Glutathione transferase T1 and M1 genotype polymorphism in the normal population of Shanghai

Glutathione transferases are known to be important enzymes in the metabolism of xenobiotics. In humans genetic polymorphisms have been reported for the hGSTM1 and hGSTT1 genes leading to individual differences in susceptibility towards toxic effects, such as cancer. This study describes the distribution of the two polymorphisms of hGSTT1 and hGSTM1 in the normal Chinese population of Shanghai. Out of 219 healthy individuals having been genotyped for GSTT1 and GSTM1, 108 (49%) were identified to be homozygously deficient for the GSTT1 gene and 107 (49%) for the GSTM1 gene.     

Cyclooxygenase isoenzyme localization and mRNA expression in rat lungs

Prostanoid generation may proceed via both isoforms of cyclooxygenase, Cox-1 and Cox-2. Cox-1 is thought to be ubiquitously expressed, whereas Cox-2 is mostly assumed to be dynamically regulated, responding to inflammatory stimuli. The cellular localization of Cox-1 and Cox-2 in the lung, an organ with high cyclooxygenase activity, is not known. In normal rat lungs the expression and localization of Cox-1 and Cox-2 were examined with immunogold-silver staining and the RT-PCR technique. Quantitative image analysis of the staining intensity was performed by measuring mean gray values of digitized epipolarization images. Expression of both Cox-1 and Cox-2 was readily detectable in rat lungs. Cox-1 immunoreactivity localized predominantly to bronchial epithelial cells, smooth muscle cells of large hilum veins, and (with lower expression) to alveolar macrophages and pulmonary artery endothelial cells. The most intense Cox-2 staining was noted in macrophage- and mast cell-like cells, detected in close vicinity to the bronchial epithelium and in the connective tissue surrounding the vessels. In addition, strong Cox-2 expression was found in smooth muscle cells of partially muscular vessels and large veins of the hilum. Bronchial epithelial cells displayed Cox-2 immunoreactivity with limited intensity. Alveolar macrophages and alveolar septal cells were only occasionally stained with anti-Cox-2 antibodies. Both Cox-1 and Cox-2 are constitutively expressed in several cell types of normal rat lung, but display clearly different patterns of cellular localization. Cox-2 may not be related only to lung inflammation, but is suggested to be implicated in regulatory processes under physiological conditions as well.     

Relaxin induces ovulations in the in-vitro perfused rat ovary

The effects of human recombinant relaxin on ovulation and ovarian steroidogenesis were investigated in vitro using a perfused rat ovary model. Ovaries of equine chorionic gonadotrophin (ECG; 20 IU)-primed Sprague-Dawley rats were perfused for 21 h. Ovarian release of oestradiol and progesterone was measured during the perfusion period and the number of ovulations was estimated by counting the released oocytes at termination of the experiment. Non-treated control ovaries did not ovulate whereas addition of ovine luteinizing hormone (LH; 100 ng/ml) resulted in a mean (+/- SEM) number of ovulations of 3.0 +/- 0.8 from all treated ovaries. Relaxin (10 micrograms/ml) induced mean (+/- SEM) number of ovulations at 2.4 +/- 0.2 in all treated ovaries but did not further increase the ovulation rate when combined with LH (mean +/- SEM 3.2 +/- 0.4). All ovulated oocytes in the groups stimulated by LH showed signs of nuclear maturation (germinal vesicle breakdown) when harvested, in contrast to ovulated oocytes in the relaxin group, which were immature (presence of germinal vesicle). Progesterone and oestradiol release was significantly increased in the LH-stimulated groups but not in the group treated only with relaxin, in comparison to the untreated control group. These results demonstrate that relaxin may have a paracrine role within the ovary and may facilitate ovulation, possibly by promoting connective tissue remodelling of the follicle wall.     

Glutathione S-transferases in human renal cortex and neoplastic tissue: enzymatic activity, isoenzyme profile and immunohistochemical localization

1. Glutathione S-transferase (GST) activity in the cytosol of renal cortex and tumours from eight men and eight women was measured using 1-chloro-2,4-dinitrobenzene (CDNB) as a substrate. GST activities ranged from 685 to 2192 nmol/min/mg protein in cortex (median 1213) and from non-detectable (minimum 45) to 2424 nmol/min/mg protein in tumours (median 469). The activities in the tumours were lower than those in the normal cortices (p < 0.05). 2. In men, the activity in the cortical cytosol was in all cases higher than that measured in the corresponding tumours (p < 0.05). In women, the difference in activity between cortices and tumours was not significantly different (p > 0.05). 3. The age of the patients ranged from 42 to 81 years (median 62) and was not found to play a role in the levels of GST activity observed in cortex or in renal tumours from either sex. 4. Immunoblotting and immunohistochemical studies confirmed that GST-alpha was the predominant form expressed both in normal cortex and tumour and probably accounted for most of the GST activity present in these samples. GST-mu and GST-phi were expressed in both tumours and normal cortex and, while in some cases the level of expression in the cortices was higher than that found in the tumours, the reverse was also observed. Within the GST-mu class, GST M1/M2 was only detected in one sample (tumour), which showed the highest overall expression of GST-mu. GSTM3 was the predominant isoenzyme of the mu class in normal and tumour tissue, whereas GTM4 and GSTM5 were not detected. 5. These differences could have functional significance where xenobiotics or cytotoxic drugs are specific substrates for the different classes of GSTs.     

Glutathione movements during cold preservation of rat hepatocytes

PURPOSE: Power-Doppler sonography is regarded as a very sensitive method for detecting low-velocity and low-volume blood flows. The purpose of our study was to investigate whether increased vascularity in breast carcinoma can be visualized by power-Doppler sonography and whether new criteria for differentiating benign and malign lesions can be found. METHOD: 315 patients were examined with a 13-MHz high-resolution linear transducer. If a suspicious lesion was found, it was evaluated further by power-Doppler sonography. Compared to normal breast parenchyma (reference structure), a focal increase in blood flow signals was registered using a 3-step grading system with a 4th step for no flow increase. RESULTS: In 97 cases the sonographic findings were correlated with histology (n = 95) or cytology (n = 2). There were 50 benign lesions, 42 cases of invasive and 5 cases of in-situ carcinoma. 73.5% benign lesions showed no or just minimal increases in flow signal. 81% of invasive cancer presented middle- or high-flow increases compared to normal breast parenchyma. The extend of flow increase was linked to tumor size in invasive cancer. In stage T1b to T4, 94.3% of invasive carcinoma presented middle or high flow increases. CONCLUSION: Power-Doppler sonography is able to visualize vascularization in breast tumors. According to first clinical results PD sonography is a promising additional diagnostic tool which seems to offer new criteria for differential diagnosis in breast tumors.     

Glutathione levels in antigen-presenting cells modulate Th1 versus Th2 response patterns

Current thinking attributes the balance between T helper 1 (Th1) and Th2 cytokine response patterns in immune responses to the nature of the antigen, the genetic composition of the host, and the cytokines involved in the early interaction between T cells and antigen-presenting cells. Here we introduce glutathione, a tripeptide that regulates intracellular redox and other aspects of cell physiology, as a key regulatory element in this process. By using three different methods to deplete glutathione from T cell receptor transgenic and conventional mice and studying in vivo and/or in vitro responses to three distinct antigens, we show that glutathione levels in antigen-presenting cells determine whether Th1 or Th2 response patterns predominate. These findings present new insights into immune response alterations in HIV and other diseases. Further, they potentially offer an explanation for the well known differences in immune responses in "Th1" and "Th2" mouse strains.     

Reanalysis of feeding patterns in the rat.

Conducted 3 experiments using 40 male Sprague-Dawley albino and hooded rats as Ss. No strict correlations between meals and intermeal intervals were observed in free-feeding patterns. Pooling of raw data into deciles did increase correlations, but also reduced data variance systematically. Reanalysis of raw data in relative terms (i.e., ratios of meals and intermeal intervals) yielded reliable meal pattern correlations, and indicated that feeding patterns are controlled by endogenous circadian signals of body nutrient depletion-repletion rather than short-term consequences of recently ingested food. Chronic treatment with insulin (which prevents mobilization of stored nutrients) appeared to attenuate the repletion signal, and termination of treatment (which leads to increased breakdown of stored nutrients) to accentuate the signal. Accordingly, the pattern of feeding may reflect lipid and/or glycogen metabolism. (19 ref.) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Glutathione oxidation and embryotoxicity elicited by diamide in the developing rat conceptus in vitro

This study was performed in the rat whole-embryo culture system to investigate the effects of glutathione oxidation by diamide, a thiol oxidant, in developing rat conceptuses during early organogenesis. The effects of diamide on reduced glutathione (GSH), glutathione disulfide (GSSG), and embryotoxicity were found to be concentration and time dependent. Diamide at concentrations of 75 and 100 microM produced abnormal axial rotation (62-89%), decreased viability (to 69% by 100 microM diamide), and reduced protein and DNA content in the embryo and visceral yolk sac (VYS) when evaluated on Day 11. High concentrations of diamide (250-500 microM) resulted in 100% mortality. GSH and GSSG levels in the conceptuses were not significantly affected during 2 hr following diamide addition at concentrations of 50 to 100 microM. At concentrations of 250 and 500 microM, rapid GSH depletion (50% of control) was seen within 5 min of exposure and was followed at 5-30 min by a significant increase in GSSG relative to control values. Diamide (500 microM) exposure for only 15 min on Gestational Day 10 was sufficient to elicit malformations (53% of exposed conceptuses with abnormal axial rotation) without significant loss of viability. After 30 min of exposure to the high concentration (500 microM), viability was decreased to 71% and defects of axial rotation increased to 87% in surviving conceptuses. This indicates that events associated with initial exposure are critical for expression of toxicity. Inhibition of glutathione disulfide reductase (GSSG reductase) activities in embryo and VYS with 1,3-bis(2-chloroethyl)-1-nitro-sourea prior to diamide addition potentiated the embryotoxicity of diamide (75 microM) and resulted in corresponding reductions in GSH/GSSG ratios as determined during the first 2 hr of exposure. Inhibition of new GSH synthesis with L-buthionine-[S,R]-sulfoximine during diamide (75 microM) exposure also exacerbated toxicity compared to diamide treatment alone. These results implicate the involvement of GSH synthesis and GSSG reductase activity in mediating the embryotoxicity of diamide.     

Protein phosphatase activity in the rat ovary throughout pregnancy and pseudopregnancy

Specific protein phosphatase activity against protein kinase C-phosphorylated substrate was measured in the rat ovary during pseudopregnancy and pregnancy. Tissues were processed in the presence of sodium fluoride and inorganic phosphate to inhibit the phosphatase and thereby prevent autodephosphorylation of the type 2A protein phosphatase (PP2A) during homogenization. Manganese was added at the time of enzyme assay to reactivate the phosphatase. The specific activity of the protein phosphatase did not vary significantly across pseudopregnancy (p > 0.05). In contrast, the specific activity of protein phosphatase decreased significantly between Day 7 and Day 10 of pregnancy (28.8 +/- 5 pmol/min x microg protein and 20.7 +/- 2 pmol/min x microg protein, respectively; p < 0.05) and remained at the decreased value for the remainder of pregnancy. To determine whether hormones of pregnancy could regulate PP2A activity in the ovaries, pseudopregnant rats were treated with prolactin (3 IU twice a day), bromocriptine (100 microg twice a day), or estradiol benzoate (50 microg). Bromocriptine and estradiol treatments caused a decrease in PP2A-specific activity, but prolactin had no effect. Bromocriptine treatment caused a decrease in the protein content of the PP2A catalytic subunit, but prolactin and estradiol treatments had no effect. The data suggest that the specific activity and protein content of PP2A in the rat ovary are hormonally regulated.     

Glutathione recycling is attenuated by acute ethanol feeding in rat liver

During chronic high-altitude (HA) exposure, basal and exercise-induced noradrenaline (NA) increases do not parallel blood pressure (BP) changes observed; unlike beta-adrenergic receptors, to our knowledge no data are available on alpha-receptors. We studied platelet alpha 2- and leucocyte beta-receptors and basal catecholamine levels in 11 trained climbers before and after they had spent a 15-day period at a height of over 4400 m. In six of the climbers we also evaluated catecholamines after maximal bicycle ergometer exercise. After chronic high-altitude exposure, a significant decrease was found in platelet alpha 2-receptor density and affinity [Bmax from 92.6 +/- 6.7 to 54.6 +/- 4.2 fmol mg-1 protein (P < 0.001) and KD from 1.271 +/- 0.034 to 1.724 +/- 0.077 nmol L-1 (P < 0.05)], although no changes to beta-receptors were observed. No changes were found in basal pre- and post-expedition NA and adrenaline (A), and there was only a slight decrease in post-expedition NA after maximal exercise. Our results suggest that prolonged exposure to hypoxia induces a down-regulation of alpha 2-receptors, which may be a contributory factor in the regulation of the physiological vascular response to acclimatization.     

The effect of photomirex on the in vitro perfused ovary of the rat

We describe the cloning of p63, a gene at chromosome 3q27-29 that bears strong homology to the tumor suppressor p53 and to the related gene, p73. p63 was detected in a variety of human and mouse tissues, including proliferating basal cells of epithelial layers in the epidermis, cervix, urothelium, and prostate. Unlike p53, the p63 gene encodes multiple isotypes with remarkably divergent abilities to transactivate p53 reporter genes and induce apoptosis. Importantly, the predominant p63 isotypes in many epithelial tissues lack an acidic N terminus corresponding to the transactivation domain of p53. We demonstrate that these truncated p63 variants can act as dominant-negative agents toward transactivation by p53 and p63, and we suggest the possibility of physiological interactions among members of the p53 family.     

Microcirculatory bed of the rat ovary in health and under laser irradiation

Personal clinical observations during the recent years allow the authors to confirm the indisputable value of surgical experience got during the Great Patriotic War (1941-1945) and in particular, the "fourfold" scheme proposed by V. I. Voiachek for the diagnosis and treatment of blind gunshot wounds to the skull base. Computed tomography considerably increases the probability of detection of the exact localization of foreign bodies in complex anatomical structures of the skull and thus facilitates choosing the most rational surgical management. The use of the electro-optical transducer for the extraction of foreign bodies from almost inaccessible areas of the skull base decreases the risk of operation.     

Differential expression of estrogen receptor-beta and estrogen receptor-alpha in the rat ovary

Immunohistochemical localization of two estrogen receptor (ER) subtypes, ER beta and ER alpha, was performed in neonatal, early postnatal, immature, and adult rats to determine whether ER alpha and ER beta are differentially expressed in the ovary. ER beta and ER alpha were visualized using a polyclonal anti-ER beta antibody and a monoclonal ER alpha (ID5) antibody, respectively. Postfixed frozen sections and antigen-retrieved paraffin sections of the ovary revealed nuclear ER beta immunoreactivity (IR) in granulosa cells, which was prevented when peptide-adsorbed antibody was used instead. In immature and adult rat ovaries, ER beta was expressed exclusively in nuclei of granulosa cells of primary, secondary, and mature follicles. Atretic follicle granulosa cells showed only weak or no staining. No specific nuclear ER beta IR was detected in thecal cells, luteal cells, interstitial cells, germinal epithelium, or oocytes. In neonatal rat ovary, no ER beta expression was found. In ovaries of 5- and 10-day-old rats, weak ER beta IR was observed in granulosa cells of primary and secondary follicles, but no staining was detected in the primordial follicles. ER alpha protein exhibited a differential distribution in the ovary with no detectable expression in the granulosa cells but evidence of ER alpha IR in germinal epithelium, interstitial cells, and thecal cells. In the oviduct and uterus, IR for ER alpha, but not ER beta, was found in luminal epithelium, stromal cells, muscle cells, and gland cells. Our present study demonstrates that ER beta and ER alpha proteins are expressed in distinctly different cell types in the ovary. The exclusive presence of ER beta in granulosa cells implies that this specific new subtype of ER beta mediates some effects of estrogen action in the regulation of growth and maturation of ovarian follicles.     

Elevated N-myristoyl transferase activity is reversed by sodium orthovanadate in streptozotocin-induced diabetic rat

N-Myristoyl transferase (NMT) is the enzyme that covalently modifies several proteins important in signal transduction. Streptozotocin-induced diabetes resulted in a 2-fold increase in NMT activity from rat liver as compared to control animals. Administration of sodium orthovanadate to the diabetic rats reduced the activity of the NMT to 75-120% of the control values. Elevated NMT activity was observed with both cAMP-dependent protein kinase-derived and pp60src-derived peptide substrates. No significant change in the apparent Km was observed with the cAMP-dependent protein kinase-derived peptide substrate. Unlike in rat brain, in all conditions highest NMT activity was observed in the particulate fraction of rat liver.     

Relations between feeding and sleep patterns in the rat.

A concomitant analysis of sleep and feeding patterns in 11 male Wistar rats was carried out over 8 days, using continuous EEG recording. The proportion of slow-wave sleep and paradoxical sleep within an intermeal interval was constant and varied only in relation to the time of the day. During the dark period only, there were significant correlations between meal size and amount of time spent in both stages of sleep in the following intermeal interval. These correlations were even stronger between meal size and sleep duration in the intermeal interval that followed the next meal. Circadian variations in satiety ratios (i.e., units of subsequent intermeal interval and sleep per units of food ingested) and in deprivation ratios (i.e., units of feeding per units of prior intermeal interval and sleep) suggest that division of 24-hr data into 3 8-hr periods rather than 2 12-hr periods reveals distinct correlations between meal size and pre- and postmeal sleep and might reflect more accurately the underlying metabolic events. (34 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Classical conditioning of hippocampal theta patterns in the rat.

Electrical stimulation (88 Hz) of the lateral hypothalamus elicited a sustained theta response at hippocampal recording sites in 11 male Sprague-Dawley rats immobilized with succinylcholine. By pairing this UCS with a 10-sec presentation of a light, conditioned theta responses were demonstrated in as few as 40 trials. Spectral analysis of hippocampal bioelectric patterns during acquisition, extinction, and reconditioning indicated that the earliest change as a result of conditioning was a loss of power in EEG frequency below 8 Hz, followed by the development of a peak at 8 Hz with further conditioning. Extinction was associated with an increase in power of the frequencies below 8 Hz. When the conditioned Ss were tested in the absence of the neuromuscular blocking agent, the CS elicited a theta response that was associated with slow motor activity on 70% of the trials. (16 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)