Effect of plasma TNF-alpha on filgrastim-stimulated hematopoiesis in mice and humans

Marshall syndrome is a rare, autosomal dominant skeletal dysplasia that is phenotypically similar to the more common disorder Stickler syndrome. For a large kindred with Marshall syndrome, we demonstrate a splice-donor-site mutation in the COL11A1 gene that cosegregates with the phenotype. The G+1-->A transition causes in-frame skipping of a 54-bp exon and deletes amino acids 726-743 from the major triple-helical domain of the alpha1(XI) collagen polypeptide. The data support the hypothesis that the alpha1(XI) collagen polypeptide has an important role in skeletal morphogenesis that extends beyond its contribution to structural integrity of the cartilage extracellular matrix. Our results also demonstrate allelism of Marshall syndrome with the subset of Stickler syndrome families associated with COL11A1 mutations.     

Cytokine priming reduces dependence on TNF-R2 for TNF-alpha-mediated induction of macrophage nitric oxide generation

In the presence of interferon-gamma (IFN-gamma), human tumor necrosis factor-alpha (Hu-TNF-alpha), which binds to murine TNF-alpha receptor type 1 (TNF-R1) but not to murine TNF-R2, was effective in inducing nitric oxide (NO) production in spleen-derived macrophages (M phi), albeit at concentrations 12.5-fold greater than those required by murine TNF-alpha (Mu-TNF-alpha), to achieve the same result. Addition of anti-TNF-R1 completely inhibited the Mu-TNF-alpha-mediated induction of NO, demonstrating that TNF-R1 is critical to the IFN-gamma-dependent TNF-alpha-mediated induction of M phi effector function. However, treatment with anti-TNF-R2 resulted in a partial inhibition of M phi activation. Spleen-derived M phi were more dependent on TNF-R2 than RAW 264.7 or peritoneal M phi based on their responsiveness to Hu-TNF-alpha. Priming of spleen-derived M phi with either IFN-gamma or granulocyte-macrophage colony-stimulating factor (GM-CSF) heightened the maximal responses to both TNF-alpha species and increased the overall effectiveness of Hu-TNF-alpha without increasing expression of either TNF-alpha receptor. The dependence of spleen-derived M phi on both TNF-alpha receptors for signaling the induction of effector function supports an active signaling role for TNF-R2 in its synergy with TNF-R1 rather than a passive ligand passing role.     

Sphingosine-1-phosphate as a ligand for the G protein-coupled receptor EDG-1

The sphingolipid metabolite sphingosine-1-phosphate (SPP) has been implicated as a second messenger in cell proliferation and survival. However, many of its biological effects are due to binding to unidentified receptors on the cell surface. SPP activated the heterotrimeric guanine nucleotide binding protein (G protein)-coupled orphan receptor EDG-1, originally cloned as Endothelial Differentiation Gene-1. EDG-1 bound SPP with high affinity (dissociation constant = 8.1 nM) and high specificity. Overexpression of EDG-1 induced exaggerated cell-cell aggregation, enhanced expression of cadherins, and formation of well-developed adherens junctions in a manner dependent on SPP and the small guanine nucleotide binding protein Rho.     

Prostaglandins suppress an outward potassium current in embryonic rat sensory neurons

One of the challenges in understanding ciliary and flagellar motility is determining the mechanisms that locally regulate dynein-driven microtubule sliding. Our recent studies demonstrated that microtubule sliding, in Chlamydomonas flagella, is regulated by phosphorylation. However, the regulatory proteins remain unknown. Here we identify the 138-kD intermediate chain of inner arm dynein I1 as the critical phosphoprotein required for regulation of motility. This conclusion is founded on the results of three different experimental approaches. First, genetic analysis and functional assays revealed that regulation of microtubule sliding, by phosphorylation, requires inner arm dynein I1. Second, in vitro phosphorylation indicated the 138-kD intermediate chain of I1 is the only phosphorylated subunit. Third, in vitro reconstitution demonstrated that phosphorylation and dephosphorylation of the 138-kD intermediate chain inhibits and restores wild-type microtubule sliding, respectively. We conclude that change in phosphorylation of the 138-kD intermediate chain of I1 regulates dynein-driven microtubule sliding. Moreover, based on these and other data, we predict that regulation of I1 activity is involved in modulation of flagellar waveform.     

Segregation analysis of breast cancer in a population-based sample of postmenopausal probands: The Iowa Women's Health Study

Inheritance of a major susceptibility gene for breast cancer has been primarily investigated in families with early-onset disease. However, familial clustering of late-onset breast cancer is well documented, and genetic factors may also be relevant. In the Iowa Women's Health Study, we evaluated evidence for a major gene after allowing for measured environmental risk factors. Two hundred sixty-five incident breast cancer probands were identified from a prospective cohort study of 41,837 women aged 55 to 69 years at baseline in 1986. A pedigree development form was mailed to the probands to ascertain all first-degree female relatives. A questionnaire and body measurement protocol were mailed to identified living relatives or surrogates. Segregation analyses were conducted on a total of 1,145 women in 251 families using regressive models as implemented in S.A.G.E. Mendelian codominant inheritance of an allele that produced an earlier-age-at-onset provided the best fit to the data. Incorporation of measured environmental risk factors as covariates yielded no significant improvements in the likelihoods. Approximately 50% of this population could be expected to carry a late-onset breast cancer susceptibility gene, and 23% of the population is susceptible because of the environment in which they live. Homozygous gene carriers are predicted to have a mean age-at-onset of 48 years, over 20 years earlier than heterozygotes; few cases would be expected among non-gene carriers. In conclusion, the transmission pattern of late-onset breast cancer may be determined by a common susceptibility gene.