OBJECTIVETo study the effect on synthesis of nitric oxide in myocardium by local cryoablation and to investigate its mechanism.

METHODSMyocardium was cryoablated locally by a probe cooled to -60 degrees C and rewarmed by normal salt solution, nitric oxide and its synthesis enzyme were measured before and after cryoablation. L-arginine or methylene blue was added before and during cryoablation and the effect of these drugs on synthesis of nitric oxide was studied.

RESULTSNitric oxide and its synthesis enzyme decreased after cryoablation; L-arginine preserved the synthesis of nitric oxide and methylene blue inhibited the synthesis of nitric oxide. However, nitric oxide in serum did not change.

CONCLUSIONNitric oxide and its synthesis enzyme in myocardium decrease after cryoablation.

Animals ; Cryosurgery ; Myocardium ; metabolism ; pathology ; Nitric Oxide ; biosynthesis ; Nitric Oxide Synthase ; metabolism ; Rabbits

OBJECTIVESTo determine the expression of nitric oxide synthase (NOS) in testis and to investigate the effects of NO on the reproductive function of testis.

METHODSTestes of adult male Sprague-Dawley rats were fixed in 4% paraformaldehyde. The paraffin sections were made as routine. Immunohistochemical ABC method was used to observe the localization of NOS.

RESULTSEndothelia NOS (eNOS), neuronal NOS (nNOS) and inductive NOS (iNOS) were all expressed in Leydig cells. Only eNOS was expressed in peritubular myoid cells, endothelial and smooth muscle cells of blood vessel, while only nNOS expressed in tunica adventitia of testicular blood vessels. The reactive substance distributes in cytoplasm with negative nuclei. Immunoreactivity for eNOS, nNOS and iNOS in all spermatogenic cells was negative.

CONCLUSIONSThree kinds of NOS were all expressed in testis and the distribution of different NOS had a little difference.

Animals ; Immunohistochemistry ; Male ; Nitric Oxide ; biosynthesis ; Nitric Oxide Synthase ; analysis ; Rats ; Rats, Sprague-Dawley ; Testis ; enzymology

The efflux of nitro oxide (NO) in the duration of storing red blood cells (RBCs) was the main reason resulting in decrease and even loss of vasodilatory activity, cell deformability and ability of carrying oxygen (O2) in the stored RBCs. The deep understanding physical functions and acting ways of NO in circulatory system, as well as transformations and balance control of S-Nitrosohemoglobin (SNO-Hb) has an important significance for ensuring sure safety and efficacy of transfusion. In this article, the physical functions, acting ways, retaining and transferring form of nitro oxide, and SNO-Hb adjusting, as well as effects of SNO-Hb concentration on change on stored red blood cells were reviewed.
Erythrocytes ; metabolism ; physiology ; Hemoglobins ; biosynthesis ; Humans ; Nitric Oxide ; metabolism

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

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 effect of herbal compound 861 (HB861) on expression and activity of nitric oxide synthase (NOS) in hepatic stellate cells (HSC), and to explore the feasibility of its application in preventing and treating the early portal hypertension.

METHODSHSC of HSC-T6 cell line (1 x 10(5)/ml) were cultured in dish with 95% O2 plus 5% CO2 under 37 degrees C for 24 hrs, then divided into 5 groups, 6 dishes in each group. Group A was the blank control group. To Group B-E, HB861 2 mg/ml, HB861 4 mg/ml, HB861 8 mg/ml, HB861 4 mg/ml + NW-Nitro-L-Arginine Methyl Ester (L-NAME)4 mg/ml were added separately, and continuously cultured for 24 hrs. NOS activity was measured using colorimetry, NO level was determined by nitrate reductase technique. The cells were fixed by 4% paraformaldehyde for 2 hrs for test HSC-T6 iNOS expression by immunocyto-chemical method.

RESULTSHB861 in 2 mg/ml, 4 mg/ml and 8 mg/ml could increase HSC-T6 NOS activity from 1.7 +/- 0.1 to 2.5 +/- 0.3, 3.5 +/- 0.4 and 3.7 +/- 0.9 respectively (P < 0.01), the NO levels in supernatant were increased in parallel from 56.1 +/- 4.8 to 90.7 +/- 4.6, 99.7 +/- 4.1 and 109.0 +/- 2.7 respectively (P < 0.01). L-NAME could not inhibit the effect of HB861 in increasing the synthesis and secretion of NO by activated HSC-T6. Immuno-cyto-chemical study showed that there was iNOS expression in cytoplasm, and which could be increased by HB861.

CONCLUSIONThe activated HSC-T6 showed positive iNOS expression, suggesting it could produce NO. HB861 could markedly increase HSC-T6 iNOS expression and NOS activity, enhance the NO synthesis and secretion, it also could inhibit the contractility of activated HSC by way of increase HSC to secrete NO, so as to lower the resistance in hepatic sinusoid, therefore would play important role in preventing and treating of early portal hypertension.

Animals ; Cells, Cultured ; Drug Combinations ; Drugs, Chinese Herbal ; pharmacology ; Hepatocytes ; enzymology ; Hypertension, Portal ; prevention & control ; Nitric Oxide ; biosynthesis ; Nitric Oxide Synthase ; biosynthesis ; metabolism ; Rats ; Rats, Sprague-Dawley

OBJECTIVESTo investigate the cause of spermatogenic cells decreasing after spinal cord injury.

METHODSImmunohistochemical S-P method was used to detect the expression of iNOS and Bcl-2 protein in the testis of spinal cord injured and sham-operated adult rats in second week and fourth week after operation.

RESULTSSecond week and fourth week after operation, the expression of iNOS of operated rats increased significantly (P < 0.05). Fourth week after operation, the number of Bcl-2 positive cells in spinal cord injured group decreased significantly (P < 0.05).

CONCLUSIONSThe expression veition of iNOS and Bcl-2 is the cause that inducing the decrease of spermatogenic cells in testis of spinal cord injured rats.

Animals ; Male ; Nitric Oxide Synthase ; biosynthesis ; genetics ; Nitric Oxide Synthase Type II ; Proto-Oncogene Proteins c-bcl-2 ; biosynthesis ; genetics ; Rats ; Rats, Sprague-Dawley ; Spinal Cord Injuries ; metabolism

OBJECTIVETo detect the protective effect of ligustrazine hydrochloride on homocysteine-injured ECV304 cells.

METHODIn the in vitro study, human umbilical vein endothelial cells were selected as objects, with homocysteine as the molding agent, to judge the injury degree by monitoring NOS and NO contents. Based on that, the best homocysteine concentration in ECV304 cells, the best reaction time could be determined, and an endothelial cell injury model was established. After adding ligustrazine hydrochloride, NOS and NO contents in injured endothelial cells were determined to observe the protective effect of ligustrazine hydrochloride.

RESULTIt was proved that the optimal concentration of homocysteine on injured ECV304 cell was 1 mmol x L(-1), the best reaction time was 48 h. An injured endothelial cell model was established. At the same time, positive drug nitroglycerin and ligustrazine hydrochloride displayed a protection effect on injured ECV304 cells, NOS and NO formation were significantly increased compared with the model group.

CONCLUSIONLigustrazine hydrochloride has a protective effect on homocysteine-injured ECV304 cells. The model established in this study can be used to screen anti-myocardial ischemia drugs targeting at an endothelial cell protective agent.

Cytoprotection ; drug effects ; Homocysteine ; adverse effects ; Human Umbilical Vein Endothelial Cells ; cytology ; drug effects ; metabolism ; Humans ; Nitric Oxide ; biosynthesis ; Nitric Oxide Synthase ; biosynthesis ; Pyrazines ; pharmacology

To explore the effects of bilirubin on alveolar macrophages (AM) and expression of iNOS and NO in them in emphysema model, the rats were pretreated with bilirubin before exposed to smoke. AM were isolated from bronchoalveolar lavage fluid (BALF) and cultured. Pathological microscopic examination of AM and immunohistochemical analysis of iNOS were performed. Nitric oxide (NO) content in the samples was determined by nitrate reductase technique. The results showed both alveoli and alveolar septum appeared normal in size and shape in normal group. AM showed kidney-shaped nucleus and were rich in Golgi complexes and primary lysosomes in the cytoplasm. The inner membrane of mitochondrion was continuous. Most cristae of the mitochondria were intact. In model group, the alveoli were expanded, ruptured and bullaes were formed. Both the population and sizes of AM increased significantly. Secondary lysosomes were rich in the cytoplasm. Deformation and pyknosis of the nucleus, swelling of the mitochondrions and rupture of the inner mitochondrial membrane could also be seen. At high magnification, most of the mitochondrial cristae were broken, or completely lost at certain points. In bilirubin group, alveoli partly expanded and the population of AM also increased, with morphological changes being slighter than that in model group. Both NO contents and expression of iNOS in model group were higher than those in normal group (P<0.05). In bilirubin group the two indice were lower than those in model group (P<0.05). Our findings suggested that high expression of iNOS and high NO content in AM accelerate the development of emphysema associated with smoking in rats. Bilirubin may exert protective effects on AM and retards the development of emphysema in rats.
Animals ; Antioxidants ; pharmacology ; Bilirubin ; pharmacology ; Cells, Cultured ; Emphysema ; metabolism ; pathology ; Female ; Macrophages, Alveolar ; drug effects ; metabolism ; pathology ; Male ; Nitric Oxide ; biosynthesis ; Nitric Oxide Synthase ; biosynthesis ; Nitric Oxide Synthase Type II ; Rats ; Rats, Wistar