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Biomedical subjects

L Mo

Publications and source records attributed to L Mo.

10 recordsLinked to original sources
PubMed1999

Comparison of amphibian and human ClC-5: similarity of functional properties and inhibition by external pH.

Loss of function mutations of the renal chloride channel, ClC-5, have been implicated in Dent's disease, a genetic disorder characterized by low weight proteinuria, hypercalciuria, nephrolithasis and, in some cases, eventual renal failure. Recently, our laboratory used an RT-PCR/RACE cloning strategy to isolate an amphibian cDNA from the renal epithelial cell line A6 that had high homology to human ClC-5. We now report a full-length native ClC-5 clone (xClC-5, containing 5' and 3' untranslated regions) isolated by screening a cDNA library from A6 cells that was successfully expressed in Xenopus oocytes. In addition, we compared the properties of xClC-5 and hClC-5 using isogenic constructs of xClC-5 and hClC-5 consisting of the open reading frame subcloned into an optimized Xenopus expression vector. Expression of the full-length "native" xClC-5 clone resulted in large, strongly rectifying, outward currents that were not significantly affected by the chloride channel blockers DIDS, DPC, and 9AC. The anion conductivity sequence was NO-3 > Cl- = I- > HCO-3 >> glutamate for xClC-5 and NO-3 > Cl- > HCO-3 > I- >> glutamate for hClC-5. Reduction of the extracellular pH (pHo) from 7.5 to 5.7 inhibited outward ClC-5 currents by 27 +/- 9% for xClC-5 and 39 +/- 7% for hClC-5. The results indicate that amphibian and mammalian ClC-5 have highly similar functional properties. Unlike hClC-5 and most other ClC channels, expression of xClC-5 in oocytes does not require the removal of its untranslated 5' and 3' regions. Acidic solutions inhibited both amphibian and human ClC-5 currents, opposite to the stimulatory effects of low external pH on other ClC channels, suggesting a possibly distinct regulatory mechanism for ClC-5 channels.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1997

Osmotic regulation of Na+ transport across A6 epithelium: interactions with prostaglandin E2 and cyclic AMP.

Previous work from this laboratory has shown that apical membrane sodium channel activity is stimulated by serosal hyposmotic solutions (Wills, Millinoff & Crowe, 1991). In the present study, we determined whether this stimulation of sodium transport is additive with the actions of prostaglandin E2 (PGE2) or cyclic AMP (cAMP). Addition of exogenous PGE2 (100 nM; serosal bath) to isosmotic solutions led to large increases in the amiloride-sensitive short-circuit current (Isc) and transepithelial conductance (Gt), whereas no significant effects of PGE2 were observed in hyposmotic serosal solutions. Subsequent addition of mucosal amiloride reduced Isc by approximately 95% and Gt by approximately 60%. Inhibition of endogenous PGE2 production by blockers of phospholipase A2 activity (quinacrine or 3[4-octadecyl]-benzoylacrylic acid; OBBA), or inhibition of cyclooxygenase activity by indomethacin reduced the stimulation of Isc and Gt by hyposmotic solutions. Addition of forskolin (FSK) or 3-Isobutyl-1-methylxanthine (IBMX) also resulted in approximately twofold increases in the amiloride-sensitive Isc and Gt and abolished the effects of subsequent hyposmotic challenge. The effects of forskolin, PGE2, and hyposmotic challenge were diminished by pretreatment with H89, a protein kinase A (PKA) inhibitor. We conclude that osmotic regulation of sodium channel activity interacts with multiple intracellular signaling pathways, specifically the arachidonic acid metabolic pathway and the cAMP/PKA intracellular messenger cascade.

1-Methyl-3-isobutylxanthine↗
PubMed1997

Generation and analysis of an IgG anti-platelet autoantibody reveals unusual molecular features.

Although serum transfer studies implicate IgG anti-platelet autoantibodies in the premature platelet destruction of idiopathic thrombocytopenic purpura (ITP), many characteristics of these putative pathogenic autoantibodies remain unclear. The inability to obtain relevant monoclonal autoantibodies from patients has prevented their molecular, genetic and functional studies as a homogenous population. We have generated a monoclonal IgG anti-platelet alpha IIb beta 3 autoantibody (termed G1) from an ITP patient. G1 binds human platelets (both resting and activated) and purified alpha IIb beta 3 with a Kd of 1.57 x 10(-8) M. G1 utilizes VH4 and V lambda 2 genes. The G1 VH region apparently has a 30 nucleotide insertion in its second complementarity determining region (CDR). Notably, somatic CDR insertion in the VH region has been observed only in one IgG rheumatoid factor, and not in any characterized polyreactive human autoantibodies reported in the literature. Combined these data suggest G1 may be a disease-relevant autoantibody. Further generation and study of monoclonal IgG anti-platelet antibodies are warranted to determine the significance of such unusual autoantibodies in the immunopathogenesis of chronic ITP.

Adult↗
PubMed1996

The frequency of homozygous deletion of a developmentally regulated Vh gene (Humhv3005) is increased in patients with chronic idiopathic thrombocytopenic purpura.

Little is known of the genetic factors that may contribute to the development of chronic idiopathic thrombocytopenic purpura (cITP). We have previously shown that a developmentally regulated Vh gene (Humhv3005) is absent in 10/41 (24%) of patients with systemic lupus erythematosus while it is absent in only 7/88 (8%) of normal controls. This finding suggests that a homozygous deletion of an Ig variable (V) gene may alter the immune system and thus predispose the host to an autoimmune disorder. We have analyzed the same gene in 44 patients with cITP and found that Humhv3005 and like genes were absent in a higher percentage of patients (14 of 44, 31.8%) than they were absent in either normals (7/88, 8%, p = 0.002) or thrombocytopenic patients without cITP (6/53, 11.3%, p = 0.042); the hv3005 deletion frequency in the latter group did not differ from that in normals (P = 0.74). These data suggest that deletions of Humhv3005 and/or highly homologous Vh genes may predispose individuals to the development of cITP, and may contribute toward production of pathogenic antiplatelet antibodies.

Chronic Disease↗
PubMed1996

The reverse transcriptase assay: problems and practical solutions.

The detection of retroviruses has become critical for addressing the safety concerns associated with biologically derived products, including those derived from blood and cell line substrates, and gene therapy based systems. Most, if not all, retroviruses encode a unique enzyme called reverse transcriptase whose RNA-dependent DNA polymerase activity can be used as a marker for detecting retroviral contamination in test material. In this presentation we document some practical concerns when using the reverse transcriptase assay for detection of retroviruses. We also illustrate important aspects in the assay design by presentation of results that needed supplemental testing to enable accurate assessment of retroviral contamination.

Animals↗
PubMed1990

Development of methods to analyse transcranial Doppler ultrasound signals recorded in microgravity.

During space flights, several clinical syndromes may be the result of changes in cerebral circulation. The purpose of the paper is to describe the development and initial evaluation of a system for recording, processing and displaying transcranial Doppler ultrasound (TCD) waveforms from the middle cerebral artery (MCA) in microgravity. Volunteers were repeatedly subjected to 15-20 s intervals of microgravity ('near zero gravity') during flights on the KC-135 military aircraft. Continuous TCD recordings from the MCA were stored on magnetic tape. The paper describes the system that was developed to digitise the Doppler ultrasound data and markers that corresponded to the various levels of microgravity, obtain the maximum and mean Doppler waveforms, identify the waveforms and quantify them. The results demonstrate the feasibility of making TCD recordings in a microgravity environment and illustrate excellent performance of the system and its ease of operation. Quantitative waveform analysis of the recordings from the first subject studied in the supine position showed statistically significant changes in MCA velocity waveforms during microgravity.

Cerebral Arteries↗
PubMedDate not supplied

Comparison of tomographic and planar radionuclide ventriculography in the assessment of regional left ventricular function in patients with left ventricular aneurysm before and after surgery.

METHODS AND RESULTS: To compare tomographic and planar radionuclide ventriculography (RNVG) in assessing regional left ventricular (LV) function and predicting improvement in LV ejection fraction (LVEF) after operation in patients with LV aneurysm, 18 patients with aneurysm underwent both tomography and planar RNVG 1 month before and 3 weeks to 6 months after aneurysmectomy and coronary artery bypass grafting. All patients also underwent preoperative contrast angiography at catheterization. The percent shortening of the apical, anterior, lateral, inferior, and basal segments was calculated from tomographic long-axis and short-axis slices and corresponding planar images (anterior and 30- and 70-degree left anterior oblique views). No significant differences in anterior, apical, and lateral percent shortening were apparent before aneurysmectomy between tomographic and planar studies. However, preoperative basal percent shortening was 47% +/- 13% from tomographic and 28% +/- 14% from planar images (p < 0.001). Preoperative tomography generally agreed better with contrast angiographic results than did planar imaging. After aneurysmectomy, basal function improved to 57% +/- 12% (p < 0.01) by tomography. For all patients, LVEF increased from 29% +/- 8% before to 38% +/- 9% (p < 0.01) after aneurysmectomy. However, the greatest improvement (31% +/- 11% to 41% +/- 9%; p < 0.01) was in the 15 patients with greater than 30% basal shortening by tomography before aneurysmectomy; in contrast, no change of LVEF occurred in the three patients with lesser preoperative basal percent shortening. Moreover, greater than 30% basal percent shortening by tomography before aneurysmectomy identified the group most likely to have an increase in LVEF of 5% or more from before to after aneurysmectomy. Prediction of postoperative results was not possible from preoperative planar data. Thus in patients with LV aneurysm, tomographic RNVG appears to provide information that is different and more accurately predictive of results after aneurysmectomy than that available from planar imaging.

Adult↗
PubMedDate not supplied

Drug allergy: identification and characterization of IgE-reactivities to aspirin and related compounds.

Twenty-seven patients with aspirin (ASA) sensitivity were studied. 14 patients had naso-ocular-bronchial reactions after taking ASA while others had cutaneous and gastrointestinal reactions. The oral challenges with salicylic acid (SA), O-methylsalicylic acid (OMSA), ASA, and the determination of IgE antibodies specific to salicyloyl, O-methylsalicyloyl, acetylsalicyloyl using correspondent disks by RAST, RAST inhibition and RAST crossinhibition assays were performed. The findings suggest that OMSA seems to be the main offender responsible for cutaneous and gastrointestinal reactions, whereas ASA is responsible for naso-ocular-bronchial reactions. The clinical crossreactions between ASA and ASA-like drugs (nonsteroidal anti-inflammatory drugs and pyrazolone drugs) are probably due to "inborn errors of metabolism". The results indicate that genetic factors, mast-cell heterogeneity, and the interindividual variability in drug metabolism, combined with immunological background should be considered as underlying mechanisms.

Adolescent↗