Morphological and metabolic characterization of a new species of strictly anaerobic rumen fungus: Neocallimastix joyonii

A new strain of strictly anaerobic fungi was isolated from the rumen of sheep. This strain is characterized by a polycentric thallus, an extensive and polynuclear rhizomycelium, polyflagellated zoospores with gamma particle-like bodies. We propose to assign this strain in a new species: Neocallimastix joyonii.     

Role of phosphatidylinositol in cardiac sarcolemmal membrane sodium-calcium exchange

The purpose of this investigation was to study the effects of a distinct type of phospholipase C on sarcolemmal Na+-Ca2+ exchange. With this phospholipase C (Staphylococcus aureus), treatment of cardiac sarcolemmal vesicles resulted in a specific hydrolysis of membrane phosphatidylinositol. This hydrolysis of phosphatidylinositol also released two proteins (110 and 36 kDa) from the sarcolemmal membrane. Phospholipase C pretreatment of the sarcolemma resulted in an unexpected stimulation of Na+-Ca2+ exchange. The Vmax of Na+-Ca2+ exchange was increased but the Km for Ca2+ was not altered. This stimulation was specific to the Na+-Ca2+ exchange pathway. ATP-dependent Ca2+ uptake was depressed after phospholipase C treatment, but passive membrane permeability to Ca2+ was unaffected. Sarcolemmal Na+,K+-ATPase activity was not altered, whereas passive Ca2+ binding was modestly decreased after phospholipase C pretreatment. The stimulation of Na+-Ca2+ exchange after phosphatidylinositol hydrolysis was greater in inside-out vesicles than in a total population of vesicles of mixed orientation. This finding suggests that the cardiac sarcolemmal Na+-Ca2+ exchanger is functionally asymmetrical. The results also suggest that membrane phosphatidylinositol is inhibitory to the Na+-Ca2+ exchanger or, alternatively, this phospholipid may anchor an endogenous inhibitory protein in the sarcolemmal membrane. The observation that a transsarcolemmal Ca2+ flux pathway may be stimulated solely by phosphatidylinositol hydrolysis independently of phosphoinositide metabolic products like inositol triphosphate is novel.     

Platelet-derived growth factor and transforming growth factor-beta enhance tissue repair activities by unique mechanisms

Platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGF-beta) markedly potentiate tissue repair in vivo. In the present experiments, both in vitro and in vivo responses to PDGF and TGF-beta were tested to identify mechanisms whereby these growth factors might each enhance the wound-healing response. Recombinant human PDGF B-chain homodimers (PDGF-BB) and TGF-beta 1 had identical dose-response curves in chemotactic assays with monocytes and fibroblasts as the natural proteins from platelets. Single applications of PDGF-BB (2 micrograms, 80 pmol) and TGF-beta 1 (20 micrograms, 600 pmol) were next applied to linear incisions in rats and each enhanced the strength required to disrupt the wounds at 5 d up to 212% of paired control wounds. Histological analysis of treated wounds demonstrated an in vivo chemotactic response of macrophages and fibroblasts to both PDGF-BB and to TGF-beta 1 but the response to TGF-beta 1 was significantly less than that observed with PDGF-BB. Marked increases of procollagen type I were observed by immunohistochemical staining in fibroblasts in treated wounds during the first week. The augmented breaking strength of TGF-beta 1 was not observed 2 and 3 wk after wounding. However, the positive influence of PDGF-BB on wound breaking strength persisted through the 7 wk of testing. Furthermore, PDGF-BB-treated wounds had persistently increased numbers of fibroblasts and granulation tissue through day 21, whereas the enhanced cellular influx in TGF-beta 1-treated wounds was not detectable beyond day 7. Wound macrophages and fibroblasts from PDGF-BB-treated wounds contained sharply increased levels of immunohistochemically detectable intracellular TGF-beta. Furthermore, PDGF-BB in vitro induced a marked, time-dependent stimulation of TGF-beta mRNA levels in cultured normal rat kidney fibroblasts. The results suggest that TGF-beta transiently attracts fibroblasts into the wound and may stimulate collagen synthesis directly. In contrast, PDGF is a more potent chemoattractant for wound macrophages and fibroblasts and may stimulate these cells to express endogenous growth factors, including TGF-beta, which, in turn, directly stimulate new collagen synthesis and sustained enhancement of wound healing over a more prolonged period of time.     

Allowing for random errors in radiation dose estimates for the atomic bomb survivor data

The presence of random errors in the individual radiation dose estimates for the A-bomb survivors causes underestimation of radiation effects in dose-response analyses, and also distorts the shape of dose-response curves. Statistical methods are presented which will adjust for these biases, provided that a valid statistical model for the dose estimation errors is used. Emphasis is on clarifying some rather subtle statistical issues. For most of this development the distinction between radiation dose and exposure is not critical. The proposed methods involve downward adjustment of dose estimates, but this does not imply that the dosimetry system is faulty. Rather, this is a part of the dose-response analysis required to remove biases in the risk estimates. The primary focus of this report is on linear dose-response models, but methods for linear-quadratic models are also considered briefly. Some plausible models for the dose estimation errors are considered, which have typical errors in a range of 30-40% of the true values, and sensitivity analysis of the resulting bias corrections is provided. It is found that for these error models the resulting estimates of excess cancer risk based on linear models are about 6-17% greater than estimates that make no allowance for dose estimation errors. This increase in risk estimates is reduced to about 4-11% if, as has often been done recently, survivors with dose estimates above 4 Gy are eliminated from the analysis.     

Intracytoplasmic neuronal inclusions in woodchuck brain stem

Homogenous eosinophilic intracytoplasmic inclusion bodies were found within the large reticular neurons of the brain stems of 57 captive woodchucks (Marmota monax). Light microscopy was consistent with a proteinaceous nature, while electron microscopy suggested a non-viral origin. The woodchucks with inclusions were older than the general population that was studied. It is hypothesized that the neuronal inclusions in the brain stem are indicative of nonspecific ageing changes.     

Evidence for Distinct 5-Hydroxytryptamine2 Binding Site Subtypes in Cortical Membrane Preparations

5-Hydroxytryptamine (5-HT) displays a sixfold higher affinity for 5-HT2 binding sites labeled by [3H]ketanserin in rat (IC50 = 200 +/- 40 nM) and human (IC50 = 190 +/- 50 nM) cortex than for 5-HT2 sites in bovine cortex (IC50 = 1,200 +/- 130 nM). The Hill slopes of the 5-HT competition curves are 0.67 +/- 0.04 in rat, 0.69 +/- 0.08 in human, and 0.96 +/- 0.02 in bovine cortex. Scatchard analysis of (+/-)-[3H]4-bromo-2,5-dimethoxyamphetamine ([3H]DOB) binding in the rat indicates a population of binding sites with a KD of 0.38 +/- 0.04 nM and a Bmax of 1.5 +/- 0.05 pmol/g tissue. In contrast, specific [3H]DOB binding cannot be detected in bovine cortical membranes. These data indicate that species variations exist in 5-HT2 binding site subtypes and that [3H]ketanserin appears to label a homogeneous population of 5-HT2 binding site subtypes in bovine cortex.     

Mechanisms by which EGF receptor and TGF alpha contribute to malignant transformation

Alterations affecting the epidermal growth factor/transforming growth factor alpha-responsive mitogenic pathway are frequently detected in malignancies. In particular, the epidermal growth factor receptor has been found overexpressed in a number of human tumors. Production and secretion of transforming growth factor type alpha has also been shown in several tumor cells but not in their normal counterparts. In this review we describe the establishment of in vitro model systems to study the transforming potential of these molecules and summarize our current understanding of the mechanisms involved in transformation by genes encoding a growth factor and a growth factor receptor.     

Genomic organization of a mouse MHC class II region including theH2-M andLmp2 loci

The region encompassing theMa, Mb1, Mb2, andLmp2 genes of the mouse class II major histocompatibility complex (MHC) was sequenced. Since this region contains clusters of genes required for efficient class I and class II antigen presentation, it was interesting to search for putative additional genes in the 21 kilobase gap between theMb1 andLmp2 genes. Computer predictions of coding regions and CpG islands, exon trapping experiments, and cross-species comparison with the corresponding human sequence indicate that no additional functional gene is present in that stretch. However, computer analysis revealed the possible existence of an alternative 3 exon forMb1. Except for the fact that the mouse MHC contains twoMb genes, the genomic organization of theH2-M loci was found to be almost identical to the organization of the humanHLA-DM genes. The promoter regions of theMa andMb genes also resemble classical class II promoters, containing typical S, X, and Y boxes. Like the human genes, the threeH2-M genes displayed very limited polymorphism when we compared the cDNA sequences from six haplotypes. Finally, comparison ofDMB withMb1 andMb2, both at the genomic level and in their coding regions, suggests that theMb gene was recently duplicated, probably only in certain rodents.     

Molecular cloning of the lectin and a lectin-related protein from common Solomon's seal (Polygonatum multiflorum)

The most prominent protein ofPolygonatum multiflorum (common Solomon's seal) rhizomes has been identified as a mannose-binding lectin. Analysis of the purified lectin demonstrated that it is a tetramer of four identical subunits of 14 kDa. Molecular cloning further revealed that the lectin from this typical Liliaceae species belongs to the superfamily of monocot mannose-binding proteins. Screening of cDNA libraries constructed with RNA isolated from buds, leaves and flowers ofP. multiflorum also yielded cDNA clones encoding a protein, which contains two tandemly arranged domains with an obvious sequence homology to the mannose-binding lectins. Molecular modelling of thePolygonatum lectin and lectin-related protein indicated that the three-dimensional structure of both proteins strongly resembles that of the snowdrop lectin. In addition, this approach suggested that the presumed carbohydrate-binding sites of the lectin can accommodate a mannose residue whereas most of the carbohydratebinding sites of the lectin-related protein cannot.Abbreviations GNA Galanthus nivalis agglutinin - HCA hydrophobic cluster analysis - LECPMA cDNA clone encoding PMA - PM30 30 kDa protein isolated fromPolygonatum multiforum - PMA Polygonatum multiflorum agglutinin - PMLRP Polygonatum multiflorum lectin-related protein