alpha4beta3delta GABA(A) receptors characterized by fluorescence resonance energy transfer-derived measurements of membrane potential.

Selective modulators of gamma-aminobutyric acid, type A (GABA(A)) receptors containing alpha(4) subunits may provide new treatments for epilepsy and premenstrual syndrome. Using mouse L(-tk) cells, we stably expressed the native GABA(A) receptor subunit combinations alpha(3)beta(3)gamma(2,) alpha(4)beta(3)gamma(2), and, for the first time, alpha(4)beta(3)delta and characterized their properties using a novel fluorescence resonance energy transfer assay of GABA-evoked depolarizations. GABA evoked concentration-dependent decreases in fluorescence resonance energy transfer that were blocked by GABA(A) receptor antagonists and, for alpha(3)beta(3)gamma(2) and alpha(4)beta(3)gamma(2) receptors, modulated by benzodiazepines with the expected subtype specificity.

Effect of alpha subunit on allosteric modulation of ion channel function in stably expressed human recombinant gamma-aminobutyric acid(A) receptors determined using (36)Cl ion flux.

Inhibitory gamma-aminobutyric acid (GABA)(A) receptors are subject to modulation at a variety of allosteric sites, with pharmacology dependent on receptor subunit combination. The influence of different alpha subunits in combination with beta3gamma2s was examined in stably expressed human recombinant GABA(A) receptors by measuring (36)Cl influx through the ion channel pore. Muscimol and GABA exhibited similar maximal efficacy at each receptor subtype, although muscimol was more potent, with responses blocked by picrotoxin and bicuculline.

Gastric Cancer.

The best treatment for gastric cancer remains early detection and prompt surgical removal. Currently, the majority of gastric carcinomas diagnosed in western centers are late-stage tumors with poor prognoses. Endoscopic ultrasound and diagnostic laparoscopy aid our ability to accurately stage patients with gastric cancers, and may prevent unnecessary laparotomies in asymptomatic patients.

Preferential coassembly of alpha4 and delta subunits of the gamma-aminobutyric acidA receptor in rat thalamus.

Pharmacological study of rat thalamic gamma-aminobutyric acidA (GABAA) receptors revealed the presence of two distinct populations, namely, diazepam-sensitive and diazepam-insensitive [3H]Ro15-4513 binding sites accounting for 94 +/- 2% (1339 +/- 253 fmol/mg protein) and 6 +/- 2% (90 +/- 44 fmol/mg protein) of total sites, respectively. Thalamic diazepam-insensitive sites exhibited a pharmacology that was distinct from diazepam-sensitive sites but comparable to that of the alpha4beta3gamma2 subtype of the GABAA receptor stably expressed in L(tk-) cells. Immunoprecipitation experiments with a specific anti-alpha4-antiserum immunoprecipitated 20 and 7% of total thalamic [3H]muscimol and [3H]Ro15-4513 sites, respectively.

In vitro modulation of AL-amyloid formation by human mesangial cells exposed to amyloidogenic light chains.

We have shown in vitro AL-amyloid formation by human mesangial cells (HMCs). AL-amyloid formation may require lysosomal processing of the light chains (LCs) by HMCs for amyloidogenesis to occur. Chloroquine inhibits lysosomal activity.

Operative treatment of tertiary hyperparathyroidism: a single-center experience.

Objective: To review the experience with the operative treatment of tertiary hyperparathyroidism (TH) from a single renal transplant center.

Summary Background Data: Most patients with chronic renal failure show evidence of secondary hyperparathyroidism by the time maintenance hemodialysis begins. Persistent secondary hyperparathyroidism (i.

Molecular conjugate-mediated gene transfer into isolated human and transplanted rat kidneys.

Technically, renal transplantation has been feasible for over four decades. However, immunological injury to the transplanted kidney continues to be the leading cause of graft loss. While current immunosuppressive protocols yield a 1-year graft survival of > 90%, the trade off is increased risks from nephrotoxicity to manifestations of long-term immunosuppression.

Adenoviral vector infection of the human exocrine pancreas.

Objective: To determine if a viable cadaveric pancreas might be used to study viral transfection efficacy in a manner precisely mimicking in vivo human studies.

Design: Ex vivo gene transfer to an intact human pancreatic duct.

Setting: Molecular biology laboratory and organ procurement center.

Immunolocalization of acidic fibroblast growth factor and receptors in the tubulointerstitial compartment of chronically rejected human renal allografts.

Tubular damage and loss associated with interstitial inflammation and fibrosis may be the most important determinants in chronic renal allograft rejection. To elucidate potential pathophysiologic mechanisms associated with tubulointerstitial lesions, we examined the expression of a fibrogenic cytokine, acidic fibroblast growth factor (FGF-1) and its high-affinity receptors, in both relevant renal transplant controls (n=5) and tissue from patients (n=19) who underwent nephrectomy after graft loss, secondary to chronic rejection. In situ hybridization and immunohistochemical analyses demonstrated minimal expression of FGF-1 mRNA and protein in the tubulointerstitial compartment of the normal human kidney.

Nitration and inactivation of manganese superoxide dismutase in chronic rejection of human renal allografts.

Inflammatory processes in chronic rejection remain a serious clinical problem in organ transplantation. Activated cellular infiltrate produces high levels of both superoxide and nitric oxide. These reactive oxygen species interact to form peroxynitrite, a potent oxidant that can modify proteins to form 3-nitrotyrosine.

Economics of pancreatoduodenectomy in the elderly.

Background: Managed care and the increasing percentage of surgical procedures performed in the elderly have renewed the focus on hospital charges and expenditures. The objective of this study was to determine whether septuagenarians and octogenarians accrue more hospital charges or have a higher risk of morbidity and death.

Methods: We retrospectively reviewed the charges and pertinent clinical outcomes data that were available on 70 of the last 100 pancreatoduodenectomies performed at our institution (1989 to 1994).

In vivo gene transfer to the human biliary tract.

The human biliary tract offers an excellent model for gene transfer studies for a variety of diseases localized to the liver. The aim of this study was to determine if a viable liver might be employed to study viral transfection of the human biliary system in order to mimic in vivo human experiments. Using a normal human liver initially procured for transplantation, but subsequently found unsuitable, and with an intact biliary tree, the hepatic vascular supply was accessed for continuous perfusion.

Immunolocalization of FGF-1 and receptors in human renal allograft vasculopathy associated with chronic rejection.

Despite recognition of chronic vasculo-occlusive disease in solid organ transplantation, the exact pathophysiologic events resulting in neointima formation remain to be elucidated. Since acidic fibroblast growth factor (FGF-1) is an established modulator of vascular cell function, we examined the expression of this growth factor and its high affinity receptors in both relevant renal transplant controls (n = 5) and tissue from patients (n = 19) who underwent nephrectomy following graft loss secondary to chronic rejection. In situ hybridization and immunohistochemical studies demonstrated minimal vascular expression and distribution of FGF-1 and FGF high affinity receptors in the normal human kidney.

Immunolocalization of FGF-1 and receptors in glomerular lesions associated with chronic human renal allograft rejection.

Glomerular lesions are considered one of the more detrimental pathologic changes associated with chronic rejection of renal allografts. To elucidate potential pathophysiologic mechanisms associated with transplant glomerulopathy, we examined the expression of acidic fibroblast growth factor (FGF-1) and its high-affinity receptors (FGFR) in both relevant renal transplant controls (n=5) and tissue from patients (n=19) who underwent nephrectomy following graft loss secondary to chronic rejection. In situ immunohistochemical analyses demonstrated minimal staining and distribution of FGFR and FGF-1, which was localized to the mesangial matrix in glomeruli from normal human kidneys.

Differential effect of trilostane on the progestin milieu in the pregnant mare.

Trilostane, a competitive inhibitor of 3 beta-hydroxysteroid dehydrogenase, was administered intravenously to pregnant mares (n = 3) between day 277 and day 282 of gestation to determine its effect on the progestin milieu. In addition, placental tissue from mares at mid-gestation (150-300 days) (n = 4) were exposed to either deuterium-labelled pregnenolone alone or deuterium-labelled pregnenolone and trilostane to examine the effect of trilostane on placental metabolism of pregnenolone. Blood samples were collected from indwelling jugular catheters at frequent intervals for 48 h after infusion.

Molecular conjugate-mediated gene transfer into isolated human kidneys.

Advances in systemic immunosuppressive therapy for solid organ transplantation have done little to decrease the percentage of allografts that eventually will develop chronic rejection. However, one of the promises of modern molecular biology includes the ability to introduce new genetic information into mammalian hosts. The ability to deliver genes and control their expression in the adult kidney has been described in appropriate animal models.

In vitro AL-amyloid formation by rat and human mesangial cells.

AA-amyloid has been produced experimentally in animal models, allowing the study of mechanisms involved in AA-amyloidogenesis, but those involved in renal AL-amyloidogenesis have not been adequately investigated due, in part, to lack of appropriate in vitro models. Rat and human mesangial cells were grown on a human extracellular matrix (Amgel) derived from normal tissues and on coverslips in the presence of 10 microliters of amyloid enhancing factor (AEF) per milliliter of media and 10 micrograms/ml monoclonal lambda light chains (LCs) obtained from two patients with AL-amyloidosis. Two additional lambda LCs derived from the urine of patients with myeloma and tubulointerstitial renal disease were used as controls.

Pharmacological properties of recombinant human N-methyl-D-aspartate receptors comprising NR1a/NR2A and NR1a/NR2B subunit assemblies expressed in permanently transfected mouse fibroblast cells.

The pharmacological properties of two recombinant human N-methyl-D-aspartate (NMDA) receptor subtypes, comprising either NR1a/NR2A or NR1a/NR2B subunits permanently transfected into mouse L(tk-) cells, have been compared using whole-cell voltage-clamp electrophysiology. Glutamate was a full agonist at both receptors, having a modestly but statistically significant (p < 0.002) higher affinity for the NR2B- than the NR2A-containing receptor (microscopic Kd [mKd] = 0.

Quantitative comparison of the stress response of striped bass (Morone saxatilis) and hybrid striped bass (Morone saxatilis x Morone chrysops and Morone saxatilis x Morone americana).

Striped bass (Morone saxatilis) exposed to a standardized confinement stress had markedly different clinical and endocrinologic responses, compared with hybrid striped bass exposed to the same stress. Plasma cortisol concentration increased at a faster rate and appeared to reach a higher value in striped bass than in hybrid bass. Mean plasma cortisol concentration was 742 +/- 43 ng/ml in striped bass, compared with 490 +/- 37 and 531 +/- 40 ng/ml in striped bass x white perch (M americana) and striped bass x white bass (M chrysops) hybrids, respectively, after a 45-minute net confinement.

An expert system for assessing safety and security of heterogeneous public water sources.

We have implemented a diagnostic system designed to advise on the likely causes of sanitary problems with public water sources. The approach to the problem makes extensive use of rule based expert systems and multi media information (maps, data, text, expert knowledge). The rules were based on actual water survey data, with a weighting scheme designed to highlight causes of health risks approximately in reverse order of importance (where such order may be presumed to exist).

An expert system for water quality modelling.

The RAISON-micro (Regional Analysis by Intelligent System ON a micro-computer) expert system is being used to predict the effects of mine effluents on receiving waters in Ontario. The potential of this system to assist regulatory agencies and mining industries to define more acceptable effluent limits was shown in an initial study. This system has been further developed so that the expert system helps the model user choose the most appropriate model for a particular application from a hierarchy of models.

Derivation of osteoclasts from hematopoietic colony-forming cells in culture.

The osteoclast is known to be derived from the hematopoietic stem cell, but its lineage remains controversial. There is evidence that osteoclastic differentiation is induced through a contact-dependent interaction between bone marrow stromal cells and hematopoietic precursors. To analyze osteoclastic lineage, colonies were generated in semisolid medium from mouse spleen cells in the presence of Wehi-conditioned medium, interleukin-3 (IL-3), granulocyte-macrophage colony-stimulating factor (GM-CSF), or macrophage colony-stimulating factor (M-CSF) with or without erythropoietin (epo).