ObjectiveTo study the expression of adhesion mol ecules (CD 54 、CD 62p ) on diabetic neuropathy in the STZ induced rats and the effect of cilostazol treatment.MethodsSD rats were divided into four groups: normal control, diabetes control, insulin and cilostaz ol treated group. Sciatic nerve conductive velocity, the level of CD 54 /CD 62p on the surface of mononuclear cell/platelet were examined, and ultrast ructure of sciatic nerve was observed.ResultsCilostazol incr eased sciatic nerve conductive velocity significantly [DM=(20.3?2.2) m/s vs ci lostazol=(28.9?7.9)m/s,(P

According to the sequence of SmLEA gene,a DNA fragment of 1 038 bp upstream of the coding sequence of SmLEA gene was amplified by DNA walking with the genomic DNA of Salvia miltiorrhiz as the template.Sequence analysis showed that the fragment contained some putative cis-elements relating to abiotic stress,ABA,seed specific expression.So the S.miltiorrhiz seedings was treated with 100?mol/L ABA,200mmol/L NaCl,4℃,and subjected to dehydration.The real-time PCR showed that expression levels of SmLEA was increased obviously,which was in accordance with the sequence analysis.

The tumor microenvironment (TME), a dynamic and complex local environment, interacts with the tumor cells and is closely related to tumor growth, metastasis, immune escape and drug resistance. Thus, targeting the TME has been a worldwide focus in cancer therapy. Many natural products possess the advantages of multiple targets, multiple pathways and wide pharmacological functions, and are the main source of antitumor drugs. In recent years studies have found that some natural products had advantageous effects on the TME. In this review, we summarize the components and functions of the TME and some natural products that target the TME, with references to the drug therapy of cancer.

Objective To study to the effect of 1064-nm Q-switched Nd:YAG laser irradiation on the melanogenesis in a human epidermal melanocyte line PIG. Methods Cultured PIG cells were irradiated with 1064-nm Q-switched Nd:YAG laser (Medlite C6) at different energy densities for 10 times. After additional culture for various durations, cell viability was detected by MT assay, tyrosinase activity by dopa oxidation assay, mRNA and protein expressions of tyrosinase, tyrosinase related protein (TRP)-1 and TRP-2 by real-time quantitative fluorescent RT-PCR and Westen blotting respectively, Results The irradiation with Q-switched Nd:YAG laser at energy densities from 1 to 3 J/cm2 had no obvious effect on the viability of PIG cells. After irradiation with Nd:YAG laser at 1 J/cm2, PIG cells showed a significant increase in the tyrosinase activity,mRNA expressions of tyrosinase and TRP-1 compared with unirradiated cells (0.563 ± 0.014 vs 0.501 ±0.019, 1.40±0.11 vs 1.0, 1.28 ± 0.03 vs 1.0, all P＜ 0.05), but both the mRNA (0.91 ± 0.17 vs 1.0, P＞0.05) and protein expressions of TRP-2 experienced no significant changes before and after the irradiation.However, a significant decrease was noted in PIG cells irradiated with Nd:YAG laser at 3 J/cm2 in tyrosinase activity, mRNA and protein expressions of tyrosinase (0.70 ± 0.02 vs 1.0, 0.64 ± 0.05 vs 1.0, both P ＜ 0.05),TRP-1 (0.73±0.04 vs l.0, 0.86±0.17 vs l.0, both P＜0.05) andTRP-2 (0.68±0.04 vs l.0,0.69±0.11vs 1.0, both P ＜0.05) in comparison with unirradiated PIG cells. Conclusions The 1064-nm Q-switched Nd:YAG laser irradiation may affect the melanogenesis in PIG cells. With no influence on cell viability, the 1064-nm Q-switched Nd:YAG laser at 1 J/cm2 could enhance melanogenesis, while that at 3 J/cm2 could suppress melanogenesis, in PIG cells.

ObjectiveTo clarify the influence of human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein 120 V4 region with mutations at amino acid locuses on its abilities to enter target cells.Methods Based on the facts that ADA strains was a CCR5-tropic strain,only had the ability to infect CCR5 cells; that HXB2 strains was a CXCR4-tropic strain,only had the ability to infect CXCR4 cells,serial glycoprotein 120 mutants with alanine substitution in V4 region of ADA and HXB2 strains,were constructed by overlaping PCR.Eukaryotic expression vectors of mutants and expression vectors of HIV framework gene with luciferase reporter gene were cotransfected into eukaryotic cells to produce pseudoviruse.Concentration of HIV-1 gag P24 in pseudoviruses was detected by enzyme-linked immunosorbent assay(ELISA).U87.CD4.CCR5 and U87.CD4.CXCR4 cells were infected with 20 and 40 ng pseudoviruses,with wild ADA and HXB2 strains as control groups,respectively.The ability to infect cells of pseudovirus of each mutant with HIV-1 V4- region mutated at amine acid locuses 386-417 was measured by detecting the luciferase activity (relative light unit,RLU).ResultsTen mutants with alanine substitution in V4 region of HIV-1 ADA and HXB2 strains were successfully constructed,respectively.Mutants of pseudoviruse with 20 ng and 40 ng at locuses 389-391 and 414-417 with alanine substitution of V4 region in both ADA and HXB2 strains lost completely the abilities to enter CCR5 and CXCR4 expressing cells[ (0 ± 0)％].It was found that introduction of alanine to ADAs 400-403 and ADAs 408-410 increased the ability to infect cells to (124 ± 35)％,(182 ± 29)％ and (127 ± 8)％,( 134 ± 16)％ with pseudoviruse of 20 ng and 40 ng,respectively.Likewise,the ability to infect CXCR4 expressing cells also increased to (144 ± 42 )％ and (121 ± 18 )％ with pseudoviruse of 20 ng and 40 ng,respectively by introduction of alanine to HXB2s 395-397.However,other mutants in V4 region of ADA and HXB2 only maintained partial entry abilities( 15％- 84％).ConclusionsMutants of V4 region of HIV-1 envelope glycoprotein 120 with alanine substitution at locuses 389-391 and 414-417 in both ADA and HXB2 strains have been constructed successfully.They completely lost the ability to enter target cells.

The effects of application of in-process quality control in Guizhi Fuling capsule production were evaluated by 192 batches data analysis. Using a statistical analysis method, each batch of data were to be counted to research for the difference between 96 samples adopting the technologies of in-process control or not. According to quality standards of Guizhi Fuling capsule, all measurements of the 192 batches of the drugs before and after the application of process control technology were analyzed, and they were within the rules. There was a significant difference between adopting the technologies of process control or not. Application of in-process control technology can improve the uniformity of lot-to-lot for Guizhi Fuling capsule.
Capsules ; chemistry ; Chemistry, Pharmaceutical ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; chemistry ; Quality Control

Objective To study the influence of fluoride on the expression of Runx2 in suckling rat osteoblasts. Methods Osteoblasts obtained from calvarium of suckling Wistar rats were cultured in the media supplemented with NaF at different doses(0, 1,2 and 4 rag/L), and Runx2 Mrna expression and protein expression were evaluated by RT-PCR and ELISA, respectively. Results Runx2 Mrna expression in suckling rat osteoblasts cultured in vitro significantly increased after exposure to NaF for 48 h at different doses (0.613±0.055, 0.773±0.070 and 0.775±0.070 for 1,2 and 4 mg/L,respecfively) compared to the control (0.482±0.043 ,P< 0.05). Runx2 Mrna expression further increased after 72 h exposure to NaF(0.969±0.048,1.229±0.061,1.255± 0.063 for 1,2 and 4 mg/L, respectively) ,which is significantly higher than the control(0.724±0.036,P<0.05) and corresponding groups at 48 h. NaF doses and exposure time exhibited a significant synergistic effect on Runx2 Mrna expression (P<0.05). Similarly, NaF also enhanced Bunx2 protein expression in suckling rat osteoblasts cultured in vitro. Significant differences were observed between groups exposed to NaF (1,2 and 4 rag/L) and control at48 h post-exposure (0.141±0.007, 0.143±0.008, 0.143±0.011 vs 0.129±0.012, P<0.05) as well as 72 h post-expesure(0.156±0.014, 0.168±0.018, 0.162±0.0100 vs 0.137±0.016, P<0.05). In addition, Runx2 protein expression at 72 h post-exposure was significantly higher than that at 48 h. Conclusions The results suggested that NaF could increase Runx2 expression in suckling rat osteoblasts with a synergistic effect between the doses and exposure time.

CC chemokine receptor 4 (CCR4) is a kind of G-protein-coupled receptor, which plays a pivotal role in allergic inflammation. The interaction between 2-(2-(4-chloro-phenyl)-5-{[(naphthalen-1-ylmethyl)-carbamoyl]-methyl}-4-oxo-thiazolidin-3-yl)-N-(3-morpholin-4-yl-propyl)-acetamide (S009) and the N-terminal extracellular tail (ML40) of CCR4 has been validated to be high affinity by capillary zone electrophoresis (CZE). The S009 is a known CCR4 antagonist. Now, a series of new thiourea derivatives have been synthesized. Compared with positive control S009, they were screened using ML40 as target by CZE to find some new drugs for allergic inflammation diseases. The synthesized compounds XJH-5, XJH-4, XJH-17 and XJH-1 displayed the interaction with ML40, but XJH-9, XJH-10, XJH-11, XJH-12, XJH-13, XJH-14, XJH-3, XJH-8, XJH-6, XJH-7, XJH-15, XJH-16 and XJH-2 did not bind to ML40.Both qualification and quantification characterizations of the binding were determined. The affinity of the four compounds was valued by the binding constant, which was similar with the results of chemotactic experiments. The established CEZ method is capable of sensitive and fast screening for a series of lactam analogs in the drug discovery for allergic inflammation diseases.

Objective To investigate the effect of curcumin (Cur)on AngⅡ-induced proliferation and oxidative stress of vascular smooth muscle cells (VSMCs).Methods Primary rat VSMCs were cultured and divided into control group,AngⅡ group,AngⅡ+Cur 5μmol/L group,AngⅡ+Cur 10μmol/L group,AngⅡ+Cur 20μmol/L group,and Cur 20μmol/L group.The proliferation of AngⅡ-induced VSMCs was measured by MTT assay.The mRNA and protein expressions of inducible nitric oxide synthase (iNOS)and p47phox were detected by real-time PCR and Western blot.Nitric oxide (NO)production was measured by Griess reaction.Production of intracellular reactive oxygen species (ROS)was measured by DCFH-DA staining,and the activities of superoxide dismutase (SOD)and glutathione peroxidase (Gpx)were detected by xanthine oxidase assay and visible spectrophotometer. small interfering RNA (siRNA)was used to silence the expression of p47phox to further explore the mechanism for Cur inhibiting the proliferation of AngⅡ-induced VSMCs and oxidative stress.Results VSMCs activities were not significantly affected by Cur at the concentration between 0 and 80μmol/L.Cur (5,10 and 20μmol/L)significantly inhibited AngⅡ-induced proliferation of VSMCs.Cur had an inhibitory effect on the overexpression of NO,iNOS, p47phox and ROS in VSMCs and upregulated the activities of SOD and Gpx in a concentration-dependent manner. AngⅡ-induced ROS production in VSMCs was significantly attenuated by pretreatment with p47phox specific siRNA.Conclusion Cur can inhibit the proliferation and oxidative stress of AngⅡ-induced VSMCs.

Objective To investigate DJ-1 expression and protective effect against H2O2-induced oxidative stress in primary human melanocytes.Methods The expression and location of DJ-1 in primary melanocytes were identified by immunofluorescence.After cultured melanocytes were exposed to H2O2 for 24 hours,modified MTT assay was used to assess the proliferation of cells and to choose the suitable concentration of H2O2 for the following experiment.Western blot was used to detect D J-1 expression in melanocytes after being treated with 0.5 mmol/L H2O2 for 24 hours.Some melanocytes were divided into 3 groups to be reversely transfected with PBS(mock control group),non-targeting siRNA(negative control group)and DJ-1 targeting siRNA(DJ-1 group).Optical microscopy was utilized to observe the morphologic changes of transfected melanocytes.At 48 hours after the transfection,the melanocytes were stimulated with H2O2 for 24 hours.Subsequently,modified MTT assay,2′,7′-dichlorofluorescein diacetate(DCFH-DA)and annexin Ⅴ-fluorescein isothiocyanate/propidium iodide were used to determine cell viability,intracellular reactive oxygen species (ROS)level and apoptosis rate respectively.Results DJ-1 was expressed in both melanocyte nucleus and cytoplasm,and predominantly in the nucleus.H2O2 inhibited the cell viability in a dose dependent manner.After treatment with H2O2 of 0.5 mmol/L for 24 hours,the cell viability began to decrease in melanocytes with the expression level of DJ-1 being 2.23 times that in the untreated melanocytes(both P ＜ 0.05).Compared with the mock control group,the dendrites of melanocytes in DJ-1 group were obviously shortened with cytoplasm vacuolization.After 24-hour treatment with H2O2 of 0.5 mmol/L,the cell viability in the DJ-1 group dropped to 35％ of that in the mock control group(P ＜ 0.05),while the intracellular ROS fluorescence intensity( FI) and apoptosis rate were higher in the DJ-1 group than in the mock control group (902 ± 40 vs.529± 32,58q％ ± 6.1％ vs.30% ± 3.8％,both P ＜ 0.05).Conclusion DJ-1 can protect melanocytes against H2O2induced oxidative stress likely by decreasing intracellular ROS production and inhibiting ceU apoptosis.