In order to explore the effect and mechanisms of tumor necrosis factor-alpha (TNF-alpha) on the activity of the acyl coenzyme A: cholesteryl acyltransferase (ACAT), THP-1 monocytes were cultured and induced to differentiate into macrophages with phorbol ester. TNF-alpha (60 ng/mL) was added at different time points into the macrophage-containing medium and the ACAT enzyme activity was measured by quantifying the incorporation of [1-(14)C] oleoyl CoA into cholesteryl esters. The expression of ACAT-1 protein and mRNA was respectively detected by Western blotting and RT-PCR in THP-1 macrophages 24 h after treatment with TNF-alpha (60 ng/mL). The results indicated that ACAT activity in THP-1 macrophages treated with TNF-alpha was increased in a time-dependent manner. The expression levels of ACAT-1 protein and mRNA were significantly increased in THP-1 macrophages after treatment with TNF-alpha (P<0.05). It was suggested that TNF-alpha could increase the activity of ACAT in THP-1 macrophages by up-regulating the expression of ACAT-1 gene.

Drug resistance of Acinetobacter baumannii ( Ab) in surgical ICU from January 2008 to December 2009 was investigated retrospectively. Total 114 clinical strains of Ab were isolated from surgical ICU and 92 strains were from respiratory tract (80.7% ). The prevalence rate of multiresistant Ab in 2009 was higher than that in 2008 (75. 7% vs 50. 0% , x2 = 7. 703, P = 0. 006). The results indicate that to monitor and control drug resistant of Ab constantly is important for the prevention of nosocomial infection.

At present,transplantation has been the predominant way to solve most of the end-stage diseases,ensued by the use of immunosuppressive drugs.Since the immunosuppressive drugs have narrow therapeutic index,the blood drug concentration is needed to mornior.Pharmacogenetics is one subject which focuses on the interaction between gene and the metabolism of the drug,providing great help for designing the regime of achieving the target drug concentration.Meanwhile,it facilitates the realization of individual therapy.This review thus focuses on the latest advancement on the pharmacogenetics of those immunosupprressants,hoping to provide help for the treatment.

OBJECTIVEThe study was aimed to investigate the inhibitory effect and mechanism of astragaloside IV (ASI) on the activation of microglial cells.

METHODAfter pre-incubated with ASI for 2 h, microglial cells BV-2 were stimulated with interferon-γ (IFN-γ) for 1. 5 h and 24 h, respectively. Secretion of nitric oxide (NO) in the medium was measured by Griess method. Production of tumor necrosis factor alpha (TNF-α) was detected by ELISA approach. Cellular gene expressions of CD11b, TNF-α, interleukin 1β (IL-1β) and induced nitric oxide synthase (iNOS) were examined by quantitative-PCR analysis. Total and phosphorylation of STAT1, IκB and NF-κB was analyzed by Western blot method.

RESULTASI could significantly inhibit the increased secretion of TNF-α and NO from BV-2 cells upon IFN-γ stimulation (P < 0.001). Further study showed that ASI significantly down-regulated gene expression of IL-1β and TNF-α (P < 0.01, P < 0.05) and exhibited a trend to reduce that of iNOS. IFN-γ and ASI have no obvious effect on gene expression of CD11b. Moreover, ASI inhibited the phosphorylation of STAT1, IκB and NF-κB elicited by IFN-γ stimulation.

CONCLUSIONASI could restrain microglial activation through interfering STAT1/IκB/NF-κB signaling pathway, reducing gene expres- sion of IL-1β and TNF-α, and thus inhibiting the production of proinflammatory mediators such as NO and TNF-α.

Animals ; Astragalus Plant ; chemistry ; Drugs, Chinese Herbal ; pharmacology ; I-kappa B Proteins ; genetics ; metabolism ; Interferon-gamma ; genetics ; metabolism ; Mice ; NF-kappa B ; genetics ; metabolism ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase Type II ; genetics ; metabolism ; STAT1 Transcription Factor ; genetics ; metabolism ; Saponins ; pharmacology ; Signal Transduction ; drug effects ; Triterpenes ; pharmacology

Objective To investigate the effects of lipopolysaccharide(LPS) on myelin phagocytosis during Wallerian degeneration after early peripheral nerve injury in rats.Methods Fifty male Wistar rats were recruited and randomly divided into LPS group(n=20),model group(n=20) and sham group(n=10).The right sciatic nerves of rats in the LPS and model groups were cut and sutured end-to-end,while the sciatic nerve of sham group rats were only exposed.Immediately after surgery,the rats in LPS group were given microinjections of LPS(2 g/L) into the surgical site in a final volume of 1 μL,and the rats in other two groups were injected with the same volume of saline.The sciatic nerves were taken at 1.5 h,24 h and 7d after surgery.Real-time quantitative PCR(qRT-PCR) was applied to detect the dynamic expressions of IL-1β mRNA and MCP-1 mRNA.Immunofluorescence staining was used to test the expression of CD68+ macrophages in sciatic nerves.HE staining was used to observe the pathological alterations of sciatic nerves tissue.ORO staining was used to observe sciatic nerves demyelination.LFB staining was used to detect the sciatic nerves myelin.Sciatic function index was used to evaluate the recovery of motor function in rats.Results Compared with the model group,qRT-PCR indicated that the expression of IL-1β and MCP-1 from LPS group were increased at 1.5 h and 24 h after surgery(P<0.001, P<0.001),respectively.Compared with the model group,the expression of CD68+ cells was increased significantly at 7th day after surgery(P<0.05).Histological examination showed that compared with the model group,a lot of inflammatory cells and Schwann cells were found at sciatic nerve stump in the LPS group at 7th day after operation.ORO staining showed that the degree of demyelination in the LPS group was higher than that in the model group.LFB staining showed that the sciatic nerve stump demyelination appeared in both model group and the LPS group at 7th day after operation,but compared with the model group,myelin debris clearance in the LPS group was significantly accelerated(P<0.05).Finally,compared with the model group,the SFI in the LPS group was increased significantly at 20 d after surgery(P<0.05).Conclusions The results confirm that LPS is possible to manipulate the innate immune response to accelerate myelin clearance during Wallerian degeneration after early peripheral nerve injury in rats.

Objective To summarize the clinical characteristics of female pseudohermaphroditism (FPH) in the publication from 1994 to 2009 and the cases diagnosed and treated in our hospital from 2000.Methods We employed Chinese and English name of female pseudohermaphroditism as a key word separately to retrieve published articles of FPH during 1994 to 2009.A meta-analysis was performed together with the ten cases in our hospital.Results Among 342 cases involved in this study,only twenty-seven children were diagnosed immediately after birth and the other 315 cases at the age of 1 month to 59 years with median age of 29.54 years.The etiological factors were mostly congenital adrenal hyperplasia (70.47%).Clinical manifestations were advanced bone age (95.52%),clitoromegaly (84.14%) and early virilization (77.24%).The treatment mainly included drug and operation.Early diagnosis and available therapy were very important for the prognosis.Conclusion FPH is not rare,so we should improve our knowledge for its early diagnosis,good intervention and final better prognosis.

Objective To explore the factors influencing the recovery of ability in the activities of daily living (ADL) after intracerebral hemorrhage. Methods A total of 108 patients with intracerebral hemorrhage admitted for rehabilitation to the rehabilitation medicine department of Huashan Hospital between January 2007 and June 2011 were studied.Twelve items of clinical data were collected with regard to the patients' medical history,physical status,modified Barthel index (MBI) score and Brunnstrom stage at admission.Functional status was classified according to the MBI scores and Brunnstrom stages assessed at admission and before discharge.Linear regression analysis was used to relate the variables. Results After rehabilitation,the MBI scores and Brunnstrom stages had improved relative to the scores at admission.Factors influencing the MBI improvements included the intervention timing of rehabilitation and the course of therapy employed. Conclusions It is very important to comprehend the factors influencing the recovery of ADL ability after cerebral hemorrhage in order to design effective rehabilitation strategies,better predict functional outcomes and improve patients' ADL ability effectively.

Background Arresting the overexpression of vascular endothelial growth factor (VEGF) will be a new approach to the inhibition of neovascularization.RNA interference (RNAi) can inhibit the expression of specific gene,and its application in eye has little interference to other gene expression.Objective This study was to investigate the effect of small interference RNA (siRNA) targeting VEGF on the expression of VEGF and retinal neovascularization in oxygen-induced retinopathy (OIR) model.Methods psi-HITM/enhanced green fluorescent protein (EGFP)/VEGF siRNA was designed and prepared in vitro.Mouse endothelioma (EOMA) were cultured in DMEM without antibiotic and divided into 5 groups.The cells were incubated in DMEM only in the blank control group;while 1 μl of LipofectamineTM 2000 + psi-HITM/EGFP,1 μl LipofectamineTM 2000 + 40,50 or 60 nmol/L of psi-HITM/EGFP/VEGF siRNA was added into DMEM in the negative control group and siRNA groups,respectively.The expression of VEGF mRNA and protein was detected by real time PCR (RT-PCR) and Western blot.The optimal effective concentration of VEGF siRNA was assessed.OIR models were established in 48 7-day-old C57BL/6J mice by raising them at an oxygen concentration of (75±2) ％ for 5 days and then to normal air.The mice were randomized into the model group,null vector group and VEGF siRNA group.1 μl of a mixture of psi-HITM/EGFP or VEGF siRNA (60 nmol/L) and LipofectamineTM 2000 was intravitreally injected,respectively,in the null vector group and VEGF siRNA group.The normal mice were used as the normal control group.Expression of VEGF mRNA and protein in the mouse retinas was detected by RT-PCR and Western blot,respectively,and FITC-dextran stretched retinal preparation was examined to evaluate the neovascularization,and retinal sections were examined to quantify the number of vascular endothelial cell nuclei extending beyond the internal limiting membrane (ILM).Results The in vitro transfection test showed that the expression of VEGF mRNA and protein in the EOMA cells was significantly different among the blank control group,negative control group and 40,50,60 nmol/L VEGF siRNA groups (F =148.890,P ＜ 0.001; F =306.960,P ＜ 0.001),and the expression of VEGF mRNA was lower in different concentrations of VEGF siRNA groups than that in the blank control group (t=73.950,119.890,156.480,all at P＜0.001).Also,the expression of VEGF protein was less in different concentrations of VEGF siRNA groups than that in the blank control group (t =15.452,23.344,42.119,all at P＜0.001).The optimal inhibitory concentration of VEGF siRNA was 60 nmol/L.In vivo study determined that compared to the model group and null vector group,the non-perfusion zones and neovascular net in the retina were decreased,and the number of vascular endothelial cell nuclei extending beyond the ILM was less in the VEGF siRNA group.The relative expression level of VEGF mRNA in the retinas was 1.23±0.18,4.02±0.16,3.98±0.19 and 1.98±0.12 in the normal control group,model group,null vector group and VEGF siRNA group,respectively,with a significant difference among them (F=369.780,P＜0.001),and the relative expression levels of VEGF mRNA in the model group and null vector group were higher than that in the normal control group (t=37.880,37.336,both P＜0.001),and the expression of VEGF mRNA in the VEGF siRNA group declined by 50.8％ (t=10.183,P＜0.001).The difference in the expression levels of VEGF protein also was assayed among the various groups (F=408.980,P＜0.001),and VEGF level in the retina was lowered by 68.0％ in the VEGF siRNA group compared to the model group (t =11.473,P＜0.001).However,VEGF level in the VEGF siRNA group remained at a high level in comparison with the normal control group (t =2.422,P＜0.001).Conclusions Intravitreal injection of VEGF siRNA can attenuate retinal neovascularization by effectively downregulate the expression VEGF mRNA and protein in the retina.

OBJECTIVETo optimize the synthetic pathway and fermentation process of yeast cell factories for production of oleanoic acid.

METHODUsing the DNA assembler method, one copy of Glycyrrhiza glabra beta-amyrin synthase (GgbAS), Medicago truncatula oleanolic acid synthase (MtOAS) and Arabidopsis thaliana cytochrome P450 reductase 1 (AtCPR1) genes were introduced into Saccharomyces cerevisiae strain BY-OA, resulting in strain BY-20A. YPD medium with different glucose concentration were then used to cultivate strain BY-2OA.

RESULTIncreasing gene copies of GgbAS, MtOAS and AtCPR1 resulted in increased beta-amyrin and oleanolic acid production. The strain BY-2OA produced 136.5 mg x L(-1) beta-amyrin and 92.5 mg x L(-1) oleanolic acid, which were 54% and 30% higher than the parent strain BY-OA. Finally, the titer of oleanolic acid increased to 165.7 mg x L(-1) when cultivated in YPD medium with 40 mg x L(-1) glucose.

CONCLUSIONProduction of oleanoic acid increased significantly in the yeast strain BY-2OA, which can provide the basis for creating an alternative way for production of oleanoic acid in place of extraction from plant sources.

Biomass ; Biotechnology ; methods ; Dose-Response Relationship, Drug ; Fermentation ; Glucose ; pharmacology ; Oleanolic Acid ; biosynthesis ; Saccharomyces cerevisiae ; cytology ; drug effects ; metabolism

AIMTo explore the effect of cerebellar fastigial nuclei (FN)on lymphocyte function and the pathway mediating the effect.

METHODSKainic acid (KA) was microinjected into bilateral FN of rats to destroy neuronal bodies in the FN. On the eighth day after the surgery, lymphocyte percentage in the peripheral blood and level of sheep red blood cell(SRBC)-specific IgM antibody in the serum were measured by using blood corpuscle counter and enzyme-linked immunosorbent assay (ELISA), respectively.A technology of electrolytic lesion was used to destroy the projections of cerebellar FN neurons to hypothalamus in decussation of superior cerebellar peduncle(xscp).

RESULTSOn the eighth day after the microinjection of KA into the bilateral FN of rats, the Nissl-stained neuronal bodies in the FN disappeared and glia could proliferated within the damaged FN. In the nuclei close to FN, the interposed nuclei and the dentate nuclei, Nissl-stained neurons still could be seen. On the control cerebellar sections, in which FN was infused with saline, we could see the normal Nissl-stained neurons in the FN and the other two nuclei.On day 8 following the effective FN lesions, both the lymphocyte percentage in the peripheral blood and the level of anti-SRBC IgM antibody in the serum were significantly increased in comparison with those of control rats infused with saline in the FN. On the eighth day after electrolytic lesion of the fibres in xscp, the FN-hypothalamic projections were damaged and there were no visible BDA-positive endings in hypothalamus. Meanwhile, both the lymphocyte percentage in the peripheral blood and the level of anti-SRBC IgM antibody in the serum were remarkably enhanced relative to those of control rats with sham lesion of xscp.

CONCLUSIONThe electrolytic lesion of the FN-hypothalamic projections in xscp causes an enhancement of lymphocyte function similar to that of KA lesions of neuronal soma in the FN. These findings suggest that the cerebellohypothalamic projections participate in mediating the modulation of lymphocyte function by the cerebellum.

Animals ; Cerebellar Nuclei ; immunology ; injuries ; Female ; Hypothalamus ; immunology ; physiology ; Kainic Acid ; Lymphocyte Count ; Lymphocytes ; cytology ; immunology ; Male ; Neural Pathways ; immunology ; physiology ; Neuroimmunomodulation ; physiology ; Rats ; Rats, Sprague-Dawley