EFFECT OF NAC ON MOUSE GV OOCYTE SURVIVAL AND SUBSEQUENT EMBRYONIC DEVELOPMENT FOLLOWING VITRFICATION.

BACKGROUND: Oocytes that survive cryopreservation may accumulate ROS which are known to bring harmful effects on embryonic development. NAC is an antioxidant which can be a supplement to reduce oxidative stress. However, whether NAC can improve the developmental competence of vitrified GV-oocytes remains unclear. OBJECTIVE: The study was to investigate the effect of NAC on subsequent embryonic developmental competence of mice vitrified GV-oocytes. MATERIALS AND METHODS: This study compared the effects of different concentration of NAC on the cleavage and blastocyst rates of mice vitrified GV-oocytes. Then the effects of NAC on mitochondria distribution, ROS level and embryonic development of vitrified oocytes were tested. RESULTS: ROS activity of vitrified oocytes was significantly annihilated and mitochondrial distribution pattern was improved by 1.5 mM NAC (P<0.05). NAC supplementation throughout vitrification/warming and IVM media significantly improved the developmental competence of vitrified oocytes. CONCLUSION: Supplementation of NAC could partially overcome the damages by vitrification and improve the development ability of mice vitrified GV-oocytes.

Inhibition of the growth of human gastric carcinoma in vivo and in vitro by swainsonine.

In Europe, swainsonine has been studied widely for prevention of metastasis and cancer therapy. In order to investigate the effects and mechanisms of swainsonine on the human gastric carcinoma SGC-7901 cell, we carried out in vivo and in vitro experiments. After treatment with swainsonine, an effective dose and IC50 value of swainsonine for SGC-7901 cells were examined by MTT assay. Cell-cycle distribution and apoptotic rates were analyzed using FCM, and [Ca2+]i was measured using LSCM. The expression of p53, c-myc and Bcl-2 were determined using an immunocytochemical method. Simultaneously, 50 mice were divided randomly into five groups. Three groups were administrated swainsonine at dose of 3, 6 and 12 mg/kg body wt., two control groups were administrated N.S. 20 ml/kg body wt. and 5-Fu 20 mg/kg body wt., respectively, by intraperitoneal injection. The inhibition rate was calculated and pathological sections were observed. The growth of SGC-7901 cell is inhibited by swainsonine in vitro, with an IC50 value at 24 h of 0.84 microg/ml, and complete inhibition concentration is 6.2 microg/ml. After treatment with swainsonine at the concentrations of 0.5, 1.5 and 4.5 microg/ml for 24 h, the expression of apoptosis inhibiting gene p53 and bcl-2 decreases, and the apoptotic trigger gene c-myc increases markedly (p<0.05), as well as [Ca2+]i overloading, SGC-7901 cell is induced to apoptosis in the end. It is also found that the percentages of S phase are 38.8%, 39.7% and 29.6%, respectively (20.0% in control group and 23.2% in 5-Fu group). The rates of inhibition were 13.2%, 28.9%, 27.3%, respectively, when the nude mice were administered swainsonine (p<0.05 or 0.01). The structure of the tumor showed hemorrhage, necrosis and inflammatory cell infiltration. We therefore conclude that swainsonine could inhibit cell proliferation in vitro and the growth of human gastric carcinoma in vivo. The mechanisms of swainsonine-induced apoptosis may relate to [Ca2+]i overloading and the expression of apoptosis-related genes.

Pharmacological studies of the large-scaled purified genistein from Huaijiao (Sophora japonica-Leguminosae) on anti-osteoporosis.

In this report, we used genistein that was extracted from a Chinese herbal medicine Huaijiao (Sophora japonica-Leguminosae) to evaluate its pharmacological function on anti-osteoporosis. This genistein is purified in a large-scale production from Huaijiao by a state-of-art method as described by Tian et al. [2004. The preparation of genistein and LC-MS/MS on-line analysis. Drug Devel. Res. 61, 6-12]. Chemical structure of the isolated genistein was examined by using various techniques including nuclear magnetic resonant spectrum, infrared absorption spectrum, ultraviolet absorption spectrum and mass spectrum, and was proved to be identical to those purified from soybean in a small scale as previously reported. We randomly divided female SD rats into 6 groups, including control, ovariectomized model, Nilestriol-treated, and three level of dosages of genistein-treated. We evaluated the pharmacological effects of genistein against osteoporosis by measuring the bone density of femur and bone mineral group including calcium, phosphorous, and magnesium. The consequences of genistein treatment on bone histology and morphology were also determined by measuring the trabcular area, thickness and number. Our results indicated that treatment with a 4.5 or 9 mg/kg dosage of genistein could also prevent osteoporosis significantly at the 4th week after treatment. In comparison with the anti-osteoporosis effects of soybean genistein, the genistein extracted from Huaijiao has the same beneficial effect on anti-osteoporosis.

Simulating bioremediation of uranium-contaminated aquifers; uncertainty assessment of model parameters.

Bioremediation of trace metals and radionuclides in groundwater may require the manipulation of redox conditions via the injection of a carbon source. For example, after nitrate has been reduced, soluble U(VI) can be reduced simultaneously with other electron acceptors such as Fe(III) or sulfate to U(IV), which may precipitate as a solid (uraninite). To simulate the numerous biogeochemical processes that will occur during the bioremediation of trace-metal-contaminated aquifers, a time-dependent one-dimensional reactive transport model has been developed. The model consists of a set of coupled mass balance equations, accounting for advection, hydrodynamic dispersion, and a kinetic formulation of the biological or chemical transformations affecting an organic substrate, electron acceptors, corresponding reduced species, and trace metal contaminants of interest, uranium in this study. This set of equations is solved numerically, using a finite difference approximation. The redox conditions of the domain are characterized by estimating the pE, based on the concentration of the dominant terminal electron acceptor and its corresponding reduced species. This pE and the concentrations of relevant species are then used by a modified version of MINTEQA2, which calculates the speciation/sorption and precipitation/dissolution of the species of interest under equilibrium conditions. Kinetics of precipitation/dissolution processes are described as being proportional to the difference between the actual and calculated equilibrium concentration. A global uncertainty assessment, determined by Random Sampling High Dimensional Model Representation (RS-HDMR), was performed to attain a phenomenological understanding of the origins of output variability and to suggest input parameter refinements as well as to provide guidance for field experiments to improve the quality of the model predictions. By decomposing the model output variance into its different input contributions, RS-HDMR can identify the model inputs with the most influence on various model outputs, as well as their behavior pattern on the model output. Simulations are performed to illustrate the effect of biostimulation on the fate of uranium in a saturated aquifer, and to identify the key processes that need to be characterized with the highest accuracy prior to designing a uranium bioremediation scheme.

[Effect of aloe-emodin on c-myc gene expression of cultured vascular smooth muscle cells of injured artery].

OBJECTIVE: To study the molecular mechanism of aloe-emodin (AE) in inhibiting smooth muscle cells' (SMC) proliferation. METHODS: Deendothelialization was performed in abdominal aorta of Japanese white ear rabbits by using 3F Fogarty arterial embolectomy catheter. The media of abdominal aorta was isolated 48 hrs later for performing primary SMC culture. Cells were synchronized with G0/G1 phase by serum starvation, the AE was then added to the culture medium of experimental group, at a concentration of 20 micrograms/ml, and to the control group, equal volume of culture solution was added instead. The c-myc mRNA and protein expressions were examined 3 hrs later by using techniques of Northern blotting, Western blotting and immunocytochemistry respectively. RESULTS: In comparing with the control group, neither the expression of c-myc mRNA nor the expression of c-myc protein was changed after addition of AE. CONCLUSION: The inhibition of AE on SMC might not be due to influencing c-myc expression, but via other pathway.

Effects of evodiamine on the secretion of testosterone in rat testicular interstitial cells.

Evodiamine, a bioactive component isolated from the Chinese medicine Wu-chu-yu, exhibits vasodilative and antianoxic action. Although evodiamine indeed has many biological effects, its effects on the endocrine system are not clear. The present study explored the effects of evodiamine on testosterone secretion in vitro. Rat collagenase-dispersed testicular interstitial cells (TICs) were incubated with evodiamine (0 to 10(-4) mol/L) in the presence or absence of human chorionic gonadotropin (hCG), forskolin, 8-bromo-adenosine 3':5'-cyclic monophosphate (8-Br-cAMP), or steroidogenic precursors (including 25-hydroxycholesterol, pregnenolone, progesterone, 17alpha-hydroxyprogesterone, and androstenedione) at 34 degrees C for 1 hour. The testosterone concentration in the media samples was measured by radioimmunoassay. Evodiamine 10(-4) mol/L was effective to reduce both basal and hCG-stimulated testosterone secretion in rat TICs after 1, 2, or 4 hours of incubation. The stimulatory effect of forskolin on testosterone release in TICs was prevented by administration of evodiamine. Evodiamine 10(-4) mol/L also decreased 8-Br-cAMP- and androstenedione-stimulated testosterone secretion. These results suggest that evodiamine reduces testosterone secretion in rat TICs via a mechanism involving reduced activity of cAMP-related pathways and 17beta-hydroxysteroid dehydrogenase (17beta-HSD).

Effects of methanol extract of Chansu on hypothalamic-pituitary-testis function in rats.

Chansu, a galenical preparation of the dried white venom of Chinese Bufo bufo gargarizans, is one of the major components of Kyushin, a traditional Chinese medicine. Kyushin is reported to have a cardiotonic effect that has been suggested to be due to the action of bufadienolides such as bufalin and cinobufagin. Recently, we found that administration of bufalin in male rats diminished the luteinizing hormone (LH) response to gonadotropin-releasing hormone (GnRH) and the secretion of testosterone both in vivo and in vitro. These observations suggest that Chansu may possess hypogonadal effects in male rats. In the present study, the effects of the methanol extract of Chansu on hypothalamic-pituitary-testicular function in male rats were examined. Crude Chansu was extracted by methanol and purified by a Sep-Pak C18 column. No activity of bufalin, cinobufagin, estradiol, or digoxin in purified methanol extract was detected; all Chansu used in this study was the purified methanol extract. A single intravenous injection of Chansu resulted in a decrease of the basal (20% to 55%) and human chorionic gonadotropin (hCG)-induced (35% to 40%) levels of plasma testosterone and the GnRH-induced level of plasma LH (25% to 30%). Administration of Chansu in vitro decreased basal and hCG-stimulated testosterone production by 60% to 70% and 40% to 60%, respectively, as well as spontaneous and forskolin- or 3-isobutyl-1-methylxanthine (IBMX)-induced accumulation of adenosine 3',5'-cyclic monophosphate (cAMP) by 30% to 45% in rat testicular interstitial cells. Although LH release by rat anterior pituitary glands was diminished, GnRH release by the rat mediobasal hypothalamus was enhanced by administration of Chansu in vitro. These results suggest that the bufalin-free extracts of Chansu inhibit testosterone secretion in rats, in part, due to (1) a decreased production of testicular cAMP, (2) a decreased response of testosterone to gonadotropin, and (3) a reduction of the LH response to GnRH.

Effects of dietary fat on food intake and brain uptake and oxidation of fatty acids.

The present study was designed to investigate (a) whether dietary fat manipulation, quantitatively and/or qualitatively, may influence the transport of fatty acids into the brain and oxidation of fatty acids in the hypothalamus; and (b) if an inhibitor of fatty acid oxidation changed food intake on these diets. Rats were fed for 4 wk 5% or 34% corn oil or tallow diets that were isocaloric and isonitrogenous. It was found that rats fed corn oil diets had significantly higher brain uptake index of palmitate than did rats fed tallow diets. In a second experiment, rats were fed either 30% corn oil or tallow diets and injected with either saline or mercaptoacetate (MA). Both saturated fat diets and mercaptoacetate injection reduced lateral hypothalamic, but not ventromedial hypothalamic fatty acid oxidation. Mercaptoacetate increased food intake only in the corn oil fed rats. It is proposed that these uptake and metabolic changes contribute to alterations in either energy expenditure or feeding behavior.

Effects of thyroidectomy on the metabolic clearance rate of prolactin and prolactin release in the rat.

The effects of thyroidectomy on both the secretion of rat prolactin (PRL) in vivo and the release of PRL from rat anterior pituitary glands (Aps) in vitro in the absence of ovarian steroids were studied. Rats were ovariectomized (Ovx) 42 days after thyroidectomy or sham-thyroidectomy. On day 15 following ovariectomy the rats were either decapitated for examining the in vitro release of PRL from AP or anesthetized for measuring the metabolic clearance rate (MCR) of rat PRL. After decapitation, blood samples were collected and APs were incubated with or without the extract of rat medial basal hypothalamus (rHE) at 37 degrees C for 4 h. The MCR of iodinated rat PRL was determined by a single injection method. Blood samples were collected by heart puncture at frequent intervals. Serum PRL concentrations of decapitated thyroidectomized-ovariectomized (TX-Ovx) rats were 56% lower than those found in Ovx rats. Thyroidectomy in Ovx rats resulted in a significant reduction of the spontaneous release of PRL from AP. The rHE decreased PRL levels in the medium after incubation with APs from sham Tx rats, but not those from Tx rats. MCR of PRL was reduced in Ovx rats by thyroidectomy. These results indicate that thyroidectomy reduces the spontaneous release of rat PRL by AP and secretion rate of rat PRL in the absence of ovarian steroids.

Cytotoxicity and sister chromatid exchanges induced in vitro by six anticancer drugs developed in the People's Republic of China.

Growth inhibition in the Chinese hamster cell line V79 and in the human lymphoid cell line Raji and induction of sister chromatid exchange(s) (SCE) in V79 cells after treatment with six anticancer drugs [harringtonine (HRT), homoharringtonine (HHRT), camptothecin (CPT), hydroxycamptothecin (HCPT), lycobetaine (LBT), and oxalysine (OXL)] developed in the People's Republic of China were studied. OXL is a new antibiotic; all other drugs are plant extracts. All drugs caused a dose-dependent growth inhibition in both cell types, as evidenced by decreases in plating efficiencies of V79 cells and in viable cell counts of Raji. However, the degree of inhibition differed widely among the drugs. HRT, HHRT, CPT, and HCPT were the most potent growth inhibitors, LBT was next, and OXL was the least effective inhibitor. SCE analyses were made in V79 cells treated with a drug in the presence or absence of the metabolic activation system S9 mixture (S9 mix), except for the HRT assay in which the S9 mix was not used. CPT, HCPT, and LBT induced a dose-dependent increase in SCE frequencies, while HRT, HHRT, and OXL caused no SCE induction at any dose level used. CPT was the most powerful SCE inducer. HCPT induced SCE but at a much reduced rate when compared to that of CPT. LBT was a weak SCE inducer; SCE induction was seen only in cultures treated with 40 micrograms or more LBT/ml. Addition of the S9 mix did not alter SCE frequencies, indicating that the drugs were direct-acting agents. HRT and HHRT were highly toxic, but they induced no increases in SCE frequency, indicating that cytotoxicity of a compound does not necessarily correlate with SCE induction.