Department of Biochemistry, College of Pharmacy, Minia University, Minia, Egypt.
J Membr Biol. 2010 Mar;234(1):29-34. doi: 10.1007/s00232-010-9236-7. Epub 2010 Feb 23.
Five fluorouracil (5-FU) is extensively used in the treatment of hepatocellular carcinoma (HCC). It is well documented that 5-FU and its metabolites inhibit DNA synthesis through inhibition of thymidylate synthetase. Little is known about additional pathways for 5-FU in managing HCC. The present experiment was mainly designed to study possible biochemical pathways that can be added to 5-FU's mechanisms of action. Four groups of rats constituted a control group (given saline only), a trichloroacetic acid group (TCA, 0.5 g/kg/day for 5 days, orally), a 5-FU-positive group (75 mg/kg body weight, intraperitoneally, once weekly for 3 weeks) and a TCA-treated with 5-FU group (24 h from last TCA dose). We executed both biochemical-serum alpha-fetoprotein (AFP), liver tissue contents of total glycosaminoglycan (TGAGs), collagen (represented as hydroxyproline), total sialic acid (TSA), free glucosamine (FGA) and proteolytic enzyme activity (as pepsin and free cathepsin-D-and histological examinations of the liver tissue. The results revealed histological changes such as central vein congestion and irregularly shaped, substantially enlarged, vesiculated and binucleated hepatocytes. The nuclei were mostly polymorphic and hyperchromatic, and several vacuolation was noticed in the cytoplasm encircling the nucleus with masses of acidophilic material. 5-FU greatly corrected these changes, except that some necrotic and cytotoxic effects of 5-FU were still shown. AFP was significantly elevated in TCA-intoxicated, but reversed in 5-FU-treated, groups. Increased proteolytic activity by TCA was reversed by 5-FU, which also restored TGAG contents to normal; but both TCA and 5-FU depleted collagen content. TCA significantly elevated FGA but depressed TSA; this action was reversed by 5-FU treatment. In conclusion, it is possible that proteolytic activity, expressed as upregulated pepsin and free cathepsin-D activities, is increased in HCC. This is accompanied by extracellular matrix macromolecular disturbance, manifested as decreased TGAGs, collagen and TSA, with marked increase in FGA liver tissue content. The elevated FGA with depressed TSA content of liver tissue may be attributed to a cancer-hampered N-acetylation of FGA into SA. 5-FU administration markedly depressed hepatic tissue proteolysis, possibly reactivated N-acetylation of FGA into SA and elevated TGAGs without stopping tissue fibrosis as collagen was not affected. This study explores additional pathways for the mechanism of action of 5-FU, through conservation of extracellular matrix composition in situ, inhibiting invasion and metastasis in addition to its DNA-disturbing pathway.
氟尿嘧啶(5-FU)在肝癌(HCC)的治疗中被广泛应用。有充分的证据表明,5-FU 及其代谢物通过抑制胸苷酸合成酶来抑制 DNA 合成。关于 5-FU 在治疗 HCC 中的其他途径知之甚少。本实验主要设计用于研究可能的生化途径,这些途径可以补充 5-FU 的作用机制。四组大鼠构成一个对照组(仅给予生理盐水)、三氯乙酸(TCA)组(0.5 g/kg/天,连续 5 天,口服)、5-FU 阳性组(75 mg/kg 体重,每周一次,连续 3 周,腹腔内注射)和 TCA 处理后加用 5-FU 组(最后一次 TCA 剂量后 24 小时)。我们进行了生化-血清甲胎蛋白(AFP)、肝组织总糖胺聚糖(TGAGs)、胶原(表示为羟脯氨酸)、总唾液酸(TSA)、游离氨基葡萄糖(FGA)和蛋白水解酶活性(如胃蛋白酶和游离组织蛋白酶-D)以及肝组织的组织学检查。结果显示出中央静脉充血和不规则形状、明显增大、囊泡化和双核肝细胞等组织学变化。细胞核大多呈多形性和深染性,细胞质中围绕着细胞核可见多个空泡,细胞质中含有大量嗜酸性物质。5-FU 极大地纠正了这些变化,但仍显示出 5-FU 的一些坏死和细胞毒性作用。TCA 中毒组 AFP 显著升高,但经 5-FU 治疗后逆转。TCA 引起的蛋白水解活性增加被 5-FU 逆转,这也使 TGAG 含量恢复正常;但 TCA 和 5-FU 均耗尽了胶原含量。TCA 显著升高 FGA,但降低 TSA;5-FU 治疗可逆转这一作用。总之,肝癌中可能存在蛋白水解活性升高的情况,表现为胃蛋白酶和游离组织蛋白酶-D 活性上调。这伴随着细胞外基质大分子的紊乱,表现为 TGAGs、胶原和 TSA 减少,肝组织 FGA 含量显著增加。肝组织中升高的 FGA 和降低的 TSA 含量可能归因于 FGA 的 N-乙酰化受阻,转化为 SA。5-FU 给药显著抑制肝组织蛋白水解,可能通过重新激活 FGA 的 N-乙酰化生成 SA,同时升高 TGAGs,而不阻止组织纤维化,因为胶原不受影响。本研究通过原位保持细胞外基质组成,抑制侵袭和转移,除了其 DNA 干扰途径外,还探讨了 5-FU 作用机制的其他途径。