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Biomedical subjects

K J Hardy

Publications and source records attributed to K J Hardy.

At least 19 recordsLinked to original sources
PubMed1992

A protein kinase C-activating phorbol ester accelerates the T cell antigen receptor-stimulated phosphatidylinositol cycle in normal human CD4+ T cells.

Ligation of the TCR on Jurkat T lymphoblastoid cells causes an 1,4,5-inositol trisphosphate-dependent rise in intracellular cytoplasmic calcium that is inhibited by PMA, a potent activator of protein kinase C. Consequently, protein kinase C is widely believed to mediate feedback inhibition of TCR-activated phospholipase C. We have now extended these studies to normal unblasted human CD4+ T lymphocytes, examining the PMA sensitivity of both the TCR complex-mediated release of total inositol-phosphates and the resynthesis of the parent phosphoinositides. In contrast to Jurkat, in which PMA inhibited release of 1,4,5-inositol trisphosphate by 60% and total inositolphosphates by 40% (50% inhibitory concentration, 5.6 nM), normal cells displayed a marked increase in anti-CD3-induced phosphatidylinositol (PI) cycling in the presence of PMA. Both total inositolphosphate release and PI resynthesis were maximally elevated (88% and 342%, respectively) by a PMA concentration that also optimally supported a subsequent proliferative response; the ED50 was at least 11.7-fold lower than that for the inhibitory effect of PMA on breakdown of total Jurkat PI. A PKC nonactivating phorbol ester had no effect. If anti-CD3 was replaced by the mitogenic lectin PHA, PI resynthesis was similarly up-regulated by PMA in these highly purified cells. The PMA up-regulatory phenomenon was not a simple consequence of cell blastogenesis, inasmuch as there was no early effect on the non-signaling-associated phosphatidylethanolamine compartment after CD3 stimulation. Thus, PKC activation appears to accelerate TCR-linked PI metabolism in normal Th cells, in contrast to the feedback inhibitor paradigm observed in Jurkat and other tumor cell systems.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1992

Detection of colour vision abnormalities in uncomplicated type 1 diabetic patients with angiographically normal retinas.

Colour vision function was assessed in 38 non-complicated type 1 diabetic patients in whom fluorescein angiography was normal, and was compared with that in 36 age-matched, non-diabetic controls. All of the patients were healthy and none were taking medication except insulin. The eye examination, which was normal in every patient, included the Ishihara and City University tests, measurement of Snellen acuity, slit-lamp examination, tonometry, and fundal photography as well as fluorescein angiography. Colour discrimination ability was measured with the Farnsworth-Munsell 100-hue test. Mean (SE) 100-hue test error score for the diabetic group was 86.8 (8.1) compared with 28.2 (3.3) for controls, p<<0.001. There was no relation between colour vision abnormalities and diabetes duration (r = 0, p>0.05), blood glucose at the time the colour tests were performed (r = 0.4, p > 0.05), most recent glycated haemoglobin result (r = 0.3, p>0.05), or the mean of all previous glycated haemoglobin results (r = 0, p>0.05). It is concluded that colour discrimination may be abnormal in uncomplicated type 1 diabetic patients before the onset of retinopathy, and that colour discrimination losses in diabetes may not be of vascular aetiology.

Adult↗
PubMed1992

Ontogeny of gastrin, somatostatin, and the H+/K(+)-ATPase in the ovine fetus.

In the term human and ovine fetus, plasma gastrin is elevated, but gastric acid secretion is below adult levels, suggesting a developmentally related immaturity in gastrin and gastric acid regulation. This study investigated a number of elements of the gastric acid regulatory system: gastrin and its glycine-extended precursor, somatostatin, and the H+/K(+)-ATPase. Measurements were made in blood, antrum, and fundus of the ovine fetus during the last half of gestation, of 15-day-old lambs, and of adult sheep at the level of mRNA synthesis, tissue storage, and secretion. Plasma amidated gastrin (gastrin-amide) was elevated at or above adult values from 125 days (term is 145 days) and steadily increased with development, peaking in the lamb. Similar changes occurred with plasma glycine-extended gastrin (gastrin-gly). The peak concentration of antral gastrin-amide was present in the lamb, while the maximum antral gastrin-gly level occurred 1 week before birth. Gastrin mRNA paralleled the changes in antral gastrin-gly. The proportion of higher mol wt species of gastrin decreased during gestation in both plasma and antrum. Low amounts of mRNA for the H+/K(+)-ATPase was present from at least 120 days of gestation and antedated gastric acid secretion. However, there was a 3-fold increase in H+/K(+)-ATPase mRNA from the 140-day-old fetus to the lamb, the period when the greatest reduction in gastric pH occurred (pH 5 to 2). Antral and fundic somatostatin increased rapidly in the fetus at 120 days gestation and were above adult values at term and in the lamb. Somatostatin mRNA changed in parallel to somatostatin peptide. Somatostatin-14 was the major species in antrum and fundus throughout development. The increase in circulating and antral gastrin-amide after birth may be the result of increased amidation of gastrin-gly as well as increased expression of gastrin mRNA. Amidation of gastrin may be a regulatory step in the production of biologically active gastrin during development. The major increase in gastrin and the H+/K(+)-ATPase that occurs in the week before and after gestation correlated with the onset of increased gastric acidity.

Adenosine Triphosphatases↗
PubMed1992

Tumor recognition and lytic competence of IL-2-activated lymphocytes: regulation of both antibody-independent and -dependent cellular cytotoxicity via P75 IL-2 receptor.

Fc receptor-positive lymphocytes (FcR+) contain lymphokine-activated killer cell (LAK) precursors that in response to IL-2 develop potent antitumor cytotoxicity. These FcR+ cells are also capable of antibody-dependent cytotoxicity (ADCC), which can be detected using fresh human peripheral blood lymphocytes (PBL) directed to murine targets, however, PBL-mediated ADCC to human tumors usually is very low, requiring a stimulation of the PBL, which also can be accomplished with IL-2. Using human melanoma tumor target cells, with and without the 14G2a monoclonal antibody, we examined in parallel the role of p75 IL-2 receptor for regulation of the induction of both LAK and ADCC forms of antitumor cytotoxicity. Enrichment of FcR+ cells from fresh peripheral blood by elutriation and flow cytometry, followed by varying periods of IL-2 culture, revealed a differential kinetics of activation. ADCC was detectable after PBL exposure to IL-2 for as short as the 4 h cytotoxicity assay, while LAK activation required more than 24 h of exposure. Elimination of the FcR+ cells by magnetic bead depletion from large granular lymphocyte populations (LGL) resulted in a loss of both LAK and ADCC. Addition of antibody known to block the binding of IL-2 to the p75 molecule of the IL-2 receptor complex (Mik-beta 1) to activation cultures at zero time resulted in abrogation of both cytotoxicities. These results suggest that differentiation and maturation of the ADCC effectors occurs in response to IL-2 via the p75 molecule, as also does LAK activation.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Monoclonal↗
PubMed1991

Disposition of the diastereoisomers quinine and quinidine in the ovine fetus.

The disposition of the diastereoisomers quinine and quinidine was investigated in the near-term pregnant ewe. Five sheep were administered quinine and quinidine separately in random order by a combination of bolus and 30-h iv infusion. On a subsequent occasion, four of the five sheep were also administered the two drugs simultaneously. After separate dosage, systemic clearance of quinine tended to be greater than that of quinidine (714 +/- 299 versus 422 +/- 146 mL/min, p = 0.08). Maternal renal clearance exhibited no stereoselectivity and represented less than 2% of total clearance. Simultaneous administration did not alter the disposition of either drug in the mother. After separate dosage, fetal total concentrations (Cf) of quinine and quinidine were substantially lower than maternal total concentrations, as reflected in Cf:Cm ratios of 0.15 +/- 0.06 versus 0.10 +/- 0.08, respectively. Similarly, fetal unbound concentrations (Cfu) were substantially lower than maternal unbound concentrations (Cmu; Cfu/Cmu = 0.46 +/- 0.09 for quinine and 0.23 +/- 0.09 for quinidine). This indicates the presence of fetal elimination of both isomers. Fetal renal clearances of quinine and quinidine were similar (0.34 +/- 0.24 mL/min versus 0.38 +/- 0.24 mL/min) and less than that of endogenous creatinine, indicating the absence of net renal tubular secretion. After simultaneous dosage of quinine and quinidine, Cf:Cm (0.48 +/- 0.24 and 0.31 +/- 0.19, respectively) and Cfu:Cmu (0.73 +/- 0.14 and 0.52 +/- 0.20, respectively) were greater than for separate dosages. Fetal renal clearance of both drugs was unchanged, suggesting that the higher Cfu:Cmu ratios after simultaneous dosage were due to mutual inhibition of the fetal metabolism of these drugs.

Animals↗
PubMed1991

Laparoscopic cholecystectomy: initial experience.

Twenty-five consecutive patients underwent percutaneous laparoscopic cholecystectomy (PCC). The gallbladder was removed successfully in 18 patients. The mean postoperative hospital stay was 1.4 days and patients returned to normal activity at a mean 8 days after operation. Postoperative pain was minimal. Formal laparotomy was performed in 7 patients due to: bleeding (3 patients), stone spillage (3 patients) and exploration of the common bile duct (1 patient). Complications were reduced with experience and strict adherence to the described operative technique. With obvious advantages for the patient, hospitals and the community an increased demand for PCC is inevitable. However, its role in the management of cholelithiasis and overall safety have yet to be determined. There is a significant learning curve and proper training is necessary. The widespread introduction of PCC has immediate implications for surgical training.

Adolescent↗
PubMed1991

Deterioration in blood glucose control in females with diabetes changed to a basal-bolus regimen using a pen-injector.

Although the basal-bolus regimen of insulin delivery is more physiological than conventional twice daily insulin it does not usually improve blood glucose control. Having recently withdrawn 3 patients from pen-injector therapy for deteriorated blood glucose control we examined the records of all our diabetic patients on a pen-injector for at least 1 year. Of the 51 patients identified, 3 had died, 2 moved away, and 4 patients (3 females) had pen-injector therapy withdrawn for poor control; 3 patients had inadequate records. The remaining 21 males and 18 females were well matched for age, duration of diabetes, and duration of pen-injector treatment. Blood glucose control deteriorated significantly during pen-injector therapy in female patients (9.6 +/- 1.2 to 10.7 +/- 1.4%, p = 0.02), but not in the males (9.3 +/- 1.3 vs 9.4 +/- 1.4%, NS). Weight and insulin dose did not change significantly in any patient. Blood glucose control may deteriorate in young women with diabetes changed to insulin therapy with a pen-injector.

Adult↗
PubMed1990

A model of human cytokine regulation based on transfection of gamma interferon gene fragments directly into isolated peripheral blood T lymphocytes.

An approach has been optimized permitting measurement of human cytokine reporter gene expression after transient transfection directly into purified human peripheral blood T lymphocytes. Comparing the expression of interleukin 2 (IL-2) CAT with a series of specially engineered gamma interferon (IFN-gamma) constructs, a fundamental difference in the molecular mechanisms regulating these two cytokines has been suggested. A potent, tissue-specific, constitutive-acting positive regulatory element was located between sequences -215 and -53 in the human IFN-gamma gene. Deletion analyses suggested that sequences slightly upstream, between positions -251 to -215, exerted a powerful dominant suppressive influence over that positive element. Negative elements appear to play a major role in controlling the regulation of human IFN-gamma gene expression. We thus propose a model of cytokine gene regulation in which selective derepression may be an important fundamental mechanism of induction and/or positive modulation.

Base Sequence↗
PubMed1990

Identification of enhancer-like elements in human IFN-gamma genomic DNA.

We have previously shown that transfection of a plasmid clone containing full length human IFN-gamma genomic DNA into a murine T-lymphoblastoid line is followed by basal expression of the transfected gene, with increased transcription occurring upon stimulation of the cells with either phorbol ester or IL-2. In addition, upon transfection of this DNA into murine fibroblasts, high level constitutive transcription was observed. In contrast to the results obtained under tissue culture conditions, introduction of the same DNA into the mouse germline resulted in tissue-specific expression of the transgene. We now report identification of a region 500-bp 5' of the human IFN-gamma TATAA box that has strong, PMA-inducible, enhancer-like activity when linked to a reporter gene (CAT) and transfected into a murine T cell line. However, when the same region of IFN-gamma genomic DNA was introduced into NIH-3T3 cells, no enhancer activity was detected either in the presence or absence of PMA. We have further found that an intronic region of the IFN-gamma genomic DNA (nucleotides 405-674) also contains enhancer activity that is functional in either fibroblasts or T cells. Enhancer activity of the intronic region is also PMA-inducible in the mouse T cells but constitutive in fibroblasts. Collectively, our observations suggest that control of human IFN-gamma gene expression is complex, involving noncontiguous regulatory domains in both 5' flanking and intronic regions of that gene.

Base Sequence↗