诱导型一氧化氮合酶与疾病

迟至 2 0世纪 80年代末 ,一氧化氮 (ｎｉｔｒｉｃｏｘｉｄｅ ,ＮＯ)才被认识到是人体内一种生物学活性分子。人体内的ＮＯ有两个来源。一为非酶生 (ｎｏｎ -ｅｎ ｚｙｍｓｔｉｇｅｎｅｓｅ) ,来自体表或者摄入的无机氮的化学降解 /转化 ;一为酶生 (ｅｎｚｙｍｓｔｉｇｅｎｅｓｅ) ,由ＮＯ合酶(ＮＯｓｙｎｔｈａｓｅＮＯＳ)所合成。现已认识到ＮＯ既是一个有细胞毒性效应器作用的分子 ,是组织损伤的诱发因子和各种病变的增强因子 ,有其病理上不好的一面 ;又是生物体许多部分的信号 (ｓｉｇｎａｌ)分子 ,是先天性免疫应答的调节性效应分子 ,作…     

Interleukin-2 receptor expression in benign and malignant melanocytic skin lesions

The immunophenotype of the lymphocytic reaction of melanocytic skin lesions (12 cases of cutaneous malignant melanoma (CMM), three of Hutchinson's melanotic freckle (HMF), and eight naevocellular naevi) has been studied using monoclonal antibodies. CMM was associated with the most intense lymphocytic reaction, the lymphocytes being T cells of both CD-4 T helper/inducer and CD-8 suppressor/cytotoxic subsets which were present in varying proportions. Although less marked, the lymphocytic reaction to HMF and benign naevi showed similar features. An antibody to the interleukin-2 receptor (IL-2R; Tac, CD-25) was included in the panel and the earlier findings of positively stained cells in association with CMM were confirmed. In addition, the novel finding of these cells in association with HMF and naevocellular naevi is reported. The number of these CD-25 positive cells was extremely variable but they appeared most prominent in association with CMM. The findings presented here indicate that it is not possible to infer that CD-25 positive lymphocytes present in the host response to CMM necessarily indicates tumour specific activation of the cells of the host immune system.     

Inducible nitric oxide synthase expression in human urinary bladder cancer

Nitric oxide (NO) is generated by a family of enzymes, nitric oxide synthases (NOS), in a wide range of mammalian cells. NO produced by the inducible NOS isoform (iNOS) has been suggested to play an important role in tumor biology with both tumor promoter and anti-tumor activity. Here, the cellular localization of iNOS in tissue of 100 cases of urinary bladder cancer was assessed immunohistologically using a commercially available antiserum. Positive iNOS immunostaining was detected in all samples of tumor tissue, whereas nonmalignant tissue adjacent to malignant areas did not show any iNOS positivity. The tumor tissue revealed a highly inhomogeneous staining pattern. In addition to uniformly stained tumor specimens, we also found markedly iNOS-positive tumor islets in the midst of unstained tumor tissue and scattered individual tumor cells expressing marked staining. In some cases, the tumor tissue showed no or only weak staining intensity. In some instances, the superficial epithelial layer of papillary carcinomas was extremely immunoreactive, in other cases it was not. Thus we were unable to show a clear correlation to tumor grade or stage. Further studies with a diversity of tumor markers including molecular genetics techniques will be necessary to elucidate how and to what extent NO and bladder cancer of different grades and stages are functionally interrelated.     

Inducible nitric oxide synthase in human diseases

Oestrogen has the capacity to suppress T cell-dependent DTH. To explore the mechanisms whereby oestrogen exerts its effects on the immune system we have used SCID mice which are largely devoid of functional T and B lymphocytes, hence being unable to raise DTH, but display intact antigen-presenting capacity. Transfer of lymphocytes to SCID mice restores the DTH capacity. In order to analyse if oestrogen down-regulates DTH by a direct action on T cells we reconstituted SCID mice with either splenocytes or thymocytes from congenic C.B-17 or allogeneic B6 donor mice. Either donor or recipient mice were exposed to estradiol before cell transfer. DTH response was registered in recipient SCID mice 1 and 3 weeks after challenge with oxazolone (OXA). SCID mice receiving estradiol-exposed spleen cells from congenic or allogeneic donor mice displayed lower DTH responses compared with control mice. In contrast, SCID mice receiving estradiol-exposed thymocytes from congenic donor mice showed no significant difference in DTH response compared with control mice. Estradiol-treated SCID mice, transferred with either spleen cells or thymocytes from congenic, hormonally non-treated donors, displayed a significantly lower DTH response compared with control mice. In contrast, estradiol-treated SCID mice receiving hormonally non-treated allogeneic spleen cells showed no difference in DTH response compared with control mice. The results show that T lymphocytes are not the target cell population for estradiol-mediated suppression of DTH in reconstituted female SCID mice.     

Inducible nitric oxide synthase expression and its prognostic significance in colorectal cancer

Zafirellis K, Zachaki A, Agrogiannis G, Gravani K. Inducible nitric oxide synthase expression and its prognostic significance in colorectal cancer. APMIS 2010; 118: 115–24. Nitric oxide synthases (NOS) are expressed in colorectal cancer. The aim of this study was to examine the inducible NOS (iNOS) expression in colorectal cancer and to investigate its prognostic relevance. Tissue sections of primary tumors from 132 patients undergoing curative resection for colorectal cancer were immunohistochemically examined for iNOS expression. The expression pattern of iNOS was correlated with various clinicopathological characteristics and survival. iNOS immunoreactivity was observed in the cytoplasm of tumor epithelial cells in 60 patients (45.5%) and positively correlated with lymph node involvement (p = 0.019). No significant correlation was found between iNOS expression and various clinicopathological characteristics, including age, gender, tumor location, tumor size, tumor grade, T stage, and Union International Contra la Cancrum (UICC) stage. Survival analysis showed a significant correlation between iNOS‐positive tumors and poor disease‐specific survival (p < 0.0001), with independent prognostic significance in multivariate analysis (HR = 4.42; p < 0.0001). Patients with stage II disease and iNOS‐positive tumors had significantly worse disease‐specific survival than those with iNOS‐negative tumors (p < 0.0001). In addition, patients with stage III disease and iNOS‐positive tumors had significantly worse disease‐specific survival than those with iNOS‐negative tumors (p = 0.001). The ability of iNOS to predict outcome in colorectal cancer patients may be independent of other known prognostic factors, providing a new molecular marker with significant potential for clinical utility.     

p16ink4a expression in benign and malignant melanocytic conjunctival lesions

Acquired conjunctival melanocytic lesions include nevi, primary acquired melanoses (PAMs), and melanomas. Conjunctival melanoma is a malignant melanocytic neoplasm with a high metastasis and mortality rate. Usually, the diagnosis can be achieved only with routine microscopic analysis, but in some cases, the samples are small or have artifacts. In these cases, complementary studies will be helpful, but currently, there are no well-understood or studied complementary methods. Objective. To analyze the immunohistochemical expression of p16 in conjunctival melanocytic lesions and to assess its potential for differentiating between benign and malignant melanocytic lesions. Methods. Immunohistochemical study against p16ink4a (p16) was performed on paraffin-embedded sections on 45 melanocytic lesions (9 melanomas, 19 nevi, and 2 PAMs with atypia and 15 without atypia). Expression was scored according to the German immunoreactive score (IRS). Results. Expression of p16 IRS differed between nevi, PAMs, and melanomas. The mean IRS for melanomas was 3.3 ± 1.8 and was lower than those for nevi (7.63 ± 3.24; P < .05), PAM with atypia (12 ± 0; P < .05), and PAM without atypia (11 ± 1.69; P < .05). Lesions with infiltration depths lower than 2 mm showed higher levels of p16. There were no differences between favorable and unfavorable locations. Conclusion. p16 Expression in conjunctival melanocytic lesions showed an expression similar to that in skin and seems to be a good marker to differentiate nevi and PAMs from melanomas. However, additional studies of larger series and follow-up are needed to confirm these findings.     

诱导型一氧化氮合酶与动脉血压的调节

目的:探讨诱导型一氧化氮合酶在血液动力学调控中的作用。方法:本实验所用动物为健康、雄性、Dahl 盐敏感(DS)及Dahl盐不敏感(DR)大鼠,体重(190±4) g。通过慢性血液动力学实验观察静脉输入诱导型一氧化氮合酶(iNOS)抑制剂AG对大鼠平均动脉压的影响。此外,本实验还测定了一氧化氮(NO)终产物NO₂^-及NO₃^-含量以及钙依赖及钙不依赖的NOS活性以了解iNOS的功能。结果:(1)iNOS特异抑制剂AG对正常血压(包括高盐或低盐输入的Dahl盐抵抗大鼠及低盐输入的Dahl盐敏感大鼠)没有明显影响,但能明显放大高钠引起的DS大鼠的血压上升效应;(2)高盐在导致的DS大鼠血压升高的同时,能引起iNOS活性的明显升高。结论:iNOS是参与血液动力学调控的重要组成部分,尤其是在血压处于较高水平时,iNOS具有不容忽视的调节作用。     

Correlated expression of inducible nitric oxide synthase and P53, bax in benign and malignant diseased gallbladder

Our goal has been to investigate the expression and correlated significance of inducible nitric oxide synthase (iNOS) and P53, Bax in benign and malignant gallbladder diseases. We detected the expression of iNOS, P53 and Bax in the gallbladder wall by SP immunohistochemistry in 16 cases of chronic cholecystitis, 11 cases of chronic cholecystitis with adenomyoma and 24 cases of gallbladder adenocarcinoma. The percentage of positively marked tumor cells was counted under microscope and the intensity of immunoreactivity was graded. SPSS10.0 statistical software was applied for statistical analysis. In this study, we found that: (1) Both benign and malignant diseased gallbladder wall expressed iNOS and Bax. Compared to benign diseased gallbladders, their expression in adenocarcinoma was decreased (p < 0.05), P53 was expressed strongly only in nuclei of adenocarcinoma cells of some cases. (2) In benign and malignant diseased gall-bladders, iNOS expression was related positively to Bax (p < 0.01), the expression of P53 and Bax had a negative relationship (p < 0.01). The results suggested that both chronic cholecystitis and chronic cholecystitis with adenomyoma carry the risk of becoming malignant, especially the latter. NO is an important mediated molecule in cancer, there are intimate relationships between gallbladder cancer and apoptosis.     

E-cadherin/catenin complex in benign and malignant melanocytic lesions

E-cadherin is a calcium-dependent cell-cell adhesion molecule expressed by melanocytes and responsible for their adhesion to keratinocytes in vitro. In this study, the expression of E-cadherin and its associated cytoplasmic proteins alpha-, beta-, and gamma-catenin was evaluated in melanocytic lesions by immunohistochemistry. E-cadherin expression was evaluated in 70 malignant melanomas and the catenins in 35 of these specimens. Twenty benign melanocytic naevi were also evaluated for E-cadherin and catenin expression. In normal epidermis, E-cadherin/catenin immunostaining was localized at the intercellular borders. In melanomas, a differential loss of E-cadherin expression was observed. Membranous E-cadherin staining was absent in dermal nests of melanomas in their radial growth phase and in Clark level II and III lesions, whereas it was present in a high proportion of melanomas in their vertical growth phase, in Clark level IV and V lesions and in metastasizing melanomas. In contrast, superficial compartments of naevi showed membranous E-cadherin immunoreactivity and junctional naevus cell nests displayed heterogeneous or diffuse cytoplasmic staining. Cytoplasmic alpha- and beta-catenin, but not gamma-catenin staining were detected in all benign and malignant lesions. These findings indicate that qualitative changes in the expression and cellular localization of E-cadherin and of alpha-, beta-, and gamma-catenin occur in melanocytic lesions and may reflect different stages in their progression.     

Inducible nitric oxide synthase expression is increased in the brain in fatal cerebral malaria

AIMS: Nitric oxide (NO) has been hypothesized to play a major role in the pathogenesis of cerebral malaria caused by P. falciparum infection. NO may act as a local neuroactive mediator contributing to the coma of cerebral malaria (CM). We hypothesized that increased expression of inducible nitric oxide synthase (iNOS) may cause increased release of NO, and examined the expression and distribution of iNOS in the brain during CM. MATERIAL AND RESULTS: Brain tissues from fatal cases of cerebral malaria in Thai adults were examined using immunohistochemical staining to detect iNOS. The distribution and strength of staining was compared between 14 patients with CM, three of whom were recovering from coma, and controls. iNOS expression was found in endothelial cells, neurones, astrocytes and microglial cells in CM cases. There was also strong staining in macrophages surrounding ring haemorrhages. iNOS staining was decreased in recovering malaria cases compared to acute CM, and was low in controls. Quantification showed a significant association between the intensity and number of iNOS positive vessels with the severity of malaria related histopathological changes, although the total number of cells staining was not increased compared to recovering CM cases. CONCLUSIONS: This study indicates that an acute induction of iNOS expression occurs in the brain during CM. This occurs in a number of different cells types, and is increased in the acute phase of CM compared to cases recovering from coma. As NO may activate a number of secondary neuropathological mechanisms in the brain, including modulators of synaptic function, induction of iNOS expression in cerebral malaria may contribute to coma, seizures and death.     

Fatty acid synthase expression in cutaneous melanocytic neoplasms.

Mammalian fatty acid synthase is a multifunctional enzyme complex involved in de novo synthesis of saturated fatty acids, and inhibitors of fatty acid synthase are being evaluated as potential therapeutic agents. Increased fatty acid synthase expression has been demonstrated in subsets of malignancies, including colon, breast, endometrium, prostate and ovarian carcinomas, and recently malignant melanomas. We evaluated the immunohistochemical expression of fatty acid synthase in 155 cutaneous melanocytic lesions. They included 30 congenital nevi, 19 compound nevi, 40 Spitz nevi, 48 primary melanomas, and 18 metastatic melanomas. Fatty acid synthase expression was stronger in malignant melanomas in comparison to conventional nevi and Spitz nevi, and was the highest for metastatic melanoma. Of the primary malignant melanomas, mean fatty acid synthase scores were significantly greater for Clark levels IV and V compared to Clark levels I and II (P<0.001). In addition, melanomas with Breslow thickness 0.75-1.50 mm and >1.50 mm showed significantly higher mean fatty acid synthase scores compared with those with Breslow thickness <0.75 mm (P=0.013 and <0.001, respectively). Of interest, congenital melanocytic nevi also showed strong fatty acid synthase expression, similar to that seen in metastatic melanoma. This may represent persistence of or regression to a fetal phenotype since normal fetal tissues are known to express high levels of fatty acid synthase.     

Inducible nitric oxide synthase expression in human colorectal cancer: correlation with tumor angiogenesis

To investigate the potential involvement of the nitric oxide (NO) pathway in colorectal carcinogenesis, we correlated the expression and the activity of inducible nitric oxide synthase (iNOS) with the degree of tumor angiogenesis in human colorectal cancer. Tumor samples and adjacent normal mucosa were obtained from 46 surgical specimens. Immunohistochemical expression of iNOS, vascular endothelial growth factor (VEGF), and CD31 was analyzed on paraffin-embedded tissue sections. iNOS activity and cyclic GMP levels were assessed by specific biochemical assays. iNOS protein expression was determined by Western blot analysis. iNOS and VEGF mRNA levels were evaluated using Northern blot analysis. Both iNOS and VEGF expressions correlated significantly with intratumor microvessel density (r(s) = 0.31, P = 0.02 and r(s) = 0.67, P < 0.0001, respectively). A significant correlation was also found between iNOS and VEGF expression (P = 0.001). iNOS activity and cyclic GMP production were significantly higher in the cancer specimens than in the normal mucosa (P < 0.0001 and P < 0.0001, respectively), as well as in metastatic tumors than in nonmetastatic ones (P = 0.002 and P = 0.04, respectively). Western and Northern blot analyses confirmed the up-regulation of the iNOS protein and gene in the tumor specimens as compared with normal mucosa. NO seems to play a role in colorectal cancer growth by promoting tumor angiogenesis.     

Inducible nitric oxide synthase is expressed in synovial fluid granulocytes

The objective of the study was to evaluate the NO-producing potential of synovial fluid (SF) cells. SF from 15 patients with arthritis was compared with blood from the same individuals and with blood from 10 healthy controls. Cellular expression of inducible nitric oxide synthase (iNOS) was analysed by flow cytometry. High-performance liquid chromatography was used to measure l-arginine and l-citrulline. Nitrite and nitrate were measured colourimetrically utilizing the Griess' reaction. Compared to whole blood granulocytes in patients with chronic arthritis, a prominent iNOS expression was observed in SF granulocytes (P < 0.001). A slight, but statistically significant, increase in iNOS expression was also recorded in lymphocytes and monocytes from SF. l-arginine was elevated in SF compared to serum (257 +/- 78 versus 176 +/- 65 micro mol/l, P = 0.008), whereas a slight increase in l-citrulline (33 +/- 11 versus 26 +/- 9 micro mol/l), did not reach statistical significance. Great variations but no significant differences were observed comparing serum and SF levels of nitrite and nitrate, respectively, although the sum of nitrite and nitrate tended to be elevated in SF (19.2 +/- 20.7 versus 8.6 +/- 6.5 micro mol/l, P = 0.054). Synovial fluid leucocytes, in particular granulocytes, express iNOS and may thus contribute to intra-articular NO production in arthritis.     

Inducible and endothelial nitric oxide synthase expression in human atherogenesis: an immunohistochemical and ultrastructural study

BackgroundNitric oxide has been proven to play an important role in the maintenance of vascular tone and structure. Impairment of nitric oxide production is an early indicator of atherosclerosis, but not much is known about the real mechanisms underlying this phenomenon.MethodsIn the present study, immunocytochemical methods have been used to analyze the patterns of expression of endothelial nitric oxide synthase and inducible nitric oxide synthase proteins in healthy and atherosclerotic human aortae using both confocal laser scanning microscopy and electron microscopy.ResultsInduction of the expression of endothelial nitric oxide synthase and inducible nitric oxide synthase proteins was observed in smooth muscle cells of atherosclerotic human aortae. Altered nitric oxide synthase expression was reported in atheromatous plaques and in apparently normal vascular tissues adjacent to the lesions.ConclusionsOur data confirm and extend previous findings of a direct relationship between dysregulation of nitric oxide pathway and atherosclerosis, suggesting another possible mechanism by which nitric oxide synthase system abnormalities may promote vascular dysfunction during human atherogenesis. Changes in nitric oxide production might be the primary step in the development of atheroma.     

Significance of inducible nitric oxide synthase expression in benign and malignant breast epithelium: an immunohistochemical study of 151 cases

The role of calcium independent inducible nitric oxide synthase (iNOS) in breast carcinoma is controversial, and the implications of iNOS expression on prognosis are not known. In this study, we aimed to investigate the significance of immunohistochemical iNOS expression in 100 invasive ductal carcinomas. In addition, 11 normal breast tissues, 20 cases of usual ductal hyperplasias (UDHs) and 20 fibroadenomas were included. We found that 78% of malignant and 75% of benign cases showed iNOS immunoreactivity. However, the intensity and the quantity of iNOS expression were significantly higher in the cancer group when compared with benign breasts (P<0.001), suggesting a role of iNOS in breast carcinogenesis. We were unable to show a correlation between iNOS expression and tumor grade, axillary lymph node status, and estrogen receptor expression. In 50 axilla negative cases having 5–12 years follow-up, disease free survival (DFS) rate was significantly lower in cases showing strong iNOS expression (P=0.05). As strong iNOS expression was correlated with short DFS, we concluded that further studies would be necessary to elucidate if iNOS expression might be a useful prognostic marker in breast carcinoma, especially in the axilla negative group.Part of this study was presented at XXIVth International Congress of the International Academy of Pathology, Amsterdam, The Netherlands on 5–10 October 2002 as a poster presentation.     

Inducible and endothelial nitric oxide synthase genes polymorphism in inflammatory bowel disease

To assess the influence of inducible and endothelial nitric oxide synthase gene (NOS2A and NOS3) polymorphisms in susceptibility to Crohn's disease (CD) and ulcerative colitis (UC). A total of 505 inflammatory bowel disease (IBD) patients (221 with UC and 284 with CD) and 332 ethnically matched controls were studied. Patients and controls were genotyped by polymerase chain reaction -based techniques for a multiallelic (CCTTT)(n) repeat and biallelic marker (TAAA)(n) in the promoter region of the NOS2A gene and for a T/C polymorphism at position -786 in the promoter region and a polymorphism in exon 7(298Glu/Asp) of the NOS3 gene. There was not association between NOS2A and NOS3 genotypes, alleles or haplotypes frequencies with UC, CD and controls. Our data suggest that NOS2A and NOS3 polymorphisms do not play a major role in IBD predisposition.     

Expression of inducible nitric oxide synthase in skin lesions of patients with american cutaneous leishmaniasis

Cytokine-inducible (or type 2) nitric oxide synthase (iNOS) is indispensable for the resolution of Leishmania major or Leishmania donovani infections in mice. In contrast, little is known about the expression and function of iNOS in human leishmaniasis. Here, we show by immunohistological analysis of skin biopsies from Mexican patients with local (LCL) or diffuse (DCL) cutaneous leishmaniasis that the expression of iNOS was most prominent in LCL lesions with small numbers of parasites whereas lesions with a high parasite burden (LCL or DCL) contained considerably fewer iNOS-positive cells. This is the first study to suggest an antileishmanial function of iNOS in human Leishmania infections in vivo.