Suppression of the p66shc adapter protein by protocatechuic acid prevents the development of lung injury induced by intestinal ischemia reperfusion in mice.

BACKGROUND: Intestinal ischemia/reperfusion (I/R) causes severe histological injury, reactive oxygen species activation, and cell apoptosis in the lung. In this study, we investigated, using a murine intestinal I/R model, the effect of a polyphenolic compound, protocatechuic acid (PCA), in modulation of ShcA and in protection of the lung from I/R-induced injury. METHODS: Fifty ICR mice were randomly divided into five groups, including a control group, intestinal I/R group, control + PCA group, I/R + PCA low-dose group, and I/R + PCA high-dose group. The I/R and I/R + PCA groups were subjected to mesenteric arterial ischemia for 45 minutes and reperfusion for 90 minutes. The control and control + PCA groups underwent a surgical procedure that included isolation of the superior mesenteric artery without occlusion. In all PCA-pretreated groups, the mice received intraperitoneal PCA administration for three consecutive days. Serum specimens were collected for measuring tumor necrosis factor-α and interleukin 6, while lung tissues were harvested for histopathologic assessment including glutathione (GSH) and GSH peroxidase assay. Lung expression of p66shc, phosphorylated p66shc, manganese superoxide dismutase, caspace-3, and Bcl-xL were determined by Western blotting for protein level and semiquantitative reverse transcription-polymerase chain reaction analysis for mRNA level. RESULTS: PCA pretreatment markedly reduced I/R-induced lung injury as indicated by histological alterations; the decreases in tumor necrosis factor-α, interleukin 6, and caspase-3 expression levels; and the increases in GSH, GSH peroxidase, manganese superoxide dismutase, and Bcl-xL levels in the lung. Moreover, PCA treatment down-regulated p66shc expression and phosphorylation. CONCLUSION: PCA has a significant protective effect in lung injury induced by intestinal I/R. The protective effect of PCA may be attributed to the suppression of p66shc and the modulation of downstream antioxidative/antiapoptotic factors.

Microbial transformation of deoxyandrographolide by Alternaria alternata AS 3.4578.

Biotransformation of deoxyandrographolide (1) by Alternaria alternata AS 3.4578 gave five derivatives identified by spectral methods including 2D NMR as the known dehydroandrographolide (2) and 9beta-hydroxy-dehydroandrographolide (3) and the new compounds 9beta-hydroxy-deoxyandrographolide (4), 3alpha,17,19-trihydroxy-8,13-ent-labdadien-15,16-olide (5) and 3-oxo-9beta-hydroxy-deoxyandrographolide (6).

Protective effect of carnosol on lung injury induced by intestinal ischemia/reperfusion.

PURPOSE: Carnosol is a phenolic diterpene that has potent antioxidant and anti-inflammatory activities. The purpose of this study was to investigate the preconditioning effects of carnosol on lung injury induced by intestinal ischemia/reperfusion (II/R). METHODS: Rats were divided into control, II/R, and carnosol groups. The II/R model was established by clamping the superior mesenteric artery for 1 h and reperfusion at 2, 4, and 6 h after ischemia. The carnosol group received 3 mg/kg carnosol intraperitoneally 1 h before the operation. The rats were then euthanized, and blood and lung specimens were obtained for analysis. RESULTS: The II/R induced lung injury, characterized by histological changes and significant increasing of bronchoalveolar lavage fluid protein. The activity of lung tissue superoxide was weakened, the tissue myeloperoxidase activity and serum interleukin-6 level increased significantly in II/R groups. A strong positive expression of lung intercellular adhesion molecule-1 (ICAM-1) and nuclear factor kappa B (NF-kappaB) were observed. Pretreatment with carnosol markedly reduced lung injury by increasing the tissue superoxide activity and decreasing the myeloperoxidase activity and interleukin-6 level, which was parallel to the decreased expression of ICAM-1 and NF-kappaB. CONCLUSION: Carnosol was able to ablate lung injury induced by II/R, partly attributed to the inhibition of NF-kappaB activation.

Preparative separation of four major bufadienolides from the Chinese traditional medicine, Chansu, using high-speed counter-current chromatography.

A preparative, high-speed, counter-current chromatographic (HSCCC) method for the isolation and purification of bufadienolides from Chansu was successfully developed by using stepwise elution with a two-phase solvent system composed of n-hexane: chloroform: methanol: water (4:1:2.5:5 and 4:1:4:5, v/v). A total of 7.5 mg of cinobufotalin (1), 8.0 mg of bufalin (2), 14.0 mg of cinobufagin (3) and 9.5 mg of resibufogenin (4) were obtained in a one-step separation from 80 mg of the crude extract with purities of 93.2%, 98.7%, 99.2%, and 99.4%, respectively. The chemical structures were determined from 1H NMR and 13C NMR spectroscopic data.

Efficient isolation and purification of five products from microbial biotransformation of cinobufagin by high-speed counter-current chromatography.

An efficient separation method of using high-speed counter-current chromatography was successfully established to directly purify cytotoxic transformed products of cinobufagin by Cordyceps militaris. The two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water (4:6:3:4, v/v) was used in high-speed counter-current chromatography. A total of 9 mg of 4beta,12alpha-dihydroxyl-cinobufagin (1), 15 mg of 12beta-hydroxyl-cinobufagin (2), 8 mg of 5beta-hydroxyl-cinobufagin (3), 12 mg of deacetylcinobufagin (4) and 6 mg of 3-keto-cinobufagin (5) were obtained in a one-step separation from 400 mg of the crude extract with purity of 98.7, 97.2, 90.6, 99.1 and 99.4%, respectively, as determined by HPLC. Their chemical structures were identified on the basis of (1)H-NMR and (13)C-NMR technology. All products (1-5) showed the potent activities against human carcinoma cervicis (Hela) and malignant melanoma (A375) cells in vitro.

Structural determination of two new triterpenoids biotransformed from glycyrrhetinic acid by Mucor polymorphosporus.

Five hydroxylated derivatives of glycyrrhetinic acid by Mucor polymorphosporus were isolated. Among them, 6beta, 7beta-dihydroxyglycyrrhentic acid (2) and 27-hydroxyglycyrrhentic acid (3) are new compounds. Their chemical structures were identified by spectral methods including 2D-NMR.

Protection of Veratrum nigrum L. var. ussuriense Nakai alkaloids against ischemia-reperfusion injury of the rat liver.

AIM: To investigate the protective effects and possible mechanisms of Veratrum nigrum L.var. ussuriense Nakai alkaloids (VnA) on hepatic ischemia/reperfusion (I/R) injury in rats. METHODS: Forty male Wistar rats were randomly divided into four experimental groups (n = 10 in each): (A) Control group (the sham operation group); (B) I/R group (pretreated with normal saline); (C) Small-dose (10 microg/kg) VnA pretreatment group; (D) Large-dose (20 microg/kg) VnA pretreatment group. Hepatic ischemia/reperfusion (Hepatic I/R) was induced by occlusion of the portal vein and the hepatic artery for 90 min, followed by reperfusion for 240 min. The pretreatment groups were administered with VnA intraperitoneally, 30 min before surgery, while the control group and I/R group were given equal volumes of normal saline. Superoxide dismutase (SOD) activity, myeloperoxidase (MPO) activity and nitric oxide (NO) content in the liver tissue at the end of reperfusion were determined and liver function was measured. The expression of intercellular adhesion molecule-1 (ICAM-1) and E-selectin (ES) were detected by immunohistochemical examinations and Western blot analyses. RESULTS: The results showed that hepatic I/R elicited a significant increase in the plasma levels of alanine aminotransferase (ALT: 74.53 +/- 2.58 IU/L vs 1512.54 +/- 200.76 IU/L, P < 0.01) and lactic dehydrogenase (LDH: 473.48 +/- 52.17 IU/L vs 5821.53 +/- 163.69 IU/L, P < 0.01), as well as the levels of MPO (1.97 +/- 0.11 U/g vs 2.57 +/- 0.13 U/g, P < 0.01) and NO (69.37 +/- 1.52 micromol/g protein vs 78.39 +/- 2.28 micromol/g protein, P < 0.01) in the liver tissue, all of which were reduced by pretreatment with VnA, respectively (ALT: 1512.54 +/- 200.76 IU/L vs 977.93 +/- 89.62 IU/L, 909.81 +/- 132.76 IU/L, P < 0.01, P < 0.01; LDH: 5821.53 +/- 163.69 IU/L vs 3015.44 +/- 253.01 IU/L, 2448.75 +/- 169.4 IU/L, P < 0.01, P < 0.01; MPO: 2.57 +/- 0.13 U/g vs 2.13 +/- 0.13 U/g, 2.07 +/- 0.05 U/g, P < 0.01, P < 0.01; NO: 78.39 +/- 2.28 micromol/g protein vs 71.11 +/- 1.73 micromol/g protein, 68.58 +/- 1.95 micromol/g protein, P < 0.05, P < 0.01). The activity of SOD (361.75 +/- 16.22 U/mg protein vs 263.19 +/- 12.10 U/mg protein, P < 0.01) in the liver tissue was decreased after I/R, which was enhanced by VnA pretreatment (263.19 +/- 12.10 U/mg protein vs 299.40 +/- 10.80 U/mg protein, 302.09 +/- 14.80 U/mg protein, P < 0.05, P < 0.05). Simultaneously, the histological evidence of liver hemorrhage, polymorphonuclear neutrophil infiltration and the overexpression of ICAM-1 and E-selectin in the liver tissue were observed, all of which were attenuated in the VnA pretreated groups. CONCLUSION: The results demonstrate that VnA pretreatment exerts significant protection against hepatic I/R injury in rats. The protective effects are possibly associated with enhancement of antioxidant capacity, reduction of inflammatory responses and suppressed expression of ICAM-1 and E-selectin.