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Biomedical subjects

D Freedman

Publications and source records attributed to D Freedman.

At least 19 recordsLinked to original sources
PubMed1996

p53 binds to a novel recognition sequence in the proximal promoter of the rat muscle creatine kinase gene and activates its transcription.

The rat muscle creatine kinase (CKM) gene promoter is unusual since it is one of the few cellular promoters containing a p53 response element which is located proximally (bp -168 to -57) to the transcription start site. We have previously shown that p53wt transactivates transcription in vivo of rat CKM, in CV-1 monkey kidney cells, through this 112 bp promoter-proximal fragment which contains at least five degenerate p53-binding elements. In this report, we employed the gel-shift assay and demonstrated that recombinant, immunoaffinity-purified mouse p53wt binds to this 112 bp CKM sequence and activates the in vitro transcription of the proximal CKM promoter by nuclear extracts from CV-1 cells. Also, a competitor plasmid containing this 112 bp CKM fragment interferred with the in vivo transactivation of CKM by p53. This CKM fragment, when cloned upstream of the rat brain creatine kinase (CKB) promoter, mediated the p53 transactivation of CKB. Analyses of p53wt and a series of missense mutants (altered in conserved region II of p53) showed that binding of p53 to the CKM promoter was required but was not sufficient for transactivation. The results are discussed in relation to the possible role of p53wt in the expression of CKM in cell types which may not express the myogenic transcription factors.

Animals↗
PubMed1996

Differential effects of physiological versus pathophysiological plasma concentrations of epinephrine and norepinephrine on ketone body metabolism and hepatic portal blood flow in man.

Few studies that have examined the effects of catecholamines on ketogenesis have considered the effects of catecholamines on hepatic portal blood flow. Since hepatic blood flow is a major determinant of hepatic ketogenesis (via modification of free fatty acid availability), interpretation of these studies is difficult. To better define the relative contributions of these variables, we studied the effects of physiological and pathophysiological plasma concentrations of epinephrine and norepinephrine on plasma ketone body concentrations and hepatic portal blood flow in controlled paired studies in young healthy male volunteers. To assess the effects of physiological catecholamine concentrations, each of eight subjects received 60-minute sequential infusions of epinephrine (10 ng/kg/min) and norepinephrine (32.5 ng/kg/min) together with a control infusion of heparin (0.4 U/kg/min) separated by 60-minute washout periods. Similar increments in plasma nonesterified fatty acid ([NEFA] to approximately 1 mmol/L) were observed during each infusion. The ketotic ratios, calculated as the ratio of plasma ketone bodies to fatty acids integrated above baseline for 90 and 120 minutes, respectively, for epinephrine and norepinephrine infusions were both significantly greater (P < .005 for each) than for the heparin control infusion. To assess the effects of pathophysiological plasma catecholamine concentrations, each of eight subjects also received sequential 60-minute infusions of epinephrine 60 ng/kg/min, norepinephrine 80 ng/kg/min (plus heparin 0.1 U/kg/min), and a separate control infusion of heparin with or without Intralipid (KabiVitrum, Alameda, CA). Whereas integrated plasma fatty acid levels were approximately twofold greater than those observed in the physiological protocol, the absolute integrated ketone body response to the pathophysiological concentration of epinephrine was significantly lower than that observed for the physiological dose of the hormone (P < .05). In contrast, the ketotic ratio for norepinephrine was significantly greater (P < .005) than for both epinephrine and the control infusion of heparin with or without Intralipid. Significant (P < .01) increases above baseline fasting levels were observed in plasma glucose and immunoreactive insulin concentrations during infusion of pathophysiological concentrations of epinephrine. Because of the technical difficulties of simultaneously measuring portal blood and sampling blood frequently, studies were repeated in six additional subjects using noninvasive image-guided flowmetry to measure percentage changes in hepatic portal blood flow during catecholamine infusion. Norepinephrine reduced hepatic portal blood flow significantly at the low-physiological concentration by 12% (P < .05) and at the pathophysiological concentration by 18% (P < .05). In summary, (1) both epinephrine and norepinephrine were associated with significant ketotic effects at physiological plasma concentrations; and (2) when infused at pathophysiological concentrations, only norepinephrine exerted a significant additional ketotic effect. Since norepinephrine has a significant simultaneous effect of reducing hepatic portal blood flow, we conclude that previous studies may have underestimated the effect of norepinephrine on hepatic ketogenesis.

Adult↗
PubMed1995

Post-operative problems following anterior cruciate ligament reconstruction.

Seventy adult patients were studied during the postoperative rehabilitation period following anterior cruciate ligament reconstruction in order to investigate the role of pre-, intra-, and postoperative factors in range of motion and graft problems. A standard bone-patellar tendon-bone autograft was used for the reconstruction. Pre- and intraoperative factors such as concomitant injuries, time from injury to surgery, age, sex, and tunnel placement were recorded. Tunnel placement was recorded on intraoperative radiographs of the final guide pin placement and compared to pin placement on cadaver knees. The results indicated a significant relation between early reconstruction (< 1 month) following the injury and range of motion problems during the early rehabilitation period (P < 0.001). This relation disappeared by the end of the first postoperative year. Prolonged surgery was also associated with early motion problems (P < 0.05). Graft laxity or failure was correlated with an earlier return of range of motion (P < 0.05). We hypothesized that graft failure can have a biologic cause rather than a mechanical one since intraoperative X-rays indicated a near-anatomic tunnel placement in this patient group when compared to ideal placement in the cadaver knees.

Adolescent↗
PubMed1994

PTR1: a reductase mediating salvage of oxidized pteridines and methotrexate resistance in the protozoan parasite Leishmania major.

Trypanosomatid protozoans are pterin auxotrophs, a finding noted decades ago which heralded the discovery of key metabolic roles played by pteridines in eukaryotes. We have now identified the enzyme mediating unconjugated pteridine salvage in the human parasite Leishmania major, PTR1 (pteridine reductase 1, formerly hmtxr or ltdh). PTR1 is the gene in the amplified H region responsible for methotrexate (MTX) resistance, and belongs to a large family of oxidoreductases with diverse substrates and roles. We generated Leishmania lacking PTR1 by homologous gene targeting, and these ptr1- mutants required reduced biopterin (dihydro- or tetrahydrobiopterin) for growth. PTR1 purified from engineered Escherichia coli exhibited a MTX-sensitive, NADPH-dependent biopterin reductase activity. PTR1 showed good activity with folate and significant activity with dihydrofolate and dihydrobiopterin, but not with quinonoid dihydrobiopterin. PTR1 thus differs considerably from previously reported pteridine reductases of trypanosomatids and vertebrates. Pteridine reductase activity was diminished in ptr1- Leishmania and was elevated in transfected parasites bearing multiple copies of PTR1; correspondingly, ptr1- was MTX-hypersensitive whereas the multicopy transfectant was MTX-resistant. The concordance of the biochemical and genetic properties of PTR1 suggests that this is the primary enzyme mediating pteridine salvage. These findings suggest several possible mechanisms for PTR1-mediated MTX resistance and should aid in the design of rational chemotherapy.

Animals↗
PubMed1994

Sexually transmitted diseases from the tropics. An overview.

The tropical ecosystem as related to sexually transmitted diseases and their epidemiology is outlined in this article. The predominance of first-generation bacterial STDs with more florid presentations, together with a back-ground of more covert infections, such as with Chlamydia and human papillomavirus, are noted. Presentations of infections in people returning from the tropics are highlighted, and particular features of the various infections relevant to their tropical origin are discussed. The overview can only hint at the wide diversity of infections and presentations from the tropics and the challenge in both diagnosis and control that they present.

Chlamydia Infections↗
PubMed1993

[Sexually transmitted diseases in the united Europe at the threshold of the year 2000].

The potential for changes in the epidemiology and prevalence of Sexually Transmitted Diseases in the next decade are reviewed against the emerging demographic and political trends. Potential development of the specialty, together with increasing availability of diagnostic and screening techniques, will require that the principles of management of Sexually Transmitted Diseases are preserved and enhanced. The computer will become an important tool for all areas of physician activity. "Europe sans frontières" will be an opportunity for enhanced contact tracing facilities across a larger geographic area and provide a forum for enhanced public health education.

European Union↗
PubMed1992

Modified CelliGen-packed bed bioreactors for hybridoma cell cultures.

This study describes two packed bed bioreactor configurations which were used to culture a mouse-mouse hybridoma cell line (ATCC HB-57) which produces an IgG1 monoclonal antibody. The first configuration consists of a packed column which is continuously perfused by recirculating oxygenated media through the column. In the second configuration, the packed bed is contained within a stationary basket which is suspended in the vessel of a CelliGen bioreactor. In this configuration, recirculation of the oxygenated media is provided by the CelliGen Cell Lift impeller. Both configurations are packed with disk carriers made from a non-woven polyester fabric. During the steady-state phase of continuous operation, a cell density of 10(8) cells per cm3 of bed volume was obtained in both bioreactor configurations. The high levels of productivity (0.5 gram MAb per 1 of packed bed per day) obtained in these systems demonstrates that the culture conditions achieved in these packed bed bioreactors are excellent for the continuous propagation of hybridomas using media which contains low levels (1%) of serum as well as serum-free media. These packed bed bioreactors allow good control of pH, dissolved oxygen and temperature. The media flows evenly over the cells and produces very low shear forces. These systems are easy to set up and operate for prolonged periods of time. The potential for scale-up using Fibra-cel carriers is enhanced due to the low pressure drop and low mass transfer resistance, which creates high void fraction approaching 90% in the packed bed.

Animals↗
PubMed1991

A proline-rich structural protein of the surface sheath of larval Brugia filarial nematode parasites.

Both cDNA and genomic DNA sequences have been isolated which encode a proline-rich precursor protein of the sheath from microfilariae, the first stage larvae of the filarial nematode parasites Brugia pahangi and Brugia malayi. This 22-kDa protein is soluble only under reducing conditions and is extensively cross-linked by both disulfide and nonreducible bonds. Immunogold electron microscopy shows that the protein is localized exclusively in the sheath, a vestigial remnant of the eggshell, which is retained by and encloses the mature microfilaria. Analysis by Western blotting confirms that the protein is expressed only in microfilariae and adult female worms, although transcripts are detectable only in adult females. The deduced amino acid sequence contains a short N-terminal hydrophobic putative leader sequence, a central repetitive domain that contains 14 copies of a degenerate 5-amino acid repeat with the consensus sequence Met-Pro-Pro-Gln-Gly, and a C-terminal proline-rich domain flanked by clusters of cysteine residues. These clusters can be aligned with cysteine residues implicated in cross-linking of a family of cuticular collagens originally identified in Caenorhabditis elegans but which extends to other nematodes.

Amino Acid Sequence↗
PubMed1991

'A lovely bunch of coconuts'.

Aspiration and instillation of Sodium tetradecyl sulphate into the sac was used as definitive treatment for 106 patients with hydroceles and epididymal cysts. 83 of these (78%) were cured by a single treatment. Complications were uncommon and mainly consisted of mild pain and swelling.

Adult↗
PubMed1990

The association of waist hip ratio and angiographically determined coronary artery disease.

Body fat distribution as measured by the ratio of waist circumference to hip circumference (WHR) is now accepted as an important risk factor for a number of diseases. This study evaluated the association of WHR and coronary artery disease (CAD). Measurements included the subjects' height, weight, waist girth, hip girth, significant CAD on coronary angiography, and cholesterol levels. A history of myocardial infarction, angina, diabetes or hypertension was obtained from subject interviews. The subjects were analyzed in two age groups: younger than age 60 (88 men and 39 women) and age 60 or older (85 men and 63 women). For older women the relative odds of CAD comparing women at the 75th percentile of WHR to women at the 25th percentile was 3.67 (P = 0.003), with a 95 percent confidence interval of 1.57-8.57. The relative odds was reduced to 2.80 after adjusting for all other risk factors. WHR was significantly associated with angiographic evidence of CAD in all women combined after adjusting for age (P = 0.0004), but it was not significantly associated with CAD in younger women or in men. The results suggest that in older women the risk of CAD increases with a greater percentage of body fat in the abdomen.

Adipose Tissue↗
PubMed1989

Usefulness of nicardipine as monotherapy for chronic, stable angina.

Using a double-blind, Latin square protocol designed to detect dose response, nicardipine hydrochloride, a new calcium antagonist, was studied as monotherapy for stable exertional angina. Eighty-one patients were enrolled in the trial and 62 patients were included in greater than or equal to 1 primary efficacy analyses. Patients received 1 to 2 weeks of placebo run-in, then 5 weeks of treatment with placebo and with 10, 20 and 30 mg of nicardipine given 3 times daily. Patients completed symptom diaries, were monitored with 24-hour electrocardiographic Holter monitors and underwent serial exercise treadmill tests. By 1 hour, 10, 20 and 30 mg of nicardipine administered 3 times daily produced statistically significant, dose-related improvements in all key exercise parameters, which persisted at the 4-hour evaluation. The systolic blood pressure at rest and during exercise decreased, but the pulse slightly increased. The peak rate-pressure product was unchanged. The side effects were not severe. Nicardipine hydrochloride is an effective, well-tolerated medication for the treatment of stable exertional angina, and is a good alternative to currently available calcium antagonists.

Adult↗
PubMed1989

Nicardipine and hydrochlorothiazide in essential hypertension.

Nicardipine is an investigational dihydropyridine calcium channel blocking agent. One hundred fifty-one patients with hypertension received either 30 mg nicardipine t.i.d. or 25 mg hydrochlorothiazide b.i.d. in a double-blind, randomized, multicenter trial. After 4 weeks of therapy and at the end of the dosing interval, nicardipine reduced arterial pressure by 10/6 mm Hg and 12/6 mm Hg in the supine and standing positions, respectively (all p less than 0.01). In the hydrochlorothiazide group, the reductions were 12/6 mm Hg and 14/6 mm Hg, respectively (all p less than 0.01). The maximum reduction in blood pressure of 16/14 mm Hg supine and 20/15 mm Hg standing occurred within 1 hour after administration of nicardipine. The mean reduction in the hydrochlorothiazide group after 1 hour was 14/11 mm Hg supine and 16/12 mm Hg standing. Neither drug affected autonomic reflexes associated with maximum exercise. Nicardipine increased urinary sodium excretion during the 4-hour period after the first dose. Adverse effects of nicardipine were primarily extensions of its vasodilator effect and included flushing, headache, and edema.

Adult↗
PubMed1989

Acquired immunodeficiency syndrome.

A report entitled "Kaposi's Sarcoma and Pneumocytosis Carinii Pneumonia among homosexual men in New York City and California" in the MMWR in July 1981 alerted the world to the appearance of a completely new disease. The opportunistic infections and cancers occurring in these patients had previously only been seen in patients who were immunosuppressed. Homosexual men were the first as a major risk group to be identified. Others quickly followed. The pattern of occurrence clearly indicated an infectious agent as the likely cause, and within two years the virus had been identified in Europe and the USA. In Europe it was named Lymphadenopathy Associated Virus (LAV) by Montagnier its discover, and in the USA, Human T cell Lymphotrophic Virus III (HTLV III). It is now known as Human Immunodeficiency Virus (HIV).

Acquired Immunodeficiency Syndrome↗
PubMed1988

Vanadate down-regulates cell surface insulin and growth hormone receptors and inhibits insulin receptor degradation in cultured human lymphocytes.

Insulin is able to down-regulate its specific cell surface receptor in cultured human lymphocytes. The effect of vanadate, a known insulinomimetic agent, was examined to determine whether it could mimic insulin to down-regulate the insulin receptor. Exposure of cultured human lymphocytes (IM-9) to vanadate (0-200 microM) resulted in a time- and dose-dependent decrease in cell surface insulin receptors to 60% of control, while insulin (100 nM) down-regulated to 40%. The vanadate effect, in contrast to the rapid effect of insulin, was slow to develop (4-6 h). Surface receptor recovery after 18 h exposure was rapid after vanadate removal (20 min), but it required hours after insulin suggesting the presence of an intracellular (cryptic) pool of receptors after vanadate treatment. Insulin binding to Triton X-100-solubilized whole cells after 18 h treatment revealed that total cell receptors had decreased to 50% of control after insulin but increased to 120 and 189% of control after 100 and 200 microM vanadate, respectively. Furthermore, vanadate inhibited the insulin-mediated loss of total cell receptors from 50 to 28%. Removal of cell surface receptors by trypsin before cell solubilization revealed that 100 microM vanadate increased insulin binding to 321% of control indicating an accumulation of intracellular receptors. Labeling of cell surface proteins with Na125I and lactoperoxidase followed by immunoprecipitation of solubilized receptors with anti-receptor antibody after incubation for various times up to 20 h and quantitation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that, while insulin shortened t1/2 from 7.3 to 5.3 h, vanadate prolonged receptor t1/2 to 14 h. No effect of vanadate was detected on insulin receptor tyrosine kinase activity with up to 4 h incubation at the vanadate concentrations used in this study. Furthermore, human growth hormone surface receptors were similarly down-regulated by vanadate. We conclude that 1) vanadate has an apparent insulin-like effect to down-regulate cell surface insulin receptors in cultured human lymphocytes; 2) in contrast to insulin-induced down-regulation which is associated with receptor degradation vanadate causes an accumulation of intracellular (cryptic) receptors and inhibits insulin receptor degradation; and 3) these effects of vanadate may be exerted on other cell surface receptors.

Binding, Competitive↗