Rationale We previously reported that type VI collagen deposition increases in the infarcted myocardium in vivo. To date, a specific role for this non-fibrillar collagen has not been explored in the setting of myocardial infarction (MI). Objective To determine whether deletion of type VI collagen in an in vivo model of post-MI wound healing would alter cardiac function and remodeling in the days to weeks after injury. Methods and Results Wild type (WT) and Col6a1-/- mice were subjected to MI followed by serial echocardiographic and histological assessments. At 8 weeks post-MI, infarct size was significantly reduced, ejection fraction was significantly preserved (43.9 ± 3.3% vs. 29.1 ± 4.3% for WT) and left ventricular (LV) chamber dilation was attenuated in the Col6a1-/- MI group (25.8 ± 7.9% increase vs. 62.6 ± 16.5% for WT). The improvement in cardiac remodeling was evident as early as 10 days post-MI in the Col6a1-/- mice. Myocyte apoptosis within the infarcted zones was initially greater in the Col6a1-/- group 3 days post-MI but by day 14 this was significantly reduced. Collagen deposition was also reduced in the infarcted and remote areas of the Col6a1-/- hearts. The reductions in chronic myocyte apoptosis and fibrosis are critical events leading to improved long-term remodeling and functional outcomes. Conclusions These unexpected results demonstrate for the first time that deletion of type VI collagen in this knockout model plays a critical protective role following MI by limiting infarct size, chronic apoptosis, aberrant remodeling and fibrosis leading to preservation of cardiac function.
We determined if prolonged isoproterenol (Iso) infusion in rats impaired the ability of the β2-adrenergic agonist terbutaline to increase alveolar liquid clearance (ALC). We infused rats with Iso (at rates of 4, 40, or 400 μg · kg−1 · h−1) or vehicle (0.001 N HCl) for 48 h using subcutaneously implanted miniosmotic pumps. After this time, the rats were anesthetized, and ALC was determined (by mass-balance after instillation of Ringer lactate containing albumin into the lungs) under baseline conditions and after terbutaline administration. Baseline and terbutaline-stimulated ALC in vehicle-infused rats averaged, respectively, 19.6 ± 1.2% (SE) and 44.7 ± 1.5%/h. The ability of terbutaline to increase ALC was eliminated at 400 μg · kg−1 · h−1 Iso, inhibited by 26% at 40 μg · kg−1 · h−1 Iso, and was not affected by 4 μg · kg−1 · h−1 Iso. β-adrenergic receptor (βAR) density of freshly isolated alveolar epithelial type II (ATII) cells from Iso-infused rats was reduced by the 40 and 400 μg · kg−1 · h−1 infusion rates. These data demonstrate that prolonged exposure to β-agonists can impair the ability of β2-agonists to stimulate ALC and produce ATII cell βAR downregulation.
We previously found that prolonged isoproterenol (Iso) infusion in rats impaired the ability of beta-adrenoceptor (beta-AR) agonists to increase alveolar liquid clearance (ALC). Here, we determined if postreceptor defects in beta-AR signaling contribute to this impairment. Iso was infused using subcutaneous miniosmotic pumps (4, 40, or 400 microg. kg-1. h-1) in rats for 48 h. At this time, forskolin-stimulated ALC was measured by mass balance. Forskolin-stimulated ALC [33.4 +/- 2.1%/h (mean +/- SE) in vehicle-infused rats] was reduced by 25 and 38%, respectively, after the 40 and 400 microg. kg-1. h-1 Iso infusions. The ability of forskolin to increase cAMP was reduced by 70% in alveolar type II (ATII) cells isolated from rats infused with 400 microg. kg-1. h-1 Iso. Additionally, the ability of the stable cAMP analog 8-bromoadenosine-3',5'-cyclic monophosphorothioate, Sp-isomer, to increase ALC (48.7 +/- 3.0% in vehicle-infused rats) was reduced by 25 and 51%, respectively, after the 40 and 400 microg. kg-1. h-1 infusions. Finally, the ability of cAMP to increase protein kinase A activity was eliminated in ATII cells isolated from rats infused with Iso at 400 microg. kg-1. h-1. These data demonstrate that prolonged beta-AR agonist exposure can impair alveolar epithelial beta-AR signaling downstream of the beta-AR.
SUMMARYInternalization of herpes simplex virus type 1 (HSV) KOS strain by HEp-2 cells was reversibly inhibited by pretreatment of cells with cytochalasins B and D. Internalization of virus following preincubation at 4 °C and temperature shift to 37 °C was normally preceded by a 5 to 8 min lag period and was complete within 20 to 30 min. A similar lag period followed HSV addition at 37 °C. Cytochalasin D was fivefold more active on HSV entry than cytochalasin B, with 50~ inhibition at 2 ~tM and 10 ~tM respectively. Inhibition was completely reversible, such that all cell-bound infectious virus was recovered upon removal of cytochalasin. In conjunction with previous reports, the activity of cytochalasin on HSV entry suggests that a change in cytoskeletal structure following virus attachment triggers a microfilament activity important for internalization of HSV by HEp-2 cells.
Hepatocellular carcinoma (HCC), one of the most lethal cancers, results in more than one million fatalities worldwide every year. In view of the limited therapeutic alternatives and poor prognosis of liver cancer, preventive control approaches, notably chemoprevention, have been considered to be the best strategy in lowering the present prevalence of the disease. Resveratrol, a naturally occurring antioxidant and antiinflammatory agent found in grapes and red wine, inhibits carcinogenesis with a pleiotropic mode of action. Recently, we have reported that dietary resveratrol significantly prevents chemically-induced liver tumorigenesis in rats. One of the mechanisms of resveratrol-mediated chemoprevention of hepatocarcinogenesis could be related to its antiinflammatory action through hepatic cyclooxygenase (COX-2) inhibition. Although several COX-2 inhibitors are known to exert chemopreventive efficacy, not all are considered ideal candidates for chemoprevention due to the risk of adverse cardiovascular events. Accordingly, the objective of the present study was to evaluate the role of resveratrol on cardiac performance during experimental hepatocarcinogenesis initiated with diethylnitrosamine and promoted by phenobarbital. Rats had free access to diet supplemented with resveratrol four weeks before the carcinogen injection and 14 weeks thereafter. The cardiotoxicity of resveratrol was assessed by monitoring the cardiac function using transthoracic echocardiography as well as Western blot analysis of cardiac tissue. Long-term dietary administration of resveratrol dose-dependently suppressed hepatic tumor multiplicity, the principal endpoint for evaluating the chemopreventive potential of a candidate agent. The chemopreventive effects of resveratrol were also reflected in histopathological assessment of hepatic tissues. Resveratrol did not exhibit any cardiotoxicity but rather improved the cardiac function in a dose-responsive fashion. Our results indicate that resveratrol-mediated chemoprevention of rat liver carcinogenesis is devoid of any adverse cardiovascular events. Resveratrol may be developed as a chemopreventive as well as therapeutic drug for human HCC.
SUMMARYPenetration of the KOS strain of herpes simplex virus type 1 (HSV-1) and the MS and 333 strains of herpes simplex virus type 2 (HSV-2) into HEp-2 cells at pH 6.3 was at least 100-fold less efficient than at pH 7-4. Penetration of two low passage clinical isolates was completely blocked at pH 6.3. The syncytium-forming HSV-1 strains GC and MP were less sensitive than KOS to the mild acidic conditions. The inhibition was completely reversed upon neutralization of the medium. Penetration was assayed by plaque production following protection from acid inactivation upon virus entry. Penetration of HSV-1 KOS into Vero and HEL diploid fibroblast cells was similarly inhibited. HSV-1 KOS grown in 2-deoxy-D-glucose and monensin was also extensively inhibited at pH 6.3 but virus grown in 2-deoxy-D-glucose penetrated more slowly than normal virus at pH 7.4. Electron microscopy of HSV-1 KOS infection indicated that fusion and endocytosis occur at both pH 7.4 and 6.3 but that fusion predominates at pH 7-4 and endocytosis predominates at pH 6.3. These results indicate that fusion at the plasma membrane is the major route of productive entry for HSV, that Strains of HSV can differ in their pH dependence for penetration and this may determine whether virus infection can occur following endocytic uptake.
A rapid means of screening drugs for toxicity and anti-herpes simplex virus activity was developed based on the flow cytometric detection of HSV induced changes in cellular DNA content. Subconfluent monolayers of human diploid fibroblasts (HEL 299) were assayed for DNA content with propidium iodide 24 h after infection with HSV-1 (multiplicity of infection 1-10) and treatment with the drug to be tested. Infection was detected by a broadening of the normal diploid and tetraploid peaks and presence of greater than 4-n DNA staining. Inhibition of viral DNA synthesis and maintenance of the normal growth pattern of control cells was indication of antiviral activity. Toxicity of the compound was indicated by the loss of S phase and tetraploid cell populations. Using this assay, we evaluated the activities of one experimental and two established antiviral agents.
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