Effects of endogenous nitric oxide induced by 5-fluorouracil and L-Arg on liver carcinoma in nude mice.
OBJECTIVE
To study the effects of endogeous nitric oxide induced by 5-fluorouracil (5-FU) and L-arginine (L-Arg) on the human liver carcinoma model in nude mice.


METHODS
The human liver carcinoma model in nude mice was established with BEL-7402 cells and normal saline (NS), 5-FU and 5-FU + L-Arg injected intraperitoneally. The tumor size was measured. The necrotic degree and range were observed under microscope. The apoptosis of cancer cell was detected by turmina deoxynucleotidyl transferanse mediated dUTP nick end labeling (TUNEL) method. Immunohistochemical method was performed to determine the expression of iNOS, P16, BAX. The chemical colorimetry was used to test the activity and nitrate reductase method was adopted to test the concentration of nitric oxide (NO) in the tumor tissue. The BI2000 pathological image analyzer was used to analyze the result of immunohistochemistry.


RESULTS
5-FU combined with L-Arg could inhibit the tumor growth apparently. In NS, 5-FU and 5-FU+L-Arg groups, the changes of tumor volumes were 257.978 +/- 59.0, 172.232 +/- 66.0 and 91.523 +/- 26.7 mm(3), respectively (P < 0.05 5-FU vs 5-FU + L-Arg group; P < 0.05 NS vs 5-FU + L-Arg group; P < 0.05, NS vs 5-FU group). The necrotic range and apoptosis index were significantly increased after the drug injection. The necrotic range was biggest in 5-FU + L-Arg group (c2 = 15.963, P < 0.05). The apoptosis indexes were as follows: NS, 17.4% +/- 6.19%; 5-FU, 31.3% +/- 12.3%; and 5-FU + L-Arg, 46% +/- 15.24% (P < 0.05, 5-FU vs 5-FU + L-Arg; P < 0.05, NS vs 5-FU + L-Arg; P < 0.05, NS vs 5-FU). The expression and activity of iNOS were increased in the tumor tissue. The concentration of NO was also increased. F of optical density of iNOS, iNOS activity and NO concentration are 31.693, 21.949, and 33.909, respectively, P < 0.05. The concentration of NO was related to the expression of P16 and BAX. The correlation coefficient was 0.764 and 0.554.


CONCLUSIONS
5-FU combined with L-Arg can inhibit the growth of tumor in nude mice. The effect may be related to inducing the synthesis and increasing the activity of iNOS. The production of NO is increased, and it can enhance the expression of apoptosis-related gene and antioncogene.
Yin
Xiao-Yan
XY
Endoscopy Center, Affilated Hospital of Medical College, Qingdao University, Qingdao 266003, Shandong Province, China.
Jiang
Jun-Mei
JM
Liu
Ji-Yong
JY
Zhu
Ju-Ren
JR
D000818:Animals; D000963:Antimetabolites / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D001120:Arginine / Q000494:pharmacology*; D006528:Carcinoma, Hepatocellular / Q000378:metabolism* / Q000473:pathology*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000378:metabolism; D004195:Disease Models, Animal; D005472:Fluorouracil / Q000494:pharmacology*; D006801:Humans; D008113:Liver Neoplasms / Q000378:metabolism* / Q000473:pathology*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D009569:Nitric Oxide / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism; D023041:Xenograft Model Antitumor Assays; D051028:bcl-2-Associated X Protein / Q000378:metabolism
D000963:Antimetabolites; D019941:Cyclin-Dependent Kinase Inhibitor p16; D051028:bcl-2-Associated X Protein; D009569:Nitric Oxide; D001120:Arginine; D052247:Nitric Oxide Synthase Type II; D005472:Fluorouracil