Objective To observe the clinical efficacy ofXing Nao Kai Qiao (brain-awakening and orifice-opening) needling plus thunder-fire moxibustion in treating vertebrobasilar ischemia.Method Fifty-five patients with vertebrobasilar ischemia were randomized into a treatment group of 26 cases and a control group of 29 cases. The treatment group was intervened byXing Nao Kai Qiao needling plus thunder-fire moxibustion, while the control group was by oral administration ofYangxue Qingnao granules. The blood flow of vertebrobasilar arteries were observed by using ultrasonic Doppler blood-flow detector before and after intervention, and the clinical efficacies were compared between the two groups.Result The recovery and markedly-effective rate and total effective rate were respectively 61.5% and 96.2% in the treatment group, versus 34.5% and 89.7% in the control group, and the differences were statistically significant (P<0.01,P<0.05). The VS and VD of vertebral arteries and VS of basilar arteries were significantly changed after intervention in the treatment group (P<0.05). The VS of basilar arteries was significantly changed after intervention in the control group (P<0.05). After treatment, The VS of vertebral and basilar arteries in the treatment group was significantly different from that in the control group (P<0.05).ConclusionXing Nao Kai Qiao needling plus thunder-fire moxibustion is an effective approach in treating vertebrobasilar ischemia.

AlM: To investigate the relationship among carotid atherosclerosis, homocysteine ( Hcys ) and anterior ischmic optic neuropathy( AlON) .METHODS: Sixty-five AlON patients enrolled in AlON group and 70 non - AlON controls enrolled in control group. All the participants in 2 groups were examined by carotid artery color doppler flow imaging to evaluate the incidence of carotid atherosclerosis. Plasma levels of Hcys, vitamin B12 and folate were measured by automatic biochemical analyzer. The incidence of carotid atherosclerosis, plasma levels of Hcys, vitamin B12 and folate were compared statistically between AlON and control group.RESULTS: Fifty - six AlON patients ( 86%) exhibited carotid atherosclerosis, which was found in 43 control subjects ( 61%) . Fifty - nine AlON patients ( 91%) exhibited increased plasma levels of Hcys compared with that in 38 control subjects ( 54%) . Lowered vitamin B12 was found in 37 patients with AlON ( 57%) and in 43 control subjects ( 61%) . Lowered folate was found in 54 AlON patients ( 83%) and in 32 control subjects ( 46%) . The incidence of atherosclerosis in carotid artery ( P =0. 001), mean Hcys level (P=0. 0005) and lowered folate ( P = 0. 0006 ) were different statistically between AlON group and control group. Mean vitamin B12 level was indifferent between AlON group and control group ( P=0. 0722). Logistic regression analysis showed that carotid atherosclerosis was an a risk factor for AlON (OR=1. 312, 95%CI:0. 927~1. 772), whereas elevated Hcys level (OR=2. 378, 95% CI: 1. 479 ~ 3. 821 ) was also shown to be significant risk factor for AlON. CONCLUSlON: Carotid atherosclerosis and elevated Hcys were related with AlON. Reducing above-mentioned risk factors related to thrombus may be crucial to the prevention and cure of AlON.

Objective: To investigate the influence of high-fat diet on glucose metabolism and fatty acid composition of serum phospholipid in OLETF rats, a model of spontaneous type 2 diabetes mellitus. Method: Twenty male OLETF rats of 14 w age were divided into 2 groups randomly. The dietary fat in two groups contained identical percentages of saturated fatty acids, monounsaturated fatty acids and polyunsaturated fatty acids. Oral glucose tolerance test and measurement of insulin level were performed after 10 w treatment. The fatty acid composition of serum phospholipid in rats was observed as well. Results:The food intakes were comparable in 2 groups. The body weight gain in high-fat group was higher than standard group during the experiment. The result of oral glucose tolerance test and measurement of insulin level showed that the glucose metabolism in two groups had no significant difference. Only the percentage of total n-6 polyunsaturated fatty acids, 18:3(n-3) of serum phospholipid were comparable, percentages of other fatty acids were significantly different in two groups. Conclusion:With the comparable fatty acid composition, higher dietary fat lead to more body weight gain, but had no significant influence on glucose metabolism in OLETF rats. The amount of dietary fat had significant effect on fatty acids composition of serum phospholipid in OLETF rats.

Objective:To investigate the effect of Na + , K + -ATPase on the increase of cytosolic Ca 2+ level induced by hypoxia in rat cortical neurons. Methods: Using confocal laser scanning microscope and video based motion edge detection system, we measured the intracellular Ca 2+ concentration (\ i ) and Na + concentration (\ i ) in cortical neurons exposed to hypoxia for different time periods and at presence of different concentrations of dihydroouabain (DHO, an inhibitor of Na + , K + -ATPase). We also observed the effect of Na + , K + -ATPase on the increase of cytosolic Ca 2+ level induced by hypoxia in rat cortical neurons. Results: DHO (10 -9 -10 -3 mol/L) dose-dependently and hypoxia(4-20 min) time-dependently increased the levels of \ i and \ i in cultured cortical neurons. \ i and \ i increased significantly 4 min after cortical neurons were exposed to hypoxia, and addition of DHO (10 -3 mol/L) further increased their concentrations. While 15 min after hypoxia exposure, different concentrations of DHO did not further increase \ i and \ i. We also found that hypoxia exposure did not enhance DHO pretreatment-induced \ i increase. Conclusion: Inhibition of Na + , K + -ATPase activity is one of the mechanisms responsible for hypoxia-induced elevation of \ i in cortical neurons.

Objective To investigate the safety and clinical response of dendritic cells (DC)vaccine loaded with HLA-A2-restricted peptides MAGE-3、 Tyr、 MART-1 and GP-100 against malignant melanoma. Methods Twenty three HLA-A2 positive patients with malignant melanoma were enrolled.Peripheral blood mononuclear cells were separated into adherent cells for induction of DC loaded with four peptides. DC vaccine was administered subcutaneously once a week in inguinal region. The immunological responses and clinical responses were evaluated. Results Dendritic cell vaccination were well tolerated in all patients and there was no toxicity observed. Serum levels of IL-2, IL-12 and IFN-γincreased significantly after first vaccination and fourth vaccination. The positive rate of DTH test was 50% (5/10), and 2 fold increase of CD8+ IFN-γ+ cells were observed in 6 of 10 patients. Stable disease was observed in 11 of 23 patients, one patient had a complete metastasis regression, four patients had 50% regression of metastasis,four patients had a minor response, and disease progressed in three patients. Conclusion DC vaccines loaded with peptides MAGE-3、 Tyr、 MART-1 and GP-100 can elicit non-specific and specific immune responses, leading disease control. DC vaccine is considered one of safe and effective treatments for malignant melanoma.

In this study, hydrophilic matrix sustained release tablets of total lactones from Andrographis paniculata were prepared and the in vitro release behavior were also evaluated. The optimal prescription was achieved by studying the main factor of the type and amount of hydroxypropyl methylcellulose (HPMC) using single factor test and evaluating through cumulative release of three lactones. No burst drug release from the obtained matrix tablets was observed. Drug release sustained to 14 h. The release mechanism of three lactones from A. paniculata was accessed by zero-order, first-order, Higuchi and Peppas equation. The release behavior of total lactones from A. paniculata was better agreed with Higuchi model and the drug release from the tablets was controlled by degradation of the matrix. The preparation of hydrophilic matrix sustained release tablets of total lactones from A. paniculata with good performance of drug release was simple.
Andrographis ; chemistry ; Delayed-Action Preparations ; chemistry ; Drug Compounding ; methods ; Drugs, Chinese Herbal ; chemistry ; Hydrophobic and Hydrophilic Interactions ; Kinetics ; Lactones ; chemistry ; Tablets ; chemistry

Objective To construct the eukaryotic expression plasmid containing glyceraldehydes-3-phosphate dehydrogenase (GAPDH) and cysteine protease inhibitor ( CPI ) gene from periodic Brugia malayi (Bm) and to lay foundation for studying multivalent vaccines. Methods Total RNA was extracted from periodic Bin. The BmGAPDH and BmCPI genes were amplified by RT-PCR. The PCR product was cloned and then subeloned into eukaryotic recombinant plasmid vector pcDNA3.1 (+). pcDNA3.1 (+)/BmGAPDH/BmCPI was constructed. The recombinant plasmids were screened and identified by digestion with restriction enzyme and PCR amplification, and were transformed into HeLa cell subsequently. The transient expression of BmGAPDH and BmCPI were examined by RT-PCR. The expressed protein was identified by sodium dodeeylsulphate-polyacrylamide gel electrophoresis(SDS-PAGE). Results Two specific bands of around 877 bp of BmGAPDH and 621 bp of BmCPI were amplified, consistent with the expected value. The same bands were obtained by double restriction enzyme digestion of recombinant plasmids or PCR using recombinant plasmid as template. BmGAPDH and BmCPI mRNA were highly expressed in transfeeted HeLa cell. The relative molecular mass (Mr) of the recombinant protein was about 54 × 103. Conclusion The recombinant eukaryotic expression plasmid pcDNA3.1 (+)/BmGAPDH/BmCPI has been constructed successfully and the protein is expressed correctly in mammalian cell.

Objective To detect the expression of chemokine receptors CXCR3 mRNA in the peripheral blood mononuclear cells (PBNCs) of patients with Rheumatoid Arthritis (RA) and to analyze the relationship between the expression and the disease activity. Methods mRNA was extracted from PBNCs and the expression of CXCR3 mRNA was detected by real-time fluorescence quantitative PCR (RFQ-PCR) in 51 RA patients and 32 controls. T-test, x2-test, ANOVA were used for statistial analysis. Results Comparison between the two groups had shown that the expression levels of CXCR3 mRNA in clinical active RA group were higher than those of the RA patients in remission and healthy controls (P<0.05). The expression levels of CXCR3 mRNA were positively correlated with serum levels of ESR and CRP in clinical active RA group (r=0.824, r=0.765, P<0.05). In addition, RF titer, APF, AKA, and anti-CCP had no significant correlation with the expression levels of CXCR3 mRNA in RA patients (P>0.05). Conclusion RFQ-PCR is a sensitive,reproducible and practical test. The mRNA expressions of CXCR3 are significantly elevated in RA patients,which suggest that CXCR3 may be involved in the pathogenesis of RA. The mRNA expressions of CXCR3 in active RA patients are higher than those of RA patients in remission. These results indicate that CXCR3 may play an important role in the pathogenesis and progression of RA, and CXCR3 may be considered as an indicator for disease activity, therapeutic efficacy and prognosis of RA.

Objective To observe the effects of rosuvastatin on the homocysteine (Hcy)-induced expression of matrix metalloproteinase 2 (MMP 2) and cell migration in rat vascular smooth muscle cells (VSMCs), and to explore the possible mechanism of Hcy-induced atherosclerosis and the role of statins in reversing atherosclerosis. Methods In one cell culture plate, the cultured rat VSMCs were incubated with different concentrations of Hcy (0, 50, 100, 500, 1000 μmol/L and 5000 μmol/L) in vitro for 24 h, 48 h and 72 h. And in another cell culture plate, the different concentrations of rosuvastatin (10-9, 10-8, 10-7, 10-6, 10-5 mol/L and 0 mol/L) were added to the cultured rat VSMCs (while the concentration of Hcy was 1000 μmol/L). The MMP 2 expression and enzyme activity were determined by gelatin zymography and Western blotting. The effects of Hcy and rosuvastatin on cell migration and invasiveness of VSMCs were observed. Results Hcy (50-5000 μmol/L) increased the protein expression, and Hcy (50-1000 μmol/L) increased enzyme activity of MMP 2 significantly. But Hcy (5000 μmol/L) inhibited activity of MMP 2 (F=9.31, 6.44 and 5.97, all P＜0.05). Rosuvastatin (10-9-10-5 mol/L) inhibited Hcy-induced expression and enzyme activity increasing of MMP 2. The counts of cell migration of VSMCs were 18.32±2.17, 32.68±4.34, 44.75±4.08, 61.39±5.21, 79.74±5.54 and 90.78±5.83, while the concentration of Hcy was 0, 50, 100, 500, 1000 μmol/L and 5000 μmol/L respectively (F=5.31, P＜0.05). The counts of cell migration of VSMCs were 79.74±5.54, 62.53±6.41, 48.37±5.66, 31.41±4.79, 19.27±3.62 and 11.17±2.33, while the concentration of rosuvastatin was 10-9, 10-8, 10-7, 10-6 and 10-5 mol/L respectively (F=4.99, P＜0.05). Rosuvastatin could decrease the stimulation of Hcy-induced migration of VSMCs. Conclusions Hcy can influence the MMP 2 protein expression/activity in VSMCs, and rosuvastatin can inhibit augmentation of Hcy-induced MMP 2 expression/activity and migration of VSMCs. It may be one of the multiple-effects of rosuvastatin reducing atherosclerosis.

AIM: To explore the effect of Snail1 siRNA on high-glucose induced tubular epithelial-to-mesenchymal transition (TEMT). METHODS: Subconfluent renal tubular epithelial cells were incubated in serum-free DMEM for 24 h to arrest and synchronize the cell growth. Then cells were treated with normal glucose (5.5 mmol/L D-glucose) or high glucose (25 mmol/L D-glucose) for 72 h. Meanwhile 19.5 mmol/L D-manntiol was used as high osmotic control. Snail1 siRNA was transfected into tubular epithelial cells. In parallel, cells were transfected with non-specific siRNA which served as the control data sets. Cells were then treated with 25 mmol/L D-glucose for 72 h. RNA and cell lysates were collected to determine the protein and mRNA levels of Snail1, TGF-β_1, α-SMA, vimentin and E-cadherin. RESULTS: Transfection caused the decreases in Snail1 at mRNA and protein levels by 62% and 68% respectively as compared to those in untransfected cells cultured in high glucose medium. Western blotting exhibited that Snail1 siRNA transfection restored E-cadherin protein expression by 61% compared to that in high-glucose-treatment cells, whereas it inhibited high-glucose-induced induction of α-SMA protein by 58%. Similarly, RT-PCR revealed that Snail1 siRNA transfection dramatically suppressed the high-glucose-induced mRNA expressions of α-SMA and vimentin by 72% and 61%, respectively, while E-cadherin mRNA increased by 53%. CONCLUSION: Our study provides direct evidence that Snail1 is able to control TEMT.