Steroidogenesis in the desensitized rat corpora-lutea

Earlier workers have observed that in the leydig cell desensitization brings results in addition to down regulation of receptors, in leisons in the steroidogenlc pathway. In the present study immature rats having heavily leutinized ovaries were given 50 iu hCG and the desenasitized CL removed 48h later were used. At that time no change in the 5 3MSD activity and CAMP binding activity(a measure of CAMP dependent protein kinase) was observed.Followlng desensitization however,l)a significent increase in phosphodiestrase activity,ii)a 50% reduction in total mitochondrial cholesterol level, iii)a significant reduction in its ability to utilize cholesterol or hydrolyse its ester and iv)a significant lowering(by 66%)in cholesterol side chain clean age activity(by measuring pregnanalone formed) was observed. Pregnanalone production was restored to normalcy if exogenous cholesterol was added to the mitohondrial preparation. The results suggest that luteal desensitization is due in addition to down regulation of LH receptors, to a marked reduction in available cholesterol pool in the mitochondrial compartment. The increase in phosphodiestrase activity, though probably a secondary effect,might effectively contribute to the overall reduction in the steroid out-put by increasing the catabolism of CAMP.(Aided by grants from ICMR,New Delhi and WHO, Geneva).

Hormonal induction of riboflavin carrier protein in the chicken oviduct and liver: a comparison of kinetics and modulation

Estrogen (E) induction of riboflavin carrier protein (RCP) in the chicken oviduct and liver was investigated to compare and contrast the kinetics, hormonal specificity and modulation of its elaboration in the 2 steroid-responsive tissues. During primary stimulation, continued daily E administration to immature female chicks elicited, after an initial lag, rapid growth and RCP content of the oviduct; neither progesterone (P) nor testosterone (T) could substitute for E in this respect. Furthermore, P given along with E curtailed tissue growth and its RCP content, whereas E + T had a synergistic effect on tissue growth only. During secondary stimulation, E administration steeply enhanced both tissue weight and RCP content without any lag. Interestingly, P (but not T) could substitute for E in augmenting magnum RCP concentration to a comparable extent while a concomitant effect on tissue growth was less marked. In contrast, hepatic induction of RCP was absolutely E-specific during both primary and secondary stimulations. Secondary stimulation with either E or P of E-primed birds enhanced the rates of RCP synthesis in the oviduct relative to that of total protein, whereas in the liver only E was effective in this regard. The absolute rate of E-induced RCP synthesis in both the steroid-stimulated tissues was significantly higher than that of general protein elaboration.

Inhibition of monkey liver serine hydroxymethyltransferase by Cibacron Blue 3G-A

Cibacron Blue 3G-A inhibited monkey liver serine hydroxymethyltransferase competitively with respect to tetrahydrofolate and non-competitively with respect to L-serine. NADH, a positive heterotropic effector, failed to protect the enzymes against inhibition by the dye and was unable to desorb the enzyme from Blue Sepharose CL-6B gel matrix. The binding of the dye to the free enzyme was confirmed by changes in the dye absorption spectrum. The results indicate that the dye probably binds at the tetrahydrofolate-binding domain of the enzyme, rather than at the 'dinucleotide fold'.

Studies on follicular atresia: role of tropic hormone and steroids in regulating cathepsin-D activity of preantral follicles of the immature rat

The ovary of the immature female rat is comprised of primary and medium-sized preantral follicles. Upon stimulation with FSH or PMSG, the cathepsin-D activity, a representative lysosomal enzyme of granulosa cells, is reduced by 50% (P < 0.01). 17β-Estradiol at the doses tried was unable to mimic this effect. Blockade of steroidogenesis with cyanoketone also had no effect on the cathepsin-D activity of isolated granulosa cells. Dihydrotestosterone (DHT), however, at a dose of 1 mg/rat was able to inhibit PMSG's tropic action. It brought about an increase in cathepsin-D activity and reduction in steroidogenic activity of isolated granulosa cells. The atretogenic activity of DHT could be relieved by supplementation with exogenous FSH. DHT was observed to significantly reduce (P < 0.01) endogenous FSH and LH levels within 12–18 h of its injection suggesting that its atretic effect was due to its action at the pituitary rather than the gonad. In addition to the above the ability of 15 IU of PMSG to reduce cathepsin-D activity of granulosa cells was also significantly reduced (P < 0.01) if endogenous FSH was neutralized by a specific FSH antiserum. The present study suggests that as far as small and medium-sized primary and preantral follicles are concerned, FSH lack is the essential signal for onset of atresia.

Biosynthesis of Long-Chain Alcohols by Developing and Regenerating Rat Sciatic Nerve

Cell-free preparations of rat sciatic nerve were found to catalyze the reduction of fatty acid to alcohol in the presence of NADPH as reducing cofactor. The reductase was membrane-bound and associated primarily with the microsomal fraction. When fatty acid was the substrate, ATP, coenzyme A (CoA), and Mg2+ were required, indicating the formation of acyl CoA prior to reduction. When acyl CoA was used as substrate, the presence of albumin was required to inhibit acyl CoA hydro-lase activity. Fatty acid reductase activity was highest with palmitic and stearic acids, and somewhat lower with lauric and myristic acids. It was inhibited by sulfhydryl reagents, indicating the participation of thiol groups in the reduction. Only traces of long-chain aldehyde could be detected or trapped as semicarbazone. Fatty acid reductase activity in rat sciatic nerve was highest between the second and tenth days after birth and decreased substantially thereafter. Microsomal preparations of sciatic nerve from 10-day-old rats exhibited about four times higher fatty acid reductase activity than brain or spinal cord microsomes from the same animals. Wallerian degeneration and regeneration of adult rat sciatic nerve resulted in enhanced fatty acid reductase activity, which reached a maximum at about 12 days after crush injury.

Induction of riboflavin-carrier protein in the immature male rat by estrogen: kinetic and hormonal specificity

The kinetics of estrogen-induced accumulation of riboflavin-carrier protein in the plasma was investigated in immature male rats using a specific and sensitive homologous radio-immunoassay procedure developed for this purpose. Following a single injection of the steroid hormone, plasma riboflavin-carrier protein levels increased markedly after an initial lag period of approximately 24 h, reaching peak levels around 96 h and declining thereafter. A 1.5 fold amplification of the inductive response was evident on secondary stimulation with the hormone. The magnitude of the response was dependent on hormonal dose, whereas the initial lag phase and the time of peak riboflavin-carrier protein induction were unaltered within the range of the steroid doses (0.1–10 mg/ kg body wt.) tested. Simultaneous administration of progesterone did not affect either the kinetics or the maximum level of the protein induced. The hormonal specificity of this induction was further adduced by the effect of administration of antiestrogens viz., En and Zu chlomiphene citrates, which effectively curtailed hormonal induction of the protein. That the induction involvedde novo-protein synthesis was evident from the complete inhibition obtained upon administration of cycloheximide. Passive immunoneutralization of endogenous riboflavin-carrier protein with antiserum to the homologous protein terminated pregnancy in rats confirming the earlier results with antiserum to chicken riboflavin-carrier protein.

An oxygenase from guinea-pig liver which catalyses sulphoxidation

A mono-oxygenase catalysing the conversion of 2-ethyl-4-thioisonicotinamide (ethionamide) into its sulphoxide was purified from guinea-pig liver homogenates. The enzyme required stoicheiometric amounts of oxygen and NADPH for the sulphoxidation reaction. The purified protein is homogeneous by electrophoretic, antigenic and chromatographic criteria. The enzyme has mol.wt. 85000 and it contains 1g-atom of iron and 1mol of FAD per mol, but not cytochrome P-450. The enzyme shows maximal activity at pH7.4 in a number of different buffer systems and the Km values calculated for the substrate and NADPH are 6.5×10-5m and 2.8×10-5m respectively. The activation energy of the reaction was calculated to be 36kJ/mol. Under optimal conditions, the molecular activity of the enzyme (mol of substrate oxidized/min per mol of enzyme) is calculated to be 2.1. The oxygenase belongs to the class of general drug-metabolizing enzymes and it may act on different compounds which can undergo sulphoxidation. The mechanism of sulphoxidation was shown to be mediated by superoxide anions.

Effect of antiserum to luteinizing hormone (LHAS) on the physiology of the epididymis and accessory glands in the albino rat

Rabbit antiserum specific to ovine luteinizing hormone free of contaminating antibodies to nonspecific proteins and FSH was administered to adult, intact rats at a dose of 0.1 and 0.2 ml/day for five days. LHAS had no effect on the weights of the epididymis but decreased their secretory activity to castrate level. Administration of 0.2 ml of LHAS or castration resulted in a marked and comparable reduction in the weights and secretory activity of the accessory glands. LHAS, even at a lower dose (0.1 ml/day), caused a significant reduction in the content of sialic acid in the vas deferons and Cowper's glands. These results are discussed in relation to the factors that regulate the functions of the epididymis and accessory glands.

Biochemical effects of 3,5-diethoxycarbonyl-1, 4-dihydrocollidine in mouse liver

3,5-Diethoxycarbonyl-1,4-dihydrocollidine (DDC) is a porphyrinogenic agent and is a powerful inducer of δ-aminolaevulinate synthetase, the first and rate-limiting enzyme of the haem-biosynthetic pathway, in mouse liver. However, DDC strikingly inhibits mitochondrial as well as microsomal haem synthesis by depressing the activity of ferrochelatase in vivo. The drug on repeated administration to female mice has been found to elicit hypertrophic effects in the liver microsomes initially, but the effects observed at later stages denote either hyperplasia or increase in polyploidal cells. The microsomal protein concentration shows a striking decrease with repeated doses of the drug. The rate of microsomal protein synthesis in vivo as well as in vitro shows an increase with two injections of DDC but decreases considerably with repeated administration of the drug. The activities of NADPH-cytochrome creductase and ribonuclease are not affected in the liver microsomes of drug-treated animals when expressed per mg of microsomal protein. DDC has also been found to cause degradation of microsomal haem, which is primarily responsible for the decrease in cytochrome P-450 content. The drug also leads to a decrease in mitochondrial cytochrome c levels due to inhibition of haem synthesis and also due to degradation of mitochondrial haem at later stages. The biochemical effects of the drug are compared and discussed with those reported for allylisopropylacetamide and phenobarbital.

Enzymatic acylation of 1-alkyl-, 1-alkenyl-and 1-acyl glycero-3-phosphorylethanolamine in developing rat brain

Rat brain particulate fractions were shown to acylate [32P]1-alkyl-sn-glycero-3-phosphorylethanolamine (GPE). While the main product is 1-alkyl-2-acyl GPE, about 12 per cent of the radioactivity was also found in 1-alkenyl-2-acyl GPE. The acyl transferase activity was completely dependent on added ATP and CoA and it was localized mainly in the microsomal fraction. A comparative study of acyl transferase activities to 1-alkyl-, 1-alkenyl-, and 1-acyl GPE by crude mitochondrial fraction and microsomes of 10, 16 and 22-day-old rat brains showed a progressive increase in activity with development. In the 22-day-old rat brain the order of activity towards the three substrates is as follows: 1-acyl GPE ± 1-alkenyl GPE ± 1-alkyl GPE with a crude mitochondrial fraction and 1-acyl GPE ± 1-alkyl GPE ± 1-alkenyl GPE with microsomes.

Patterns of service utilisation within Australian hepatology clinics: High prevalence of advanced liver disease

Accepted Article Abstract Background: Liver diseases in Australia are estimated to affect 6 million people with a societal cost of $51 billion annually. Information about utilization of specialist hepatology care is critical in informing policy makers about the requirements for delivery of hepatology-related health care. Aims: This study examined etiology and severity of liver disease seen in a tertiary hospital hepatology clinic, as well as resource utilisation patterns. Methods: A longitudinal cohort study included consecutive patients booked in hepatology outpatient clinics during a 3 month period. Subsequent outpatient appointments for these patients over the following 12 months were then recorded. Results: During the initial 3 month period 1471 appointments were scheduled with a hepatologist, 1136 of which were attended. 21% of patients were “new cases”. Hepatitis B (HBV) was the most common disease etiology for new cases (37%). Advanced disease at presentation varied between etiology, with HBV (5%), Hepatitis C (HCV) (31%), non-alcoholic fatty liver disease (NAFLD) (46%) and alcoholic liver disease (ALD) (72%). Most patients (83%) attended multiple hepatology appointments, and a range of referrals patterns for procedures, investigations and other specialty assessments were observed. Conclusions: There is a high prevalence of HBV in new case referrals. Patients with HCV, NAFLD and ALD have a high prevalence of advanced liver disease at referral, requiring ongoing surveillance for development of decompensated liver disease and liver cancer. These findings that describe patterns of health service utilisation among patients with liver disease provide useful information for planning sustainable health service provision for this clinical population

Effect of administration of clofibrate and clofenapate on kidney mitochondria of the rat

Abstract is not available.

Allosteric serine hydroxymethyltransferase from monkey liver: Correlation of conformational changes caused by denaturants with the alterations in catalytic activity

The far-ultraviolet region circular dichroic spectrumof serine hydroxymethyltransferase from monkey liver showed that the protein is in an α-helical conformation. The near ultraviolet circular dichoric spectrum revealed two negative bands originating from the tertiary conformational environment of the aromatic amino acid residues. Addition of urea or guanidinium chloride perturbed the characteristic fluorescence and far ultraviolet circular dichroic spectrum of the enzyme. The decrease in (θ)222 and enzyme activity followed identical patterns with increasing concentrations of urea, whereas with guanidinium chloride, the loss of enzyme activity preceded the loss of secondary structure. 2-Chloroethanol, trifluoroethanol and sodium dodecyl sulphate enhanced the mean residue ellipticity values. In addition, sodium dodecyl sulphate also caused a perturbation of the fluorescence emission spectrum of the enzyme. Extremes of pH decreased the – (θ)222 value. Plots of –(θ)222and enzyme activity as a function of pH showed maximal values at pH 7.4-7.5. These results suggested the prevalence of "conformational flexibility" in the structure of serine hydroxymethyltransferase.