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Biomedical subjects

B J Morris

Publications and source records attributed to B J Morris.

231 records · Page 13Linked to original sources
PubMedDate not supplied

Properties of inactive renin in human plasma.

1. "Inactive" renin in human plasma can be revealed by pH 3.3- or cold-mediated activation and in normal plasma represented 76% of the "total" renin. 2. Pregnancy plasma contained considerably more "inactive" renin and consisted of 93% of "total" renin. 3. "Active" renin in normal plasma had an apparent molecular weight of 43,000 compared with 60,000 for "active" renin in pregnancy plasma by gel filtration. 4. "Inactive" renin in pregnancy plasma also had an apparent molecular weight of 60,000, while in normal plasma there were two peaks of inactive renin at 62,000 and 46,000. 5. After affinity chromatography of a protein preparation from pregnancy plasma on Concanavalin A-Sepharose activation by pH 3.3 could no longer be produced, suggesting that the activating factor had been removed, as would occur if it were not a glycoprotein. When pepsinogen, in a concentration similar to that found in plasma, was added prior to dialysis to pH 3.3 activation was restored. 6. Ion-exchange chromatography demonstrated that at pH 8.4 "inactive" renin bore slightly less negative charges than "active" renin. 7. "Inactive" renin in human plasma therefore appears to be a larger molecular weight species than the "active" renin in normal plasma and is capable of activation during treatment to pH 3.3 or cold with no apparent alteration in size. The results suggest an important role of pepsin (after conversion from pepsinogen) in the activation of "inactive" renin during dialysis at pH 3.3.

Angiotensinogen↗
PubMedDate not supplied

Activation of plasma prekallikrein and inactive renin by puff adder venom.

The activation of inactive renin during incubation of human plasma with puff adder venom at pH 7.4 was found to be a complex process. Gel filtration on Sephacryl S-200 indicated that the venom contains a major peak of caseinolytic and renin-activating activity of low molecular weight. This enzyme was a metalloproteinase. During incubation with plasma the metalloproteinase formed a complex with alpha 2-macroglobulin and retained caseinolytic and reninactivation activity. Venom metalloproteinase activated prekallikrein, but not plasmin, and plasms kallikrein may account for at least part of the activation of inactive renin observed.

Caseins↗
PubMedDate not supplied

Biosynthesis of preprorenin and intracellular conversion of prorenin to renin.

1. The existence of a biosynthetic precursor of renin (prorenin) has been inferred, but never established experimentally. The aim of the present study was to provide this evidence. 2. In a continuous labelling experiment using submandibular glands of male Quackenbush mice (an extraordinarily rich source of renin) a molecule of Mr 44 500 was immunoprecipitated with antiserum to renin. 3. The Mr 44 500 protein was converted to Mr 40 000 renin in a time-dependent fashion during incubation of tissue at 37 degrees C: 15 min after addition of [35S] methionine only 16% of immunoprecipitable radioactivity was renin, whereas by 4 h this had increased to 86%. The Mr 44 500 protein would therefore appear to be a 'pro' form of renin. 4. Submandibular gland mRNA was translated in a cell-free system derived from rabbit reticulocyte lysate and a molecule of Mr 46 000 was immunoprecipitated with antirenin. This was preprorenin and the Mr of the 'pre' region would thus be 1500, which is consistent with the size generally found for the 'pre' region of secretory proteins. 5. The present study has therefore demonstrated for the first time the biosynthesis of a 'pro' form of renin (prorenin) and its conversion to renin intracellularly.

Animals↗
PubMedDate not supplied

Synthesis of mouse renin as a 2-5-33-5 kilodalton pre-pro-two-chain molecule and use of its cDNA to identify the human gene.

A DNA coding for mouse submandibular gland (MSG) renin was used in studies of the biosynthesis and processing of MSG renin and precursors and as a probe for identification of the human renin gene. A 2 kilodalton signal peptide was demonstrated by cleavage of preprorenin (45 kilodaltons) in the presence of microsomal membranes from dog pancreas. Prior selection of renin mRNA by hybridization with its cDNA obviated the possibility that a protease translated from total MSG mRNA could be activated by the microsomal membranes and then act on the primary translation product. In vitro labelling experiments with female MSG demonstrated that prorenin (43 kilodaltons) is rapidly converted to renin (38 kilodaltons) and that testosterone stimulated synthesis by increasing transcription of renin mRNA. Electrophoresis under non-reducing conditions demonstrated 38 kilodalton renin which when reduced ran as two bands of 33 and 5 kilodaltons. Thus, native MSG renin has two chains linked by disulphide bonding. Hydrolysis of the 38 kilodalton single chain occurred only slowly during in vitro labelling. Prorenin bound only weakly to a pepstatin affinity column and could be activated by adding trypsin to column fractions. Both 38 kilodalton single and two chain renin bound strongly however, suggesting that both are active. Base sequencing of MSG renin cDNA indicated an Arg-Arg residue additional to the published amino acid sequence of the heavy chain. This may account for the two forms seen on isoelectric focusing. Mouse renin cDNA was used as a hybridization probe in screening a human genomic library in order to identify the human renin gene. The DNA in positive colonies had overlapping restriction maps and the coding region was found.

Animals↗
PubMedDate not supplied

Structure of human renin and expression of the renin gene.

The amino acid sequence of human renin was identified for the first time. This was determined from the nucleotide sequence of exons in the human renin gene identified in a genomic library by recombinant DNA techniques. Examination of amino acid residues involved in the enzymatic hydrolysis by human renin of the unique Leu10-Val11 bond of human angiotensinogen revealed features peculiar to this highly specialized aspartyl protease. The expression of the renin gene was examined with a hybridization probe for renin mRNA in sections and extracts of tissues. In the submandibular gland of mice renin mRNA, like renin, increased during development and in response to testosterone in females; sodium depletion increased renin mRNA in kidney.

Amino Acid Sequence↗
PubMedDate not supplied

Isolation of genomic clones coding for the heavy chains of two human cardiac myosins.

A 14 kilo-base pair DNA clone (lambda HCMHC8) was isolated from a human genomic library by hybridization with a complementary DNA coding for a rabbit cardiac myosin heavy chain. lambda HCMHC8 hybridized to RNA isolated from cardiac but not skeletal muscle and formed heteroduplexes with a genomic clone coding for the fast type of rabbit cardiac myosin heavy chain. lambda HCMHC8 represented at least the 3' half of the gene and contained over 11 exons which together spanned 4 kb of the coding region estimated to be 6 kb. Probes made from lambda HCMHC8 were used to rescreen the library in order to isolate overlapping clones and so extend the sequence (estimated to be approximately 25 kb for the whole gene, including introns). In addition, a clone having a different restriction map was isolated suggesting that man, like rat and rabbit, has two cardiac myosin heavy chain genes. These may code for proteins having different ATPase activity and be expressed in different proportions in different cardiac states, including hypertension.

DNA↗
PubMedDate not supplied

A structural analysis of human renin.

A three-dimensional model of human renin has been used to define amino acid residues involved in interaction with substrate. The structure revealed proteolytic cleavage sites on the surface of the molecule that may help explain numerous experimental observations concerning prorenin and its activation and forms of renin differing in size.

Binding Sites↗
PubMedDate not supplied

Renin mRNA concentration in rat hypothalamus is decreased by enalapril.

1. Renin mRNA is expressed in several extrarenal tissues, including brain. The aim of the present study was to quantify renin mRNA in the hypothalamus in response to a low NaCl diet and angiotensin converting enzyme inhibitor treatment, both of which are well-known stimuli of renin mRNA in kidney. 2. Groups of six Sprague-Dawley rats were given either a normal diet, low NaCl chow (0.04% NaCl), enalapril (0.25 mg/mL in drinking water), or low NaCl + enalapril, for 7 days. Renin mRNA in the hypothalamus was quantified by a competitive reverse transcriptase-polymerase chain reaction (RT-PCR) technique. 3. Renin mRNA concentration in the hypothalamus of rats receiving a normal diet was 52 +/- 3 S.E. fg/microgram total RNA. Low NaCl had no effect (57 +/- 7), whereas values were significantly lower in rats treated with enalapril, either given alone (38 +/- 2; P = 0.002) or combined with a low NaCl diet (33 +/- 4; P = 0.003). 4. In conclusion, we have quantified renin mRNA in the rat hypothalamus and shown that it can be suppressed significantly by enalapril. This response is opposite to that seen in the kidney after enalapril.

Angiotensin-Converting Enzyme Inhibitors↗
PubMedDate not supplied

Cross-sectional study of a microsatellite marker in the low density lipoprotein receptor gene in obese normotensives.

1. The low density lipoprotein receptor is an important regulator of serum cholesterol which may have implications for the development of both hypertension and obesity. In this study, genotypes for a low density lipoprotein receptor gene (LDLR) dinucleotide polymorphism were determined in both lean and obese normotensive populations. 2. In previous cross-sectional association studies an ApaLI and a HincII polymorphism for LDLR were shown to be associated with obesity in essential hypertensives. However, these polymorphisms did not show an association with obesity in normotensives. 3. In contrast, this study reports that preliminary results for an LDLR microsatellite marker, located more towards the 3' end of the gene, show a significant association with obesity in the normotensive population studied. These results indicate that LDLR could play an important role in the development of obesity, which might be independent of hypertension.

Chi-Square Distribution↗
PubMedDate not supplied

Examination of the role of nitric oxide synthase and renal kallikrein as candidate genes for essential hypertension.

1. Nitric oxide synthase and renal kallikrein are both involved in blood pressure regulation. Genes for these enzymes may, therefore, be considered candidates for hypertension pathogenesis. 2. In the present study, genotypes for nitric oxide synthase and renal kallikrein microsatellite markers were determined in a cross-sectional association analysis of hypertensive patients and normotensive control subjects. 3. Results from this study did not indicate an association of either of the candidate gene polymorphisms with essential hypertension. Hence, findings for this study do not support a role for these genes in human hypertension.

Alleles↗
PubMedDate not supplied

No difference in the proportion of insulin receptor exon 11 +/- isoform mRNA in the liver of rats after development of hypertension.

1. There are two functionally different isoforms of the insulin receptor in humans and rats. We hypothesized that a change in their relative proportion could be of relevance to insulin resistance in hypertension. 2. A reverse-transcriptase polymerase chain reaction technique was established for the detection of mRNA for the exon 11+ and exon 11- isoforms and the proportion of each was determined in 3, 6, 9 and 12 week old spontaneously hypertensive rats and Wistar-Kyoto rats, as well as adrenocorticotrophin (ACTH)-induced hypertensive rats and controls. 3. The proportion of the exon 11+ form (approximately 95%) and exon 11- form (approximately 5%) was similar in the liver of all rats studied. 4. We conclude that there is no change in insulin receptor isoform expression in the liver in the models of hypertension studied.

Adrenocorticotropic Hormone↗
PubMedDate not supplied

A linkage and cross-sectional study of hypertension and obesity using a poly (A) Alu-repeat polymorphism at the glucagon receptor gene locus (17q25).

1. Previous glucagon receptor gene (GCGR) studies have shown a Gly40Ser mutation to be more prevalent in essential hypertension and to affect glucagon binding affinity to its receptor. An Alu-repeat poly(A) polymorphism colocalized to GCGR was used in the present study to test for association and linkage in hypertension as well as association in obesity development. 2. Using a cross-sectional approach, 85 hypertensives and 95 normotensives were genotyped using polymerase chain reaction primers flanking the Alu-repeat. Both hypertensive and normotensive populations were subdivided into lean and obese categories based on body mass index (BMI) to determine involvement of this variant in obesity. For the linkage study, 89 Australian Caucasian hypertension affected sibships (174 sibpairs) were genotyped and the results were analysed using GENEHUNTER, Mapmaker Sibs, ERPA and SPLINK (all freely available from http:@linkage.rockefeller.edu./soft/list.hmtl). 3. Cross-sectional results for both hypertension and obesity were analysed using Chi-squared and Monte Carlo analyses. Results did not show an association of this variant with either hypertension (chi(2) = 6.9, P = 0.14; Monte Carlo chi(2) = 7.0, P = 0.11; n = 5000) or obesity (chi(2) = 3.3, P = 0.35; Monte Carlo chi(2) = 3.26, P = 0.34; n = 5000). In addition, results from the linkage study using hypertensive sib-pairs did not indicate linkage of the poly(A) repeat with hypertension. Hence, results did not indicate a role for the Alu-repeat in either hypertension or obesity. However, as the heterozygosity of this poly(A) repeat is low (35%), a larger number of hypertensive sib-pairs may be required to draw definitive conclusions.

DNA↗
PubMedDate not supplied

Reassignment of human renin gene to chromosome 1q32 in studies of a (1;4)(q42;p16) translocation.

The human renin gene (REN) has been assigned to chromosome 1q42. Linkage studies are, however, inconsistent with this localization. We therefore reexamined the question of the location of REN using a patient whose distal chromosome 1q arm was translocated to chromosome 4 [(1;4)(q42;p16)]. In situ hybridization using a 3H-labelled REN probe demonstrated hybridization signals confined to the q32 band of chromosome 1, with radioactivity in the translocated 1q42 region being similar to the low levels along all other chromosomes.

Chromosome Mapping↗
PubMedDate not supplied

Insulin receptor gene in hypertension.

The first molecular genetic association with human essential hypertension (HT) involved the insulin receptor gene (INSR). This highly significant result in Caucasians was for an insertion/deletion polymorphism in intron 9. A polymorphism in exon 8 showed a weak association, but a microsatellite in intron 2 proved negative for HT, although has shown an association with plasma insulin in Japanese. A similar spectrum of genetic associations for variants spanning INSR has been noted for insulin-dependent diabetic patients with rapidly-progressing renal disease, a subgroup having a strong family history of essential HT. Association with HT has also been found for an INSR variant in CHinese. Insulin resistance secondary to an INSR 'defect', or other causes, would increase insulin, which has cardiovascular effects, and insulin can raise angiotensinogen. Also, insulin is co-secreted with amylin, which can increase renin secretion. In the spontaneously HT rat there is evidence for reduced down-regulation of INSR expression in response to NaCl-loading, consistent with a promoter effect. When combined with observations of insulin resistance in essential HT patients and their pre-HT offspring, the possibility of dys-regulation of INSR merits attention in disease etiology in a proportion of essential HT patients.

Alternative Splicing↗