A synthetic peptide homologous to the envelope proteins of retroviruses inhibits monocyte-mediated killing by inactivating interleukin 1

The synthetic peptide CKS-17 has homology to a highly conserved region of the immunosuppressive retroviral envelope protein P15E, to envelope proteins of HTLV I, II, III, and to that encoded by an endogeneous C-type human retroviral DNA. CKS-17 inhibits the immune response of lymphocytes and the respiratory burst of human monocytes. Because P15E-related antigens are present in human malignant cell lines and cancerous effusions, we sought to determine the effect of CKS-17 on monocyte-mediated tumor cell lysis. Lysis of A375 tumor cells by lymphokine or lipopolysaccharide-activated human monocytes was inhibited by 10 microM CKS-17 (control, 79%; CKS-17-treated, 19%). Another synthesized peptide, CS-2, which has partial homology to CKS-17, failed to block monocyte-mediated killing. Thus, the inhibition by CKS-17 appeared to be specific. Because interleukin 1 (IL-1) is a cytocidal factor produced by activated monocytes, we also investigated the effect of CKS-17 on IL-1 production by monocytes and on direct IL-1-mediated cytotoxicity. CKS-17 did not block production or secretion of IL-1 by lipopolysaccharide- or interferon-gamma-activated monocytes. However, the direct cytocidal activity of both recombinant IL-1 alpha and IL-1 beta against A375 tumor cells was blocked by CKS-17. The cytotoxic activity of IL-1 was inhibited by CKS-17 if (a) IL-1 was preincubated with CKS-17 for 1 hr at 37 degrees C or (b) the A375 cells were incubated with CKS-17 for 1 hr prior to the addition of IL-1. CKS-17 also blocked IL-1-induced proliferation of murine thymocytes, the D10 T cell line, and an IL-1-responsive astrocytoma cell line. These data suggest that CKS-17 may be a potent inhibitor of IL-1.     

High level of divergence of male-reproductive-tract proteins, between Drosophila melanogaster and its sibling species, D. simulans

We compared male-reproductive-tract polypeptides of Drosophila melanogaster and D. simulans by using two-dimensional gel electrophoresis. Approximately 64% of male-reproductive-tract polypeptides were identical between two randomly chosen isofemale lines from these two species, compared with 83% identity for third-instar imaginal wing-disc polypeptides. Qualitatively similar differences were found between reproductive tracts and imaginal discs when D. sechellia was compared with D. melanogaster and with D. simulans. When genic polymorphism was taken into account, approximately 10% of male- reproductive-tract polypeptides were apparently fixed for different alleles between D. melanogaster and D. simulans; this proportion is the same as that found for soluble enzymes by one-dimensional gel electrophoresis. Strikingly, approximately 20% of male-reproductive- tract polypeptides of either D. melanogaster or D. simulans had no detectable homologue in the other species. We propose that proteins of the Drosophila male reproductive tract may have diverged more extensively between species than have other types of proteins and that much of this divergence may involve large changes in levels of polypeptide expression.      

Monocyte-conditioned medium, interleukin-1, and tumor necrosis factor stimulate the acute phase response in human hepatoma cells in vitro

Human hepatoma cells mimic the acute phase response after treatment with monocyte-conditioned medium. Levels of secreted fibrinogen, alpha-1 acid glycoprotein, C-reactive protein, haptoglobin, and the third component of complement were elevated compared with control levels after 48 h of incubation with conditioned supernatant medium from an enriched fraction of normal peripheral monocytes. Albumin levels declined and alpha-1 antitrypsin remained unchanged. Levels of specific mRNA were measured by hybridization to slot blots and Northern blots and changed in correspondence with protein alterations. Interleukin-1 and tumor necrosis factor stimulated the third component of complement, but did not elevate any other member of the acute phase group and were therefore only partially active in this system. The identification of an in vitro model of the human acute phase response will permit analysis of the molecular basis for coordinate regulation of this group of facultative genes.     

Microelectrode studies of the active Na transport pathway of frog skin

When the outer surface of short-circuited frog skin was penetrated with microelectrodes, stable negative potentials that averaged near -100 mV were recorded consistently, confirming the results of Nagel (W. Nagel. 1975. Abstracts of the 5th International Biophysics Congress, Copenhagen. P-147.). The appearance of these stable potentials, V(O), concurrent with the observations that (a) a high resistance outer barrier R(O) accounting for approximately 75 percent or more of the transcellular resistance of control skins had been penetrated and that (b) 10(-5) M amiloride and reduced [Na] outside caused the values of V(O) to increase towards means value near -130 mV while the values of percent R(O) increased to more than 90 percent. It was of relationships were the same as the values of E(1) observed in studies of the current-voltage relationships were the same as the values of E’(1) defined as the values of voltage at the inner barrier when the V(O) of the outer barrier was reduced to zero by voltage clamping of the skins. Accordingly, these data are interpreted to mean that the values of E(1), approximately 130 mV, represent the E(Na) of the sodium pump at the inner barrier. 2,4-DNP was observed to decrease the values of transepithelial voltage less than E(1) the V(O) was negative. These data can be interpreted with a simple electrical equivalent circuit of the active sodium transport pathway of the frog skin that includes the idea that the outer membrane behaves as an electrical rectifier for ion transport.     

Synthesis of model compounds relevant to the active-site-directed inactivation of L-asparaginase by 5-diazo-4-oxo-L-norvaline.

Earlier work has shown that 5-diazo-4-oxo-L-norvaline (DONV) irreversibly inactivates the L-asparaginase from E. coli by formation of a covalent bond in the region of the active site. Model compounds have been prepared to study this acid-labile covalent bond tentatively assigned to a serine or possibly a threonine residue in a decapeptide isolated from 14C-DONV-inactivated enzyme. Appropriately blocked DONV was found to alkylate methanol, and the hydroxyl function of blocked serine or threonine in the presence of boron trifluoride. The labile beta-ketoethers thus formed were reduced to the more stable beta-hydroxyethers. Facile lactonization of these 5-substituted-4-hydroxy-L-norvalines was observed. The diastereoisomers of both the lactonized and open forms of 5-methoxy-4-hydroxy-L-norvaline and related 4-hydroxy-L-2-amino acids of similar length were distinguishable on the amino acid analyzer. The beta-hydroxyethers derived from serine and threonine were hydrolyzed with acid and yielded the expected cleavage products. When the beta-ketoether was reduced by sodium borohydride prior to deblocking, in addition to the beta-hydroxyether, N-blocked amino alcohols were also formed, yielding a complex mixture of products.     

A novel dysmorphic syndrome with open calvarial sutures and sutural cataracts maps to chromosome 14q13-q21

We describe a new dysmorphic syndrome in an inbred Saudi Arabian family with 21 members. Five males and one female have similar craniofacial features including wide open calvarial sutures with large and late-closing anterior fontanels, frontal bossing, hyperpigmentation with capillary hemangioma of the forehead, significant hypertelorism, and a broad and prominent nose. In addition, these individuals have Y-shaped sutural cataracts diagnosed by 1-2 years of age. No chromosomal or biochemical abnormalities were identified. A genome-wide scan was performed, and two-point LOD score analysis, assuming autosomal recessive inheritance, detected linkage to chromosome 14q13-q21. The highest LOD scores were obtained for marker GATA136A04 (LOD=4.58 at theta=0.00) and for the adjacent telomeric marker D14S1048 (LOD=4.32 at theta=0.00). Multipoint linkage analysis resulted in a maximum LOD score of 5.44 between markers D14S1048 and GATA136A04. Model independent analysis by SIBPAL confirmed linkage to the same chromosomal region. Haplotype analysis indicated that all affected individuals were homozygous for the interval on chromosome 14q13-q21 with two recombinants for D14S1014 (centromeric) and one recombinant for D14S301 (telomeric). These recombinations limit the disease locus to a region of approximately 7.26 Mb. Candidate genes localized to this region were identified, and analysis of PAX9 did not identify mutations in these patients. The unique clinical phenotype and the mapping data suggest that this family represents a novel autosomal recessive syndrome.     

Direct intracardiac injection of umbilical cord-derived stromal cells and umbilical cord blood-derived endothelial cells for the treatment of ischemic cardiomyopathy

The development of new therapeutic strategies is necessary to reduce the worldwide social and economic impact of cardiovascular disease, which produces high rates of morbidity and mortality. A therapeutic option that has emerged in the last decade is cell therapy. The aim of this study was to compare the effect of transplanting human umbilical cord-derived stromal cells (UCSCs), human umbilical cord blood-derived endothelial cells (UCBECs) or a combination of these two cell types for the treatment of ischemic cardiomyopathy (IC) in a Wistar rat model. IC was induced by left coronary artery ligation, and baseline echocardiography was performed seven days later. Animals with a left ventricular ejection fraction (LVEF) of ≤40% were selected for the study. On the ninth day after IC was induced, the animals were randomized into the following experimental groups: UCSCs, UCBECs, UCSCs plus UCBECs, or vehicle (control). Thirty days after treatment, an echocardiographic analysis was performed, followed by euthanasia. The animals in all of the cell therapy groups, regardless of the cell type transplanted, had less collagen deposition in their heart tissue and demonstrated a significant improvement in myocardial function after IC. Furthermore, there was a trend of increasing numbers of blood vessels in the infarcted area. The median value of LVEF increased by 7.19% to 11.77%, whereas the control group decreased by 0.24%. These results suggest that UCSCs and UCBECs are promising cells for cellular cardiomyoplasty and can be an effective therapy for improving cardiac function following IC.     

Most nuclear systemic autoantigens are extremely disordered proteins: implications for the etiology of systemic autoimmunity

Patients with systemic autoimmune diseases usually produce high levels of antibodies to self-antigens (autoantigens). The repertoire of common autoantigens is remarkably limited, yet no readily understandable shared thread links these apparently diverse proteins. Using computer prediction algorithms, we have found that most nuclear systemic autoantigens are predicted to contain long regions of extreme structural disorder. Such disordered regions would generally make poor B cell epitopes and are predicted to be under-represented as potential T cell epitopes. Consideration of the potential role of protein disorder may give novel insights into the possible role of molecular mimicry in the pathogenesis of autoimmunity. The recognition of extreme autoantigen protein disorder has led us to an explicit model of epitope spreading that explains many of the paradoxical aspects of autoimmunity – in particular, the difficulty in identifying autoantigen-specific helper T cells that might collaborate with the B cells activated in systemic autoimmunity. The model also explains the experimentally observed breakdown of major histocompatibility complex (MHC) class specificity in peptides associated with the MHC II proteins of activated autoimmune B cells, and sheds light on the selection of particular T cell epitopes in autoimmunity. Finally, the model helps to rationalize the relative rarity of clinically significant autoimmunity despite the prevalence of low specificity/low avidity autoantibodies in normal individuals.     

Bent bone dysplasia-FGFR2 type, a distinct skeletal disorder, has deficient canonical FGF signaling

Fibroblast growth factor receptor 2 (FGFR2) is a crucial regulator of bone formation during embryonic development. Both gain and loss-of-function studies in mice have shown that FGFR2 maintains a critical balance between the proliferation and differentiation of osteoprogenitor cells. We have identified de novo FGFR2 mutations in a sporadically occurring perinatal lethal skeletal dysplasia characterized by poor mineralization of the calvarium, craniosynostosis, dysmorphic facial features, prenatal teeth, hypoplastic pubis and clavicles, osteopenia, and bent long bones. Histological analysis of the long bones revealed that the growth plate contained smaller hypertrophic chondrocytes and a thickened hypercellular periosteum. Four unrelated affected individuals were found to be heterozygous for missense mutations that introduce a polar amino acid into the hydrophobic transmembrane domain of FGFR2. Using diseased chondrocytes and a cell-based assay, we determined that these mutations selectively reduced plasma-membrane levels of FGFR2 and markedly diminished the receptor's responsiveness to extracellular FGF. All together, these clinical and molecular findings are separate from previously characterized FGFR2 disorders and represent a distinct skeletal dysplasia.     

EFFECTS OF HUMAN ACTIVITIES ON STRUCTURE AND COMPOSITION OF WOODY SPECIES OF THE NOKREK BIOSPHERE RESERVE OF MEGHALAYA,NORTHEAST INDIA

Aims Our study was conducted in the Nokrek Biosphere Reserve (NBR) in the Garo hills districts of Meghalaya, Northeast India. Our aim was to assess the effects of human activities on plant diversity,population structure and regeneration.Methods We selected a representative 1.2 hm2 stand in both the core and buffer zones of NBR. Structure and composition were determined by randomly sampling square quadrats, population structure was assessed by determining age structure, and regeneration was assessed by measuring densities of seedling, sapling and adult trees.Important findings More woody species were recorded from the core zone than the buffer zone (87 vs. 81 species), and there were a large number of tropical, temperate, and Sino-Himalayan, Burma-Malaysian and Malayan elements, primitive families and primitive genera. The trees were distributed in three distinct strata,canopy, subcanopy and sapling. Subcanopy and sapling layers had the highest species richness (81％ -88％ ). Lauraceae and Euphorbiaceae were the dominant families in terms of the number of species, and a large number of families were represented by single species. Most woody species (57 ％ - 79 ％ ) were contagiously distributed and had low frequency ( ＜ 20％ ). Although stand density was high in the buffer zone, its basal area was low compared to the stand in the core zone. Low similarity and high β-diversity indicate marked differences in species composition of the stands. Shannon diversity index was high in both the stands, while Simpson dominance index was low. The diameter-class distribution for dominant species revealed that the most had a large number of young individuals in their populations. Preponderance of tree seedlings, followed by a steep decline in population density of saplings and adult trees, indicated that the seedling to sapling stage was the most critical in the life cycle of the tree populations. Most species (42 ％ - 48 ％ ) had no regeneration,25 ％ - 35 ％ had good/fair regeneration, and the rest had poor regeneration or reoccurred as immigrants.