Discovery of novel dihydro-9,10-ethano-anthracene carboxamides as glucocorticoid receptor modulators

A series of dihydro-9,10-ethano-anthracene-11-carboxamides as novel glucocorticoid receptor modulators is reported. SAR exploration identified compounds from this series displaying a promising dissociation profile in discriminating between transrepression and transactivation activities. 17a is a partial agonist of GR-mediated transactivation which elicits potent and efficacious transrepression in reporter gene assays. A hypothetical binding mode is provided which accounts for the induction of functional activity by a bridgehead methyl group.     

Prevalence of Antibodies against Avian Influenza A (H5N1) Virus among Cullers and Poultry Workers in Ho Chi Minh City, 2005

Background

Between 2003 and 2005, highly pathogenic avian influenza A (H5N1) viruses caused large scale outbreaks in poultry in the Ho Chi Minh City area in Vietnam. We studied the prevalence of antibodies against H5N1 in poultry workers and cullers who were active in the program in Ho Chi Minh City in 2004 and 2005.

Methodology/Principal Findings

Single sera from 500 poultry workers and poultry cullers exposed to infected birds were tested for antibodies to avian influenza H5N1, using microneutralization assays and hemagglutination inhibition assay with horse blood. All sera tested negative using microneutralization tests. Three samples showed a 1∶80 titer in the hemagglutination inhibition assay.

Conclusions/Significance

This study provides additional support for the low transmissibility of clade 1 H5N1 to humans, but limited transmission to highly exposed persons cannot be excluded given the presence of low antibody titers in some individuals.     

Dissociated Fear and Spatial Learning in Mice with Deficiency of Ataxin-2

Mouse models with physiological and behavioral differences attributable to differential plasticity of hippocampal and amygdalar neuronal networks are rare. We previously generated ataxin-2 (Atxn2) knockout mice and demonstrated that these animals lacked obvious anatomical abnormalities of the CNS, but showed marked obesity and reduced fertility. We now report on behavioral changes as a consequence of Atxn2-deficiency. Atxn2-deficiency was associated with impaired long-term potentiation (LTP) in the amygdala, but normal LTP in the hippocampus. Intact hippocampal plasticity was associated behaviorally with normal Morris Water maze testing. Impaired amygdala plasticity was associated with reduced cued and contextual fear conditioning. Conditioned taste aversion, however, was normal. In addition, knockout mice showed decreased innate fear in several tests and motor hyperactivity in open cage testing. Our results suggest that Atxn2-deficiency results in a specific set of behavioral and cellular disturbances that include motor hyperactivity and abnormal fear-related behaviors, but intact hippocampal function. This animal model may be useful for the study of anxiety disorders and should encourage studies of anxiety in patients with spinocerebellar ataxia type 2 (SCA2).     

In vitro culture and differentiation of osteoblasts from human umbilical cord blood

It is well accepted that human umbilical cord blood (UCB) is a source of mesenchymal stem cells (MSCs) which are able to differentiate into different cell phenotypes such as osteoblasts, chondrocytes, adipocytes, myocytes, cardiomyocytes and neurons. The aim of this study was to isolate MSCs from human UCB to determine their osteogenic potential by using different kinds of osteogenic medium. Eventually, only those MSCs cultured in osteogenic media enriched with vitamin D₂ and FGF9, were positive for osteocalcin by RT-PCR. All these cells were positive for alizarin red, alkaline phosphatase and Von Kossa. The results obtained from RT-PCR have confirmed that osteogenesis is complete by expression of the osteocalcin marker. In conclusion, vitamin D₂, at least in vitro, may replace vitamin D₃ as an osteogenic stimulator factor for MSC differentiation.     

Toxoplasma gondii protease TgSUB1 is required for cell surface processing of micronemal adhesive complexes and efficient adhesion of tachyzoites

Host cell invasion by Toxoplasma gondii is critically dependent upon adhesive proteins secreted from the micronemes. Proteolytic trimming of microneme contents occurs rapidly after their secretion onto the parasite surface and is proposed to regulate adhesive complex activation to enhance binding to host cell receptors. However, the proteases responsible and their exact function are still unknown. In this report, we show that T. gondii tachyzoites lacking the microneme subtilisin protease TgSUB1 have a profound defect in surface processing of secreted microneme proteins. Notably parasites lack protease activity responsible for proteolytic trimming of MIC2, MIC4 and M2AP after release onto the parasite surface. Although complementation with full‐length TgSUB1 restores processing, complementation of Δsub1 parasites with TgSUB1 lacking the GPI anchor (Δsub1::ΔGPISUB1) only partially restores microneme protein processing. Loss of TgSUB1 decreases cell attachment and in vitro gliding efficiency leading to lower initial rates of invasion. Δsub1 and Δsub1::ΔGPISUB1 parasites are also less virulent in mice. Thus TgSUB1 is involved in micronemal protein processing and regulation of adhesive properties of macromolecular adhesive complexes involved in host cell invasion.     

Effect of hypercapnia on intracellular pH regulation in a rainbow trout hepatoma cell line,RTH 149

Fish exposed to elevated water CO₂ experience a rapid increase in blood CO₂ levels (hypercapnia), resulting in acidification of both intra- and extra-cellular compartments. While the mechanisms associated with extracellular pH regulation have been well explored, much less is known about intracellular pH (pH_i) regulation. There is great interest in developing non-animal models for research. One such model is the rainbow trout hepatoma cell line (RTH 149), which has been used to study a wide range of topics; however, no studies have investigated its potential use in pH_i regulation. Employing the pH-sensitive fluoroprobe BCECF, the present study examined pH_i regulation in RTH 149 under normocapnia and during extracellular acidification induced by either elevated CO₂ or 1 M HCl. During exposure to hypercapnia, RTH 149 cells were acidified without recovery as long as the elevated CO₂ was maintained. In addition, rates of pH_i recovery from NH₄Cl-induced acidosis were significantly lower in cells exposed to hypercapnia or HCl compared to that in normocapnic cells, indicating that elevated CO₂ indirectly impeded pH_i recovery through a reduction in pH_e and/or pH_i. Moreover, pH_i regulation in RTH 149 was EIPA-sensitive, suggesting that an NHE may be involved. Overall, RTH 149 may have the potential for identifying transporters likely to play a role in pH_i regulation in fish. However, it should not be used as a complete replacement for in vivo studies, especially to quantify acid–base regulatory ability at whole animal level, since RTH 149 appeared to have enhanced pH_i recovery rates relative to primary hepatocytes.     

The dynamics of interleukin‐8 and its interaction with human CXC receptor I peptide

Interleukin‐8 (CXCL8, IL‐8) is a proinflammatory chemokine important for the regulation of inflammatory and immune responses via its interaction with G‐protein coupled receptors, including CXC receptor 1 (CXCR1). CXCL8 exists as both a monomer and as a dimer at physiological concentrations, yet the molecular basis of CXCL8 interaction with its receptor as well as the importance of CXCL8 dimer formation remain poorly characterized. Although several biological studies have indicated that both the CXCL8 monomer and dimer are active, biophysical studies have reported conflicting results regarding the binding of CXCL8 to CXCR1. To clarify this problem, we expressed and purified a peptide (hCXCR1pep) corresponding to the N‐terminal region of human CXCR1 (hCXCR1) and utilized nuclear magnetic resonance (NMR) spectroscopy to interrogate the binding of wild‐type CXCL8 and a previously reported mutant (CXCL8M) that stabilizes the monomeric form. Our data reveal that the CXCL8 monomer engages hCXCR1pep with a slightly higher affinity than the CXCL8 dimer, but that the CXCL8 dimer does not dissociate upon binding hCXCR1pep. These investigations also showed that CXCL8 is dynamic on multiple timescales, which may help explain the versatility in this interleukin for engaging its target receptors.