Effect of acute heat stress on heat shock protein 70 messenger RNA and on heat shock protein expression in the liver of broilers

1. The synthesis of heat shock protein 70 (Hsp70) mRNA and the expression of Hsp70 in the liver of broiler chickens submitted to acute heat stress (35 degrees C for 5 h) was investigated.2. Hsp70 expression was detected by SDS-PAGE and Western blot analysis using a polyclonal antiserum against Hsp70 of Blastocladiella emersonii. The specific signal of Hsp70 mRNA was analysed by Northern blot using as probe a Hsp70 cDNA of B. emersonii.3. An increase in the amount of Hsp70 was detected from the first up to the fifth hour of acute heat exposure. This increase in the amount of Hsp70 was accompanied by an increase in Hsp70 mRNA which peaked at 3 h.4. This study shows that the heat induced increase in Hsp70 mRNA and protein in broiler liver, in vivo, are time dependent, similar to that in mammals.

CHOLESTEROL MODULATION OF LIPID PROTEIN INTERACTIONS IN LIVER MICROSOMAL MEMBRANE - A SPIN LABEL STUDY

ESR spectra of spin probes were used to monitor lipid-protein interactions in native and cholesterol-enriched microsomal membranes. In both systems composite spectra were obtained, one characteristic of bulk bilayer organization and another due to a motionally restricted population, which was ascribed to lipids in a protein microenvironment. Computer spectral subtractions revealed that cholesterol modulates the order/mobility of both populations in opposite ways, i.e., while the lipid bilayer region gives rise to more anisotropic spectra upon cholesterol enrichment, the spectra of the motionally restricted population become indicative of increased mobility and/or decreased order. These events were evidenced by measurement of both effective order parameters and correlation times. The percentages of the motionally restricted component were invariant in native and cholesterol-enriched microsomes. Variable temperature studies also indicated a lack of variation of the percentages of both spectral components, suggesting that the motionally restricted one was not due to protein aggregation. The results correlate well with the effect of cholesterol enrichment on membrane-bound enzyme kinetics and on the behavior of fluorescent probes [Castuma & Brenner (1986) Biochemistry 25, 4733-4738]. Several hypothesis are put forward to explain the molecular mechanism of the cholesterol-induced spectral changes.

Factors affecting metabolic and electrolyte changes after reperfusion in liver transplantation

Background. The metabolic and electrolyte changes were evaluated after various durations of cold and warm ischemia times to correlate ASA status with hemodynamic changes that may affect the severity of the reperfusion syndrome.Patients and methods. Sixty-one patients who underwent liver transplantation (OLT) were monitored by arterial pH, PaO2, PaCO2, HCO3, BE, K+, Ca2+, Na+, GL, and serial Ht at three specific times: after the skin incision (baseline), 10 minutes before reperfusion (T-2), and 10 minutes after reperfusion (T-3). Changes in metabolic parameters were correlated with ASA status, hemodynamic changes, time of OLT, as well as cold and warm ischemia times.Results. The pH in ASA IV patients was significantly lower at T-1 and T-3, and PCO2 higher in ASA V at T-1. A significant correlation was observed between pH, PaCO2, HCO3 BE, Na+, Ca2+, and glucose with the phase of the procedure. The pH and HCO3 decreased significantly from T-1 and T-2, increasing during T-3. Ca2+ fell from T-1 to T-2 increasing in T-3. Mean glucose and sodium levels increase from T-1 to T-3. Mean BE dropped from T-1 to T-2 and increased at T-3 without a significant correlation between the metabolic parameters in any phase of the study and the cold or warm ischemia times. Patients with a high ASA status showed an increased risk for cardiovascular collapse after reperfusion.Conclusions. Patients with advanced ASA status are more prone to metabolic and acid-base disturbances during reperfusion, without any relation to the cold or warm ischemia times. High ASA status shows an increased risk for cardiovascular collapse after reperfusion.

Placental glutathione S-transferase correlates with cellular proliferation during rat tonguie carcinogenesis induced by 4-nitroquinoline 1-oxide

Taking into consideration that glutatione S-transferase (GST) and cellular proliferation play a crucial role during carcinogenesis, the goal of this study was to investigate the expression of placental GST, called GST-P, and proliferating cellular nuclear antigen (PCNA) by means of immunohistochemistry during rat tongue carcinogenesis induced by 4-nitroquinoline 1-oxide (4NQO). This is a useful model for studying oral squamous cell carcinoma phase by phase. Male Wistar rats were distributed into three groups of 10 animals each and treated with 50 ppm 4NQO solution by drinking water for 4, 12 or 20 weeks. Ten animals were used as negative control. GST-P positive foci were detected in non-neoplastic oral cells at 4 weeks of 4NQO administration. In the same way, GST-P positive cells were detected in pre-neoplastic lesions and squamous cell carcinomas induced after 12 and 20 weeks-treatment, respectively. None of the control animals expressed GST-P positive cells. Regarding cellular proliferation, PCNA positive nuclei were higher at 12 and 20 weeks following 4NQO exposure (p < 0.05) when compared to negative control. These results suggest that the expression of GST-P is correlated with cellular proliferation, in which GST-P is associated with risk and progression of oral cancer, whereas PCNA is closely involved during neoplastic conversion. (c) 2007 Published by Elsevier GmbH.

Liver lesions produced by aflatoxins in Rana catesbeiana (bullfrog)

This study describes alterations induced in Rana catesbeiana (bullfrog) liver after extended dietary exposure to aflatoxins (AFs). Bullfrogs of both sexes were fed for 120 days a commercial chow blended with a rice bran-based mixture of Al's containing 667.0, 11.65, 141.74, and 3.53 mg/kg of AFs B1, B2, G1, and G2, respectively. Animals were sacrificed on study days 45, 90, and 120. Severe and progressive liver lesions with structural collapse, increased hepatocyte and biliary duct cell proliferation, appearance of basophilic hepatocytes, and diffuse scarring, were observed at all time points. There were no quantitative alterations in the liver melanomacrophage centers of the AFs-exposed animals. Increased amounts of lipid hydroperoxides, indicative of ongoing oxidative stress, were more evident in the Addutor magnum muscle than in the AFs-damaged livers. No tumors were found in the R. catesbeiana livers after 120 days of exposure to relatively high doses of AFs. (c) 2006 Elsevier B.V. All rights reserved.

ALTERED FOCI OF HEPATOCYTES IN RATS INITIATED WITH DIETHYLNITROSAMINE AFTER PROLONGED FASTING

The influence of fasting on the potential of diethylnitrosamine (DEN) to initiate liver cucinogenesis was tested in a medium-term assay using the development of putative preneoplastic altered foci of hepatocytes (AFH) as the endpoint. Male Wistar rats fasted for 48 hr were given a single ip injection of DEN (200 mg/kg body weight). Partial hepatectomies were carried out at wk 3 and the rats were killed at wk 8. Fasted rats exhibited a small increase in the numbers of AFH with glutathione S-transferase in the placental form and eosinophilic AFH when compared with non-fasted animals. However, after a 6-wk exposure to 0.05% sodium phenobarbital in the diet, there were no differences in the numbers of AFH between fasted and non-fasted animals. Fasting also increased DEN-dependent centrilobular cell necrosis and specifically drug metabolism as indicated in vivo by a decreased time of paralysis of the lower limbs induced by zoxazolamine (40 mg/kg body weight, ip) and by an unaltered sleeping time induced by sodium pentobarbital (40 mg/kg body weight, ip). The results indicate that although fasting during the initiation stage of carcinogenesis increases DEN hepatotoxicity, it does not interfere quantitatively with the development of liver preneoplastic lesions.

Purification and partial characterization of cathepsin D from porcine (Sus scrofa) liver using affinity chromatography

Cathepsin D, a lysosomal aspartic protease, has been purified from porcine liver using a combination of pepstatin-A agarose and Affi-Gel Blue affinity chromatography, followed by size-exclusion chromatography. The purified protein consists of two polypeptide chains of 15 and 30 kDa, and has an isoelectric point of 6.8. Porcine liver cathepsin D has maximum activity at pH 2.5-3.0 as determined by its activity against hemoglobin, with a K-cat of 14.3 s(-1) and a k(cat)/K-M of 2.70 x 10(6) s(-1) M-1 as determined by the hydrolysis of a fluorogenic peptide substrate.