Objective To study whether p38 mitogen-activated protein kinases (p38MAPK) signal pathway were activated in the process of tumor necrosis factor-like weak inducer of apoptosis (TWEAK) which then induces rheumatoid arthritis (RA) fibroblast-like synoviocyte (FLS) to synthesize matrix metalloproteinase-1 (MMP-1) and look for the relative mechanisms of how TWEAK was involved in the destruction of articular bones and cartilage. Methods RA FLS were primarily cultured and stimulated with TWEAK. FLS were pretreated with SB203580 or not. ELISA was used to detect the concentration of MMP-1 in cell-cultured supernatant. Western blotting was used to detect the expression of p-p38MAPK and P65 in RA FLS. Results TWEAK (100 ng/ml) could induce RA FIS to synthesize MMP-1. SB203580 could partially inhibite the expression of MMP-1 producted by RA FLS which was induced by TWEAK. TWEAK could make p38MAPK phosphorylated and increase the expression of P65 protein in the cell nucleus. Conclusion TWEAK induces RA FLS to synthesize MMP-1. In this process, p38MAPK signal transduction pathway is activated and then induce the expression of NF-κB.

AIM: To affirm marker peaks for the fingerprint chromatography of Shengmai Injection. METHODS: LC-MS/MS method was used, with a Waters symmetryshield TM RP_ 18 column(4.6 mm?250 mm; 5.0 ?m), acetonitrile-water as a mobile phase, The detection wavelength was at 203 nm. Ion trap mass spectrum. RESULTS: Affirming marker peaks for fingerprint chromatography of Shengmai Injection and 10 marker peaks were affirmed. CONCLUSION: The method can affirm marker peaks for the fingerprint chromatography of Shengmai Injection. It is simple, accurate, and has practicality.

AIM: To determine 5 different aphrodisiacs,including apomorphine,yohimbine,vardenafil,sildenafil and alprostadil,adulterated into Chinese proprietary medicine by LC/MS(n),and build a laboratory library including MS(n) spectra and retention time. METHODS: ZORBAX SB C_18 column(150 mm?4.6 mm,5 ?m)was applied.The mobile phase consisted of acetonitrile and 0.5% formic acid changing by linear gradient elution,and the flow rate was 1.0 mL/min.The ion trap mass spectrometer with positive ion mode electrospray(ESI?) was applied. RESULTS: In the mass spectrum apomorphine,yohimbine,vardenafil and sildenafil showed abundant pseudomolecular ion +,and alprostadil showed abundant pseudomolecular ion -.All the MS(n) spectra showed characteristic product ion. CONCLUSION: It is proved that the method is simple,reliable and suitable to determin aphrodisiac adulterated to health food.

OBJECTIVE To investigate the effect of quercetin on primary cultured newborn rat cortex neuron cell which is estrogen depletion, and discuss the possible mechanism, to provide new ideas and strategies for developing a drug of neurodegenerative disease. METHODS Rat cortex neurons were isolated from one day old Sprague Dawley rats and treated with estrogen, quercetin and estrogen receptor antagonists (ICI182,780). Cell viability was determined by MTT assay, neurite outgrowth was measured by fluorescent microsope and estrogen receptors were determine by Western blot. RESULTS Quercetin functions like estrogen to increase cortex neuronal cell viability, the Que (50, 100 μmol·L-1) group compared with the control group could significantly improve the activity of the cortical neurons(P<0.05). It can also increase neurite out growth, the Que (50,100 μmol·L-1) group significantly promoted the formation of synapse, most of the neurons were full, and the synapses of neurons became thick, growth, and connect to a dense neural network. And in the Western blot experiments, Que (50, 100 μmol·L-1) group could obviously increase the expression of estrogen receptor alpha protein, in addition, the neural protective effect of quercetin can be inhibited by ICI182,780. CONCLUSION Quercetin like estrogen can protected cortex neuronal and the effect of quercetin on cortex neuronal cells was mediated by estrogen receptor alpha.

Optically pure α-substituted propanoic acids and their derivatives represent as an important kind of organic building blocks and key intermediates,which has been widely used in the synthesis of chiral drugs.Some of them have been used directly as nonsteroidal anti-inflammatory drugs (NSAIDs),such as ibuprofen,naproxen,ketoprofen and so on.Dihydroartemisinic acid (DHAA),the same structure as the α-substituted propanic acids,is a key intermediate for the synthesis of artemisinin,the most effective and current used anti-malarial drug.Therefore,the asymmetric synthesis of α-substituted propanoic acids is always a hot topic for chemical scientists.Asymmetric catalytic hydrogenation attracts more and more attentions because of its atom economy and efficiency.This dissertation will disclose the asymmetric synthesis of α-substituted propanoic acids using transition metal-complex as a chiral catalyst.

Objective To investigate the influence of thymosin beta 4 (Tβ4) with two different dosages on the expression of transforming growth factor beta (TGF-β) and connective tissue growth factor (CTGF) in rats with renal tubular interstitial fibrosis,and to further estimate the changes of renal tubular interstitial lesions.Methods Rat models of renal tubular interstitial fibrosis were established by unilateral ureteral occlusion (UUO).The male SD rats were randomly divided into 4 groups and 15 rats in each group:sham group,model group,treatment group with 1 mg/L Tβ4 and treatment group with 5 mg/L T34.Rats in sham group and model group were poured into the same amount of saline.The renal function and renal pathological changes were observed after the second week.The mRNA and protein expression of TGF-β and CTGF in renal tissues was tested by in-situ hybridization and Western blotting.Results Compared with that in sham group,the expression of TGF-β mRNA and its protein,CTGF mRNA and its protein was significantly higher in model group (all P ＜ 0.01).Compared with rats of model group,Tβ4 treatment rats had lower mRNA and protein expression of TGF-β and CTGF (all P ＜ 0.01),and the expression in treatment group with 5 mg/L Tβ4 was lower than that in treatment group with 1 mg/L Tβ4 (P ＜ 0.05).And the expression of TGF-β mRNA was positively correlated with CTGF mRNA expression (r=0.697,P ＜ 0.01).The 24 h total urinary protein and the area of renal tubular interstitial lesion in model group were significantly more than those in sham group,and also more than those in Tβ4 treatment group (all P ＜ 0.05).Tβ4 treatment attenuated kidney damage,and the effects in treatment group with 5 mg/L Tβ4 were better than those in treatment group with 1 mg/L Tβ4.No difference in serum creatinine and blood urea nitrogen was observed among 4 groups (all P ＞ 0.05).Conclusions Tβ4 treatment can inhibit the renal TGF-β and CTGF expression of rats with tubular interstitial fibrosis in a dose-dependent manner,and play a protective role in kidney.

Objective To investigate the role of astroglial glutamate-glutamine shuttle in the development of neuropathic pain (NP) in rats. Methods Forty-eight adult male SD rats weighing 200-230 g were randomly divided into 8 groups (n =6 each): Ⅰ control group (group C);Ⅱ sham operation group (group S);group ⅢNP;Ⅳ-Ⅶ 0.01, 0.03, 0.05 and 0.10 mmol/L methionine sulfoximine (MSO, an inhibitor of glutamine synthetase (GS)) group (group M1-4 );Ⅷ MSO + glutaminate group (group MG). In group C no operation was performed. In group S the sciatic nerve was only exposed but not ligated. NP was induced by ligation of the tibial nerve and commom peroneal nerve according to the technique described by Dixon. After the establishment of the model, intrathecal PBS 50 μl was injected in group NP, IT 0.01, 0.03, 0.05 and 0.10 mmol/L MSO 50 μl was injected intrathecally in group M1-4, and 0.05 mmol/L MSO 50 μl was injected intrathecally and then 0.25 mmol/L glutamine 50 μl was injected intrathecally 15 min later in group MG. Mechanical pain threshold was measured 1 week before ligation (T0 , baseline), 1 week after ligation (T1) and 15, 30, 45 and 60 min after injection of MSO (T2-5). Then rats were killed and the lumbar segment of the spinal cord was removed for determination of the expression of glial fibrillary acidic protein (GFAP) and GS and the co-expression (GFAP/GS) in the dorsal horn.Results Mechanical pain threshold was significantly lower at T1-5 in group MG and NP and at T2-4 in group M3.4 ,and the expression of GFAP, GS and GFAP/GS was significantly higher in group MG,NP and M3 than in group S and C ( P ＜ 0.05) .Conclusion Astroglial glutamate-glutamine shuttle in the spinal cord is involved in the development of neuropathic pain in rats.

BACKGROUND: Migu tablet, a Chinese drug for kidney invigorating, is effective on preventing and treating osteoporosis, but the concrete mechanism of pharmacology is still not clear. Transforming growth factor-β1(TGF-β1) is an important cytokine, which can regulate bone resorption and formation.OBJECTIVE: To investigate the effect of kidney invigorating recipe on mRNA expression of TGF-β1 of osteoblast.DESIGN: A completely randomized controlled study was conducted.SETTING: Department of Traumatic Orthopedics, Union Hospital Affiliated to Tongji Medical College of Huazhong University of Science and Technology.MATERIALS: This experiment was conducted at the laboratory for bone metabolism of integration of Chinese and western medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology from May 2003 to April 2004. Experimental rats: Totally 16 newborn SD rats of clean degree were involved. Experimental drug: Medical liquor of Migu tablet was prepared in the Department of Traumatic Orthopedics,Union Hospital Affiliated to Tongji Medical College, Huazhong University of Science and Technology. The subscription was mainly composed of Chinese herbs such as Herba Epimedii, Cortex Eucommiae, Semen Juglandis,Radix Rehmanniae, Radix Achyranthis Bidentatae. and so on. Positive control drug: which was recombinant basic fibroblastic growth factor (rbFGF), was purchased from Beijing Banding Company. Negative control group was subdivided into negative control of probe and antibody METHODS: 100,1 000,5 000,10 000 mg/L Chinese herb Migu tablet liquor for kidney invigorating and positive control drug 5 μg/L rbFGF were added into the osteoblasts of cranial bones of newly born SD rats separated and cultured in vitro. 24 hours later, nuclear acid molecular in situ hybridization of osteoblasts were analyzed by self-made digoxin-labeled TGF-β1 cDNA probe . The mean absorbance of positive particles representing the mRNA expression of TGF-β1. A total of 40 osteoblasts were randomly chosen from each group under 200-fold amplification. The average absorbance of hybridized particles of the cells was measured with TJTY-300 automatic image analyzer.MAIN OUTCOME MEASURES: mRNA expression of TGF-β1 in osteoblasts of each group.RESULTS: Automatic image analyzer showed that the TGF-β1 mRNA expressions of Migu liquor groups whose concentration were 5 000 mg/L and 10 000 mg/L were respectively 1.18 times and 1.30 times that of control group, with a significant difference. [The mean absorbance's of hybridized particles of the cells in the 5 000,10 000 mg/L Migu liquor groups and negative control were 0.213 67±0.015 00,0.237 03±0.021 73,0.181 27±0.015 28 ,respectively, P ＜ 0.05 and P ＜ 0.01].Although the mean absorbance ( 0.254 45±0.020 81)of the hybridized particles of the cells in the 5 μg/L recombinant rbFGF was higher than those of 5 000,10 000 mg/L Migu liquor groups, but there was no significant difference(P ＞ 0.05).CONCLUSION: Migu tablet for kidney invigorating can stimulate the secretion and synthesis of TGF-β1 in osteoblasts, thus promote bone formation and inhibit bone resorption.

BACKGROUND:The therapeutic effects of donkey-hide glue reinforcing bone oral solution on osteoporosis have been determined, but the exact effective mechanism is to be approached. OBJECTIVE: To investigate the effects of donkey-hide glue reinforcing bone oral solution (DGRBOS) medicated serum on osteoprotegerin (OPG)and its ligand(OPGL)mRNAexpression of osteoblast in fetal rats and explore the molecular mechanism of treating osteoporosis with DGRBOS. DESIGN: A randomized controlled trial. MATERIALS: The experiment was carried out from June 2003 to October 2004 in Bone Metabolic Laboratory of Department of Integrative Chinese and Western Medicine, Affiliated Hospital of Tongji Medical College,Huazhong University of Technology and Science. Totally 30 3-month-oldWistar rats (15 males and 15 females) were randomly divided into 3 groups, I.e. DGRBOS group, estrogen group and control group, with 10 rats in every group. 12 clean newborn SD rats were selected to isolate and cul ture osteoblast. METHODS: ①After intragastric administration for 7 days, medicated serum was prepared respectively from the three groups. ②Skull osteoblast isolated from newborn SD rats was made into single cell suspension, then after digestion and passage, the subcultured osteoblast cell was made into cell suspension. The cultured osteoblasts were divided into 5 groups and given equal volumes of drug liquor. The DGRBOS group was given DGRBOS-medicated serum at the concentration of 100, 500 and 1 000 g/L which was diluted by nutrient solution; the estrogen group was given tibolone-medicated serum of 100 and 1 000 g/L; the control group was givenonly culture fluid. Meanwhile every group was given calf serum (100 g/L) for further culture. ③The osteoblast proliferation was measured by antigenic MTT colorimetric analysis and 3H-TdR penetration method. The in tra-cellular BGP contents were evaluated by radioimmunity .The mRNA expression of OPG and RANKL in osteoblast was analyzed by Rt-PCR. ④ One-way analysis of variance was applied to compare data among groups. MAIN OUTCOME MEASURES: mRNA expression of OPG and PAN KL in osteoblasts from fetal rats after intervention by medicated serum ofDGRBOS or Livial. RESULTS: ①The osteoblast proliferation measured by antigenic MTT colorimetric analysis and 3H-TdR penetration method showed that the proliferation in the DGRBOS group and tibolone group was enhanced moresignificantly than that in the control group (P ＜ 0.05-0.01), and reached maximal effect at the concentration of 500 g/L (P ＜ 0.01), but when the concentration was over 500 g/L, the effect tended to saturate. The medicated serum with all concentrations from DGRBOS and estrogen groups could increase the contents of BGP in osteoblasts (P ＜ 0.05). ②The mRNA expression of OPG reached the peak when the DGRBOS medicated serum was 1 000 g/L, and was obviously higher than that at the concentration of 100 and 500 g/L (P ＜ 0.05). The expression in DGRBOS group at the concentration of 1 000 g/L and in the estrogen group at the concentration of 100 and 1 000 g/L was apparently higher than that of the control group (P ＜ 0.01). ③The mRNAexpression of RANKL was the highest in DGR BOS group with 1 000 g/L concentration, and was markedly lower than that of the concentration of 100 and 500 g/L (P ＜ 0.05). The expression in DGRBOS group at the concentration of 1 000 g/L and in the estrogen group at the concentration of 100 and 1 000 g/L was noticeably lower than that in the control group (P ＜ 0.01).CONCLUSION: ①The DGRBOS could remarkably enhance osteoblast proliferation in dose-dependent and a dose-saturable manner, and the effect was close to that of tibolone. ②Partial mechanism of DGRBOS in treating osteoporosis might be promoting osteoblast proliferation and regulating OPG/RANKL expression.