Biomedical subjects
J D Clark
Publications and source records attributed to J D Clark.
Data and trends affecting Michigan physicians.
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A novel arachidonic acid-selective cytosolic PLA2 contains a Ca(2+)-dependent translocation domain with homology to PKC and GAP.
We report the cloning and expression of a cDNA encoding a high molecular weight (85.2 kd) cytosolic phospholipase A2 (cPLA2) that has no detectable sequence homology with the secreted forms of PLA2. We show that cPLA2 selectively cleaves arachidonic acid from natural membrane vesicles and demonstrate that cPLA2 translocates to membrane vesicles in response to physiologically relevant changes in free calcium. Moreover, we demonstrate that an amino-terminal 140 amino acid fragment of cPLA2 translocates to natural membrane vesicles in a Ca(2+)-dependent fashion. Interestingly, we note that this 140 amino acid domain of cPLA2 contains a 45 amino acid region with homology to PKC, p65, GAP, and PLC. We suggest that this homology delineates a Ca(2+)-dependent phospholipid-binding motif, providing a mechanism for the second messenger Ca2+ to translocate and activate cytosolic proteins.
Normoglycaemic ketoacidosis in a woman with gestational diabetes.
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Na(+)-H+ exchanger subtypes: a predictive review.
In recent years, many reports have appeared describing distinct heterogeneity of proteins that heretofore were considered to be a single species or type. The division of proteins into different classes or subtypes is aided by pharmacological tools such as selective ligands, functional measurements such as those examining kinetic or regulatory differences, and molecular biological approaches that have identified distinct genes coding for similar yet distinguishable gene products. Currently, much effort is directed toward understanding the significance of these sometimes subtle differences in terms of functional consequences for the cells in which they exist. Although most reports to date involve hormone and neurotransmitter receptor subtypes, it is also possible that other cell surface molecules such as ion transporters exist as multiple subtypes. In this paper we review the current evidence that Na(+)-H+ exchange activity is mediated by different Na(+)-H+ exchanger subtypes. Although subtypes have not been identified with certainty, we can predict certain distinguishing characteristics that these putative subtypes may have that may be of value in correlating predicted gene products obtained from cDNA cloning with previously characterized Na(+)-H+ exchangers.
Michigan's hospitals in the 1980s.
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The changing face of Michigan's smaller and rural hospitals.
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Health care in the USSR: one woman's story.
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Influence of type of enclosure on exercise fitness of dogs.
The effect of various confinement conditions on physical fitness in dogs was evaluated. Eighteen 9.5- to 10-month-old female purpose-bred Beagles were maintained individually for 3 months at a time in 1 of 6 confinement conditions: Condition A--an outdoor housing area with a conventional dog house and free access to a 6.1 x 9.1-m pen; condition B--outdoor kennel with a conventional dog house and free access to a 1.8 x 6.1-m run; condition C--indoor environmentally controlled 1.2 x 3.66-m run; condition D-0.9 x 1.2 x 0.84-m conventional laboratory cage in an indoor environmentally controlled room; condition E--0.9 x 1.2 x 0.84-m conventional laboratory cage in an indoor environmentally controlled room with treadmill exercise (7 km/h at a 10% grade) for 30 min/d, 5 d/wk; condition F--0.71 x 0.86 x 0.69-m conventional laboratory cage in an indoor environmentally controlled room. During the final week of each 3-month interval, muscle succinate dehydrogenase enzyme activities and submaximal exercise heart rates (during treadmill exercise) were determined to estimate physical fitness. Also, 5 days after being moved into a different housing condition, blood samples were collected for plasma cortisol determination. The type of confinement condition for dogs had little effect on muscle succinate dehydrogenase activity, but had a modest effect on submaximal exercise heart rates of dogs.(ABSTRACT TRUNCATED AT 250 WORDS)
Small area analysis. What is it and does it have practical value?
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Cloning, sequencing, and expression of the gene encoding the porcine alpha 2-adrenergic receptor. Allosteric modulation by Na+, H+, and amiloride analogs.
The gene for an alpha 2-adrenergic receptor has been cloned from a porcine genomic library, using as a probe a 0.95-kilobase Pst fragment of the gene for the human platelet alpha 2-adrenergic receptor. The identity of the cloned porcine gene was confirmed initially on the basis of partial amino acid sequence information obtained following cyanogen bromide digestion of homogeneous preparations of porcine brain alpha 2-adrenergic receptors. The deduced amino acid sequence for the porcine receptor, when compared to other members of the family of guanine nucleotide-binding protein-coupled receptors, shares the same overall structural characteristics and most closely resembles the human platelet C10 alpha 2-adrenergic receptor (greater than 93% homology). The putative porcine alpha 2-receptor gene was expressed in the COS-M6 cell line. Transfected cells display saturable [3H]yohimbine binding. The KD for [3H]yohimbine, determined in digitonin-solubilized preparations, is 5.8 nM. The selectivity of agonists and antagonists in competing for [3H]yohimbine binding to membranes prepared from the transfected cells is characteristic of the alpha 2A subtype of adrenergic receptors. The porcine alpha 2-receptor also was expressed permanently in LLC-PK1 porcine kidney cells at a level of 100 pmol/mg protein. The alpha 2-agonist UK14304 is able to attenuate forskolin or vasopressin-stimulated cAMP accumulation by at least 50% in these cells. Allosteric modulation of [3H] yohimbine binding by Na+, H+, and 5-amino-substituted analogs of amiloride also was demonstrated for the alpha 2-receptor expressed in COS-M6 cells. Moreover, these modulatory effects were quantitatively similar to those observed for homogeneous preparations of the alpha 2-receptor purified from porcine brain cortex. Retention of the effects of cations and amiloride analogs in transiently expressed alpha 2-receptors supports the interpretation that the allosteric sites for these agents reside in the alpha 2-receptor molecule itself.
Variation in Michigan hospital use rates: do physician and hospital characteristics provide the explanation?
Previous small area analysis studies have shown that hospital admission rates (total, medical and surgical) vary among hospital service areas. Using 1983 Michigan hospital inpatient data from 53 nonmetropolitan Detroit lower peninsula hospital service areas, one physician characteristic and 13 hospital characteristics (in the categories of resource supply, services offered and organization) were tested for their association with and explanation of 14 hospital use rates. Registered nurses per bed and the weighted proportion of board certified physicians to total physicians were inversely related to and offered significant contribution to the explanation of the variation in total use rates and in four medical causes for admission rates (circulatory, respiratory, digestive and genito-urinary). Physician and hospital variables provided significant explanation for six of the seven surgical procedure rates tested (appendectomy, hemorrhoidectomy, cholecystectomy, inguinal hernia repair, prostatectomy and hysterectomy). Four causative factors derived from the characteristics studied were postulated to influence the hospital use rates. The first factor was the small rural nature of the average high use hospital service area. High use areas had a lower proportion of board certified physicians and fewer RNs per bed, beds per hospital, and house staff per 10,000 population than did low use areas. Another factor was the inequality in the distribution of high technology diagnostic services. High use hospital service areas had fewer diagnostic services than did low use areas. The third factor was the inequality in the rural hospital environment produced by the presence or absence of medical education programs. The fourth factor was the impact of the definition and size of a hospital service area. Current small area analysis methodology assigns every small area to a hospital service area, no matter what the probability of the population using the hospital(s) within the service area. This research questions that methodology, suggests the need for hospital service area definitions based upon the specific diagnosis or procedure being studied and postulates that some rural hospital distance decay curves may turn upward at farther distances when the immediate availability of treatment is too critical to allow patients to return to distant residences.
Purification of a 110-kilodalton cytosolic phospholipase A2 from the human monocytic cell line U937.
The major dithiothreitol-resistant phospholipase A2 activity present in the cytosol of U937 cells has been purified greater than 200,000-fold by sequential chromatography on phenyl-5PW, heparin-Sepharose CL-6B, high-performance hydroxylapatite, TSK-gel G3000-SW, and Mono Q columns. This 110-kDa cytosolic phospholipase A2 is distinct from the relatively small (14-kDa) dithiothreitol-sensitive phospholipases A2 that are secreted from many cell types. This additional phospholipase A2 selectively hydrolyzes fatty acid at the sn-2 position of the glycerol and favors phospholipids containing arachidonic acid, which is the rate-limiting precursor for prostaglandin and leukotriene production. Interestingly, a greater than 5-fold increase in phospholipase A2 activity is noted as the calcium concentration increases from the levels found in resting cells to those observed in activated macrophages. We suggest that this enzyme and not the previously described secretory phospholipase A2 is activated by cytosolic effectors such as GTP-binding regulatory proteins and protein kinases to initiate the production of prostaglandins, leukotrienes, and platelet-activating factor. To distinguish this cytosolic enzyme from the previously described secretory ones, we suggest referring to it as cPLA2 for cytosolic phospholipase A2 and collectively referring to the secretory phospholipases A2 as sPLA2s.
A profile of orthodontic treatment in the general dental service in Scotland 1979-1987.
The study casts and records of two samples of patients who had received orthodontic treatment involving the use of appliances within the General Dental Service (GDS) in Scotland during 1979/81 and 1986/87 were examined to study the pattern of treatment, and to determine whether there had been any change over this period. Both samples contained a wide range of malocclusions which were treated mostly with removable appliances. Although few fixed appliances were used and few lower arch treatments were carried out there was a trend towards a greater use of these appliances and an increase in these treatments in 1986/87 compared with 1979/81. There was no change in the infrequent use of headgear while functional appliances were used in only six of the treatments in the 1986/87 sample as compared to none in the 1979/81 sample. In the more recent sample many more treatments were undertaken by 'specialist' General Dental Practitioners (i.e. those who work in the GDS, but limit their practice to orthodontics) and fewer treatments were undertaken under the direction of a Consultant Orthodontist.
Insulin requirements of diabetic women who breast feed.
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Abnormalities of thyrotrophin (TSH) evening rise and pulsatile release in haemodialysis patients: evidence for hypothalamic-pituitary changes in chronic renal failure.
Using a sensitive enzyme amplified immunoassay for TSH, the evening rise and pulsatile release of TSH were studied in 10 men with chronic renal failure treated by haemodialysis. Compared to euthyroid male controls the evening rise of TSH was attenuated (median 0.066 vs 0.195 mU/l/h, P less than 0.01) and the rate of rise correlated with the TSH response to TRH (r = 0.93, P less than 0.001). All subjects showed TSH pulsatility in at least one method of data analysis but the less sensitive incremental method showed no significant difference in pulse frequency and amplitude between the two groups. However, with time series analysis, periodicity was shorter (median 45 vs 95 min, P = 0.013) and pulse amplitude smaller (median 0.06 vs 0.175 mU/l, P = 0.017) in renal patients. Pulse amplitude, but not periodicity, correlated with the TSH response to TRH (r = 0.68, P less than 0.05). In addition, serum total thyroxine, free thyroxine and free triiodothyronine concentrations were reduced, while serum prolactin and 17 beta-oestradiol concentrations were raised. These changes in TSH evening surge and pulsatile release may contribute to the reduction in thyroidal hormone concentrations seen in renal failure and emphasize the value of sensitive methods of hormone and pulse data analysis.
Studies of the entero-insular axis following pancreas transplantation in man: neural or hormonal control?
To study the role of hormonal and neural factors in the control of the entero-insular axis the insulin, C-peptide, and glucose-dependent insulinotropic peptide (GIP) responses to oral and intravenous glucose were investigated in 5 patients who had received a combined kidney and paratopic pancreas transplant, with physiological portal venous drainage. The incremental areas under the insulin and C-peptide responses to oral glucose were significantly greater than the responses to intravenous glucose (insulin: patients 7983 +/- 1937 (+/- SE) vs 3513 +/- 2188 mU l-1 min, p less than 0.002, control subjects 5505 +/- 1035 vs 1066 +/- 484 mU l-1 min, p less than 0.004; C peptide: patients 440 +/- 80 vs 144 +/- 61 nmol l-1 min, p less than 0.01, control subjects 200 +/- 38 vs 63 +/- 16 nmol l-1 min, P less than 0.01). The incretin effects for insulin (patients 4.4 +/- 1.4, control subjects 7.7 +/- 1.8) and C-peptide (patients 4.4 +/- 0.9, control subjects 3.7 +/- 0.9) and the GIP responses to oral and intravenous glucose were not significantly different between transplant patients and control subjects. As the incretin effect was preserved, despite a denervated pancreas, hormonal rather than neural factors may be more important in mediating increased insulin secretion after oral carbohydrate. The normal GIP response is compatible with its proposed role as an insulinotropic hormone.
Malate synthase: proof of a stepwise Claisen condensation using the double-isotope fractionation test.
Although aldolase-catalyzed condensations proceed by stepwise mechanisms via the intermediacy of nucleophilic enol(ate)s or enamines, the mechanisms of those enzymes that catalyze Claisen-type condensations are unclear. The reaction pathway followed by an enzyme from this second group, malate synthase, has been studied by the double-isotope fractionation method to determine whether the reaction is stepwise or concerted. In agreement with earlier work, a deuterium kinetic isotope effect D(V/K) of 1.3 +/- 0.1 has been found when [2H3]acetyl-CoA is the substrate. The 13C isotope effect at the aldehydic carbon of glyoxylate has also been measured. For this determination, the malate product (containing the carbon of interest at C-2) was quantitatively transformed into a new sample of malate having the carbon of interest at C-4. This material was decarboxylated by malic enzyme to produce the appropriate CO2 for isotope ratio mass spectrometric analysis. The 13C isotope effect with [1H3]acetyl-CoA [that is, 13(V/K)H] is 1.0037 +/- 0.0004. By use of the known values of the intermolecular and intramolecular deuterium effects and of 13(V/K)H, the value of the 13C isotope effect when deuteriated [2H3]acetyl-CoA is the substrate [that is, 13(V/K)D] can be predicted for three possible mechanisms. If 13(V/K)H is a kinetic isotope effect and the reaction is concerted, the value of the 13C effect on deuteriation of acetyl-CoA will rise to 1.011; if 13(V/K)H is a kinetic isotope effect and the reaction is stepwise, the value of the 13C effect will fall to 1.0025; and if the 13C effect is an equilibrium isotope effect deriving from glyoxylate dehydration, the reaction is necessarily stepwise, and the value of 13(V/K)D will be 1.0037, unchanged from that of 13(V/K)H. Experimentally, the value of 13(V/K)D is 1.0037 +/- 0.0007, which requires that malate synthase follow a stepwise path. It is therefore clear that the two salient characteristics of enzymes that catalyze Claisen-like condensations, namely, the absence of enzyme-catalyzed proton exchange with solvent and the inversion of the configuration at the nucleophilic center, which had been suggestive of a concerted pathway, are not mechanistically diagnostic.