Somatic cells such as oviduct epithelial cell, uterine epithelial cell, cumulus-granulosa cell and buffalo rat river cell has been used to establish an effective culture system for bovine embryos produced in in vitro. But nitric oxide (NO) metabolites secreted from somatic cells were largely arrested the development of bovine in vitro matured/ in vitro fertilized (IVM/IVF) embryos, suggesting that NO was induced the embryonic toxic substance into culture medium. The objective of this study was to investigate whether BOEC co-culture system can ameliorate the NO-mediated oxidative stress in the culture of bovine IVM/IVF embryos. Therefore, we evaluated the developmental rate of bovine IVM/IVF embryos under BOEC co-culture system in the presence or absence of sodium nitroprusside (SNP), as a NO donor, and also detected the expression of growth factor (TGF-p , EGF and IGFBP) and apoptosis (Caspase-3, Bax and Bcl-2) genes. The supplement of SNP over 5 uM was strongly inhibited blastocyst development of bovine IVM/IVF embryos than in control and 1 uM SNP group (Table 2). The developmental rates beyond morulae stages of bovine IVM/IVF embryos co-cultured with BOEC regardless of SNP supplement (40.4% in 5 uM SNP+ BOEC group and 65.1% in BOEC group) were significantly increased than those of control (35.0%) and SNP single treatment group (23.3%, p<0.05: Table 3). The transcripts of Bax and Caspase-3 genes were detected in all experiment groups (1:Isolated fresh cell (IFC), 2:Primary culture cell (PCC), 3:PCC after using the embryo culture, 4: PCC containing 5 uM SNP and 5: PCC containing 5 uM SNP after using the embryo culture), but Bcl-2 gene was not detected in IFC and PCC (Fig. 1). In the expression of growth factor genes, TGF-p gene was found in all experimental groups, and EGF and IGFBP genes were not found in IFC and PCC (Fig. 2). These results indicate that BOEC co-culture system can increase the development beyond morula stages of bovine IVM/IVF embryos, possibly suggesting the alleviation of embryonic toxic substance like nitric oxide.

In the present study, we have demonstrated that a novel synthetic chemical JSH-21 of N¹-Benzyl-4-methylbenzene-1,2-diamine could inhibit nitric oxide (NO) production in lipopolysaccharide (LPS)-activated macrophages RAW 264.7. The JSH-21 showed an IC50 value of 9.2 uM on the LPS-induced NO production. Furthermore, JSH-21 attenuated LPS-induced mRNA and protein levels of inducible NO synthase (iNOS) in the cells, as well as inhibited LPS-induced iNOS promoter activity. These results indicates hat the compound could down-regulate iNOS expression at the transcription level. Since NF-kB activation is a key mechanism in the expression of LPS-inducible iNOS gene, we further examined whether JSH-21 could affect LPS-induced NF-kB activation. JSH-21 inhibited LPS-induced nuclear import of NF-kB p65 in macrophages RAW 264.7 and sequentially prevented NF-kB transcriptional activity. However, JSH-21 was not effective in a LPS-induced degradation of cytoplasmic IkB-alpha in the cells. These results suggest that JSH-21 could inhibit LPS-induced NF-kB activation targeting nuclear import of NF-kB, an event downstream IkB degradation. Taken together, this study may provide a pharmacological potential of JSH-21 in the NO- or NF-kB-associated inflammatory disorders.

We have examined the effect of NO donor, S-nitl‘ oso-N-acetyl-DL-penicillamine(SNAP) on heme oxygenase-1 (HQ-l) ex pression in human oral immortalized & malignant keratinocytes, and investigated in the control of keratinocyte proliferation evidence tha t HO-1 cou ld be involved in a low dose of NO, NO inhibitor, HOinducer, and HO inhibitor medi ated cytoprotect ion against cytotoxi city induced by a high dose of NO Oral keratinocyte growth inhibitory or anti-proliferative effects were exerted by with SNAP and hemin in a dose- and cul tivation time dependent manner The level of HQ-1 protein was increased in all cell types after exposure hernin dose, and the hemin induced HQ-1 protein achieved at higher maximum level by 12 hrs in all kind of cells , The pretreatment of cells with 0, 2 μ M SNAP reduced 1 mM SNAP-induced death in IHOK and HN4 cells , These cytoprotective effects on high dose of NO induced HQ-1 expresion and cell ular toxicity were blocked by low dose of SNAP, HCB, and ZnPP IX supporting the involvement of HQ-1 in high dose NO induced growth arrest or cell death, But these cytoprotection pattern is different from immortalized and malignant keratinocytes , These results indirectly demonstrate that HQ-1 could be involved in cytoprotection by NO priming against high dose NO induced cytotoxicity in immortalized and maigla nt oral keratinocytes, Thus, HQ-1 might be an important cellular target of NO donor, with clinical implications for the pre vention of inJlammatory di seases and anti-tumor immunity

Nitric oxide(NO) is a labile, uncharged, reactive radical that functions as a sensitive mediator of intercellular communication in diverse tissues. It has been reported that NO is produced by osteoblast and these results may suggest that NO is integrally involved in the regulation of osteoclast formation and osteoclast resorption activity by osteoblastic cells. We examined the effect of cytokines on NO release by mouse bone marrow cell. We also examined the effects of cytokines and sodium nitroprusside(SNP) on the formation of osteoclast-like cell from mouse bone marrow cells in culture. Cytokines stimulated NO production of mouse bone marrow cells, and N-nitro-L-arginine methyl ester, a specific inhibitor of NO synthase, suppressed the cytokine-induced NO production. SNP showed dual action in the generation of osteoclasts. The addition of (30μM)SNP inhibited the formation of tartrate resistant acid phosphatase(TRAP)(+) multinucleated cell, whereas lower concentration(30μM) of SNP enhanced it. Although the precise action of NO remains to be elucidated in detail, the action of NO in osteoclast generation in our studies seems to be associated, at least in part, with bone metabolism and bone pathophysiology.

Inflammation in the brain has known to be associated with the development of a various neurologiacal diseases. The hallmark of neuro-inflammation is the activation of microglia, brain macrophage. Pro-inflammatory compounds including nitric oxide(NO) are the main cause of neuro-degenerative disease such as Alzheimer"s disease. In the study, we examined whether Harmonia axyridis extracts inhibit the NO production by a direct method using Griess reagent, western blotting and by RT-PCR(Reverse Transcription-Polymerase Chain Reactionin) the gene expression of inducible nitric oxide synthase(iNOS). Distilled water(H₂O) and methanol(MeOH) extracts of H. axyridis inhibited the protein expression of TNF-α(Tumor Necrosis Factor) and IL-6(Interleukin) in LPS (Lipo-polysaccharide) stimulated BV -2 cells at the concentration of 100 ng/㎖. Incubation of BV-2 cells with the extracts of H₂O of MeOH inhibited the LPS induced NO and iNOS protein. And this inhibition of iNOS protein is concordant with the inhibition of iNOS mRNA expression. These data suggested that H. axyridis extracts may play a crucial role in inhibiting the NO production.

Nitric oxide (NO) has been known to inf1uence cell fate through apoptotic or necrotic cell death. Here, we investigated the role of nitric oxide on the growth and viability of immortalized human salivary gland (HSG) cells 띠 vitro. Treatrnent of HSG with a NO donor, S-nitroso-N-acetyl-DL-penκi1lamine (SNAP), significantly diminished the growth rate of HSG in a concentration dependent manner. However, this retardation of cell비ar growth rate was not corresponded to the apoptotic cell death of HSG cells, because there were no characteristic apopto디c features such as condensation of nuclear chromatin, nuclear fragmentation, and the apoptotic peak of propidium iodide (PI)-stained nuclei by flow cytome띠. 까ùs implies that HSG cells are resistant to NO-mediated 다π。to:잉city. 1n SNAP treated HSG cells, cell cycle analysis revealed that the number of G2/M phase increased markedly, according to while the percentage of cells in GO/Gl and S phases was not significantly affected. Otherwise, high concentrations of SNAP increased both P1 and annexin V positive cells. 1nterestingly, preincubation of HSG cells with iron chelator, deferoxamine (DFO), significantly diminished NO cytotoxicity more than when HSG cells are only incubated with SNAP which su잃.ests the role of iron homeostasis in NO-mediated cell death of HSG cells. 1n addi디。n ， treatrnent of HSG cells with SNAP specifically cleaved iron regulatory protein-2 (IRP2) while not affecting 1RP1. Collectively, the mπent results s멍gest that NO has a potential to control HSG cell growth through cell cycle arresting at G2/M phase. 1n addi디on ， intracellular iron homeostasis nùght play an important role in regulating cell survival of HSG cells

체외수정 후 체외배양48시간에 난구세포의 제거 유ㆍ무에 따라 CR/sub 1aa/ 배양액에 homoglobin을 0, 1, 5 ㎍/㎖를 첨가한 구의 상실배기 이상 체외발육성 적은 난구세포를 제거한 구에서 23.8%, 33.3% 및 26.8% 였으며, 난구세포를 제거하지 않은 구에서는 각각 39.5%, 54.8% 및 48.8%로서 난구세포를 제거하지 않은 구에 Hb를 1 ㎍/㎖를 첨가한 구가 여타구보다 통계적으로 높은 결과를 얻었다(P<0.05). 체외수정 후 체외배양 96시간 후 난구세포를 제거 유ㆍ무에 따라 Hb를 0, 1, 5 ㎍/㎖를 첨가하였을 때, 상실배기 이상 체외발육성적은 난구세포를 제거한 구에서 각각 28.6%, 46.2% 및 39.1%였으며, 난구세포를 제거하지 않은 구에서는 각각 33.9%, 67.2% 및 46.0%로서, 난구세포를 제거하지 않은 구에 Hb를 1 ㎍/㎖를 첨가구가 여타구에 비해 통계적으로 유의하게 높게 나타났다(P<0.05). CR/sub 1aa/ 배양액에 L-NAME를 0, 10, 50 및 100 mM을 첨가한 구에서 상실배 이상 발육된 체외발육성적은 각각 55.6%, 64.9%, 58.8% 및 66.7%로써 L-NAME 첨가구가 무첨가구에 비해 커다란 차이가 나타나지 않았다(P>0.05). 모든 처리구에 배반포까지 발육된 체외수정란의 세포수에는 커다란 차이가 인정되지 않았다.

Nitric oxide (NO) plays a key role in the processes of inflammation and carcinogenesis. Three isoforms of NO 야mthase have been identified: endothelial 띠띠c oxide 와nth앓e (NOS), neuronal NOS, and inducible NOS (이OS). The purpose of this study was to investigate the characteristics of iNOS expression in 7, 12-dimethylbenz[alanthracene (DMBA)-induced hamster buccal pouch carcinogenesis. Sixty three outbred young (6-week-old) male Syrian golden hamsters were randomly divided into three groups: DMBA treated group (n=57) and non-treated (n=3), and mineral-oil treated group (n=3). No iNOS activity could be detected in the untreated or mineral oil-treated pouches. 80th cytoplasmic and nuclear stainings were observed in the DMBA-treated pouch kera띠lCX까es. There were iNOS expression 외so in the strorna1 cells. The mean values of iNOS expression in the epithelium increased gradually from control to dysplastic lesions and more to invasive squ따nous cell carcinoma. πle clifference between iNOS expr'않sion in the normal and that the dysplastic and carcinomatous lesions is statistically significant. The mean values of iNOS expression in the stroma increased gradually from control to dysplastic lesions and more to invasive squamous cell carcinoma. The difference between iNOS expression in the normal and that the carcinomatous lesions is statistica11y si맑， ificant. In conclusion, this study has demonstrated that iNOS is expressed in DMBA-induced hamster pouch carcinomas. πlis finding suggests that iNOS expression may be associated with the development of chemically induced oral carcmomas.

In most tissues, apoptosis plays a pivotal role in normal development and in regulation of cell number. Therefore inappropriate apoptosis is revealed in a variety of diseases. This study was carried out to investigate the effects of acupuncture and needle electrode electrical stimulation on the change of caspase-3, 9 and neuronal nitric oxide synthase (nNOS) immunoreactive cells in the sprague dawley rats (SD rat). In immobilized SD rats (n=5), enhanced caspase-3 and caspase-9 expression were detected in the reticular part of substantia nigra, and enhanced nNOS was detected in the dorsolateral periaqueductal gray (DL-PAG) of midbrain and the paraventricular nucleus (PVN) of the hypothalamus using immunohistochemistry. Following the immobilization, acupuncture (n=5) and needle electrode electrical stimulation (n=5, 2 Hz) was applied at Hg (LI4) acupoint of SD rats, respectively. The stress-induced enhancement in the expression of caspase-3, 9 and nNOS were The present results demonstrate that and needle electrode electrical stimulation are effective in the modulation of expression of caspase-3, 9 and nNOS induced by immobilization.

The selective catalytic reduction(SCR) of nitric oxide by ethane in the presence of oxygen was investigated on Cu-ZSM-5, Co-ZSM-5 and Ga-ZSM-5 catalysts over a range of 400, 450 and 500℃. The catalysts were prepared by ion-exchange method. The composition of the reactant gases were 1000 ppm of NO, 1000 ppm of C2H6 and 2.5% of O2, and the reaction was conducted in a fixed-bed reactor at 1 atm. For the 20wt% Co-ZSM-5(50) catalyst, the NO conversion reached up to 100%, while the C2H6 conversion and the CO selectivity were about 50% and 25%, respectively, at 450℃. For the 20wt% Cu-ZSM-5(50) catalyst, the NO conversion and the C2H6 conversion were about 80% and 100%, respectively, but there was no CO produced. The metal ion-exchanged ZSM-5 catalysts exhibited a tendency to increase the NO conversion with the Si/Al ratio of the ZSM-5, that is, NO conversion was inversely proportional to the acidity of the catalysts. But, the effect of the acidity on NO conversion was not so large. From the XRD results of the catalysts before and after SCR reaction it was found that there was no structural change.