Effect of low ambient temperature on protein turnover and heat production in chicks

1. Effect of low ambient temperature on protein turnover in the liver and whole body was investigated in chicks together with the contribution of protein synthesis to the total heat production. 2. Both protein synthesis and degradation in the whole body were increased, the latter to a larger extent, at low ambient temperature (LT, 22 degrees C) compared with adequate temperature (AT, 30 degrees C). Liver protein synthesis was not significantly altered by the temperature treatment. 3. The total heat production of LT group was as high as 160% of the AT group. 4. The increased heat production due to enhanced whole-body protein synthesis accounted for only 1.4% of the heat increment in thermogenesis at low ambient temperature, suggesting that protein synthesis would contribute little, if any, to cold-induced thermogenesis in chicks.     

Radioimmunoassay of chenodeoxycholic acid-3-sulfate in urine

A sensitive and specific radioimmunoassay (RIA) for the measurement of urinary total chenodeoxycholic acid-3-sulfate (SCDCA) was developed and the accuracy was confirmed. SCDCA bound to bovine serum albumin as the antigen and emulsified with Freund's complete adjuvant was injected into rabbits. The antiserum obtained was capable of binding 75% of [11,12-3H]SCDCA at 1:1000 dilution. The percentage of bound radioactivity decreased linearly with logarithmic increases in unlabeled SCDCA, from 8 to 200 pmol/ml. The antiserum showed an extremely high specificity for SCDCA (free and conjugated), and the values determined by RIA indicated a close correlation with those found by gas-liquid chromatography. The daily urinary SCDCA level was determined using SCDCA-RIA in 12 disease-free humans and 74 patients with chronic liver diseases. In the normal subjects the daily urinary SCDCA level was 0.74 +/- 0.83 mg/day and increased levels were evident in all groups with chronic liver diseases. The daily urinary SCDCA level corresponds closely with the extent of hepatic dysfunction.     

Matrix Gla protein C-terminal region binds to vitronectin. Co-localization suggests binding occurs during tissue development.

Matrix Gla protein (MGP) regulates calcification in cartilage and arteries. MGP synthesis during embryonic development and its binding and regulation of growth factors and morphogens of the TGF-beta/BMP superfamily suggests that it has additional functions. Assay by far-western gel overlays and gel filtration shift shows MGP binds vitronectin. Binding is saturable and consistent with a single class of binding sites. MGP binds to vitronectin but not collagen, fibromodulin, heparin, osteocalcin, chondroitin sulfate, laminin, ovalbumin or albumin. We have identified a vitronectin binding site within a 17-amino acid peptide 61-77 near the carboxyl-terminus that corresponds to a naturally occurring MGP C-terminus. MGP and the 61-77 MGP peptide also binds to fibronectin. MGP and vitronectin are focally co-localized in embryonic tissues. Co-localization in vivo suggests that the MGP and vitronectin interactions may modify cell-matrix interactions. Alternatively, vitronectin-bound MGP may have altered function for modulating BMP2 or TGF-beta activity. The current study demonstrates that MGP has a novel binding activity for vitronectin, an extracellular protein that promotes cell-matrix interactions and regulates coagulation.     

Rate of DNA Synthesis during Meiotic Prophase in Lily Microsporocytes in the Presence of Deoxyadenosine and Nalidixic Acid

The rate and period of DNA synthesis during meiotic prophasewere examined using lily microsporocytes. Meiocytes at the earlyleptotene stage were cultured for discrete periods in the presenceof inhibitors of DNA synthesis, deoxyadenosine and nalidixicacid. Deoxyadenosine, which arrests meiotic development at theearly zygotene stage, markedly suppressed DNA synthesis to 35%of control at 2 mM. Nalidixic acid simply reduced the rate ofDNA synthesis, resulting in prolongation of the synthetic period.The relevance of DNA synthesis to meiotic development is discussed. (Received January 12, 1987; Accepted May 7, 1987)     

Influence of the gut microflora on protein synthesis in tissues and in the whole body of chicks.

1. The influence of the gut microflora on protein synthesis in individual tissues and in the whole body of young chicks was investigated by the large-dose injection of [3H]phenylalanine. 2. Growth of germ-free chicks was significantly better than that of conventional controls. Wet weights of liver, spleen, duodenum, jejunum + ileum and caeca were heavier in conventional birds than in germ-free counterparts. 3. Fractional rates of protein synthesis were higher in jejunum + ileum and whole body of conventional birds than in those of germ-free birds. Amounts of protein synthesized were larger in liver, jejunum + ileum and caeca in the presence of the gut microflora. 4. When tissues were classified into gut + liver and the remainder of the carcass, in the presence of the gut microflora an enhanced protein synthesis in fractional and absolute rate was found in the gut + liver, which is in direct contact or in close association with micro-organisms, whereas virtually no effect of the gut micro-organisms was detected in the remainder of the carcass. 5. The contribution of protein synthesis of gut + liver to that of the whole body was larger in conventional chicks than in germ-free birds, whereas the reverse was true for the remainder of the carcass.     

Molecular relationship between two types of phospholipase B from Penicillium notatum and reconstitution of active enzyme from its peptide fragments

Two types of phospholipase B of Penicillium notatum, the native type and the modified type that is thought to be generated by the introduction of some nicks into the native type of enzyme by the endogenous protease(s), were distinguished on a slab sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis under a nonreducing condition. The native form migrated with a rate corresponding to 95K Da, whereas the modified form migrated more slowly, corresponding to 106K Da, presumably because of its more extended conformation. That the "106K" protein was indeed a nicked product of the "95K" protein was confirmed by amino acid analysis, peptide mapping, N- and C-terminal sequence analyses, and immunoblotting. The peptide fragments (70K and 37K + 32K) comprising the modified protein were isolated by gel filtration in the presence of SDS and 2-mercaptoethanol (the 32K peptide was suggested to be a partial proteolytic product of the 37K peptide). When the "95K" protein was subjected to the same treatment under denaturing condition, it retained a low, but significant, enzymatic activity; in contrast, the separated peptide fragments did not show any significant activity. By a coincubation of these fragments, however, a restoration of enzymatic activity was observed through a reformation of the active complex, corresponding to the original modified protein. The enzymatic activity of this complex was further increased by a treatment with guanidine X HCl, followed by dialysis. The association of peptide fragments appears to occur through the formation of interpeptidal disulfide bonds.     

Cloning of the Membrane-Bound Aldehyde Dehydrogenase Gene of Acetobacter polyoxogenes and Improvement of Acetic Acid Production by Use of the Cloned Gene

A genomic clone bank of Acetobacter polyoxogenes NBI1028 constructed in Escherichia coli by use of the expression vector pUC18 was screened with antibody raised against membrane-bound aldehyde dehydrogenase (ALDH; 75 kilodaltons [kDa]) from A. polyoxogenes NBI1028. A clone that synthesized a 41-kDa protein cross-reactive with anti-ALDH antibody was isolated. For cloning of the full-length ALDH structural gene, a cosmid gene bank was screened by Southern blot hybridization with the cloned DNA as a probe, and subcloning from the positive cosmid clone was performed with shuttle vector pMV24. Plasmid pAL25, containing the full-length ALDH structural gene, was isolated and expressed in both E. coli and Acetobacter aceti to produce a fused protein (78 kDa) with a short NH₂-terminal β-galactosidase peptide. pAL25 conferred ALDH production on a mutant of A. aceti lacking the enzyme activity. Transformation of A. aceti subsp. xylinum NBI2099 with pAL25 caused 2- and 1.4-fold increases in the production rate and in the maximum concentration of acetic acid in submerged fermentation, respectively.     

The Engelbreth-Holm-Swarm mouse tumor produces undersulfated heparan sulfate and oversulfated galactosaminoglycans

Glycosaminoglycans were prepared from the Engelbreth-Holm-Swarm mouse tumor. Enzymatic analysis demonstrated heparan sulfate (95.8%) and chondroitinase ABC-sensitive galactosaminoglycans (4.2%). HPLC analysis of the disaccharide units showed that heparan sulfate chains were undersulfated on average, comprising approximately 30% nonsulfated and 60% N-sulfated disaccharide units with small proportions of other monosulfated and disulfated disaccharide units. In contrast, galactosaminoglycan chains were oversulfated, containing an appreciable proportion (15%) of a 4,6-disulfated (so-called E-type) disaccharide unit in addition to 51% of a 4-sulfated, 22% of a 6-sulfated, and 11% of a nonsulfated disaccharide unit. The significance of the oversulfated disaccharide structure is discussed in relation to the possible regulation of functions of hybrid proteoglycans from which the galactosaminoglycan chains are derived.     

Interleukin-1 inhibits the secretion of gastric acid in rats: possible involvement of prostaglandin

To examine the hypothesis that interleukin-1 may inhibit the secretion of gastric acid, the present study was carried out using pylorusligated rats. Based upon three lines of evidence, we report here that interleukin-1, both endogenously released and exogenously administered, suppresses gastric acid secretion and that the interleukin-1-induced inhibition of acid output is possibly mediated by prostaglandin. First, lipopolysaccharide, a potent stimulant of the release and production of endogenous interleukin-1, caused the suppression of gastric acid, and this response was dose-related. Second, the intraperitoneal injection of interleukin-1 resulted in a dose-related inhibition of gastric acid output. Third, the administration of indomethacin completely blocked the suppression of gastric acid secretion induced by interleukin-1. These results demonstrated for the first time that IL-1 might be involved in the regulation of gastric secretion.     

Recent problems in cell culture systems using normal human cells

Some remarkable studies of human cell culture have contributed to many basic and applied sciences on "normal human cells"; developments of biological products or physiologically activated substances. In this reviews, some problems concerning in vitro culture systems were discussed and recent advances on the researches of normal human cells were described.