Mechanistic studies on C-19 demethylation in oestrogen biosynthesis

Mechanistic aspects of the biosynthesis of oestrogen have been studied with a microsomal preparation from full-term human placenta. The overall transformation, termed the aromatization process, involves three steps using O₂ and NADPH, in which the C-19 methyl group of an androgen is oxidised to formic acid with concomitant production of the aromatic ring of oestrogen: [Formula: see text] To study the mechanism of this process in terms of the involvement of the oxygen atoms, a number of labelled precursors were synthesized. Notable amongst these were 19-hydroxy-4-androstene-3,17-dione (II) and 19-oxo-4-androstene-3,17-dione (IV) in which the C-19 was labelled with ²H in addition to ¹⁸O. In order to follow the fate of the labelled atoms at C-19 of (II) and (IV) during the aromatization, the formic acid released from C-19 was benzylated and analysed by mass spectrometry. Experimental procedures were devised to minimize the exchange of oxygen atoms in substrates and product with oxygens of the medium. In the conversion of the 19-[¹⁸O] compounds of types (II) and (IV) into 3-hydroxy-1,3,5-(10)-oestratriene-17-one (V, oestrone), it was found that the formic acid from C-19 retained the original substrate oxygen. When the equivalent ¹⁶O substrates were aromatized under ¹⁸O₂, the formic acid from both substrates contained one atom of ¹⁸O. It is argued that in the conversion of the 19-hydroxy compound (II) into the 19-oxo compound (IV), the C-19 oxygen of the former remains intact and that one atom of oxygen from O₂ is incorporated into formic acid during the conversion of the 19-oxo compound (IV) into oestrogen. This conclusion was further substantiated by demonstrating that in the aromatization of 4-androstene-3,17-dione (I), both the oxygen atoms in the formic acid originated from molecular oxygen. 10β-Hydroxy-4-oestrene-3,17-dione formate, a possible intermediate in the aromatization, was synthesized and shown not to be converted into oestrogen. In the light of the cumulative evidence available to date, stereochemical aspects of the conversion of the 19-hydroxy compound (II) into the 19-oxo compound (IV), and mechanistic features of the C-10–C-19 bond cleavage step during the conversion of the 19-oxo compound (IV) into oestrogen are discussed.     

Glycerol Phosphate Dehydrogenase, Glucose-6-Phosphate Dehydrogenase, and Lactate Dehydrogenase: Activities in Oligodendrocytes, Neurons, Astrocytes, and Myelin Isolated from Developing Rat Brains

Abstract: Glycerol phosphate dehydrogenase (GPDH), glucose-6-phosphate dehydrogenase (G6PDH), and lactate dehydrogenase (LDH) activities were determined in Oligodendrocytes, neurons, and astrocytes isolated from the brains of developing rats. The activity of each enzyme was significantly lower in both neurons and astrocytes than in Oligodendrocytes. The GPDH activity in Oligodendrocytes increased more than 4-fold during development, and at 120 days cells of this type had 1.4-fold the specific activity of forebrain homogenates. The G6PDH activities in Oligodendrocytes from 10-day-old rats were 1.4-fold the activities in the forebrain homogenates. The activities of this enzyme in Oligodendrocytes were progressively lower at later ages, such that at 120 days the cells had 0.8 times the specific activities of homogenates. The Oligodendrocytes had 0.6 times the homogenate activities of LDH at 10 days, and this ratio had decreased to 0.2 by 120 days. These enzymes were also measured in myelin isolated from 20-, 60-, and 120-day-old rats. By 120 days the specific activities of G6PDH and LDH in myelin were <8% of the respective activities in homogenates. The GPDH activity in myelin was, however, at least 20% the specific activity in the homogenates, even in the oldest animals. It is proposed that LDH could be used as a marker for oligodendroglial cytoplasm in subfractions of myelin and in myelin-related membrane vesicles.     

Properties of Bovine Oligodendroglia Isolated by a New Procedure Using Physiologic Conditions

A new class of procedures, previously shown to permit the isolation of pure oligodendroglia from whole rat cerebrum, has been applied with equal or greater success for the bulk isolation of this cell type from bovine white matter. Thus, the generality of this approach has been demonstrated. The bovine preparations have a purity of greater than 90% intact, phase-bright oligodendroglia and are obtained in a yield of 8 x 10(6) cells per gram of white matter. Within 1 day it is possible to obtain a preparation containing 60 mg of protein from a single cell type. These cells show a higher degree of ultrastructural preservation of all cytoplasmic constituents than previously obtained. The values for protein (33 pg/cell), DNA (5.4 pg/cell), and lipid (5-6 pg/cell) are very similar to those obtained with an earlier procedure. The cell lipids are rich in galactolipid, which comprises 20% of the total. The activity of the "myelin-specific" enzyme, 2',3'-cyclic nucleotide 3'-phosphohydrolase (EC 3.1.4.37), is 4.7 mumol/min/mg protein, similar to that obtained previously for isolated oligodendroglia and about 25-40% of that found in myelin. The activity of 5'-nucleotidase (EC 3.1.3.5) in the cells is about 10% of that in myelin or white matter.     

The effect of exhaustive exercise on expired pentane as a measure of in vivo lipid peroxidation in the rat

The concentrations of pentane and ethane in the expired breath of swimming rats were used to determine the possible occurrence of lipid peroxidation caused by strenuous exercise. Rats swum to exhaustion produced significantly more pentane but not ethane than did rats at rest. Rats fed a vitamin E-deficient diet produced slightly more pentane following exhaustive exercise than they did while at rest, but this increase was not statistically significant. Rats were also swum for prescribed lengths of time. Only rats that had swum for 20 or 40 min had significantly elevated concentrations of pentane in the breath. Rats swum for 10 or 30 min had elevated concentrations of pentane in the breath, but these increases were not statistically significant. These results suggest that lipid peroxidation is moderately increased following exhaustive exercise.     

Evolution of chromosomal abnormalities in sequential cytogenetic studies of ataxia telangiectasia

Summary Sequential cytogenetic studies of four patients with ataxia telangiectasia showed the progressive development of lymphocyte clones, each marked with a rearranged chromosome 14. Initial studies had shown random chromosomal breaks and rearrangements. Later studies in all patients showed nonrandom rearrangement of chromosome 14 with a breakpoint at 14q12 and with the distal segment translocated to either chromosome 14 or 7. The proportion of circulating lymphocytes carrying the marker tended to increase with time, accounting for the majority of the lymphocytes eventually in one case. The marked lymphocyte clones evolved further, as a result of loss of the small centric portions of the rearranged chromosome 14 (14pter14q12).Perhaps the abnormal clones in ataxia telangiectasia escape immunologic surveillance and flourish in an immunologically impaired environment. Subsequent to the loss of the centric portion of the rearranged chromosome 14, the cells may acquire additional capabilities that enhance malignant transformation.     

Evaluation of Nosema locustae (Microsporida) as a control agent of grasshopper populations in Saskatchewan

A quantitative study of the effect of a microsporidan, Nosema locustae, as a control agent against grasshopper populations in Saskatchewan, Canada, revealed that 50% of the populations of Melanoplus sanguinipes, M. packardii, and Camnula pellucida were infected between 4 and 5 weeks (400 to 424 degree-days) after application of the pathogen. Maxima of 95–100% infection were evident between 9 and 12 weeks (600–700 degree-days) after application. The percentage reduction in surviving populations of M. sanguinipes, i.e., those that did not die from natural causes, reached about 20% by the 4th week (400 degree-days) after inoculation, about 50% by the 9th week (600 + degree-days), and a maximum of about 60% by the 12th week (700 degree-days). An exponential relationship was obtained between percentage reduction and percentage infection in all three species. However, a similar percentage infection resulted in different percentage reduction in the populations. Results also revealed that the rate of reduction in populations reached its peak by about 40–80 degree-days before the maximum rate of infection was attained for each species. Egg production in the two Melanoplus species in treated plots was significantly lower (P < 0.05) than in the control plot.     

Glycogenosis in the Dog

Four cases of a generalized form of glycogenosis occurring in German Shepherd dogs, all females, are described. Symptoms could be noticed as early as the age of two months and progressed slowly for months. They appeared as dizziness, muscular weakness, and in two of the cases as poor nutritional state. The abdomen became gradually distended. The main lesion seen at postmortem was a greatly increased liver size with some moderate liver fibrosis. Heavy deposits of a granular substance behaving as glycogen in histochemical tests and at electron microscopy were found in the hepatic cells, muscle fibres of the heart, skeletal and smooth muscles, and in nerve and glial cells of the central nervous system. The substance was lying freely dispersed in the cell cytoplasm without any indication of lysosomal storage. The disease of dogs does not seem to be fully comparable with any of the types observed in man, but is probably much related to Type III or Cori's Disease. Structure analysis of the deposits and enzyme investigations have been done and are published (Čeh et al. 1976).     

Effect of Polychlorinated Biphenyl Formulations on the Growth of Estuarine Bacteria

Polychlorinated biphenyl formulations inhibited the growth of certain estuarine bacteria. The sensitive strains, although exhibiting some similar physiological characteristics, contained both gram-positive and gram-negative bacteria.