Immunohistochemical Expression Of iNOS Research Articles

Evening primrose oil attenuates oxidative stress, inflammation, fibrosis, apoptosis, and ultrastructural alterations induced by metanil yellow in the liver of rat: a histological, immunohistochemical, and biochemical study

ABSTRACT The food color metanil yellow (Myl) is hazardous to several body systems. Evening primrose oil (EPO) was reported to have anti-inflammatory and anti-oxidant properties. The present work investigated the impact of Myl on the hepatic structure and function of rats and evaluated the protective effect of EPO. Forty adult male rats were divided into four groups: control, EPO (5 g/kg/day), Myl (200 mg/kg/day), and EPO- Myl group. Myl significantly increased liver enzymes, advanced glycation end products (AGE), oxidative stress parameters, pro-inflammatory cytokines, nuclear factor kappa B (NF-κB), and inducible nitric oxide synthase (iNOS). Blood vessels in the liver were dilated and congested, with cellular infiltration around them and associated with fibrosis. The hepatocytes were vacuolated and had dark nuclei. The immunohistochemical expression of iNOS, glial fibrillary acidic protein (GFAP), and Bax was significantly elevated. Ultrastructurally, the hepatocytes showed lipid droplets, irregular condensed nuclei with widened perinuclear space, dilated rER, mitochondria with destructed cristae, and multiple vacuoles. Dilated congested blood sinusoids and collagen fiber bundles were seen between hepatocytes. Interestingly, these alterations were less pronounced in rats co-administrated with EPO and Myl. In conclusion, EPO can protect liver against the toxic effects of Myl due to its anti-inflammatory and anti-oxidant activities.

Ezetimibe alleviates acetic acid-induced ulcerative colitis in rats: targeting the Akt/NF-κB/STAT3/CXCL10 signaling axis.

Ulcerative colitis (UC) is a relapsing inflammatory health state posing significant worldwide problems. Ezetimibe is a cholesterol-lowering drug having anti-inflammatory and pleiotropic properties. Twenty-four rats were classified into four groups (n = 6). Group (I) was considered negative control. Acetic acid (AA) was instilled intrarectally in groups (II-IV). Group (II) was considered UC-control. Groups (III and IV) were orally treated with Ezetimibe (5 and 10 mg/kg/day; 14 days). AA installation resulted in severe macroscopic colonic lesions associated with elevations in the relative colon weight, the wet weight/length ratio and oxidative stress markers in the colorectum tissues. UC-control rats showed significantly elevated colorectal tissue CXCL10 and STAT3 gene expression. Akt, phosphorylated Akt, phosphorylated STAT3, TNF-α, IL-6 and NF-κB were expressively upregulated in the UC-control group. AA installation also resulted in significant histopathological alterations in the colorectum tissues of UC-control rats along with increasing the colorectal tissues' immunohistochemical iNOS expression. Collectively, these data suggest activation of the Akt/NF-κB/STAT3/CXCL10 signaling axis. Ezetimibe treatment significantly ameliorated all the aforementioned parameters. This is the first study to elucidate the modulatory actions of Ezetimibe against oxidative stress and inflammation associated with AA-induced UC in rats. Ezetimibe treatment mitigates UC via downregulation of the Akt/NF-κB/STAT3/CXCL10 signaling axis.

GRAPE SEED EXTRACT VERSUS CAPTOPRIL IN AMELIORATING 5-FLUOROURACIL-INDUCED NEPHROTOXICITY IN ADULT MALE ALBINO RATS: BIOCHEMICAL, HISTOLOGICAL AND IMMUNOHISTOCHEMICAL STUDY

Background: 5-Fluorouracil (5-FU) is one of the extensively used chemotherapeutic agents for different human malignancies. It has severe side effects and is considered a nephrotoxic agent. Captopril is one of the angiotensin-converting enzyme inhibitors (ACEI) used in treatment of hypertension and congestive heart failure. It is also an effective radical scavenger and antioxidant due to its free thiol group. Grape seeds extract is one of the effective radical scavengers with antioxidant, anti-proliferative and anti-inflammatory properties. Objective: The present study was designed to assess the probable protective role of captopril and grape seeds extract against 5-FU-induced nephrotoxicity in adult male albino rats. Materials and Methods: Forty eight adult male albino rats were divided into 8 equal groups; Group I kept as the control group, Group II (grape seeds consumed group), received aqueous grape seeds extract (500 mg/kg/day) orally by gastric tube. Group III (captopril group), injected intraperitoneally by captopril solution (60 mg/kg) once daily. Group IV (grape seeds & captopril), Group V (5-Fluorouracil group), injected intraperitoneally by 5-FU solution (20 mg/kg) once daily. Group VI (grape seeds/5-FU), Group VII (captopril/5-FU) and Group VIII (grape seeds & captopril/5-FU). Treatment was continued at the same time daily for 4 weeks. At the end of the experiment, final body weight, blood urea nitrogen (BUN), creatinine (Cr) and uric acid levels were measured. Also, specimens of right kidneys were taken for histological and immunohistochemical studies with anti-inducible nitric oxide synthase (iNOS). Results: 5-FU resulted in a significant reduction in final body weight and a significant elevation of (BUN), (Cr) and uric acid levels. Moreover, glomerular and tubular distortion, vacuolated epithelial lining, intraluminal acidophilic hyaline casts, congested peritubular capillaries, interstitial inflammatory infiltration, and increased fibrosis within the renal interstitium. The immunohistochemical expression of iNOS supported kidney impairment in the 5-FU group. Treatment with either grape seeds or captopril revealed some improvement in the final body weight, biochemical markers and histological changes. The best effect was encountered when grape seeds and captopril were combined. Conclusion: and Recommendation: Grape seeds and captopril succeeded in ameliorating 5-FU induced nephrotoxicity with a better effect by their combination. So, it is recommended to use grape seeds extract and captopril during chemotherapy with 5 fluorouracil to reduce its renal toxicity.

GRAPE SEED EXTRACT VERSUS CAPTOPRIL IN AMELIORATING 5-FLUOROURACIL-INDUCED NEPHROTOXICITY IN ADULT MALE ALBINO RATS: BIOCHEMICAL, HISTOLOGICAL AND IMMUNOHISTOCHEMICAL STUDY

Background: 5-Fluorouracil (5-FU) is one of the extensively used chemotherapeutic agents for different human malignancies. It has severe side effects and is considered a nephrotoxic agent. Captopril is one of the angiotensin-converting enzyme inhibitors (ACEI) used in treatment of hypertension and congestive heart failure. It is also an effective radical scavenger and antioxidant due to its free thiol group. Grape seeds extract is one of the effective radical scavengers with antioxidant, anti-proliferative and anti-inflammatory properties. Objective: The present study was designed to assess the probable protective role of captopril and grape seeds extract against 5-FU-induced nephrotoxicity in adult male albino rats. Materials and Methods: Forty eight adult male albino rats were divided into 8 equal groups; Group I kept as the control group, Group II (grape seeds consumed group), received aqueous grape seeds extract (500 mg/kg/day) orally by gastric tube. Group III (captopril group), injected intraperitoneally by captopril solution (60 mg/kg) once daily. Group IV (grape seeds & captopril), Group V (5-Fluorouracil group), injected intraperitoneally by 5-FU solution (20 mg/kg) once daily. Group VI (grape seeds/5-FU), Group VII (captopril/5-FU) and Group VIII (grape seeds & captopril/5-FU). Treatment was continued at the same time daily for 4 weeks. At the end of the experiment, final body weight, blood urea nitrogen (BUN), creatinine (Cr) and uric acid levels were measured. Also, specimens of right kidneys were taken for histological and immunohistochemical studies with anti-inducible nitric oxide synthase (iNOS). Results: 5-FU resulted in a significant reduction in final body weight and a significant elevation of (BUN), (Cr) and uric acid levels. Moreover, glomerular and tubular distortion, vacuolated epithelial lining, intraluminal acidophilic hyaline casts, congested peritubular capillaries, interstitial inflammatory infiltration, and increased fibrosis within the renal interstitium. The immunohistochemical expression of iNOS supported kidney impairment in the 5-FU group. Treatment with either grape seeds or captopril revealed some improvement in the final body weight, biochemical markers and histological changes. The best effect was encountered when grape seeds and captopril were combined. Conclusion: and Recommendation: Grape seeds and captopril succeeded in ameliorating 5-FU induced nephrotoxicity with a better effect by their combination. So, it is recommended to use grape seeds extract and captopril during chemotherapy with 5 fluorouracil to reduce its renal toxicity.

Protective Effect of D-Carvone against Dextran Sulfate Sodium Induced Ulcerative Colitis in Balb/c Mice and LPS Induced RAW Cells via the Inhibition of COX-2 and TNF-α.

Ulcerative colitis (UC) is an intractable ailment, in which may chronic inflammations/ulcerations may develop in the mucosal lining of the colon with multiple recurrences. Various drugs such as steroids, immunosuppressants, and antibiotics are extensively used to treat UC. The patients suffer from adverse effects of these advanced drugs. So, they need a harmless therapeutic agent from natural sources. The therapeutic D-carvone has an anti-inflammatory action against the investigational colon cancer models. Therefore, we analyzed the effect of D-carvone on dextran sulfate sodium (DSS) provoked colitis model in mice as follows: Group I: noncolitis healthy control mice; Group II: ulcerative colitis mice models; Group III: D-carvone (40 mg/kg) + ulcerative colitis models; Group IV: sulfasalazine (50 mg/kg) + ulcerative colitis models. On the 8th day, the experimental study was terminated and serum samples and colon tissues were processed for further analysis. The effect of D-carvone at different concentration was studied on the LPS challenged RAW 264.7 cell lines. The D-carvone (40 mg/kg) treatment maintained the colon length and decreased disease activity index (DAI) score in UC animals. The increased antioxidant enzymes status and decreased oxidative stress and pro-inflammatory markers were noted in the D-carvone (40 mg/ kg) + UC mice. Histopathological study of colon tissue of D-carvone (40 mg/kg) treated UC mice displayed less mucosal damage and improved crypt integrity and goblet cells compared with DSS only provoked mice. The im-munohistochemical expression of iNOS and COX-2 was drastically diminished in the D-carvone treated UC mice. D-carvone (40 mg/kg) treatment appreciably diminished the LPS provoked NO production and pro-inflammatory modulators in the RAW 264.7 macrophage cell lines. These findings proved that D-carvone has a potential therapeutic effect to prevent LPS induced inflammation in in vitro cells and chemically induced ulcerative colitis in vivo models.

The role and prognostic value of inducible nitric oxide synthase (iNOS) and interleukin-33 (IL-33) in serous and mucinous epithelial ovarian tumours.

Understanding different mechanisms contributing to the aggressive behaviour of epithelial ovarian cancer (EOC) is a large challenge. Interaction between inflammation, immunity and carcinogenesis occurs in different cancers; however, the potential roles of different molecules involved in these processes in relation to ovarian carcinogenesis were not fully investigated. Inducible nitric oxide synthase (iNOS) and interleukin-33 (IL-33) are implicated in carcinogenesis. iNOS is an NOS isoform that generates nitric oxide, which plays important roles in various stages of carcinogenesis. IL-33 is a cytokine implicated in modulation of anti-tumour immunity and tumour growth. This work aimed at studying the immunohistochemical expression of iNOS and IL-33 in serous and mucinous epithelial ovarian tumours to investigate their role and prognostic significance. Immunohistochemical expressions of iNOS and IL-33 were assessed in 90 patients with epithelial ovarian tumours (45 serous and 45 mucinous tumours, categorized as benign, borderline, and malignant tumours). iNOS and IL-33 showed significantly higher expressions in borderline and malignant serous and mucinous tumours compared to benign ones (p=0.0001). The differences between borderline and malignant tumours were statistically insignificant (p=0.2351&0.6321). iNOS showed significantly higher expression with increasing tumour grade in malignant mucinous tumours (p=0.0011). IL-33 showed significantly higher expression with increasing tumour grade in both malignant serous and mucinous tumours (p=0.0074 and 0.0007). Upregulation of iNOS and IL-33 expression in borderline and malignant epithelial ovarian tumours indicates their involvement in the development and progression of EOC, and their increased expression in less differentiated cancers suggests their association with poor prognosis in this category of tumours.

Analysis of immunohistochemical expression of inducible nitric oxide synthase for the evaluation of agonal time in forensic medicine.

Although establishing agony is crucial in forensic practice, the identification of specific signs indicative of a detailed duration of agony is however not of immediate execution. Nitric oxide (NO) is the most important messenger molecule in the modulation of vascular tone and it is produced during stress conditions by inducible nitric oxide synthase (iNOS), as occurs during agony. The aim of this study was to investigate the relationship between immunohistochemical expression of iNOS, and agonal time (T), defined as the interval between the onset of a hypoxic-ischemic injury and the death. INOS expression was evaluated by measuring the average of signal intensity (SI) from cytoplasm of 300 smooth muscle cells of sample of renal artery, performed by ImageJ software: high values of SI correspond to a low enzyme expression and vice versa. We aimed also to check if gender, age, type of death (violent or natural death), post mortem interval, and storage in cold chamber influenced SI. We assessed 50 autopsied cases, of which 28 violent and 22 natural deaths, with a well-known T in a range between 1 and 631min. Statistical analysis was performed to estimate the relationship between SI and the other variables. Results pointed out that only SI is related to T, and since data showed a bi-phase relationship between T and SI, we used a piecewise regression method for estimation of T as function of SI. The transition from the first to the second phase takes place at SI = 117.5 which corresponds to a T of 29.5min. In conclusion, the study demonstrates that iNOS is a good marker for estimating T and the final regression model can be used in many forensic activities.

Immunohistochemical expression of iNOS and eNOS in thyroid goiter

Searchable abstracts of presentations at key conferences in endocrinology ISSN 1470-3947 (print) | ISSN 1479-6848 (online)

Reappraisal of the Significance of Inducible Nitric Oxide Synthase in Colorectal Cancer

Backgrounds: Inducible nitric oxide synthase (iNOS), which produces high levels of nitric oxide (NO), is overexpressed in activated macrophages and some cancer cells. Although iNOS was thought to be involved in promoting colorectal cancer, contradictory reports supporting its tumoricidal effect exist. Methods: We first examined the iNOS enzyme activity in colorectal cancer tissue and immunohistochemical expression of iNOS in cancer cells and tumor-infiltrating macrophages. Then, association of iNOS activity or its protein expression was analyzed in relation to various clinicopathological covariates. Results: Four groups of patients were classified based on their iNOS expression status. Univariate and multivariate analyses showed that group 1 patients (low iNOS in both types of cells) and group 4 patients (high iNOS in both types of cells) had a shorter disease-free survival. Patients with extremely high or low iNOS enzyme activity tended to have a lower disease-free survival rate (p = 0.059). Conclusion: Macrophage/stroma-derived NO negatively regulates colorectal cancer development when cancer cells express low levels of iNOS, but might synergistically contribute to tumor progression in the presence of high levels of cancer-cell derived NO. The dual effects of NO should be considered in the design of anti-iNOS/NO therapy for colorectal cancer patients.

Expression of inducible nitric oxide synthase in carcinomas and sarcomas affecting the oral cavity.

Context:Inducible nitric oxide synthase (iNOS) is a cytoplasmic enzyme which plays a crucial role in the pathogenesis of oral carcinomas and sarcomas.Aims:The objective of this study was to analyze the immunohistochemical expression of iNOS in carcinomas and sarcomas affecting the oral cavity in order to understand the possible role of iNOS in their biologic behavior and to correlate iNOS expression with lymph node metastasis in carcinomas and sarcomas.Settings and Design:Patients, who attended the oral diagnosis department of Vinayaka Missions Sankarachariyar Dental College, were screened, for the purpose of the study. Besides these, paraffin-embedded tissue blocks were also retrieved from archives of the Oral and Maxillofacial Pathology Department. A total of 40 cases (20 carcinomas and 20 sarcomas) were selected for the study.Subjects and Methods:A total of 40 cases (20 carcinomas and 20 sarcomas) were selected for the study. Five apparently normal tissues were obtained from the tumor adjacent normal tissue to be used as a control. These were subjected to immunohistochemical staining using antibody to iNOS and evaluated.Statistical Analysis Used:The results were analyzed using the Chi-square test.Results:Among the 20 carcinomas 19 showed a positive immunoreactivity for iNOS and 1 case was negative. Among the 19 immunopositive iNOS cases of carcinomas, 15 cases showed positive lymph node metastasis. Among the sarcomas, positive immunoreactivity for iNOS was seen in 10 hard tissue sarcomas, while the remaining 10 soft tissue sarcomas were negative for iNOS expression. The results were analyzed using the Chi-square test.Conclusions:iNOS is a reliable marker for lymph node metastasis in carcinomas irrespective of the histologic grade. The high expression in carcinomas shows that the carcinomas elaborate more angiogenesis for growth compared with the sarcomas with the exception of hard tissue sarcomas.

Expression of inducible nitric oxide synthase in the epithelial linings of odontogenic keratocyst, dentigerous cyst and radicular cyst: a pathological insight.

Background:The present study is aimed at analyzing the expression of inducible nitric oxide synthase (iNOS) in the epithelial lining of odontogenic keratocyst (OKC), dentigerous cyst (DC), radicular cyst (RC) in order to understand the possible role of iNOS with special reference to its neoplastic nature and local aggressive of cysts.Aim:The primary aim of the following study is to analyze the immunohistochemical expression of iNOS and secondary aim is to compare the iNOS expression, pattern and intensity of staining among the epithelial linings of OKC, DC and RC.Materials and Methods:iNOS in the epithelial lining cells were analyzing in 10 OKC's, 10 DC's and 10 RC's using immunohistochemistry. The percentage of positive cells was assessed and presented as mean ± standard deviation. The correlation with respect to the intensity and percentage of staining within the epithelial linings of OKCs, DCs and RCs was carried out using (analysis of variance and Student's t-test) Chi-square test.Results:Staining intensity of iNOS portion was seen in the entire thickness of the epithelial linings of OKC, whereas in DC's only one case had entire thickness of the epithelial lining staining and in RC's none of the cases showed entire thickness of staining. On comparing the staining intensity of iNOS between OKC, DC and RC groups, using Chi-square test, there was a statistically significant difference between these groups (P < 0.01). On analyzing the immuno-reactivity of iNOS in OKC, DC and RC there was a positive variable expression iNOS between the cysts.Conclusion:iNOS was over expressed in OKCs when compared with DC and RC suggesting that iNOS may contribute to the aggressive behavior of OKC. This is yet another evidence to support that OKC is the neoplasm.

The Effects of L-Arginine on Liver Damage in Experimental Acute Cholestasis an Immunohistochemical Study

Obstructive jaundice damages critical functions in the liver. Nitric oxide modulation would influence liver damage induced by biliary obstruction, and little is known about it Acute cholestasis was induced by bile duct ligation (BDL) in two groups of male Sprague-Dawley rats. L-Arginine or serum physiologic was administered to treatment and control group. Histopathological and immunohistochemical iNOS expression was investigated in hepatic tissue. Plasma enzyme activities were increased in acute cholestasis, and that L-arginine treatment partially but significantly prevented the elevation of these markers of liver damage (P < .05). Also histopathology scoring showed that the liver injury was prevented and immunohistochemical iNOS activity was increased significantly in L-arginine group (P < .05). This study shows that, after 7 days of biliary obstruction, liver damage is well established and exogenous L-arginine treatment partially but significantly prevented the liver injury in acute cholestasis.

Protective effect of pyrrolidine dithiocarbamate on kidney tissue in streptozotocin-induced diabetic rats

Objective: In this study, we investigated protective effects of pyrrolidine dithiocarbamate (PDTC), which is an antioxidant and nuclear factor kappa B (NF-κB) inhibitor, on nephropathy in diabetic rat model. Materials and methods: Twenty-eight Sprague-Dawley rats were allocated into 3 groups. Control group (n=8) received no treatment; diabetes group (n=10) received single intraperitoneal (i.p.) injection of streptozotocin (STZ, 65 mg/kg) to induce experimental diabetes; and PDTC group (n=10) received i.p. 100 μL/day PDTC for a total of 10 weeks following diabetes induction with i.p. STZ injection. At the end of the study kidneys were excised from sacrificed rats, and glomerular and tubular changes were examined under light microscopy. Immunohistochemically, NF-κB (p65) and inducible nitric oxide synthase (iNOS) expression were evaluated in the renal cortex. Results: In diabetes group, immunohistochemical NF-κB expression was higher than control and PDTC groups (p<0.05). In PDTC group, NF-κB expression level was very similar to control group, but less than diabetes group (p<0.05). Immunohistochemical iNOS expression was less in control group. In PDTC group, iNOS expression was more intense than control group (p<0.05), but less than that in diabetes group (p<0.05). Conclusion: PDTC has a protective effect on renal injury secondary to diabetes and can be a treatment option for patients with diabetic nephropathy.

Effects of pneumonectomy on nitric oxide synthase expression and perivascular edema in the remaining lung of rats

Pneumonectomy is associated with high mortality and high rates of complications. Postpneumonectomy pulmonary edema is one of the leading causes of mortality. Little is known about its etiologic factors and its association with the inflammatory process. The purpose of the present study was to evaluate the role of pneumonectomy as a cause of pulmonary edema and its association with gas exchange, inflammation, nitric oxide synthase (NOS) expression and vasoconstriction. Forty-two non-specific pathogen-free Wistar rats were included in the study. Eleven animals died during or after the procedure, 21 were submitted to left pneumonectomy and 10 to sham operation. These animals were sacrificed after 48 or 72 h. Perivascular pulmonary edema was more intense in pneumonectomized rats at 72 h (P = 0.0131). Neutrophil density was lower after pneumonectomy in both groups (P = 0.0168). There was higher immunohistochemical expression of eNOS in the pneumonectomy group (P = 0.0208), but no statistically significant difference in the expression of iNOS. The lumen-wall ratio and pO(2)/FiO(2) ratio did not differ between the operated and sham groups after pneumonectomy. Left pneumonectomy caused perivascular pulmonary edema with no elevation of immunohistochemical expression of iNOS or neutrophil density, suggesting the absence of correlation with the inflammatory process or oxidative stress. The increased expression of eNOS may suggest an intrinsic production of NO without signs of vascular reactivity.

Overexpression of inducible nitric oxide synthase and accumulation of 8‐OHdG in nasopharyngeal carcinoma

Nitric oxide (NO), produced by inducible NO synthase (iNOS), has been suggested to cause oxidative stress, leading to 8-hydroxydeoxyguanosine (8-OHdG) accumulation and subsequent transversion mutation of DNA. The aim was to evaluate iNOS expression and the status of oxidative stress in nasopharyngeal carcinoma (NPC). Seventy-three cases of NPC were investigated to examine the immunohistochemical expression of iNOS, 8-OHdG and latent membrane protein-1 (LMP-1) and Epstein-Barr virus-encoded small RNA (EBER) expression using in situ hybridization. iNOS mRNA expression and p53 gene mutations were also assessed. Overexpression of iNOS, LMP-1 and EBER was observed in 62 (84.9%), 28 (38.4%) and 53 (72.6%) cases respectively. p53 gene mutation was found in 10 of 73 (13.7%) cases. Immunohistochemical iNOS expression was associated with the 8-OHdG labelling index, iNOS mRNA expression and p53 gene alteration (P < 0.0001, P = 0.016 and 0.0082 respectively). Our present findings suggest that the expression of iNOS induces oxidative stress in NPC. Although the presence of p53 mutation was associated with iNOS overexpression, the type of acid-base change of p53 was transition, but not transversion, which suggests that the p53 gene is not the direct target of DNA damage by 8-OHdG accumulation.

Expression of inducible nitric oxide (NO) synthase but not prevention by its gene ablation of hepatocarcinogenesis with fibrosis caused by a choline-deficient, l-amino acid-defined diet in rats and mice

Expression of inducible nitric oxide synthase (iNOS) and effects of iNOS gene ablation on the hepatocarcinogenesis associated with fibrosis caused by a choline-deficient, l-amino acid-defined (CDAA) diet, were examined in male F344 rats and C57BL/6J wild-type and iNOS−/− mice. Western blot, RT-PCR and immunohistochemical analyses revealed increased expression of iNOS protein and mRNA in the livers of rats and wild-type mice fed a CDAA diet for 12–80 weeks, associated with elevated serum NO x and liver nitrotyrosine levels. iNOS−/− mice demonstrated greater liver injury and fibrosis in the early stage than their wild-type counterparts, but this did not significantly affect the incidence and multiplicity of altered foci, adenomas and hepatocellular carcinomas in spite of immunohistochemical iNOS expression in these lesions. Results suggested no major determinant roles of the expressed iNOS in the development of liver tumors caused by the CDAA diet.

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.

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.

Immunohistochemical expression of cytokine induced nitric oxide synthase in colorectal carcinoma

Significance of expression of cytokine induced nitric oxide synthase (iNOS) in colorectal carcinoma remains controversial. The aim of the current study was to elucidate the significance of immunohistochemical expression of iNOS in colorectal carcinoma. The subjects were the 54 patients with colorectal carcinoma, including 13 with obstructing carcinoma. The immunohistochemical expression of iNOS was examined and the clinicopathological data were compared between iNOS-positive and iNOS-negative carcinomas. Of 54 colorectal carcinomas, there were 27 tumors with iNOS expression. The proportion of tumors occurring in the rectum was significantly lower in iNOS-positive carcinomas than in iNOS-negative carcinomas (p=0.028). The incidence of lymph node metastasis and lymphatic invasion in colorectal carcinomas with iNOS expression (63.0%, 17 out of 27 and 96.3%, 26 out of 27, respectively) were significantly more frequent than those in carcinomas without iNOS expression (29.6%, 8 out of 27 and 70.4%, 19 out of 27, respectively) (p=0.013 and p=0.011, respectively). The proportion of obstructing carcinoma in tumors with iNOS expression (37.0%, 10 out of 27) was significantly higher than that in tumors without iNOS expression (11.1%, 3 out of 27; p=0.020). Our results could suggest that iNOS expression can be an indicator of malignant potential in colorectal carcinoma and that it may be one of the biological features of obstructing carcinoma of the colon and rectum. Moreover, our results demonstrated that nitric oxide might be associated with the difference in the mechanism of carcinogenesis in the colon and the rectum.

Inducible nitric oxide synthase expression before and after eradication of Helicobacter pylori in different forms of gastritis

An increased expression of inducible nitric oxide synthase (iNOS) has been observed in the inflamed human gastric mucosa as well as in some tumors. This observation suggests a pathobiological role of elevated NO production. The purpose of this study was to compare the immunohistochemical iNOS expression in the different kinds of gastritis before and after the eradication of Helicobacter pylori. We performed iNOS and H. pylori immunohistochemical staining and counted iNOS positive cells. We detected elevated expression of iNOS around sites infected with H. pylori. iNOS expression in chemical gastritis was strongly elevated in mucosal glands. After treatment, we found a significant difference in iNOS expression in patients with classical H. pylori-induced antrum predominant gastritis and in patients with active autoimmune gastritis. In the special case of progressed gastritis with intestinal metaplasia we found persistence of intestinal metaplasia, and we could not find a significant difference in the number of positive iNOS cells before and after treatment. The persistence of IM as a possibly precancerous lesion is probably at least in the antrum a source of continuous iNOS induction even after H. pylori eradication.