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Biomedical subjects

G D Price

Publications and source records attributed to G D Price.

36 records · Page 2Linked to original sources
PubMed1994

Paternity by cardiac transplant recipients.

To assess whether children fathered by cardiac transplant recipients are at high risk of teratogenicity, cardiac transplant centers listed with the International Society for Heart and Lung Transplantation were surveyed. Paternities after transplantation by heart (n = 35) and heart-lung (n = 1) allograft recipients have resulted in 42 pregnancies (children's age 3.3 +/- 0.3 years). The fathers' age at conception was < 45 years in 40 (95%) and > 45 years in 2 (5%). Most fathers (86%) were enjoying an active and healthy lifestyle at the time of conception; one (2%) was on dialysis and listed for kidney transplantation due to nephrotic syndrome, 1 (2%) had asthma, 4 (10%) had allograft coronary disease (1 died while waiting for second heart transplant when the child was 2 months old), and 2 (5%) were retransplanted after the pregnancies. Immunosuppressive regimens were reported for 37 paternities; drug protocols at the time of conception were as follows: 25 (60%) CsA/prednisone/AZA, 6 (14%) CsA/prednisone, 4 (10%) CsA/AZA, and 2 (5%) AZA/prednisone. Twenty-six (62%) had received treatment for rejection episodes before conception; seven (17%) had received treatment for rejections since conception. Of the 42 children fathered by these recipients, 3 (7%) were preterm, 1 (2%) had a cleft palate and lip that have subsequently been corrected, 1 (2%) died from interruption of umbilical cord circulation at 24 weeks, and 1 (2%) whose father had familial cardiomyopathy was born with a cardiomyopathy that improved with time. Although the numbers are small, the available data suggest that paternity by cardiac transplant recipients may be safe.

Adult↗
PubMed1993

Analysis of a genomic DNA region from the cyanobacterium Synechococcus sp. strain PCC7942 involved in carboxysome assembly and function.

We report on the sequencing and analysis of a 3,557-bp genomic DNA clone that is located between 4.8 and 1.2 kilobase pairs (kb) upstream of the rbcL gene and is capable of complementing a class of cyanobacterium Synechococcus sp. strain PCC7942 mutants requiring a high level of CO2. The upstream 2,704 bp of this sequence is novel, the remaining 852 bp having been reported by other workers. Four new open reading frames (ORFs) have been identified along with putative promoter elements. These ORFs, which could code for proteins of 7, 10.9, 11, and 58 kDa in size, have been named ORF 64, ccmK, ccmL, and ccmM, respectively. The last three have been named ccm genes on the basis that insertional mutagenesis of each produces a phenotype requiring a high level of CO2 (i.e., each produces a lesion in the CO2 concentrating mechanism). The putative gene product for the large ccmM ORF has three internally repeated regions and also has two possible DNA binding motifs. Two defined mutants in the 3,557-bp region, mutants PVU and P-N, have been more fully characterized. The PVU mutant has a drug marker inserted into the ccmL gene, and it possesses abnormal rod-shaped carboxysomes. The P-N mutant is a 2.64-kb deletion of DNA from the same position in ccmL to a region closer to rbcL. This mutant, which has previously been shown to lack carboxysomes and have soluble ribulosebiphosphate carboxylase/oxygenase activity, has now been shown to have a predominantly soluble carboxysomal carbonic anhydrase activity. Both mutants were found to possess carboxysomal carbonic anhydrase activities which are below wild-type levels, and in the P-N mutant this activity appears to be unstable. The results are discussed in terms of the possible interactions of putative ccm gene products in the process of carboxysome assembly and function.

Amino Acid Sequence↗
PubMed1992

Association of Carbonic Anhydrase Activity with Carboxysomes Isolated from the Cyanobacterium Synechococcus PCC7942.

The development of a simple method for the isolation of purified carboxysomes from the cyanobacterium Synechococcus PCC7942 has made it possible to identify a specific and inducible, intracellular carbonic anhydrase (CA) activity that is strongly associated with carboxysomes. This was shown, in part, through enzyme recovery experiments that indicated that a clear majority of a CA activity that is sensitive to the CA inhibitor ethoxyzolamide (I(50) = 4 mum) copurifies with a majority of the cell's ribulose-1,5-bisphosphate carboxylase/oxygenase activity in a highly purified pelletable fraction. Electron microscopy of this pelletable fraction revealed the presence of carboxysomes that were physically intact. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of carboxysome proteins showed that the large and small subunits of ribulose-1,5-bisphosphate carbosylase/oxygenase were clearly prominent and that several other minor proteins could be distinguished. The specific location of this carboxysomal CA activity is further reinforced by the finding that a previously isolated high CO(2)-requiring mutant, Type II/No. 68 (G.D. Price, M.R. Badger [1989] Plant Physiol 91: 514-525), displayed a 30-fold reduction in carboxysome-associated CA activity when tested under optimal conditions. Carboxysomal CA has the unusual property of being inactivated by dithiothreitol. The enzyme also requires 20 mm Mg(2+) (as MgSO(4)) for near maximum activity; other divalent cations, such as Ca(2+) and Mn(2+), also stimulate carboxysomal CA activity, but to a lesser extent than Mg(2+). Results are discussed in relation to the role of carboxysomes in the CO(2)-concentrating mechanism in cyanobacteria and the role that carboxysomal CA activity appears to play in this process.

Journal Article↗
PubMed1992

Isolation of a Putative Carboxysomal Carbonic Anhydrase Gene from the Cyanobacterium Synechococcus PCC7942.

The Type II mutants of the cyanobacterium Synechococcus PCC7942 (G.D. Price, M.R. Badger [1989] Plant Physiol 91: 514-525) are able to accumulate a large pool of inorganic carbon inside the cell, but are unable to utilize it for CO(2) fixation, resulting in a high CO(2)-requiring phenotype. We have isolated a 3.5-kb BamHI clone (pT2) that complements the Type II mutants, and complementation analysis with DNA subclones indicated that the complementing region was located in the 0.75-kb XhoI-Bg/II fragment. This same region hybridized to the chloroplastic carbonic anhydrase (CA) gene from spinach on Southern blots and to a mRNA of approximate 1 kb on northern blots. Restriction mapping and sequence analysis revealed that pT2 is the same as a genomic clone (pBM3.8) that complements another high CO(2)-requiring (temperature sensitive) mutant, C3P-O (E. Suzuki, H. Fukuzawa, S. Miyachi [1991] Mol Gen Genet 226: 401-408). Recently, a 272-amino acid open reading frame showing 22% homology with pea and spinach chloroplast CA genes was identified in clone pBM3.8 (H. Fukuzawa, E. Suzuki, Y. Komukal, S. Miyachi [1992] Proc Natl Acad Sci USA 89: 4437-4441). CA activity was detected in Escherichia coli cells transformed with subclones of pT2 (pT2-A and pT2-A1) containing the HindIII-Bg/II fragment, and the expressed CA has properties similar to those of the CA activity associated with carboxysomes purified from Synechococcus PCC7942 (G.D. Price, J.R. Coleman, M.R. Badger [1992] Plant Physiol 100: 784-793). Therefore, it is reasonable to conclude that the HindIII-Bg/II fragment codes for the carboxysomal CA gene product. The result is discussed in the context of the role that carboxysomal CA plays in the operation of the CO(2)-concentrating mechanism in cyanobacteria.

Journal Article↗
PubMed1990

Carbon Oxysulfide Is an Inhibitor of Both CO(2) and HCO(3) Uptake in the Cyanobacterium Synechococcus PCC7942.

Carbon oxysulfide (COS) was reinvestigated as an inhibitor of active inorganic carbon transport in cells of Synechococcus PCC7942 adapted to growth at low inorganic carbon. COS inhibited both CO(2) and HCO(3) (-) transport processes in a reversible (in the short term) and mixed competitive manner. The inhibition of COS was established using both silicone oil centrifugation experiments and O(2)-evolution studies. The K(i) for COS inhibition was 29 micromolar for CO(2) transport and 110 micromolar for HCO(3) (-) transport. These results support a model of inorganic carbon transport with a central CO(2) pump and an inducible HCO(3) (-) utilizing accessory protein which supplies CO(2) to the primary pump.

Journal Article↗
PubMed1989

Ethoxyzolamide Inhibition of CO(2) Uptake in the Cyanobacterium Synechococcus PCC7942 without Apparent Inhibition of Internal Carbonic Anhydrase Activity.

In high inorganic carbon grown (1% CO(2) [volume/volume]) cells of the cyanobacterium Synechococcus PCC7942, the carbonic anhydrase (CA) inhibitor, ethoxyzolamide (EZ), was found to inhibit the rate of CO(2) uptake and to reduce the final internal inorganic carbon (C(i)) pool size reached. The relationship between CO(2) fixation rate and internal C(i) concentration in high C(i) grown cells was little affected by EZ. This suggests that in intact cells internal CA activity was unaffected by EZ. High C(i) grown cells readily took up CO(2) but had little or no capacity for HCO(3) (-) uptake. These cells appear to possess a CO(2) utilizing C(i) pump that has a CA-like function associated with the transport step such that HCO(3) (-) is the species delivered to the cell interior. This CA-like step may be the site of inhibition by EZ. Low C(i) grown cells possess both CO(2) uptake and HCO(3) (-) uptake activities and EZ inhibited both activities to a similar degree, suggesting that a common step in CO(2) and HCO(3) (-) uptake (such as the C(i) pump) may have been affected. The inhibitor had no apparent effect on internal CO(2)/HCO(3) (-) equilibria (internal CA function) in low C(i) grown cells.

Journal Article↗
PubMed1989

Ethoxyzolamide Inhibition of CO(2)-Dependent Photosynthesis in the Cyanobacterium Synechococcus PCC7942.

Cells of the cyanobacterium, Synechococcus PCC7942, grown under high inorganic carbon (C(i)) conditions (1% CO(2); pH 8) were found to be photosynthetically dependent on exogenous CO(2). This was judged by the fact that they had a similar photosynthetic affinity for CO(2) (K(0.5)[CO(2)] of 3.4-5.4 micromolar) over the pH range 7 to 9 and that the low photosynthetic affinity for C(i) measured in dense cell suspensions was improved by the addition of exogenous carbonic anhydrase (CA). The CA inhibitor, ethoxyzolamide (EZ), was shown to reduce photosynthetic affinity for CO(2) in high C(i) cells. The addition of 200 micromolar EZ to high C(i) cells increased K(0.5)(CO(2)) from 4.6 micromolar to more than 155 micromolar at pH 8.0, whereas low C(i) cells (grown at 30 microliters CO(2) per liter of air) were less sensitive to EZ. EZ inhibition in high and low C(i) cells was largely relieved by increasing exogenous C(i) up to 100 millimolar. Lipid soluble CA inhibitors such as EZ and chlorazolamide were shown to be the most effective inhibitors of CO(2) usage, whereas water soluble CA inhibitors such as methazolamide and acetazolamide had little or no effect. EZ was found to cause a small drop in photosystem II activity, but this level of inhibition was not sufficient to explain the large effect that EZ had on CO(2) usage. High C(i) cells of Anabaena variabilis M3 and Synechocystis PCC6803 were also found to be sensitive to 200 micromolar EZ. We discuss the possibility that the inhibitory effect of EZ on CO(2) usage in high C(i) cells of Synechococcus PCC7942 may be due to inhibition of a ;CA-like' function associated with the CO(2) utilizing C(i) pump or due to inhibition of an internal CA activity, thus affecting CO(2) supply to ribulose bisphosphate carboxylase-oxygenase.

Journal Article↗
PubMed1989

Carbonic Anhydrase Activity Associated with the Cyanobacterium Synechococcus PCC7942.

Intact cells and crude homogenates of high (1% CO(2)) and low dissolved inorganic carbon (C(i)) (30-50 microliters per liter of CO(2)) grown Synechococcus PCC7942 have carbonic anhydrase (CA)-like activity, which enables them to catalyze the exchange of (18)O from CO(2) to H(2)O. This activity was studied using a mass spectrometer coupled to a cuvette with a membrane inlet system. Intact high and low C(i) cells were found to contain CA activity, separated from the medium by a membrane which is preferentially permeable to CO(2). This activity is most apparent in the light, where (18)O-labeled CO(2) species are being taken up by the cells but the effluxing CO(2) has lost most of its label to water. In the dark, low C(i) cells catalyze the depletion of the (18)O enrichment of CO(2) and this activity is inhibited by both ethoxyzolamide and 2-(trifluoromethoxy)carbonyl cyanide. This may occur via a common inhibition of the C(i) pump and the C(i) pump is proposed as a potential site for the exchange of (18)O. CA activity was measurable in homogenates of both cell types but was 5- to 10-fold higher in low C(i) cells. This was inhibited by ethoxyzolamide with an I(50) of 50 to 100 micromolar in both low and high C(i) cells. A large proportion of the internal CA activity appears to be pelletable in nature. This pelletability is increased by the presence of Mg(2+) in a manner similar to that of ribulose bisphosphate carboxylase-oxygenase activity and chlorophyll (thylakoids) and may be the result of nonspecific aggregation. Separation of crude homogenates on sucrose gradients is consistent with the notion that CA and ribulose bisphosphate carboxylase-oxygenase activity may be associated with the same pelletable fraction. However, we cannot unequivocally establish that CA is located within the carboxysome. The sucrose gradients show the presence of separate soluble and pelletable CA activity. This may be due to the presence of separate forms of the enzyme or may arise from the same pelletable association which is unstable during extraction.

Journal Article↗
PubMed1989

Expression of Human Carbonic Anhydrase in the Cyanobacterium Synechococcus PCC7942 Creates a High CO(2)-Requiring Phenotype : Evidence for a Central Role for Carboxysomes in the CO(2) Concentrating Mechanism.

Active human carbonic anhydrase II (HCAII) protein was expressed in the cyanobacterium Synechococcus PCC7942 by means of transformation with the bidirectional expression vector, pCA. This expression was driven by the bacterial Tac promoter and was regulated by the IacIQ repressor protein, which was expressed from the same plasmid. Expression levels reached values of around 0.3% of total cell protein and this protein appeared to be entirely soluble in nature and located within the cytosol of the cell. The expression of this protein has dramatic effects on the photosynthetic physiology of the cell. Induction of expression of carbonic anhydrase (CA) activity in both high dissolved inorganic carbon (C(i)) and low C(i) grown cells leads the creation of a high C(i) requiring phenotype causing: (a) a dramatic increase in the K(0.5) (C(i)) for photosynthesis, (b) a loss of the ability to accumulate internal C(i), and (c) a decrease in the lag between the initial C(i) accumulation following illumination and the efflux of CO(2) from the cells. In addition, the effects of the expressed CA can largely be reversed by the carbonic anhydrase inhibitor ethoxyzolamide. As a result of the above findings, it is concluded that the CO(2) concentrating mechanism in Synechococcus PCC7942 is largely dependent on (a) the absence of CA activity from the cytosol, and (b) the specific localization of CA activity in the carboxysome. A theoretical model of photosynthesis and C(i) accumulation is developed in which the carboxysome plays a central role as both the site of CO(2) generation from HCO(3(-) ) and a resistance barrier to CO(2) efflux from the cell. There is good qualitative agreement between this model and the measured physiological effects of expressed cytosolic CA in Synechococcus cells.

Journal Article↗
PubMed1989

Isolation and Characterization of High CO(2)-Requiring-Mutants of the Cyanobacterium Synechococcus PCC7942 : Two Phenotypes that Accumulate Inorganic Carbon but Are Apparently Unable to Generate CO(2) within the Carboxysome.

A total of 24 high CO(2)-requiring-mutants of the cyanobacterium Synechococcus PCC7942 have been isolated and partially characterized. These chemically induced mutants are able to grow at 1% CO(2), on agar media, but are incapable of growth at air levels of CO(2). All the mutants were able to accumulate inorganic carbon (C(i)) to levels similar to or higher than wild type cells, but were apparently unable to generate intracellular CO(2). On the basis of the rate of C(i) release following a light (5 minutes) --> dark transition two extreme phenotypes (fast and slow release mutants) and a number of ;intermediate' mutants (normal release) were identified. Compared to wild-type cells, Type I mutants had the following characteristics: fast C(i) release, normal internal C(i) pool, normal carbonic anhydrase (CA) activity in crude extracts, reduced internal exchange of (18)O from (18)O-labeled CO(2), 1% CO(2) requirement for growth in liquid media, normal affinity of carboxylase for CO(2), and long, rod-like carboxysomes. Type II mutants had the following characteristics: slow C(i) release, increased internal C(i) pool, normal CA activity in crude extracts, normal internal (18)O exchange, a 3% CO(2) requirement for growth in liquid media, high carboxylase activity, normal affinity of carboxylase for CO(2), and normal carboxysome structure but increased in numbers per cell. Both mutant phenotypes appear to have genetic lesions that result in an inability to convert intracellular HCO(3) (-) to CO(2) inside the carboxysome. The features of the type I mutants are consistent with a scenario where carboxysomal CA has been mistargeted to the cytosol. The characteristics of the type II phenotype appear to be most consistent with a scenario where CA activity is totally missing from the cell except for the fact that cell extracts have normal CA activity. Alternatively the type II mutants may have a lesion in their capacity for H(+) import during photosynthesis.

Journal Article↗
PubMed1981

Quality personnel.

Assuring the competent function of personnel is a fundamental part of any clinical laboratory quality assurance program. This paper will review some of the problems associated with defining and measuring the competence of laboratory personnel and will suggest a model for a uniform, a nationwide competence measuring system.

Allied Health Personnel↗
PubMed1979

Photoreceptor membrane breakdown in the spider Dinopis: localisation of acid phosphatases.

The ultrastructural localisation of acid phosphatases (AcPhs) during the normal daily breakdown of rhabdomere membrane in Dinopis has been examined using beta-glycerophosphate and p-nitrophenyl phosphate as substrates. Results are related to the classification of organelles in the receptors given by Blest, Powell and Kao (1978). Weak and infrequent reactions are obtained in multivesicular bodies (mvbs) and multilamellar bodies (mlbs) derived from them. Residual bodies (rbs) begin to react strongly as they lyse. Source of AcPhs is endoplasmic reticulum which has barely differentiated towards the GERL configuration; it becomes reactive as it is incorporated into secondary lysosomes. GERL tubules, Y-bodies and vesicles respond erratically and weakly, and are also incorporated into rbs. No evidence was found for a significant participation of Golgi bodies in these processes, and acid phosphatase cytochemistry fails to reveal a topographical relationship between GERL in these cells and Golgi saccules. Coated vesicle clusters found in the predawn receptive segments are AcPh-negative; this implies that their previous identification as GERL-derived "Nebenkerne" carrying hydrolytic enzymes to newly-formed mvbs (Blest, Kao and Powell, 1978) is dubious. Isolation bodies and autophagic vacuoles enclosing other organelles in pathological receptors give strong reactions while adjacent secondary lysosomes derived from rhabdomere membrane and associated GERL give weak ones. It is concluded that rhabdomere-derived rb lysis is more tightly regulated than other autophagic processes, and it is suggested that a high degree of control is necessary in a receptor which may repeat the autophagy of a large mass of transductive membrane at least 60--100 times in the course of its working life.

Acid Phosphatase↗
PubMedDate not supplied

Vascular rejection and its relationship to allograft coronary artery disease.

We have prospectively monitored 268 patients by our previously described method of routine immunofluorescence of endomyocardial biopsy specimens. We have classified these patients according to their rejection pattern: cellular, vascular, and mixed. The criteria for these designations have been previously described. In this study we retrospectively reviewed coronary angiograms of these patients to assess the presence and time-course of developing allograft coronary artery disease. All available explanted hearts and postmortem hearts were also assessed by light microscopic examination for acute coronary vasculitis and allograft coronary artery disease and by immunofluorescent microscopy for vascular immune complex deposition in a manner identical to immunofluorescent microscopic examination of endomyocardial biopsy specimens. Patients were also monitored for sensitization to immunoprophylactically administered murine monoclonal CD3 antibody (OKT3) and those demonstrated to be sensitized were separately analyzed. Clinical features and treatment of patients were retrospectively reviewed. We found that 141 patients could be classified as having cellular rejection, 76 as having vascular rejection, and 52 as having a mixed rejection pattern. The allograft survival in vascular rejection patients was significantly worse than in allografts of patients with cellular or mixed rejection, confirming our earlier results. Most importantly, we found a significant difference in the time to the development of allograft coronary artery disease based on the rejection pattern. This difference existed whether or not patients sensitized to OKT3 were excluded from evaluation. Patients with mixed rejection had an intermediate time to the development of allograft coronary artery disease between that of patients with cellular and vascular rejection.(ABSTRACT TRUNCATED AT 250 WORDS)

Coronary Disease↗
PubMedDate not supplied

Immunosuppressive therapy, management, and outcome of heart transplant recipients during pregnancy.

To evaluate challenges facing heart transplant recipients who become pregnant, we surveyed 194 heart transplantation centers and reviewed the literature. Thirty-two known pregnancies in heart (n = 29) or heart-lung (n = 3) allograft recipients have resulted in 29 children, including two sets of twins. The method of delivery was most often vaginal (cesarean section rate was 33%), and premature delivery was common (41%). The onset of pregnancy from the time of transplantation was 2.6 +/- 0.3 years, with the age at conception ranging from 19 to 35 years. Hypertension (44%), premature labor (30%), and preeclampsia (22%) were the most frequent maternal complications. Four patients experienced a worsening of ongoing chronic renal insufficiency; four patients experienced infections during pregnancy, and six patients (22%) were successfully treated for rejection episodes during pregnancy by adjustments in standard immunosuppressive agents. No peripartum deaths were reported; three late deaths occurred. Of the 29 children born of heart transplant recipients who became pregnant, no fetal anomalies or neonatal deaths occurred. Prematurity (41%) and low birth weight (17%) were the most common complications. All children are reported in good health at 3.4 +/- 0.4 years of age. Most transplant recipients (59%) were being treated with triple-drug immunosuppression with azathioprine, corticosteroids, and cyclosporine during pregnancy. The most common alteration to immunosuppressive therapy during pregnancy (41%) involved increasing cyclosporine doses caused by decreasing cyclosporine levels during pregnancy. Twenty-two percent of patients underwent empiric lowering of cyclosporine doses during pregnancy; four patients continued with corticosteroid tapering during pregnancy, and four patients increased corticosteroid doses.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗