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Biomedical subjects

H M Goodman

Publications and source records attributed to H M Goodman.

At least 163 records · Page 9Linked to original sources
PubMed1983

Chromosome assignment of genes encoding the alpha and beta subunits of glycoprotein hormones in man and mouse.

The chromosomal locations of the genes for the common alpha subunit of the glycoprotein hormones and the beta subunit of chorionic gonadotropin in humans and mice have been determined by restriction enzyme analysis of DNA isolated from somatic cell hybrids. The CG alpha gene (CGA), detected as a 15-kb BamHI fragment in human DNA by hybridization to CG alpha cDNA, segregated with the chromosome 6 enzyme markers ME1 (malic enzyme, soluble) and SOD2 (superoxide dismutase, mitchondrial) and an intact chromosome 6 in human-rodent hybrids. Cell hybrids containing portions of chromosome 6 allowed the localization of CGA to the q12 leads to q21 region. The greater than 30- and 6.5-kb BamHI CGB fragments hybridizing to human CG beta cDNA segregated concordantly with the human chromosome 19 marker enzymes PEPD (peptidase D) and GPI (glucose phosphate isomerase) and a normal chromosome 19 in karyotyped hybrids. A KpnI-HindIII digest of cell hybrid DNAs indicated that the multiple copies of the CG beta gene are all located on human chromosome 19. In the mouse, the alpha subunit gene, detected by a mouse thyrotropin (TSH) alpha subunit probe, and the CG beta-like sequences (CG beta-LH beta), detected by the human CG beta cDNA probe, are on chromosomes 4 and 7, respectively.

Animals↗
PubMed1983

The effects of growth hormone on blood pressure and renin secretion in hypophysectomized rats.

The effects of hypophysectomy and GH on blood pressure and renin secretion were studied. Within 2 weeks after removal of the pituitary gland of rats, mean arterial blood pressure declined 30%, and heart rate fell 50%. No significant effect of hypophysectomy on blood volume or hematocrit was noted. Within 2 h after the iv administration of 10 micrograms ovine GH to hypophysectomized rats, blood pressure, but not heart rate, was restored to normal. Despite the hypotension, the PRA of hypophysectomized rats was not significantly greater than that of intact animals. This relatively low PRA could not be accounted for by a lack of renin per se, since kidneys of hypophysectomized rats contained at least as much renin as kidneys from intact rats. Neither the PRA of hypophysectomized rats nor the kidney renin content was significantly altered 24 h after a single injection of GH (100 micrograms, ip). When perfusions were performed at 100 mm Hg, the rate of renin secretion by isolated kidneys obtained from hypophysectomized rats was markedly lower than that of kidneys from intact rats. Reduction of mean perfusion pressure to 50 mm Hg or adding isoproterenol to the perfusate produced much smaller increases in renin secretion by kidneys of hypophysectomized rats than that observed in kidneys from intact rats. Kidneys obtained from hypophysectomized rats treated with GH 24 h earlier had a renin content similar to that of kidneys from untreated animals, but secreted renin at a much higher rate. Kidneys from hormone-treated hypophysectomized rats also exhibited a greater renin secretory response to low perfusion pressure or isoproterenol. These data indicate that hypophysectomy severely impairs the renin secretory response of the isolated kidney and that a single injection of GH can reverse this impairment. These observations suggest the possibility that the hypotension characteristic of hypophysectomized rats may, in part, reflect the lack of GH and alterations in the renin-angiotensin system.

Animals↗
PubMed1983

Relationship between binding and biological effects of human growth hormone in rat adipocytes.

Insulin-like responses to human GH (hGH) were produced in adipocytes isolated from the epididymal fat of normal rats 3 h after excision of the tissues. Insulin-like responses consisted of increased oxidation of glucose and incorporation of its carbons into total lipid, increased oxidation of L-[1-14C]leucine, and antagonism of the lipolytic actions of epinephrine. Refractoriness to these effects of hGH in the fourth hour of incubation was produced by the addition of as little as 3 ng/ml hGH as soon as possible after excision of the tissues. These cells also responded to the delayed lipolytic effect of hGH in the presence of theophylline. The cells were found to have high affinity, low capacity, specific binding sites for 125I-labeled hGH. Monoiodination of hGH did not interfere with its capacity to produce biological responses. Specific binding equilibrated rapidly and appeared to saturate at about 100 ng/ml. In cells that were capable of exhibiting an insulin-like response to hGH, rat and ovine GH successfully competed with [125I]hGH for binding sites, but porcine insulin, at a concentration of 100 mU/ml, failed to reduce the binding of [125I]hGH, indicating that GH does not produce its insulin-like effects by interacting with the insulin receptor. Binding of [125I]hGH in cells that are refractory to the insulin-like effects of GH is indistinguishable from binding in responsive cells. Scatchard analysis of the data for both responsive and refractory cells gave linear plots consistent with a single class of about 20,000 receptors/cell, which become half-saturated at a concentration of approximately 20 ng/ml. This corresponds well with 30-50 ng/ml needed for half-maximal insulin-like responses and the 3-10 ng/ml ED50 for induction of refractoriness or lipolysis. It thus appears unlikely that there are appreciable spare receptors for insulin-like responses. These findings make it likely that refractoriness to the insulin-like effects of GH occurs at a postreceptor site.

Adipose Tissue↗
PubMed1983

Growth hormone and lipolysis: a reevaluation.

A recent report claimed that the lipolytic activity found in clinical grade human GH (hGH) was entirely attributable to an acidic peptide contaminant. Using the same preparations as studied in the earlier report, we found that delayed lipolytic activity, as demonstrated in the presence of dexamethasone, was fully evident in the highly purified hGH preparation which had been reported to be devoid of lipolytic activity. We also studied the acidic peptide contaminant in which all of the lipolytic activity of clinical grade hGH was alleged to reside and found its lipolytic activity was equivalent to that of the highly purified hGH. Both compounds increased lipolysis in the presence of 0.1 micrograms/ml dexamethasone after a lag period of 1 h. The minimum effective concentration of both fell within the range of 1 and 10 ng/ml, and both were capable of producing a 2- to 3-fold increase in glycerol production. When saturating amounts of each were added simultaneously they produced no greater effect than either alone, although the lipolytic rate was far from saturated as evidenced by the more than 2-fold further increase in glycerol production when 100 ng/ml epinephrine was added along with the combined peptides. The lipolytic response to highly purified hGH was comparable to that seen with bacterially synthesized hGH which should be free of pituitary peptide contaminants. The acidic peptide appeared to be a variant of hGH as judged by its ability to produce both an insulin-like response and refractoriness. Furthermore, the acidic peptide was indistinguishable from hGH in competing for specific binding sites on adipocytes. Although an ability to increase glycerol production in rat adipose tissue in vitro thus appears to be an intrinsic property of hGH, the physiological significance of this effect remains to be established.

Adipose Tissue↗
PubMed1983

Preparation and biological reactivity of polyiodinated human growth hormone.

Studies with iodinated human GH (hGH) were undertaken to characterize the iodinated product and to evaluate the possibility that iodination might distort the hGH molecule sufficiently to interfere preferentially with binding to one or more classes of receptors on adipocytes. hGH containing an average of 1.2 atoms of 125I/molecule was subjected to cleavage with cyanogen bromide, and the cleavage products were examined by autoradiography after electrophoresis of the lyophilized reaction mixture on polyacrylamide gel. 125I was found to be distributed in the fragments according to their content of tyrosine, suggesting that there is no single tyrosine residue that is iodinated preferentially. Digestion of the iodinated hormone with pronase followed by paper chromatography revealed that 7.6 times as much iodine appeared as MIT than as DIT, suggesting that the incorporation of 125I into a tyrosine residue had little influence on the incorporation of a second iodine atom. By increasing the molar ratios of iodide and chloramine-T to hGH in the reaction mixture, increasing amounts of iodine were incorporated into hGH in a highly predictable and reproducible manner until a product containing 11-12 atoms of iodine/molecule hGH was obtained. The ratio of MIT to DIT decreased progressively with increasing incorporation of iodine. The data suggest that only six of the eight tyrosine residues of hGH are accessible for iodination. Iodinated hGH retained full biological potency with respect to stimulation of glucose oxidation (insulin-like effect) and lipolysis even after incorporation of 4.6 atoms of I/molecule hGH. Two preparations containing 9.4 and 9.6 atoms of I/molecule hGH retained full biological potency, whereas another preparation containing 9.4 atoms/molecule had approximately one third the potency of uniodinated hGH with respect to both activities. Some biological potency, at least with respect to stimulation of lipolysis, was retained even in maximally iodinated preparations. We conclude that iodination occurs randomly among the six accessible tyrosine residues, and even after polyiodination, the interaction of hGH with its receptors in adipocytes is not completely compromised.

Adipose Tissue↗
PubMed1982

Assigning the polymorphic human insulin gene to the short arm of chromosome 11 by chromosome sorting.

We have determined the subchromosomal location of the human insulin gene by analyzing DNA isolated from sorted human metaphase chromosomes. Metaphase chromosome suspensions were sorted into fractions according to relative Hoechst fluorescence intensity by the fluorescence activated chromosome sorter. The chromosomal DNA in each fraction was characterized by restriction endonuclease analysis. Initial sorts indicated that the insulin gene-containing fragment resided in a fraction containing chromosomes 9, 10, 11, and 12. Studies of cell lines that contained chromosome translocations permitted the assignment of the insulin gene to a derivative chromosome that contains portions of the short arm of chromosome II. Simultaneous sorting of the normal homolog from this small derivative chromosome separated the two different sized insulin gene-containing restriction fragments in this individual. These data indicate that the two restriction fragments represent insulin gene polymorphism and not duplicate gene loci.

Alleles↗
PubMed1982

Disproportionate expression of the two nonallelic rat insulin genes in a pancreatic tumor is due to translational control.

The expression of the two nonallelic but highly homologous rat insulin genes (1 and 2) in a transplantable beta-cell tumor is found to be 10-fold higher for rat1 insulin than rat2 insulin, while in normal pancreatic tissue there are approximately equal amounts of each protein. No large sequence rearrangements of the genes were apparent by restriction analysis of the tumor DNA, and both genes were found to be specifically hypomethylated in the tumor as compared with other nonpancreatic tissue. Equivalent amounts of both insulin 1 and 2 precursor transcripts, as well as stable, mature mRNAs were detected in the tumor. However, two-dimensional gel analysis of immunoprecipitated rat1 and rat2 preproinsulins synthesized in vitro revealed a 10:1 ratio of rat1 to rat2 proteins. A 1:1 ratio was obtained when the tumor mRNA was treated in vitro with vaccinia virus capping extract, suggesting a structural modification at the 5' terminus of the rat2 mRNA. These results are discussed in the context of insulin regulation by glucose, shown to be due to translational control.

Adenoma, Islet Cell↗
PubMed1982

A plasmid that replicates in both mouse and E. coli cells.

Cloned bovine papilloma virus (BPV) DNA induces cellular transformation when introduced into mouse cells growing in culture; the transferred viral DNA replicates as an extrachromosomal, closed circular element. BPV DNA is therefore an inviting replicon to construct a "shuttle" vector that can replicate in both mammalian cells and E. coli. Although BPV DNA devoid of bacterial plasmid sequences has been successfully employed to reintroduce cloned genes into rodent cells, construction of a true shuttle plasmid has been hampered by a disruptive influence of bacterial plasmid sequences that appear to block cellular transformation when included on the transferred molecule. We constructed a molecule, pGP, containing the transforming region of the BPV genome, the rat growth hormone gene, and bacterial plasmid pBR 327, and have found unexpectedly that the intact molecule can induce cellular transformation of mouse cells at high efficiency despite the presence of bacterial sequences in the transferred plasmid. A similar plasmid without the growth hormone segment does not transform mouse cells. The pGP molecule replicates as a stable plasmid in both mouse cells, where there are 30 to 80 monomer episomes per cell, and E. coli, and may be shuttled back and forth unaltered between the two kinds of cells. The growth hormone gene is transcribed in the mouse cells and gives rise to a transcript that is longer than authentic rat growth hormone mRNA and does not appear to be regulated by glucocorticoids. When pGP is cotransferred into mouse L-cells with herpes simplex virus tk gene, it appears to integrate, and free monomer episomes are not observed.

Animals↗
PubMed1982

Tumor induction by Agrobacterium tumefaciens: analysis of the boundaries of T-DNA.

Molecular cloning has been used to isolate the ends of that portion of the tumor-inducing (Ti) plasmid of Agrobacterium tumefaciens which has been designated T-DNA and which has been transferred to the genome of tobacco crown gall tumor cells. Analysis of the DNA sequences of the plant border clones compared with the corresponding sequences of the Ti plasmid suggests that the mechanism of transferred DNA integration and subsequent stabilization is precise at the right border and imprecise on the left. The T-DNA junction occurs within a variation of a single base pair (bp) on the right but varies over at least 70 bp on the left. In addition, there are several sequences which are repeated near the ends of the T-DNA region in the Ti plasmid. Seemingly, there is no specificity with regard to the site of integration in the plant genome.

Base Sequence↗
PubMed1982

Nopaline synthase: transcript mapping and DNA sequence.

The DNA sequence of the nopaline synthase gene (nos) from Agrobacterium tumefaciens Ti plasmid pTiT37 and adjacent regions up to the right border of the T-DNA was determined. The 5' and 3' termini of the polyadenylated nos mRNA, isolated from a T37 tobacco teratoma tumor line, were localized by S1 mapping. The final mRNA is unspliced, encoded by a region of about 1450 bp, and specifies an open reading frame of 413 amino acids. Potential transcriptional signals in the 5' flanking DNA, such as CATAAA ("TATA box") and GGTCACTAT ("CAT box"), bear close resemblance to other eukaryotic promoters. Two putative polyadenylation signals, AATAAA and AATAAT, are found about 135 and 50 bp from the 3' end, respectively. This study may provide information for the development of expression vectors for genes in plant cells; moreover, the structural gene can be used as an easy screenable marker.

Amino Acid Oxidoreductases↗
PubMed1981

Polymorphism in the 5'-flanking region of the human insulin gene and its possible relation to type 2 diabetes.

The arrangement of the human insulin gene in DNA from 87 individuals was analyzed by the Southern blot hybridization technique with a cloned genomic human insulin probe. Insertions of 1.5 to 3.4 kilobase pairs in the 5'-flanking region of the gene were found in DNA from 38 individuals. These insertions occurred within 1.3 kilobase pairs of the transcription initiation site. In contrast, no insertions were observed in the region 3' to the coding sequence. The prevalence of these insertions in type 2 diabetes was significantly greater than in the other groups (P less than .001). The limitation of this striking length polymorphism to a potential promoter region suggests that these insertions may play a role in insulin gene expression.

Base Sequence↗
PubMed1981

Human growth hormone DNA sequence and mRNA structure: possible alternative splicing.

We have determined the complete sequence of the human growth hormone (hGH) gene and the position of the mature 5' end of the hGH mRNA within the sequence. Comparison of this sequence with that of a cloned hGH cDNA shows that the gene is interrupted by four intervening sequences. S1 mapping shows that one of these intervening sequences has two different 3' splice sites. These alternate splicing pathways generate hGH peptides of different sizes which are found in normal pituitaries. Comparison of sequences near the 5' end of the hGH mRNA with a similar region of the alpha subunit of the human glycoprotein hormones reveals an unexpected region of homology between these otherwise unrelated peptide hormones.

Amino Acid Sequence↗
PubMed1981

DNA sequence of the rat growth hormone gene: location of the 5' terminus of the growth hormone mRNA and identification of an internal transposon-like element.

The present communication describes the molecular cloning and DNA sequence determination of the rat growth hormone (rGH) gene. The rGH gene was cloned on an 11 kilobase EcoRI fragment of total rat DNA; it has four intervening sequences which correspond in position to those of the human growth hormone (hGH) gene. One of the intervening sequences in the rGH gene contains a possible transposable element: a 200 base pair direct repeat that is itself flanked by an exact 15 base pair direct repeat. The DNA sequence was used to estimate the location of the 5' end of the mature growth hormone mRNA. By S1 nuclease mapping it was located approximately 25 bases "downstream" from a TATAAA sequence presumed to play a role in initiation of transcription of the rGH gene.

Animals↗
PubMed1981

L-Leucine activates branched chain alpha-keto acid dehydrogenase in rat adipose tissue.

The activity of branched chain alpha-keto acid dehydrogenase in extracts of adipose tissue was elevated after homogenization of tissue segments which had been incubated in buffer containing 0.3 mM leucine. A maximum increase (4-fold) was observed in extracts of tissues incubated in buffer containing 2.5 mM leucine, alpha-Ketoisocaproate and leucine caused maximum increases which were of similar magnitude and which required the same length of incubation of the tissue segments (5 to 15 min). The effect of leucine on branched chain alpha-keto acid dehydrogenase activity was observed both in the presence and absence of insulin, which also increased the activity of the enzyme in tissue extracts. Intact adipose tissue segments oxidized [I-14C]leucine at a maximum rate approximately 4 times that of [1-(14)C]valine. The rate of valine oxidation by intact tissue segments was doubled by addition of 0.2 to 0.5 mM unlabeled leucine, but not isoleucine, to medium containing 2 mM [1-(14)C]valine. Leucine, but not valine, also stimulated the rate of oxidation of 2 mM [U-14C]isoleucine by intact tissue segments. These results suggest that branched chain alpha-keto acid dehydrogenase activity, which is thought to limit the rate of branched chain amino acid oxidation in adipose tissue, may be sensitive to changes in the concentration of leucine in rat blood.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
PubMed1981

Effects of growth hormone on the oxidation of [1-14C]-pyruvate in adipose tissue of hypophysectomized rats.

The acute effects of growth hormone on the oxidation of [1-14C]-pyruvate to 14CO2 were studied in epididymal adipose tissue obtained from hypophysectomized rats. At concentrations ranging from 10 ng/ml to 1 microgram/ml, growth hormone increased the rate of pyruvate oxidation by 20-60%. A lag period of up to 30 min was required for the full effect of the hormone to develop. Addition of fructose to the incubation medium increased the rate of pyruvate oxidation in response to either growth hormone or insulin. The effects of 1 microgram/ml growth hormone were comparable in magnitude to those of 1 mU/ml insulin, and pyruvate oxidation in the presence of both agents was no greater than in the presence of either on its own. The enhancement of pyruvate oxidation by growth hormone, like that caused by insulin, probably results from activation of pyruvate dehydrogenase. Increased activity of pyruvate dehydrogenase was found in cell-free extracts of adipose tissue that had been exposed to either growth hormone or insulin. The response of tissue segments to growth hormone followed the same pattern as observed for other acute insulin-like effects of the hormones; it was transient and disappeared within 3 hours despite continued presence of the hormone. Previous exposure of the tissues to growth hormone made them refractory to the hormone upon reexposure.

Adipose Tissue↗