Development of sensitive esterase assays based on alpha-cyano-containing esters.

A novel approach is reported for the development of fluorogenic esterase reporters using alpha-cyano-containing esters as substrates. After ester hydrolysis, the released alcohol, a cyanohydrin, rapidly eliminates HCN to yield the corresponding aldehyde resulting in strong fluorescence. The pi conjugation of the resulting aldehyde also greatly enhances UV absorption and red shifts fluorescence emission relative to a corresponding alcohol or phenol. Two substrates, R/S-acetic acid cyano-(6-methoxynaphthalen-2-yl)-methyl ester (compound I) and trans/cis-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropanecarboxylic acid R/S-cyano-(6-methoxynaphthalen-2-yl)-methyl ester (compound II), were synthesized and evaluated as substrates. Such alpha-cyano substrates possess very low background fluorescence and are more stable under enzyme assay conditions than phenolic substrates due to the aliphatic cyano group. The higher molar absorbtivity and quantum yield of the aldehyde, along with its larger Stokes' shift combined with the increased stability and lower background signal of the cyanohydrin substrate, increases the utility and sensitivity of the resulting assays over current methods. Moreover, compound II showed high selectivity to pyrethroid-cleaving esterases and may provide a direct tool to monitor pyrethroid resistance in insects.     

Nerve growth factor and neuronal cell death

The regulation of neuronal cell death by the neuronotrophic factor, nerve growth factor (NGF), has been described during neural development and following injury to the nervous system. Also, reduced NGF activity has been reported for the aged NGF-responsive neurons of the sympathetic nervous system and cholinergic regions of the central nervous system (CNS) in aged rodents and man. Although there is some knowledge of the molecular structure of the NGF and its receptor, less is known as to the mechanism of action of NGF. Here, a possible role for NGF in the regulation of oxidant--antioxidant balance is discussed as part of a molecular explanation for the known effects of NGF on neuronal survival during development, after injury, and in the aged CNS.     

Effects of selection criteria on yield stability in chickpea (Cicer arietinum L.)

Summary Selection in the F₃ generation for seed yield, fruiting branches/plant, effective pods/plant, and seed index (100-seed weight) was carried out in two chickpea crosses. Sixty F₅ lines (15 lines/selection criterion) along with check variety were evaluated for seed yield in three distinct environments. The effects of selection criteria on yield stability was examined using linear regression approach and genotype-grouping technique. There were no differences between selection criteria for linear yield responses of F₅ lines to different environments. Within all four selection criteria the lines showed similar linear responses. The non-linear component was relatively higher for lines selected for effective pods and seed index than lines selected for yield and fruiting branches. On the basis of mean yield and coefficient of variation across environments, the seed index was the least effective selection criterion for developing high yielding and stable lines. When the results of stability parameters and genotype-grouping technique were considered together, selection for yield and fruiting branches was highly effective for isolating stable and high yielding lines.     

Serial studies of autologous antibody reactivity to squamous cell carcinoma of the head and neck

Summary In previous studies we evaluated the incidence and specificity of autologous antibody reactivity against squamous cell carcinoma of the head and neck (SCCHN). We were able to demonstrate that autologous antibody reactivity is present in native sera but was usually of too low a titer to allow further analysis. Dissociation of immune complexes by acidification and ultrafiltration of serum augmented autologous antibody reactivity in nine out of nine autologous systems tested. Native antibody and antibody derived from immune complexes produced by the host and reactive with autologous tumor cells may be directed against physiologically relevant antigens. Therefore, correlations of antibody titers with clinical course may provide insight into the nature of the host response to cancer. In the present analysis, serological studies of six patients with SCCHN were performed with serum samples obtained over many months. Results of serial serological assays were correlated to tumor progression and clinical course. Fluctuations in autologous antibody reactivity were noted over time. In four cases, rises in autologous antibody titers preceded the clinical diagnosis of recurrence by several months. Drops in autologous antibody reactivity were noted in two cases following surgery or radiation therapy. In two cases of long-term survivors, no correlation between antibody reactivity and clinical course was noted. Specificity analysis of the six autologous systems demonstrated reactivity against autologous and allogeneic SCCHN as well as melanoma cell lines. These sera did not react with glioma, neuroblastoma, renal cell, breast, bladder and colon carcinoma cell lines nor with fetal calf serum, pooled lymphocytes, red blood cells and platelets. Autologous serial serological studies may provide a means by which to evaluate the host/tumor relationship in patients with SCCHN.     

Onychomycosis caused by Scopulariopsis brumptii

Scopulariopsis brumptii was isolated from nail lesions in left hand of a 42 year-old-farmer. The direct microscopic examination of the nail samples revealed light brown, septate, branched fungal hyphae along with thick-walled spherical cells. The histopathological examination showed involvement of internal phase of the nail plate. Amongst the antimycotics tested against S. brumptii In vitro oxiconazole was found to be the most active with MIC value of 10 g/ml^–1. This report documents the first instance of onychomycosis caused by S. brumptii.     

Acclimatory responses in enzymatic activity and metabolite concentrations in the tropical scorpion Heterometrus fulvipes

1. 1.|Changes in tissue metabolite concentrations and enzyme activities in the pedipalpal (PM) and heart (HM) muscles of the tropical scorpion Heterometrus fulvipes show that the metabolism in PM and HM is fundamentally reorganized following low (18°C) and high (38°C) temperature acclimation.

2. 2.|Changes in metabolite concentrations show that metabolite biosynthesis showed increases after cold acclimation but decreases after warm acclimation.

3. 3.|Similarly, changes in enzyme activities show a preponderance of glycolysis and HMP shunt activity after cold acclimation, while after warm acclimation glycogenolysis, oxidative metabolism and gluconeogenesis predominated.

4. 4.|Higher metabolite concentrations and enzyme activities both before and after thermal acclimation in HM reflect its greater compensatory abilities.

Acute and chronic effects of insulin on lactate dehydrogenase (LDH) activity in tissues of albino rat (Wistar strain)

Male albino rats were treated with insulin for one week (acute) and four weeks (chronic). The lactate dehydrogenase (LDH) activity, lactate and pyruvate levels were estimated in the tissues of experimental and control animals. LDH activity decreased in all the tissues of acute- and chronic treated animals whereas the lactate content is elevated. Pyruvate content also showed increment except in heart and pancreas with reference to acute treatment where it is decreased. The hyperinsulinaemia effect in relation to lacticacidaemia and its influence on energy demand and ammonia secretion is discussed.     

Ultrastructural localization of an extracellular protease in Pseudomonas fragi by using the peroxidase-antiperoxidase reaction.

An extracellular protease, which previously has been found to correlate with the appearance of bleblike evaginations on the cell wall of Pseudomonas fragi ATCC 4973, was purified 38-fold by ammonium sulfate precipitation and Sephadex chromatography to yield a single band by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Polyclonal rabbit antiserum raised against the purified enzyme had an enzyme-linked immunosorbent assay titer of 4 X 10(7). The peroxidase antiperoxidase method was used to localize the neutral protease in P. fragi at the ultrastructural level. Electron microscopy of cell sections of this organism revealed that high concentrations of positive immunoperoxidase reaction product were located near the cell wall, whereas control sections stained with preimmune or heterologous serum did not show similar deposits to be present. These results are consistent with the hypothesis that blebs appearing on P. fragi contain high concentrations of neutral protease.     

A determinant of polyomavirus virulence enhances virus growth in cells of renal origin.

We have identified a strain of polyomavirus, Py(L), which is unusual in causing acute morbidity and early death after inoculation of newborn mice. We determined that these animals died of kidney failure associated with extensive, virus-mediated destruction of renal tissue. Interestingly, the Py(L) strain infects baby mouse kidney cell cultures more efficiently than do other strains.     

Ultrastructural localization of an extracellular protease in Pseudomonas fragi by using the peroxidase-antiperoxidase reaction

An extracellular protease, which previously has been found to correlate with the appearance of bleblike evaginations on the cell wall of Pseudomonas fragi ATCC 4973, was purified 38-fold by ammonium sulfate precipitation and Sephadex chromatography to yield a single band by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Polyclonal rabbit antiserum raised against the purified enzyme had an enzyme-linked immunosorbent assay titer of 4 X 10(7). The peroxidase antiperoxidase method was used to localize the neutral protease in P. fragi at the ultrastructural level. Electron microscopy of cell sections of this organism revealed that high concentrations of positive immunoperoxidase reaction product were located near the cell wall, whereas control sections stained with preimmune or heterologous serum did not show similar deposits to be present. These results are consistent with the hypothesis that blebs appearing on P. fragi contain high concentrations of neutral protease.