The expression of nitric oxide synthase (NOS) isoforms in the ovaries of pigs was examined to study the involvement of nitric oxide, a product of NOS activity, in the function of the ovary. Western blot analysis detected three types of NOS in the ovary, including constitutive neuronal NOS (nNOS), endothelial NOS (eNOS) and inducible NOS (iNOS); eNOS immunoreactivity was more intense compared with that of iNOS or nNOS. Immunohistochemical studies demonstrated the presence of nNOS and eNOS in the surface epithelium, stroma, oocytes, thecal cells, and endothelial cells of blood vessels. Positive immunoreactions for nNOS and iNOS were detected in the granulosa cells from multilaminar and antral follicles, but not in those of unilaminar follicles. iNOS was detected in the surface epithelium, oocytes, and theca of multilaminar and antral follicles. Taking all of the findings into consideration, the observed differential expression of the three NOS isoforms in the ovary suggests a role for nitric oxide in modulating reproduction in pigs.
Animals ; Blotting, Western/veterinary ; Female ; Immunohistochemistry/veterinary ; Nerve Tissue Proteins/*biosynthesis ; Nitric Oxide/metabolism ; Nitric Oxide Synthase/*biosynthesis ; Nitric Oxide Synthase Type I ; Nitric Oxide Synthase Type II ; Nitric Oxide Synthase Type III ; Ovarian Follicle/*enzymology/growth&development ; Swine/*physiology

In order to further investigate the mechanisms of action of berberine (Ber), we assessed the effects of Ber on the mRNA expression of nitric oxide synthases (NOS) in rat corpus cavernosum. After incubation with Ber for 1 or 3 h respectively, the levels of NOS mRNA were examined by reverse transcription polymerase chain reaction (RT-PCR). Our results showed that there were iNOS and eNOS mRNA expressions in rat corpus cavernosum. Ber enhanced eNOS mRNA expression in rat penis, but exhibited no effect on the expression of iNOS mRNA (P > 0.05). The present study indicated that the relaxation of Ber involved the NO-cGMP signal transduction pathway. The enhancing effect of Ber on eNOS mRNA expression might associated with its relaxation of corpus cavernosum.
Berberine/*pharmacology ; Connective Tissue/physiopathology ; Nitric Oxide Synthase/*biosynthesis ; Nitric Oxide Synthase/genetics ; Nitric Oxide Synthase Type I/biosynthesis ; Nitric Oxide Synthase Type I/genetics ; Nitric Oxide Synthase Type III/biosynthesis ; Nitric Oxide Synthase Type III/genetics ; Penile Erection/*physiology ; Penis/*metabolism ; Penis/physiology ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics

OBJECTIVELong time exhaled oxygen will induced oxygen toxicity. Some studies had found that different pathology may exised in normobaric and hyperbaric pulmonary oxygen toxicity, and nitric oxide synthase (NOS) may play a role. In this study, we discussed the change of NOS in normobaric and hyperbaric pulmonary oxygen toxicity.

METHODSSixty male SD rats were randomly divided into 6 groups (n = 10), exposed to 1 ATA (atmosphere absolute), 1.5 ATA, 2 ATA, 2.5 ATA and 3 ATA, 100% oxygen for 56, 20, 10, 8, 6 hours respectively. Rats were exposed to air as control. After exposure, the protein in bronchoalveolar lavage fluid (BALF), the wet/dry weight of lung and the expression of eNOS, nNOS in lung were defined.

RESULTSAs compared to air group, the protein in BALF, the wet/dry of lung were significantly elevated in 1.0 ATA group, while these changes were not so obviously in the other groups, and these changes in hyperbaric oxygen group (approximately 1.0 ATA) were significantly decreased as compared with nonnrmobaric oxygen group (1.0 ATA). The expression of nNOS were not changed in normobaric and hyperbaric pulmonary oxygen toxicity, while the expression of eNOS was significantly decreased in 2 ATA group, and significantly elevated in 2.5 ATA and 3 ATA group.

CONCLUSIONThe expression of eNOS can change when exposed to different pressures of oxygen.

Animals ; Disease Models, Animal ; Lung ; metabolism ; Male ; Nitric Oxide Synthase Type I ; metabolism ; Nitric Oxide Synthase Type III ; metabolism ; Oxygen ; poisoning ; Pressure ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo observe the expression of endothelial nitric oxide synthase (eNOS)and nervous nitric oxide synthase (nNOS) in rats during cerebral ischemia/reperfusion (CI/R) and study if change will be happen in subgroup between eNOS and nNOS during earlier period of CI/R.

METHODSA total of 60 Wistar rats weighting 200-280 g, supplied by Animal Center of China Medical University, were divided into 6 groups (n=10) (sham operation group; ischemia 1 h, 2 h group; reperfusion 0.5 h, 1 h, 2 h group). Female and male was half-and-half. Cerebral ischemia/reperfusion injury was induced by a 2-hour suture occlusion of the unilateral middle cerebral artery, immediately after suture withdrawal to allow reperfusion, eNOS and nNOS expressions were examined by the method of immunohistochemistry.

RESULTSeNOS expressions increased in 1-hour during ischemia, keeping up with decreasing until reperfusion 2-hour. While nNOS expressions increased in 2-hour between ischemia and reperfusion.

CONCLUSIONChanges of expression between eNOS and nNOS in rats during cerebral ischemia/reperfusion are different. This may be related with ischemia and reperfusion injury.

Animals ; Brain ; enzymology ; Brain Ischemia ; enzymology ; Female ; Male ; Nitric Oxide Synthase Type I ; metabolism ; Nitric Oxide Synthase Type III ; metabolism ; Rats ; Rats, Wistar ; Reperfusion Injury ; enzymology ; Time Factors

OBJECTIVETo study the effects of insulin-like growth factor II (IGF-II) on regulating the levels of nitric oxide (NO) and the mRNA transcriptions of inducible nitric oxide synthase (iNOS) and endothelial NOS (eNOS) in mouse osteoblast-like cells.

METHODSMouse osteoblastic cell line MC3T3-E1 was selected as the effective cell of IGF-II. After the cells were treated with IGF-II at different concentrations for different intervals of time, MTT colorimetry was used for examining the cell proliferation. Nitrate reductase method was applied for detecting the NO concentrations in cell culture supernatants and RT-PCR employed for determining the levels of cellular iNOS and eNOS mRNAs.

RESULTSAfter the MC3T3-E1 cells were treated with IGF-II at the dosages of 1 microg/L for 72 h, 10 and 100 microg/L for 24, 48 and 72 h respectively, all the MTT values increased markedly (P < 0.05 or P < 0.01). After the cells were treated for 48 and 72 h at the dosage of 100 microg/L IGF-II respectively, the levels of NO in the supernatants of cell cultures and cellular iNOS mRNA decreased significantly (P < 0.01). However, the levels of eNOS mRNA in the cells treated with any of the IGF-II dosages for the different times were stable (P > 0.05).

CONCLUSIONSIGF-II at the dosages of 1 approximately 100 microg/L showed the effects on promoting proliferation, which as probably due to the maintenance of low NO levels. Inducible NOS gene expression at the level of transcription was down regulated in the MC3T3-E1 cell treated with higher dosage of IGF-II (100 microg/L) but eNOS mRNA was not, which might be one of the mechanisms for the maintenance of low NO levels.

3T3 Cells ; Animals ; Insulin-Like Growth Factor II ; pharmacology ; Mice ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase ; biosynthesis ; genetics ; Nitric Oxide Synthase Type II ; Nitric Oxide Synthase Type III ; Osteoblasts ; cytology ; drug effects ; enzymology ; RNA, Messenger ; biosynthesis

OBJECTIVETo study the changes in the microglial cells and the activity of nitric oxide synthase (NOS), inducible nitric oxide synthase (iNOS) and constitutive nitric oxide synthase (cNOS) in the striatum of rats with methamphetamine (METH) treatment.

METHODSThe rats were randomly divided into two groups for injections with METH or saline. Specific antibody against OX-42 was used to detect the changes in the morphology and the number of microglia, and the activities of NOS, iNOS and cNOS were compared between the two groups.

RESULTSThe microglial cells were activated and their number significantly increased in the striatum of rats with METH treatment as compared with those in the saline group. The activated microglial cells showed bushy and amoeboid morphologies in the METH group. METH also significantly enhanced the activities of NOS, iNOS and cNOS in the striatum (P < 0.05).

CONCLUSIONMicroglial activation and increased NOS activity may participate in METH-induced neurotoxicity in rat striatum.

Animals ; Corpus Striatum ; enzymology ; Male ; Methamphetamine ; pharmacology ; Microglia ; metabolism ; Nitric Oxide Synthase ; metabolism ; Nitric Oxide Synthase Type II ; metabolism ; Nitric Oxide Synthase Type III ; metabolism ; Random Allocation ; Rats ; Rats, Wistar

BACKGROUNDExposure of adult mice to more than 95% O(2) produces a lethal injury by 72 hours. Nitric oxide synthase (NOS) is thought to contribute to the pathophysiology of murine hyperoxia-induced acute lung injury (ALI). Osteopontin (OPN) is a phosphorylated glycoprotein produced principally by macrophages. OPN inhibits inducible nitric oxide synthase (iNOS), which generates large amounts of nitric oxide production. However, the relationship between nitric oxide and endogenous OPN in lung tissue during hyperoxia-induced ALI has not yet been elucidated, thus we examined the role that OPN plays in the hyperoxia-induced lung injury and its relationships with NOS.

METHODSOne hundred and forty-four osteopontin knock-out (KO) mice and their matched wild type background control (WT) were exposed in sealed cages > 95% oxygen or room air for 24- 72 hours, and the severity of lung injury was assessed; expression of OPN, endothelial nitric oxide synthase (eNOS) and iNOS mRNA in lung tissues at 24, 48 and 72 hours of hyperoxia were studied by reverse transcription-polymerase chain reaction (RT-PCR); immunohistochemistry (IHC) was performed for the detection of iNOS, eNOS, and OPN protein in lung tissues.

RESULTSOPN KO mice developed more severe acute lung injury at 72 hours of hyperoxia. The wet/dry weight ratio increased to 6.85 +/- 0.66 in the KO mice at 72 hours of hyperoxia as compared to 5.31 +/- 0.92 in the WT group (P < 0.05). iNOS mRNA (48 hours: 1.04 +/- 0.08 vs. 0.63 +/- 0.09, P < 0.01; 72 hours: 0.89 +/- 0.08 vs. 0.72 +/- 0.09, P < 0.05) and eNOS mRNA (48 hours: 0.62 +/- 0.08 vs. 0.43 +/- 0.09, P < 0.05; 72 hours: 0.67 +/- 0.08 vs. 0.45 +/- 0.09, P < 0.05) expression was more significantly increased in OPN KO mice than their matched WT mice when exposed to hyperoxia. IHC study showed higher expression of iNOS (20.54 +/- 3.18 vs. 12.52 +/- 2.46, P < 0.05) and eNOS (19.83 +/- 5.64 vs. 9.45 +/- 3.82, P < 0.05) in lung tissues of OPN KO mice at 72 hours of hyperoxia.

CONCLUSIONOPN can protect against hyperoxia-induced lung injury by inhibiting NOS.

Animals ; Hyperoxia ; genetics ; physiopathology ; Immunohistochemistry ; Lung ; metabolism ; Lung Injury ; etiology ; genetics ; metabolism ; Mice ; Mice, Knockout ; Nitric Oxide Synthase ; genetics ; metabolism ; Nitric Oxide Synthase Type II ; genetics ; Nitric Oxide Synthase Type III ; genetics ; Osteopontin ; genetics ; physiology ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo study the relationship between the dynamic changes of caveolin-1 with the degrees of liver fibrosis and portal venous pressure (PVP) in the process of rat liver cirrhosis formation induced by dimethylnitrosamine (DMN); also to investigate the mechanisms of caveolin-1 in the regulation of endothelial nitric oxide synthase (eNOS).

METHODSLiver cirrhosis was induced in rats by DMN. The degrees of liver fibrosis and PVP were measured. NOS activity was assessed by citrulline generation. Protein expressions of caveolin-1, eNOS and caveolin-1-eNOS interactions were examined by Western blot and immunoprecipitation, respectively.

RESULTSFour weeks after DMN administration, liver fibrosis was at its peak and then decreased gradually. Immunoprecipitation and Western blot demonstrated that there was enhanced binding of caveolin-1 with eNOS in the process of rat liver cirrhosis. An increase in caveolin-1 expression was detected but the expression of eNOS was lower in cirrhotic tissues than in normal liver tissues. Caveolin-1 protein levels were positively correlated with the degrees of liver fibrosis and the levels of PVP (r=0.967, P < 0.01; r=0.922, P < 0.01, respectively), while NOS catalytic activity was negatively correlated with the degrees of liver fibrosis and levels of PVP (r= 0.973, P < 0.01; r=-0.947, P < 0.01) respectively.

CONCLUSIONSCaveolin-1 upregulation is associated with the development of portal hypertension in liver cirrhosis. Over-expression of caveolin-1 in perisinusoidal cells may promote caveolin-1-eNOS binding and reduce the activity of eNOS.

Animals ; Caveolin 1 ; metabolism ; Hypertension, Portal ; metabolism ; Liver Cirrhosis, Experimental ; metabolism ; Nitric Oxide Synthase Type III ; metabolism ; Rats ; Rats, Wistar

AIMTo investigate the effects of simulated microgravity on dilatory responsiveness and NOS expression of abdominal aorta in rats.

METHODSTwenty male healthy SD rats, which body weight ranged from 300 g to 330 g, were divided into control group and simulated microgravity group randomly. After 4 weeks, using isolated arterial rings from rats, arterial dilatory responsiveness of abdominal aorta were examined in vitro. And the expression of nitric oxide synthase (NOS), including endothelial NOS (eNOS) and inducible NOS (iNOS), were observed by Western blot.

RESULTSDilatory responses of arterial rings to L-Arginine (10(-8)-10(-3) mol/L), and Acetylcholine mol/L) were decreased in simulated microgravity rats compared with that of controls; but dilatory responses of isolated aortic rings to sodium nitroprusside (mol/L) and 8-bromo-cGMP(mol/L) were similar in both simulated microgravity rats and control rats. The expression of both eNOS and iNOS had not showed significant differences between two groups.

CONCLUSIONThe data indicate that endothelium dependent vasorelaxation in abdominal aortic rings are decreased by 4-week simulated microgravity, and this change may be result from altered NOS activity in endothelium.

Animals ; Aorta, Abdominal ; metabolism ; Arginine ; metabolism ; Cyclic GMP ; metabolism ; Male ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase Type II ; metabolism ; Nitric Oxide Synthase Type III ; metabolism ; Rats ; Rats, Sprague-Dawley ; Weightlessness Simulation

OBJECTIVETo explore the expressions of endothelial nitric oxide synthase (eNOS) and cytochrome P450 (aromatase) in the testis of sexually mature male SD rats and their significance.

METHODSEighteen male SD rats, 6 five-week, 6 seven-week and 6 ten-week old, were selected for this study. Paraffin sections of the left testis were made and the expressions of eNOS and P450 observed by the immunohistochemical ABC method.

RESULTSPositive expressions of eNOS and P450 were found to be + + +, + and + + in the Leydig cells of the five-week, seven-week and ten-week old rats, respectively, and they were also observed in a few spermatocytes, though with no regularity.

CONCLUSIONIn the Leydig cells of sexually mature male SD rats, eNOS and P450 are differently expressed in different stages of sexual maturation, and they are correlated as well.

Animals ; Aromatase ; metabolism ; Male ; Nitric Oxide Synthase Type III ; metabolism ; Rats ; Rats, Sprague-Dawley ; Sexual Maturation ; Testis ; metabolism