Downregulation of electroacupuncture at ST36 on TNF-α in rats with ulcerative colitis

AIM: To investigate the regulatory effect of electroacupuncture (EA) at Zusanli (ST36) on tumor necrosis factor-alpha (TNFα) in rats with ulcerative colitis (UC), and further elucidate the therapeutic mechanism of EA on UC.METHODS: Thirty-two male Sprague-Dawley (SD) rats were randomly divided into four groups (n=-8): normal control group, UC control group, UC+ST36 group and UC+nonacupoint group. A solution containing ethanol and 2,4,6-trinitrobenzenesulfonic acid (TNBS) was instilled into the distal colon in the rat (at a dose of 100 mg/kg) to set up UC rat model. Rats in wakefulness state of UC+ST36 group were stimulated at ST36 by EA once a day, while those of UC+nonacupoint group were done at 0.5 cm beside ST36. After 10d treatment, all rats were sacrificed simultaneously. Colon musocal inflammation and damage were assessed by measuring colon mass, morphologic damage score, colonic myeloperoxidase enzyme (MPO) activity, serum TNF-α and colonic TNF-α mRNA level. Morphologic damage score was examined under stereomicroscope. Colonic MPO activity was measured by spectrophotometer method. Serum TNF-αconcentration was determined by radioimmunoassay (RIA).Colonic TNF-α mRNA expression level was analyzed by semiquantitative reverse transcription polymerase chain reaction (RT-PCR).RESULTS: Ratio of colonic mass/body mass (mC/mB) and activity of colonic MPO (μkat/g tissue) markedly increased (8.5±2.6 vs 2.5±0.4; 145±25 vs 24±8, P＜0.01 vs normal control group). Compared with normal control rats, serum TNF-α and colonic TNF-α mRNA level in UC control group were increased 2.5 fold (2 278±170 vs 894±248, P＜0.01)and 4.3 fold (0.98±0.11 vs 0.23±0.11, P＜0.01)respectively. After EA at ST36, mc/mB and MPO activity were reduced significantly (5.3±2.0 vs 8.5±2.6; 104±36 vs145±25, P＜0.01, 0.05) compared with those of UC control group. Serum TNF-α and colonic TNF-α mRNA level were inhibited by EA stimulation at ST36 (P＜0.01). The inhibitory rate was 16 % and 44 % respectively.Morphologic damage score was also increased markedly in rat with UC (P＜0.01), whereas it was decreased by EA at ST36 (P＜0.05). There was no significant difference between UC control group and UC+EA at non-acupoint (P＞0.05). Furthermore, these parameters were highly correlated with each other (P＜0.01).CONCLUSION: Serum TNF-α concentration and colonic TNF-α mRNA expression level are increased significantly in UC rats in correlation with the severity of disease. It indicates that TNF-α is closely involved in the immune abnormalities and inflammatory responses in UC. EA at ST36 has therapeutic effect on UC by downregulating serum TNF-r and colonic TNF-r mRNA expression. High levels of TNF-αand its corresponding mRNA expression seem to be implicated in the pathogenesis of UC.

Antiangiogenic effect of somatostatin receptor subtype 2 on pancreatic cancer cell line:Inhibition of vascular endothelial growth factor and matrix metalloproteinase-2 expression in vitro

AIM:To investigate the anti-angiogenic effect of somatostatin receptor subtype 2 (SSTR2) gene transfer into pancreatic cancer cell line PC-3, and the mechanisms involved in this effect.METHODS: The full length human SSTR2 cDNA was introduced into pancreatic cancer cell line PC-3 by lipofectamine-mediated transfection. Positive clones were screened by G418 and stable expression of SSTR2 was detected by immunohistochemistry SABC methods and RT-PCR. Enzyme-linked immunosorbent assay (ELISA) was used to detect vascular endothelial growth factor (VEGF) levels in the cell culture supernatants of SSTR2-expressing cells, vector control and mock control cells. Furthermore, the expressions of VEGF and matrix metalloproteinase-2 (MMP-2) were detected by immunohistochemistry SABC methods and RT-PCR in these cells.RESULTS: VEGF levels in the cell culture supernatants were significantly reduced in the SSTR2-expressing cells (first week,172.63±21.2ng/L and after two months, 198.85±26.44ng/L)compared with the vector control (first week, 790.39±86.52ng/L and after two months, 795.69±72.35ng/L) and mock control (first week, 786.42±90.62ng/L and after two months,805.32±84.36ng/L) (P<0.05).The immunohistochemical assay showed a significant reduction of the integral optical density of VEGF and MMP-2 in the SSTR2-expressing cells (42.25±8.6 and 70.5±6.25, respectively) compared with the vector control (85.75±12.9 and 110.52±13.5, respectively) and mock control (82.6±9.28 and 113.56±9.62,respectively) (P<0.05).Conversely, the average gray value of VEGF and MMP-2 was significantly increased in the SSTR2-expressing cells (121.56±8.43 and 134.46±19.95, respectively) compared with the vector control (55.72±5.6 and 62.26±12.68,respectively) and mock control cells (58.48±6.2 and 65.49±9.16, respectively) (P<0.05). Moreover, the expressions of VEGF mRNA and MMP-2 mRNA were significantly reduced in the SSTR2-expressing cells (0.1384±0.017 and 0.2343±0.070, respectively) compared with the vector control (1.024±0.117 and 0.806±0.119,respectively) and mock control (1.085±0.105 and 0.714±0.079,respectively) (P<0.05).CONCLUSION: The expression of reintroduced human SSTR2 gene exerts its antiangiogenic effects by downregulating the expressions of the factors involved in tumor angiogenesis and metastasis, suggesting SSTR2 gene transfer as a new strategy of gene therapy for pancreatic cancer.

Potential inhibition of cytochrome P450 3A4 by propofol in human primary hepatocytes

AIM: Hepatic cytochrome P450 isoenzymes constitute a superfamily of hemoproteins that play a major role in the metabolism of endogenous compounds and in the detoxification of xenobiotic molecules. P450 3A4 is one of the most important forms in human being, and mediates the metabolism of around 70% of therapeutic drugs and endogenous compounds. Propofol, a widely used intravenous anesthetic drug, is known to inhibit cytochrome P450activities in isolated rat hepatocytes. The goal of this study was to evaluate the potential efficacy of propofol on P4503A4 in a dose-dependent manner to understand its drugdrug interaction.METHODS: Hepatocytes were isolated from liver specimens from hepatic angioma patients undergone hepatic surgery.Primary incubated hepatocytes were treated with 0, 0.01,0.05, 0.1, 0.5, and 1.0 mM propofol for 24 hours. P450 3A4activity was measured with Nash′s colorimetry. The protein expression was assessed by Western blot analysis.RESULTS: A dose-dependent inhibitory effect of propofol was observed in cytochrome P450 3A4 activity. A minimal dosage of propofol (0.01 mM) induced a significant inhibition of P450 3A4 activity, although its regular dosages (0.01-0.1mM) showed no inhibitory effect on the cellular protein expression of P450 3A4.CONCLUSION: Propofol may be a potential CYP3A4 inhibitor as this anesthetic can inhibit isoenzyme activity significantly and reduce the metabolic rate of CYP3A4 substrates. This inhibition occurs at post-expression level, and concentration of propofol used clinically does not affect CYP3A4 protein expression. propofol may thus induce drug interaction of cytochrome P450 3A4 activity at the dosage used clinically.