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Biomedical subjects

R J Crawford

Publications and source records attributed to R J Crawford.

At least 19 recordsLinked to original sources

The structure and activity of two cytochrome P450c21 proteins encoded in the ovine adrenal cortex.

The steroidogenic enzyme cytochrome P450c21 (CYP21A1) is synthesized in the adrenal cortex and is essential for cortisol and aldosterone production. We have studied the structure and activity of ovine P450c21 proteins by analysis and expression of the corresponding cDNAs. Two P450c21 mRNAs (2.2 and 1.7 kilobases) were detected in ovine adrenal RNA and corresponded to two types of P450c21 cDNA clones that differed in their 3' region. One clone encoded a protein similar in structure to bovine, murine, and human P450c21 proteins. The other clone contained a 3' deletion of about 500 nucleotides and encoded a P450c21 protein that was truncated by 18 residues at the carboxyl terminus. The boundaries of this deletion suggested that an additional splicing event was responsible for the shortened mRNA sequence. Detailed Southern analysis of ovine genomic DNA indicates that the two mRNAs are derived from one gene even though two P450c21 genes are present in the ovine genome. The activities of the two P450c21 proteins were determined by expressing the respective cDNA clones in COS cells. The complete P450c21 protein was an efficient catalyst of 21-hydroxylation reactions, whereas no 21-hydroxylation activity was detected in cells containing the P450c21 protein with the carboxyl-terminal deletion.

Adrenal Cortex

Evaluation of OSHA method 5 for measuring chloroform in pulp and paper industry workplace and ambient atmospheres.

As a result of the recent lowering of the Occupational Safety and Health Administration's (OSHA's) permissible exposure limit (PEL) for chloroform from 50 ppm to 2 ppm, interest has increased in measuring chloroform in pulp and paper industry workplace atmospheres. OSHA suggested that its Method 5 should be used for measuring chloroform concentrations in the range of the new PEL. There are some potential problems with using Method 5 for this purpose. Pulp and paper industry atmospheres may interfere with the measurement because of high temperature, high humidity, and the presence of chlorine. The charcoal sorbent tubes specified in OSHA Method 5 were challenged in the laboratory with a test gas containing 0.1, 0.5, 1, and 2 times the PEL of chloroform at 38 degrees C with 80% to 93% relative humidity and ca. 0.5 ppm chlorine. Application of the National Institute for Occupational Safety and Health (NIOSH) validation protocol to the results indicated a bias of -8.2% and a pooled coefficient of variation (CV) of 0.0274. The method CV is less than the critical CV; thus, the method is acceptable by the NIOSH validation protocol under these conditions. Field testing of OSHA Method 5 was successful and did not reveal any unexpected problems. Additional tests showed that with some modification, OSHA Method 5 can be used to measure chloroform concentrations in ambient atmospheres down to 0.1 ppb.

Air Pollutants, Occupational

The sequence and tissue expression of ovine renin.

The primary structure of the sheep renin precursor has been determined from its cDNA sequence. A library of cDNA clones was constructed from adrenalectomized sheep kidney poly(A)+ RNA and screened for sheep renin sequences with a cloned mouse renin cDNA probe. Of the 300,000 clones generated, 24 were hybridization positive and the nucleotide sequences of two of the longest clones were determined. These clones coded for the mature sheep renin protein and the 3'-untranslated sequence but did not extend to the amino-terminal region of preprorenin. Clones corresponding to the 5' region of renin mRNA were generated by the polymerase chain reaction and their nucleotide sequences determined. The sheep renin precursor consists of 400 amino acids with a putative leader sequence of 14 amino acids and a putative 45 or 53 amino acid prosegment. The mature sheep renin protein has a 73% sequence identity with human renin. Northern analysis demonstrated the presence of renin mRNA in the kidney but not in other tissues in the sheep. While sodium depletion of sheep caused a rise in renin mRNA in the kidney, adrenalectomy also led to a large increase in renal renin mRNA. Southern analysis of genomic DNA suggests that there is only one gene coding for renin in the sheep.

Amino Acid Sequence

The use of totally de-proteinized heterologous bone graft in non-union of the scaphoid.

In an attempt to avoid the complications associated with taking autologous bone graft for use in the treatment of non-union of the scaphoid, a clinical trial of the use of totally de-proteinized heterologous bone graft has been carried out. Ten patients with established non-union of the scaphoid were treated with internal fixation and grafting using one such commercially available material, Pyrost. The clinical and radiological results in eight patients about one year later are reported. The operative use of this material proved difficult because of its poor mechanical properties and it is not recommended as a substitute for the patient's own bone.

Adult

Imovane (zopiclone)

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Azabicyclo Compounds

Transmission of non-A, non-B hepatitis by pH4-treated intravenous immunoglobulin.

Four patients (2 with X-linked, one with common variable hypogammaglobulinaemia, and 1 with ulcerative colitis) developed non-A, non-B hepatitis (NANBH) following administration of a specific batch of intravenous immunoglobulin (IV IgG) manufactured by the Scottish National Blood Transfusion Service using the pH4/mild pepsin method. Each patient had normal serum ALT levels over a preceding period of 12-67 months, with raised values developing within 4-18 weeks of first administration of the implicated batch. Two patients had very mild symptoms of hepatitis, the other 2 being asymptomatic. Over a follow-up period of 8-12 months, ALT levels returned to normal in 3 patients, but biopsy-proven chronic NANBH developed in the fourth. The level of NANBH virus in the starting plasma used to manufacture this batch may have exceeded the capacity of the process to inactivate the virus. The transmission of NANBH by one of approximately 110 batches administered demonstrates the importance of continued close surveillance of recipients of IV IgG, even if asymptomatic, by regular monitoring of liver function tests and recording of all batches received.

Adolescent

Structure of rhesus monkey relaxin predicted by analysis of the single-copy rhesus monkey relaxin gene.

The gene encoding rhesus monkey relaxin has been investigated. A cDNA library was prepared using corpus luteal RNA from a pregnant rhesus monkey, cDNA clones encoding relaxin were isolated and the nucleotide sequence was determined. The amino acid sequence of rhesus monkey preprorelaxin, predicted from the cDNA, demonstrates that the sequence has not been strongly conserved when compared with that of man, although features characteristic of the relaxin molecule have been maintained. This structural information will allow production of rhesus monkey relaxin, leading to studies investigating the bioactivity of relaxin in a homologous primate system. Southern blot analysis indicated that there is only one relaxin gene in the rhesus monkey and baboon genomes. In this respect these primate genomes are different from the human genome which contains two relaxin genes.

Amino Acid Sequence

Segmental spinal instrumentation. A study of the mechanical properties of materials used for sublaminar fixation.

Segmental spinal instrumentation with sublaminar fixation is widely used but a variety of materials and techniques are being employed for both scoliosis and other spinal surgery. This study addresses the question as to which is best. Both the materials and the methods of securing them were studied. The materials tested were stainless steel 316L, Ortron 90 high nitrogen steel, titanium alloys, nylon straps, and Mersilene tapes. Doubled cold-worked 00.84-mm diameter stainless steel formed in a knot with a twist proved to be the strongest and most secure means of fixation with a yield value of 1,610 N. Attention to the details of technique and material such as the incorporation of a secondary twist and avoidance of hot-worked stainless steel may reduce the incidence of implant failure.

Bone Wires

Seronegative blood products prevent primary cytomegalovirus infection after bone marrow transplantation.

Seventy one patients underwent bone marrow transplantation for aplastic anaemia or haematological malignancy, 39 as allografts and 32 as autografts. All patients who were seronegative to cytomegalovirus received blood product support exclusively from seronegative community blood donors; seropositive patients received unscreened products. In no patients was there any attempt to reduce cytomegalovirus (CMV) infection by giving prophylaxis with immunoglobulin, and granulocyte transfusions were not given. The incidence of cytomegalovirus infection in the seronegative recipients (22 allograft, 15 autograft) was 0%; in the seropositive recipients 16 (63%) in allografts and 17 (18%) in autografts. These results suggest that provision of exclusively seronegative blood products is an important contribution for seronegative transplant recipients, but make little impact in autologous transplantation where the incidence of infection is low.

Adolescent

Who may give blood?

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Acquired Immunodeficiency Syndrome

The viral safety of intravenous immunoglobulin.

Human immunoglobulin for intravenous (IV) use has an established safety record with regard to transmission of hepatitis B virus. The bulk of available evidence also suggests that the human immunodeficiency virus (HIV) is not transmitted by IV immunoglobulin. There has been one report, however, of isolation of HIV from two patients with hypogammaglobulinaemia who had been treated with several immunoglobulin products. Certain IV immunoglobulin products have transmitted non-A, non-B (NANB) hepatitis but careful clinical assessment of recipients of other products suggests that non-infective preparations can be made. Interpretation of available data most likely to be correct is that contamination with NANB is reduced but not eliminated by cold-ethanol fractionation and that the use of further virucidal procedures in the finishing of immunoglobulin products will confer a higher degree of safety.

Animals

Alpha-inhibin gene expression occurs in the ovine adrenal cortex, and is regulated by adrenocorticotropin.

Inhibin is a glycoprotein hormone composed of two nonidentical subunits. It is produced by the ovary and testis and plays a vital role in gonadal function by inhibiting the secretion of FSH. More recently, additional activities associated with inhibin peptides have been identified. Inhibin heterodimers (alpha-beta) are reported to act directly on ovarian granulosa cells and inhibit estrogen production induced by FSH. Furthermore, homodimers of beta-inhibin subunits stimulate the secretion of FSH, an activity that is directly opposite to that of inhibin. Each of these inhibin-related activities are concerned with the hypothalamic-pituitary-gonadal axis. We have investigated further the complexity of inhibin activity by determining whether inhibin genes are expressed in nongonadal tissue. RNA hybridization experiments demonstrate that the alpha-inhibin gene is expressed in the sheep adrenal cortex and hybridization histochemistry shows that this gene is expressed in each of the functional zones within the cortex. Dot blot analysis showed that the level of alpha mRNA within the adrenal is influenced by ACTH, one of the major regulators of adrenal cortex function. These observations imply that there are inhibin-related peptides not directly associated with the gonads. beta-inhibin gene expression was not clearly detected in the adrenal and we conclude that if expression occurs then it does so at extremely low levels.

Adrenal Cortex