agarose gel electrophoresis and flow cytometry.The activation of caspase-3 was detected with the caspase-3 activity assay kit.RESULTS: Following treatment with H2O2 for 24 h,H2O2 induced a significant decrease in cell viability;DNA ladder was observed and apoptosis percentage was as high as 48.0%.Accumulation of intracellular ROS,increase in caspase-3 activity and the decrease in mitochondrial membrane potential as indicated with the decrease of red/green ratios(from 5.97 to 0.41) were detected.However,pretreatment with tubuloside B(1,10 or 100 mg?L-1) for 12 h exhibited cytoprotective effects in a dose-dependent manner.Tubuloside B obviously enhanced the cell viability,reduced formation of the DNA ladder,and significantly reduced the number of cells labeled with Annexin-V.The percentage of apoptosis/necrosis neurons was significantly decreased to 30.9%,18.3% and 6.2%,respectively.LSCM showed that the tubuloside B attenuated the accumulation of ROS and the H2O2-induced collapse of mitochondrial membrane potential in PC12 cells.The significant decrease in caspase-3 activity was detected,compared to the H2O2-treated cells at the same time point.CONCLUSION: Tubuloside B has the neuroprotective capacity to antagonize H2O2-induced apoptosis and injury in PC12 cells,indicating it may be useful for treating some neurodegenerative diseases.