The recombinaut porcine insulin precursor(PIP)produced by Pichia pastoris in shake-flask and 501.fermenter was investigated respectively.The results indicated that 60h induction time length and 2.0%～2.5% methanol addition every day was optimum in shake- flask.The process in 50L fermenter was consisted of batch,feed-batch and induction phases.The relationship between dry cell weight(y) and culture time (t) in growth phase(batch and feed-batch phase)could be described by model y=0.6525e~(0.1907t).Glycerol and ammonia were almost used for cell growth and maintain,and no by-product was observed in batch and fed-batch phase Only 80% ammonia and 70% methanol were used by cell in induction phase.By comparison the results of shake-flask and 50L fermenter,it was concluded that the limit- ing factor in the fermentation of shake-flask and 50L fermenter was dissolved oxygen(DO)and.carbon source,respectively.When scaling the result of shake-flask to 501.fermenter,the control strategy was adapted for 50L fermenter by increasing the feed rate of methanol and the maximum PIP concentration reached 1.72 g/L.

Objective To explore the apoptosis and proliferation of peripheral blood lymphocytes(PBLs) from primary nephrotic syndrome(PNS) and effects of dexamethasone(Dex) on them.Methods Minimal change NS(MCNS), non-minimal change NS(NMCNS) and healthy children were involved in this study. PBLs were cultured in vitro with Dex or PHA+Dex or without PHA and Dex. Apoptosis of PBLs was measured by propidium iodide(PI) staining; The effects of Dex at different concentrons on PBLs′proliferation were investigated by 3H-TdR incorporation.Results The apoptotic rate of vacuity group in MCNS was lower compared with NMCNS and healthy controls(P

Objective To study the expression of membrane glucocorticoid receptors(mGR), the correlation between mGR and glucocorticoid(GC)'s effects on apoptosis and proliferation of peripheral blood lymphocytes(PBLs) in minimal change nephrotic syn-drome(MCNS)and the influence of GC on mGR. Methods MCNS, nonminimal change NS(NMCNS) and healthy children were involved in this study. Indirect immune fluorescence and flow cytometry were used to examine the percentages of positive mGR lymphocytes; the apoptosis of PBLs was measured by propidium iodide(PI) staining and the proliferation of PBLs was investigated by fa -TdR incorporation. Results 1.mGR expression in MCNS was higher than that in NMCNS and healthy control,but it was reduced after clinical GC therapy(P

A strain of high-efficiency bioflocculant-producting bacteria,which was named TJ-3,was screened from sewage sludge.The strain was gram-negative and rod-shaped in physiological biochemical test and was identified as Pseudomonas aeruginosa by 16S rDNA Sequencing.According to the growth curve of the TJ-3 strain,the growth stabilization period is long so that Microbial Flocculants(MBF) production is stable.The MBF produced by the TJ-3 strain is able to flocculante kaolin suspension with 98.2%.The MBF activity distribution tests show that most of the active components of the bioflocculant exist in deposition after centrifufation.When pH is 8.5,1%(quality fraction) CaCl2 Solution dosing quantity is 3.5 mL,bioflocculant dosing quantity is 1.5 mL in 100 mL kaolin suspension,the bioflocculant has an optimal flocculation.

OBJECTIVETo investigate whether resveratrol (RES) plays a protective role in hypothermic preserved isolated rat hearts and whether it is mediated by regulation of silent information regulator protein-1 (Sirt-1) expression.

METHODSThe Langendorff model of isolated rat heart was used. After stored in different Celsior solution at 4 degrees C for 9 h, SD rat hearts were randomly divided into 7 groups: blank control group;9 h group (soley hypothermic preservation for 9 h); RES group (3, 10, 30 micromol/L RES treatment plus hypothermic preservation for 9 h ), niacinamide (NAM) group (40 micromol/L NAM added in Celsior solution plus hypothermic preservation for 9 h), RES + NAM group (30 micromol/L RES and 40 micromol/L NAM were added in Celsior solution plus hypothermic preservation for 9 h). The morphological changes of cardiomyocytes were detected by the HE staining with the light microscope. The mRNA and protein expression levels of Sirt-1 were detected by Real-Time PCR and Western blot respectively.

RESULTS(1) Compared with the blank control group, myocardiocytes were injured remarkably in the 9 h group and the Sirt-1 mRNA and protein expression levels were decreased significantly (P < 0.01); (2) Compared with the 9 h group, rat myocardial injury was alleviated gradually in 3, 10, 30 micromol/L RES group and the Sirt-1 mRNA and protein expression levels were increased in a dose-dependent manner (P < 0.05); (3) The above protective effects of RES were attenuated by Sirt-1 inhibitor NAM.

CONCLUSIONRES can protect myocardiocytes from injury caused by long range hypothermic preservation and this protective effect maybe mediated by upregulation of Sirt-1 expression.

Animals ; Cryopreservation ; Heart ; drug effects ; Male ; Organ Preservation ; Rats ; Rats, Sprague-Dawley ; Sirtuin 1 ; metabolism ; Stilbenes ; pharmacology

Background: We found microRNA (miR)-1246 to be significantly differentially expressed between severe active alopecia areata (AA) patients and healthy individuals. Objective: To explore the role and mechanism of miR-1246 in severe AA. Methods: Expression of miR-1246, dual-specific tyrosine phosphorylation-regulated kinase 1A (DYRK1A), and nuclear factor of activated T cells 1c (NFATc1) in peripheral CD4+ T cells and in scalp tissues of patients were detected using RT-qPCR, Western blot, and immunohistochemistry assays. Peripheral CD4+ T cells from the AA patients were transfected with lentiviral vectors overexpressing miR-1246. RT-qPCR and Western blot analysis were used to measure mRNA or protein expression of retinoic-acid-receptor-related orphan nuclear receptor gamma (ROR-γt), interleukin (IL)-17, DYRK1A, NFATc1, and phosphorylated NFATc1. Flow cytometry was used to assay the CD4+ IL-17+ cells proportion. ELISA was used to measure cytokine levels. Results: miR-1246 levels decreased and DYRK1A and NFATc1 mRNA levels significantly increased in the peripheral CD4+ T cells and scalp tissues of severe active AA samples.NFATc1 protein expression was also significantly increased in the peripheral CD4+ T cells but not in the scalp tissues. NFATc1 positive cells were mainly distributed among infiltrating inflammatory cells around hair follicles. In peripheral CD4+ T cells of severe active AA, overexpression of miR-1246 resulted in significant downregulation of DYRK1A, NFATc1, ROR-γt, and IL-17 mRNA and phosphorylated NFATc1 protein, as well as a decrease in the CD4+ IL-17+ cells proportion and the IL-17F level. Conclusion miR-1246 can inhibit NFAT signaling and Th17 cell activation, which may be beneficial in the severe AA treatment.

Eleven compounds were isolated from the culture of Streptomyces sp. CPCC 202950 by a combination of various chromatographic techniques including column chromatography over macroporous resin HP-20, MCI, and reversed-phase HPLC. Their structures were identified as 1H-pyrrole-2-carboxamide(1),5'-deoxy-5'-methylthioinosine(2), vanillamide(3), trans-3-methylthioacrylamide(4), 1,2,3,4-Tetraydro-1H-pyrido[3,4-b]indole-3-carboxylic acid(5), cyclo(L-pro-L-tyr) (6), N-[2-(4-hydroxyphenyl)]ethylacetamide(7), benzamide (8), cyclo ('L-leucyl-trans-4-hydroxy-L-proline)(9), cyclo-(Phe-Gly) (10), and tryptophan (11). Among them, compounds 1 and 2 were new natural products. In the preliminary assays, none of the compounds exhibited obvious inhibition of HIV-1 protease activity (IC50 > 10 micromol x L(-1)).
Culture Media ; chemistry ; metabolism ; HIV Protease ; analysis ; HIV Protease Inhibitors ; chemistry ; isolation & purification ; Molecular Structure ; Spectrometry, Mass, Electrospray Ionization ; Streptomyces ; chemistry ; metabolism

By using a cell-based high throughput screening model for the CLA-1 up-regulator, Streptomyces 203909 was found to produce up-regulator of CLA-1. A novel trichostatin analogue was isolated from the rice fermentation of Streptomyces sp. CPCC 203909by a combination of various chromatographic techniques including column chromatography (CC) over silica gel, flash C18 CC, and reversed-phase HPLC. Its structure was identified as (-)-(R,2E,4Z)-7-[(4'-dimethylamino) phenyl]-4,6-dimethyl-7-oxohepta-2,4-dienoyl-L-glutamine (1) by the spectroscopic and chemical methods, and combination with the CD spectroscopy and Marfey's method. In the prelimi- nary assays, Compound 1 showed cytotoxicity against human embryonic kidney 293 cell line with IC50 value 35.3 [µmol · L(-1).
Cell Survival ; drug effects ; Fermentation ; Hep G2 Cells ; Humans ; Hydroxamic Acids ; chemistry ; isolation & purification ; metabolism ; pharmacology ; Molecular Structure ; Streptomyces ; chemistry ; metabolism

Objective To retrospectively study revisions for infected total hip replacements in 30 cases and discuss the bacteriological characters of the infected total hip replacements,difficulties and strategies in the revision.Methods Thirty revisions of infected total hip replacements were reviewed retrospectively.There were 12 males and 18 females,with mean age of 62.5 years(31-86 years)at revi- sion surgery.Infection was presented one month to four years(mean seven months)after THA operation. The diseases for initial operation included femoral neck fractures in 12 cases,femoral head necrosis in 11,hip osteoarthritis in five and rheumatoid arthritis in two.Twelve eases were treated by one-stage revi- sion and 18 by two-stage revision.Results Before the revision operation,the hip infection were diag- nosed by bacterial culture in 18 cases including five with Staphylococcus epidermidis,four with Staphylo- coccus aureus and nine with other bacteria.Bacteria growth appeared on the specimens from 23 hip joints during the revision surgery but not on the specimens from seven hip joints.Of 12 one-stage revisions,10 cases were followed for mean 16 months,which showed infection recurrence in two eases.Of 18 two-stage revisions,13 cases were followed for mean 20 months,which showed one case with infection recurrence. The mean Harris hip score was improved from preoperative 44 to 84 at follow-up.Conclusions 1) The main bacteria in the infected hip are antibiotic resistant Staphylococcus.2)Because the revision op- eration is difficult,careful preparation before revision is important for success.The fresh surgeon should not attempt.3)The revision strategies should vary according to specific status of the cases.The infection recurrence rate is lower when using a two-stage revision strategy.4)Application of antibiotic bone cement can help improve treatment effect and facilitate functional recovery of the joints.5)The scientific rehabil- itation after operation is very important to functional recovery.

Objective To report the experience of the application of microsurgery in joint replace- ment.Methods There were 22 cases,10 cases with segmental acetabular defects treated with the pedicle sartorius muscle iliac bone grafting,5 cases with vascular repair following major vascular injury of extremity during operation,6 cases with neural repair following neural injury during operation,1 case with serious injury reconstruction by elbow joint replacement and free flap.Results The operations succeeded in 22 cases without any postoperative infection.The mean follow-up was 40.1 months (3-72 months) in 22 cases,in which the joint function improved and the operative result was satisfactory with no joint pain.Conclusion Microsurgical technique can reconstruct bone and tissue defect effectively in joint replacement.