TNF-alpha induced DNA damage in primary murine hepatocytes

Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide, usually arising from a background of chronic inflammatory disease. Tumor necrosis factor alpha (TNF-alpha) is a pro-inflammatory cytokine produced in response to tissue injury, endotoxin exposure or infection and TNF-alpha signalling in hepatocytes is associated with an increase in oxidative stress. DNA is vulnerable to reactive oxygen species (ROS)-induced damage, which is highly mutagenic. Cells respond to DNA damage through the stabilisation of the tumor suppressor p53, which maintains genomic fidelity through induction of a cell cycle arrest in order to allow repair or elimination of the damaged cell through apoptosis. This study was carried out to determine if TNF-alpha caused oxidative DNA damage in primary cultures of murine hepatocytes and whether any damage would result in the induction of the tumor suppressor p53 and cell-cycle arrest. Using a modified Comet assay, to measure DNA damage we have demonstrated that TNF-alpha causes the formation of 8-oxo-deoxyguanosine (8-oxodG), an established marker of oxidative DNA damage, and a lesion associated with chronic hepatitis in human livers. In addition, the increase in DNA damage did not result in p53 stabilisation and TNF-alpha caused an increase in cell-cycle progression. We believe that this study indicates a possible putative role for TNF-alpha in the early stages of malignant transformation of hepatocytes.     

Effects of controlled mechanical ventilation on respiratory muscle contractile properties in rabbits

OBJECTIVE: We examined in rabbits the effects of more than 48 h of mechanical ventilation on the contractile properties and fiber type adaptations of the respiratory muscles. DESIGN AND SETTING: Experimental prospective study in a university laboratory. ANIMALS AND INTERVENTIONS: Nineteen rabbits were randomly allocated to two groups: control (n=10) or mechanically ventilated (MV; n=9) for 51+/-3 h. MEASUREMENTS AND RESULTS: Respiratory muscles contractile properties were analyzed before and after a fatigue protocol using in vivo isometric 1-s tetanic contraction characteristics in both muscles: peak tetanic force, contraction time, relaxation time, and total contraction time. Both muscle fiber type proportions, diameter, and cross-sectional areas were measured using ATPase staining. The MV rabbits showed significant weight loss in both muscles, accompanied by a reduced peak tetanic force (9.96+/-3.2 vs. 7.44+/-2.2 N for diaphragm of control and MV animals respectively), fatigue resistance index, and increased relaxation time (57.5+/-8.7 vs. 85.8+/-9.4 ms for diaphragm of control and MV animals) and contraction time. These impairments in the MV group worsened after the fatigue runs. Both muscle showed a significant atrophy of type IIa and IIb fibers but a stability in type I fibers cross-sectional area. CONCLUSIONS: Mechanical ventilation in rabbits produces alterations in contractile properties of the diaphragm and 5th external intercostal muscle, increases both muscles fatigue, and promotes atrophy of type II fibers.     

Effects of angiotensin II type 1 receptor blockade in ApoE-deficient mice with post-ischemic heart failure

We investigated in mice whether atherosclerosis exacerbates the development of post-ischemic heart failure and alters the beneficial effects of long-term angiotensin II type 1 receptor blockade in this model. ApoE-deficient (ApoE(-/-)) and C57BL/6J (C57) mice with myocardial infarction (coronary ligation) received vehicle (C57 and ApoE(-/-)) or irbesartan (Ir, 50mg/kg/d orally, C57-Ir and ApoE(-/-)-Ir). Ten months post myocardial infarction, survival rates were similar in C57 (58%) and ApoE(62%). Atherosclerosis induced no significant alteration in blood pressure, cardiac output (fluospheres), total peripheral resistance, or shortening fraction (echocardiography) but increased renal resistance (+50%, P<0.05). Chronic Ir treatment significantly improved survival to a similar extent in both C57-Ir (85%) and ApoE(-/-)-Ir (86%). It also decreased blood pressure to a similar extent in both strains (-16% and -18%, both P<0.05). In C57-Ir mice, Ir did not modify cardiac output or total peripheral resistance, but it decreased renal resistance (-28%, P<0.001) and left-ventricular weight (-28%, P<0.05). In ApoE(-/-)-Ir mice, Ir limited atherosclerotic lesions (-13%, P<0.05), increased cardiac output (+28%, P<0.05) and shortening fraction (+24%, P<0.05), and decreased total peripheral resistance (-33%, P<0.01), renal resistance (-61%, P<0.001), and left-ventricular weight (-27%, P<0.001). In conclusion, atherosclerosis does not worsen heart failure development in mice and, although the beneficial cardiovascular effects of AT1 receptor blockade are greater in ApoE(-/-) than in C57, reduction in mortality is similar in both strains.     

Non-AIDS-related malignancies: prognostic and treatment

Malignant tumors currently rank among the leading cause of morbidity and mortality in patients infected with HIV. The survival is increased for patients who benefits from better control of viral replication and from progress in prevention of opportunistic infections. These patients are thus exposed to an increased risk of cancer. The most frequent malignant tumors not associated with AIDS stage are Hodgkin disease, germ-cell tumors, cutaneous cancer, head and neck cancer and lung cancer. The goal of this chapter is to describe the clinical and histological pattern of these malignant tumors, and to propose therapeutic guidelines for HIV patients with cancer.     

Decreased arteriolar density in endothelial nitric oxide synthase knockout mice is due to hypertension, not to the constitutive defect in endothelial nitric oxide synthase enzyme

BACKGROUND : Hypertension in endothelial nitric oxide synthase knockout (eNOS-/-) mice is believed to be partly due to altered vasodilatation. However, nitric oxide (NO) is also known to play an important part in angiogenesis. OBJECTIVE : To investigate whether capillary and arteriolar density were impaired in eNOS-/- mice, as this could account for increased vascular resistance and hypertension. METHODS : Using immunohistochemistry with mouse monoclonal smooth muscle alpha-actin antibody to detect arterioles and rabbit polyclonal fibronectin antibody to detect capillaries, we quantified arteriolar and capillary density in the left ventricle and in the gracilis muscle from eNOS-/- mice compared with those in C57BL6J littermates (n = 6-8) in 8- and in 12-week-old mice. In a second set of experiments, we treated 8-week-old normotensive eNOS-/- mice with the antihypertensive vasodilator, hydralazine, for 1 month. RESULTS : Eight-week-old eNOS-/- mice were normotensive and presented similar arteriolar and capillary densities in cardiac and skeletal muscles compared with those in eNOS+/+ mice. Twelve-week-old eNOS/- mice were hypertensive (mean arterial pressure 118 +/- 21 mmHg compared with 64 +/- 2 mmHg; P < 0.05). Capillary densities were similar in eNOS-/- mice and eNOS+/+ mice in the heart (4154 +/- 123 and 4051 +/- 247/mm2, respectively) and in skeletal muscle (961 +/- 40 and 1025 +/- 41/mm2, respectively). Arteriolar densities were 15% lower in skeletal muscle and in the heart in eNOS-/- mice than in the eNOS+/+ control group (P < 0.05). Hydralazine prevented hypertension and arteriolar rarefaction in eNOS-/- mice, whereas capillary density was unaffected by treatment with the vasodilator. CONCLUSION : In young non-hypertensive eNOS-/- mice, the lack of eNOS did not affect microvascular densities in either of the muscles studied. In adult hypertensive eNOS-/- mice, we observed a lower arteriolar density, but a similar capillary density compared with controls. Hydralazine prevented hypertension and arteriolar rarefaction in adult mice, suggesting a non-NO-dependent pathway. Capillary density was not affected by hydralazine.     

In vitro increase in chloroquine accumulation induced by dihydroethano- and ethenoanthracene derivatives in Plasmodium falciparum-parasitized erythrocytes

The effects of a series of dihydroethano- and ethenoanthracene derivatives on chloroquine (CQ) accumulation in CQ-susceptible strain 3D7 and CQ-resistant clone W2 were assessed. The levels of CQ accumulation increased little or none in CQ-susceptible strain 3D7 and generally increased markedly in CQ-resistant strain W2. At 10 microM, 28 compounds yielded cellular accumulation ratios (CARs) greater than that observed with CQ alone in W2. At 10 microM, in strain W2, 21 of 31 compounds had CQ CARs two or more times higher than that of CQ alone, 15 of 31 compounds had CQ CARs three or more times higher than that of CQ alone, 13 of 31 compounds had CQ CARs four or more times higher than that of CQ alone, and 9 of 31 compounds had CQ CARs five or more times higher than that of CQ alone. At 1 microM, 17 of 31 compounds had CQ CARs two or more times higher than that of CQ alone, 12 of 31 compounds had CQ CARs three or more times higher than that of CQ alone, 6 of 31 compounds had CQ CARs four or more times higher than that of CQ alone, and 3 of 31 compounds had CQ CARs five or more times higher than that of CQ alone. At 1 microM, 17 of 31 compounds were more potent inducers of CQ accumulation than verapamil and 12 of 31 compounds were more potent inducers of CQ accumulation than promethazine. The nature of the basic group seems to be associated with increases in the levels of CQ accumulation. At 1 and 10 microM, 10 of 14 and 13 of 14 compounds with amino group (amines and diamines), respectively, had CARs >or=3, while at 1 and 10 microM, only 1 of the 13 derivatives with amido groups had CARs >or=3. Among 12 of the 31 compounds which were more active inducers of CQ accumulation than promethazine at 1 microM, 10 had amino groups and 1 had an amido group.     

Assessment of the cytoprotective role of adenosine in an in vitro cellular model of myocardial ischemia

This work aimed to detect functional adenosine receptors in isolated rat cardiomyocytes and to study the influence of stimulation of these receptors in an in vitro model of ischemia. Cultures of cardiomyocytes were prepared from newborn rat ventricles. The contractions were photometrically monitored. In this preparation, adenosine induced a positive chronotropic response. This effect was reproduced by CGS 21680 (2-(4-[2-carboxyethyl]-phen-ethyl-amino) adenosine-5'N-ethylunosamide), a specific adenosine A(2) receptor agonist, and antagonized by DMPX (3,7-dimethyl-1-propargylxanthine), an adenosine A(2) receptor antagonist. However, R-PIA (R-N(6)-(2-phenylisopropyl)-adenosine; a specific adenosine A(1) receptor agonist) induced a negative chronotropic effect that was abolished by its corresponding adenosine A(1) antagonist DPCPX (1,3-dipropyl-8-cyclo-pentyl-adenosine). Substrate-free hypoxia, as simulation of ischemia, induced a progressive decrease and then arrest of spontaneous cell contractions. The spontaneous rhythmic contractile activity was restored during reoxygenation following simulated ischemia. Adenosine A(1) receptor stimulation with R-PIA induced a decrease of hypoxia-induced damage. This effect was antagonized by DPCPX, an adenosine A(1) receptor antagonist. Conversely, the cells treated with CGS 21680 did not display complete recovery after reoxygenation. In addition, this effect was abolished by DMPX, since the cells recovered normal function after reoxygenation. To conclude, it appeared that cardiomyocytes possess both functional adenosine A(1) and A(2) receptors and that only the activation of adenosine A(1) receptor had a cytoprotective effect against simulated ischemia-induced cardiac cell injury.