Research communications in chemical pathology and pharmacology最新文献

Species-specific differences in the inflammatory response, specifically with regard to mast cells, have been proposed to explain the phenotypic variation among dystrophin-deficient humans, and mdx mice (Gorospe et al., 1994). To test this hypothesis we have intramuscularly injected a mast cell secretogogue into both dystrophin-negative mdx and dystrophin-positive normal mice. Mast cell activity was determined by measuring the activity of mast cell tryptase, while creatine kinase activity was used to determine the course of muscle damage in vivo. Area of damage around the injection site was measured at autopsy, and used as an indication of relative sensitivity to the secretogogue effect of compound 48/80. Mdx mice exhibited more damage in response to intramuscular injection than normal control mice. In addition, mdx mice showed a substantial increase in plasma tryptase activity, followed by a large increase in muscle creatine kinase activity. On the other hand, dystrophin-positive normal controls injected with 48/80 liberated little CK or tryptase activity. These results are consistent with the hypothesis that species-specific differences in mast cell activity, or sensitivity to mast cell products could account for the variation in pathology seen in dystrophin-deficient animals.

Primary biliary cirrhosis (PBC) patients showed increased ceruloplasmin activity and Cu content in the serum as compared to the control group. Moreover, Cu, Fe, and Zn contents were measured in the livers of autopsied cases. The PBC patients showed increased Cu content and reduced Fe and Zn levels. In PBC cases, Cu transportation via bile is disturbed due to collapse of the interlobular bile ducts and consequently Cu accumulates in the liver. Thus, it is likely that serum ceruloplasmin activity increases to metabolize the accumulated hepatic Cu and that Cu content increases in the serum as a ceruloplasmin-bound form. Furthermore, reduction of Fe and Zn contents in the liver of the PBC patients indicated the possible relationship of ceruloplasmin to Fe and Zn metabolism as well as Cu metabolism.

We measured serum levels of endotoxin, cytokines, and eicosanoids and investigated their relationship to serum complement levels in patients with sepsis. Serum endotoxin (Et) levels (5.3 +/- 2.4 pg/ml) were within the normal range, but levels of tumor necrosis factor-alpha (TNF-alpha, 114 +/- 104.94 pg/ml), interleukin 6 (IL-6, 86.7 +/- 50.9 pg/ml), interleukin 8 (IL-8, 86.8 +/- 49.7 pg/ml), type-II phospholipase A2 (type II PLA2, 211.3 +/- 193.9 ng/ml), leukotriene B4 (LTB4, 88.7 +/- 27.2 pg/ml), thromboxane B2 (TXB2, 58.7 +/- 50.9 pg/ml) and 6-keto-prostaglandin F1 alpha (PGF1 alpha, 21.0 +/- 11.0 pg/ml) levels were above normal. Levels of C3a (1088.4 +/- 83.8.7 ng/ml) and C4a (1951.5 +/- 1697.8 ng/ml) were also above normal; C3 (66.0 +/- 25.6 mg/dl) and C4 (23.6 +/- 5.3 mg/dl) were within the normal range, and C5a was lower than the detectable limit in all but one of the subjects. Serum TNF-alpha was significantly correlated with C3a (p < 0.001). Serum IL-6 had a significant negative correlation with C3 (p = 0.002) and C4 (p = 0.010). Type II PLA2 was significantly correlated with C3a (p < 0.001). There were no significant correlations between serum Et or IL-8 and serum C3, C4, C3a or C4a. Our findings suggest that increased levels of TNF-alpha, IL-6, and Type II PLA/ in patients with sepsis contribute to activation of the complement system.

We investigated the effects of bradykinin on intracellular oxidative stress in bovine aortic endothelial cells using a hydroperoxide-sensitive fluorescent dye, 2',7'-dichlorofluorescein (DCFH), and a laser scanning confocal microscope. Bradykinin induced an immediate increase in intracellular Ca2+ concentration, and stimulated the oxidation of DCFH in cultured endothelial cells. This bradykinin-induced oxidation of DCFH was inhibited by pretreatment with N-(2-mercaptopropionyl)-glycine (MPG) and 1,3-dimethyl-thiourea (DMTU), scavengers of hydroxyl radical, and the removal of extracellular Ca2+ but was unaffected by NG-nitro-L-arginine or NG-monomethyl-L-arginine, both inhibitors of nitric oxide (NO) synthase. On the other hand, pretreatment with indomethacin and aspirin, inhibitors of cyclooxygenase, inhibited bradykinin-induced oxidation of DCFH. These findings suggest that bradykinin increases intracellular Ca2+ and stimulates the generation of hydroxyl radical-like reactive oxygen species (scavenged by MPG or DMTU) via the cyclooxygenase pathway but not via the reaction of NO and superoxide anion.

Long-Evans Cinnamon (LEC) mutant rat, which possesses a genetic defect of the block of copper exclusion pathway in the hepatocytes, is a good model for studying the mechanism of cellular copper toxicity. We have examined effects of copper toxicity on DNA synthesis and protein phosphorylation upon growth stimulation by treating LEC rat primary-cultured hepatocytes with EGF and insulin. The proportion of DNA-synthesizing S-phase cells in LEC rat markedly decreased when compared to that of normal rat. However, the electrophoretic pattern and the signal intensity of 32P-labeled nuclear proteins in LEC rat were indistinguishable from those of normal rat. These results suggest that a cellular event other than protein phosphorylation required for the initiation of DNA synthesis upon growth stimulation is impaired by copper cytotoxicity.

Clonidine, an alpha 2-agonist, caused a concentration-dependent vasodilation of the mesenteric arterial beds in both normotensive and hypertensive rats. The clonidine-induced vasodilation was inhibited by NG-nitro-L-arginine, and the inhibition was reversed by L-arginine. The concentration-dependent vasodilation was not significantly different between normotensive and hypertensive rats. These results suggest that clonidine has an endothelium-dependent vasorelaxant action in the resistance artery such as rat mesenteric arterial bed, and the endothelium-dependent vasodilator effects of clonidine in the mesentery may be involved in the depressor effect of the drug.

We determined the plasma levels of type-II phospholipase A2 (type II PLA2), platelet-activating factor acetylhydrolase (PAFAH) leukotriene B4 (LTB4) and of several complements (C3a, C4a, and C5a), which are considered to be among the cytokines and eicosanoids involved in vascular endothelial disorders and that vary in concentration during sepsis. We investigated the relationship between those levels and those of ET-1 and TM levels in plasma. Plasma levels of type II PLA2, PAFAH, LTB4, C3a, C4a, ET-1, and TM at the time that sepsis was diagnosed in 30 patients were 218.3 +/- 179.9 ng/ml, 23.92 +/- 9.66 nmol/min/ml, 90.35 +/- 31.49 pg/ml, 838.73 +/- 2.30 pg/ml, 1951.46 +/- 1697.78 pg/ml, 6.98 +/- 4.08 pg/ml and 7.80 +/- 3.34 ng/ml, respectively. The C5a plasma level was below the limit of detection in all cases. There were significant correlations between type II PLA2 and ET-1 plasma levels (r = 0.39, p = 0.032) and C3a and ET-1 plasma levels (r = 0.60, p = 0.03). There were also significant correlations between type II PLA2 and TM levels in plasma (r = 0.76, p = 0.0017), PAFAH and TM plasma levels (r = 0.53, p = 0.037), LTB4 and TM plasma levels (r = 0.46, p = 0.016) and C4a and TM plasma levels (r = 0.58, p = 0.037). Results suggest that the elevation of type II PLA2, PAFAH, LTB4 and complement in plasma is involved in vascular endothelial disorders in patients with sepsis.

To investigate the roles of tumor necrosis factor-alpha (TNF-alpha), endothelin-1 (ET-1) and thrombomodulin (TM) in the plasma of patients with sepsis, plasma levels of endotoxin (Et), TNF-alpha, ET-1 and TM were determined in 30 such patients. Plasma levels of Et, TNF-alpha, ET-1 and TM at the time sepsis was diagnosed were 4.0 +/- 6.7 pg/ml, 98.5 +/- 92.1 pg/ml, 7.0 +/- 4.1 pg/ml and 7.8 +/- 3.3 ng/ml, respectively. There was no significant difference in the plasma Et level between the group of patients that survived (n = 13) or died (n = 17). Plasma levels of TNF-alpha, Et-1 and TM were significantly higher in the group that died than in the surviving group (TNF-alpha, p < 0.0001; ET-1, p = 0.028; TM, p = 0.0004). There were significant correlations between the plasma levels of TNF-alpha and ET-1, of TNF-alpha and TM, and of ET-1 and TM (r = 0.37, p < 0.046; r = 0.61, p = 0.0008; r = 0.63, p = 0.004), respectively. Results suggest that TNF-alpha is involved in the production of ET-1 and TM. Both of those substances appear to be involved in the morbidity of sepsis, and their plasma levels reflect its severity.

To examine the relationship between arachidonic acid metabolism and sepsis, we determined the blood concentration of platelet-activating factor acetylhydrolase (PAFAH) and of arachidonic acid cascade substances, leukotriene B4 (LTB4), thromboxane B2 (TXB2), and 6-keto-prostaglandin Fla (PGF1a), in patients with clinical sepsis. Blood concentrations of PAFAH, LTB4, TXB2 and PGF1a at the time of the initial diagnosis of sepsis were 23.9 +/- 9.7 nmol/min/mL, 90.4 +/- 31.5 pg/mL, 55.3 +/- 44.1 pg/mL and 22.3 +/- 11.9 pg/mL, respectively. There was a significant correlation between the blood PAFAH and LTB4 concentration (r = 0.520, p = 0.0033) and the diagnosis of sepsis. There also was a significant correlation between the blood PAFAH and TXB2 concentration (r = 0.460, p = 0.0311). There was no correlation between the blood PAFAH and PGF1a concentrations. Elevated LTB4 and TXB2 concentrations showed a significant correlation with the PAFAH concentration in patients with sepsis, suggesting a direct relationship between these substances.