3-nitrotyrosine and Neoplasm-Metastasis

The cells of a weakly tumorigenic and non-metastatic murine fibrosarcoma (QR-32) are converted into highly malignant tumors (acquiring metastatic potential) once they have grown in vivo after being co-implanted with gelatin sponge which induces inflammation. In the present study, we examined whether nitric oxide (NO) is involved in the inflammation-based tumor progression by administrating a specific inhibitor to inducible nitric oxide synthase, aminoguanidine (AG). First, we co-implanted 1 x 10(5) QR-32 cells with gelatin sponge (10 x 5 x 3 mm piece) into a subcutaneous space in C57BL6 mice. Administration of AG in drinking water (1%) had started 2 days before the tumor implantation and continued until the termination of the experiment. The incidence of tumor formation and the tumor growth did not differ between AG-treated group and -untreated group. On day 28, we excised the arising tumors to establish culture cell lines for evaluation of their acquisition of metastatic phenotype in other normal mice. Metastasis incidence and the number of metastatic colonies were significantly reduced in the tumor cell lines obtained from AG-treated mice compared to those from non-treated mice (p < 0.05). Immunohistochemical analysis demonstrated that inducible nitric oxide synthase and nitrotyrosine in the inflamed lesion were reduced in the AG-administered mice. However, intensity of 8-hydroxy-2-deoxyguanosine was not different between the groups. These results showed that nitric oxide and its reactive nitrogen oxide species cooperatively play a pivotal role in the progression of benign tumor cells in inflamed lesions.

Topics: Animals; Cell Movement; Cell Proliferation; Disease Progression; Female; Fibrosarcoma; Gelatin Sponge, Absorbable; Guanidine; Guanine; Inflammation; Mice; Mice, Inbred C57BL; Neoplasm Metastasis; Neoplasm Transplantation; Nitric Oxide Synthase Type II; Oxidation-Reduction; Reactive Nitrogen Species; Tyrosine

Nitric oxide (NO) is a small molecule produced during the conversion of L-arginine to L-citrulline by NO synthase (NOS). Several isoforms of NOS exist, of which the Ca2+-independent, inducible NOS (iNOS or NOS2) is most prominently expressed by macrophages. iNOS activity and increased levels of iNOS have been found in various tumours and tumour cell lines but not in normal tissues; however, the precise role of NO in tumour progression has yet to be elucidated. We studied the expression of iNOS in paraffin sections of 41 benign naevi and 52 primary malignant melanomas (MM) of the skin, as well as in 13 metastatic MM. In addition, nitrotyrosine, indicative of NO production and formation of peroxynitrite, was studied in frozen sections of 13 naevi and 30 MM. Virtually all naevi expressed iNOS, but very few expressed nitrotyrosine, indicating either that iNOS in naevi is functionally inactive, or that naevus cells lack reactive oxygen radicals and thus do not form peroxynitrite. Normal melanocytes in adjacent uninvolved skin were unreactive for both markers. In MM, iNOS was most frequently expressed in the 'pure' and 'invasive' radial growth phase (RGP), whereas expression in the vertical growth phase (VGP) and metastatic phase occurred only in 76% of cases; moreover, in these latest phases of tumour progression, iNOS staining was weak and focal. We conclude that iNOS is expressed de novo in most benign pigment cell lesions. In MM (iNOS-generated) NO appears to play an important part in the early steps of invasion (i.e. the 'invasive' RGP), where it may stimulate neo-angiogenesis and may be a prerequisite for further tumour progression; this view is also supported by the finding of iNOS in the associated blood vessels in the papillary dermis. Finally, our data suggest that (iNOS-generated) NO plays a less significant part in the VGP and in metastatic melanoma.

Topics: Biomarkers, Tumor; Disease Progression; Enzyme Induction; Humans; Isoenzymes; Macrophages; Melanoma; Neoplasm Metastasis; Nevus; Nitric Oxide Synthase; Skin Neoplasms; Tyrosine

Despite recognition of the malignant potential of human melanomas, the mechanisms responsible for the pathobiological characteristics contributing to tumor growth, vascular invasiveness, and distant organ metastasis remain undefined. Recent studies have shown that various human tumors express an inducible form of nitric oxide synthase (iNOS) and nitrotyrosine (NT), which suggests a mechanistic role of tumor-associated nitric oxide (NO) in tumorigenesis. We investigated iNOS and NT expression by immunohistochemistry in 20 human metastatic melanoma tissue specimens specifically with respect to iNOS-expressing cell types in the tumor area, pathological and clinical response to systemic therapy, potential role as a prognostic indicator, and NT formation. Our results showed that melanoma cells from 12 of 20 tumors express iNOS, yet the expression of this molecule in the tumor did not correlate with pathological or clinical response to therapy. More importantly, iNOS and NT expression by the melanoma cells strongly correlated with poor survival in patients with stage 3 disease (P < 0.001 and P = 0.020, respectively), suggesting a pathway whereby iNOS might contribute to enhanced tumor progression. In conclusion, our findings strongly suggest that iNOS expression has potential to be considered as a prognostic factor and NO as a critical mediator of an aggressive tumor phenotype in human metastatic melanomas.

Topics: Adolescent; Adult; Female; Humans; Immunohistochemistry; Male; Melanoma; Middle Aged; Neoplasm Metastasis; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Prognosis; Time Factors; Treatment Outcome; Tyrosine