Effects and mechanisms of action of sildenafil citrate in human chorionic arteries

Objectives  

Sildenafil citrate, a specific phosphodiesterase-5 inhibitor, is increasingly used for pulmonary hypertension in pregnancy. Sildenafil is also emerging as a potential candidate for the treatment of intra-uterine growth retardation and for premature labor. Its effects in the feto-placental circulation are not known. Our objectives were to determine whether phosphodiesterase-5 is present in the human feto-placental circulation, and to characterize the effects and mechanisms of action of sildenafil citrate in this circulation.     

Characterization of Ross River virus tropism and virus-induced inflammation in a mouse model of viral arthritis and myositis

Mosquito-borne alphaviruses are a significant cause of both encephalitic and arthritic disease in humans worldwide. In contrast to the encephalitic alphaviruses, the pathogenesis of alphavirus-induced arthritic disease is not well understood. Utilizing a mouse model of Ross River virus (RRV) disease, we found that the primary targets of RRV infection are bone, joint, and skeletal muscle tissues of the hind limbs in both outbred CD-1 mice and adult C57BL/6J mice. Moreover, histological analyses demonstrated that RRV infection resulted in severe inflammation of these tissues. Characterization of the inflammatory infiltrate within the skeletal muscle tissue identified inflammatory macrophages, NK cells, and CD4+ and CD8+ T lymphocytes. To determine the contribution of the adaptive immune system, the outcome of RRV-induced disease was examined in C57BL/6J RAG-1(-/-) mice, which lack functional T and B lymphocytes. RAG-1(-/-) and wild-type mice developed similar disease signs, infiltration of inflammatory macrophages and NK cells, and muscle pathology, suggesting that the adaptive immune response does not play a critical role in the development of disease. These results establish the mouse model of RRV disease as a useful system for the identification of viral and host factors that contribute to alphavirus-induced arthritis and myositis.     

Experimental infection magnifies inbreeding depression in house mice

It is often assumed that inbreeding reduces resistance to pathogens, yet there are few experimental tests of this idea in vertebrates, and no tests for the effects of moderate levels of inbreeding more commonly found in nature. We mated wild-derived mice with siblings or first cousins and compared the resistance of their offspring to Salmonella infection with outbred controls under laboratory and seminatural conditions. In the laboratory, full-sib inbreeding reduced resistance to Salmonella and survivorship, whereas first-cousin inbreeding had no detectable effects. In competitive population enclosures, we found that first-cousin inbreeding reduced male fitness by 57% in infected vs. only 34% in noninfected control populations. Our study provides experimental evidence that inbreeding reduces resistance and ability to survive pathogenic infection, and moreover, it shows that even moderate inbreeding can cause significant fitness declines under naturalistic conditions of social stress, and especially with exposure to infectious agents.     

Female infertility and disrupted angiogenesis are actions of specific follistatin isoforms

The circulating and tissue-bound forms of follistatin (FST315 and FST288, respectively) modulate the actions of activins. FST knockout (KO/null) mice, lacking both isoforms, die perinatally with defects in lung, skin, and the musculoskeletal system. Using constructs of the human FST gene engineered to enable expression of each isoform under the control of natural regulatory elements, transgenic mouse lines were created and crossed with FST null mice to attempt to rescue the neonatal lethality. FST288 expression alone did not rescue the neonatal lethality, but mice expressing FST315 on the KO background survived to adulthood with normal lung and skin morphology and partial reversal of the musculoskeletal defects noted in FST KO mice. The FST315 rescue mice displayed a short period of neonatal growth retardation, impaired tail growth, and female infertility. The latter may be due to failure of corpus luteum formation, a decline in the ovarian follicular population, and an augmented uterine inflammatory response to mating. Failure of corpus luteum formation and impaired tail growth indicate abnormal vascularization and suggest that FST288 is required for the promotion of angiogenesis. The augmented uterine inflammatory response may result from the failure of FST315 to modulate the proinflammatory actions of activin A in the uterus or may result from the altered steroid milieu associated with the ovarian abnormalities. Although we cannot definitively conclude that the remaining defects are due to the absence of a particular isoform or due to variable expression of each, these models have demonstrated novel physiological processes that are influenced by FST.     

MyD88 Is Required for Protection from Lethal Infection with a Mouse-Adapted SARS-CoV

A novel human coronavirus, SARS-CoV, emerged suddenly in 2003, causing approximately 8000 human cases and more than 700 deaths worldwide. Since most animal models fail to faithfully recapitulate the clinical course of SARS-CoV in humans, the virus and host factors that mediate disease pathogenesis remain unclear. Recently, our laboratory and others developed a recombinant mouse-adapted SARS-CoV (rMA15) that was lethal in BALB/c mice. In contrast, intranasal infection of young 10-week-old C57BL/6 mice with rMA15 results in a nonlethal infection characterized by high titer replication within the lungs, lung inflammation, destruction of lung tissue, and loss of body weight, thus providing a useful model to identify host mediators of protection. Here, we report that mice deficient in MyD88 (MyD88^−/−), an adapter protein that mediates Toll-like receptor (TLR), IL-1R, and IL-18R signaling, are far more susceptible to rMA15 infection. The genetic absence of MyD88 resulted in enhanced pulmonary pathology and greater than 90% mortality by day 6 post-infection. MyD88^−/− mice had significantly higher viral loads in lung tissue throughout the course of infection. Despite increased viral loads, the expression of multiple proinflammatory cytokines and chemokines within lung tissue and recruitment of inflammatory monocytes/macrophages to the lung was severely impaired in MyD88^−/− mice compared to wild-type mice. Furthermore, mice deficient in chemokine receptors that contribute to monocyte recruitment to the lung were more susceptible to rMA15-induced disease and exhibited severe lung pathology similar to that seen in MyD88^−/−mice. These data suggest that MyD88-mediated innate immune signaling and inflammatory cell recruitment to the lung are required for protection from lethal rMA15 infection.     

A combination of proteasome inhibitors and antibiotics prevents lethality in a septic shock model

Our recent studies with lactacystin, a prototype proteasome inhibitor, have suggested that the proteasome is a key regulator of LPS-induced signaling pathways contributing to the inflammatory process. Moreover, lactacystin protects animals from LPS-induced shock. Therefore, we sought to identify other less toxic compounds that would block the chymotrypsin-like activity of the proteasome or LPS-induced nitric oxide (NO). After screening over 100 natural compounds (based on chemistry and inhibition of LPS-induced biological activities), we now report for the first time that quercetin, like lactacystin (the prototype proteasome inhibitor), and mevinolin are also inhibitors of the chymotrypsin-like activity of the cellular proteasome within living cells. In addition, this study also suggests that mevinolin and quercetin both have relatively potent anti-inflammatory effects on LPS-treated macrophages in vitro. Interestingly, both of these compounds behave like lactacystin in that they block LPS-induced NO to a greater extent than TNF-alpha. The results of our experiments clearly suggest that mevinolin, in combination with the antibiotic imipenem, can provide protection against polymicrobial septic lethality induced by cecal-ligation and puncture in mice. Collectively, these studies strongly support the conclusion that therapeutic targeting of cellular proteasomes, in conjunction with standard antimicrobial therapy, may be of considerable survival benefit in the treatment of septic shock.