mdr1启动子调控CD∷UPP基因对紫杉醇耐药卵巢癌细胞的杀伤作用

目的:探讨腺病毒介导的mdr1启动子调控胞嘧啶脱氨酶∷尿嘧啶磷酸核糖转移酶(CD∷UPP)融合基因联合5-氟胞嘧啶(5-FC)对紫杉醇耐药卵巢癌细胞的特异性杀伤作用。方法:扩增、纯化含有mdr1-CD∷UPP基因的重组腺病毒,转染人卵巢癌紫杉醇耐药细胞株A2780/Taxol和亲本细胞株A2780,RT-PCR检测mdr1和CD∷UPP基因的表达水平;之后加入5-FC,MTT法检测细胞抑制情况及旁观者效应,并观察腺病毒转染后裸鼠移植瘤的生长情况。结果:mdr1和CD∷UPP基因在A2780/Taxol细胞中可稳定表达,转染后A2780/Taxol组的细胞生长明显低于A2780组;转基因的A2780/Taxol细胞联合5-FC后可通过旁观者效应杀伤周围未转基因的耐药细胞;耐药组移植瘤生长明显受到抑制,肿瘤体积为(569.10±187.93)mm3,对照组肿瘤体积为(2111.98±230.82)mm3,差异有统计学意义(P<0.01)。结论:mdr1启动子可调控CD∷UPP基因特异性表达并特异性杀伤紫杉醇耐药卵巢癌细胞。     

Levels and expressions of orotate phosphoribosyltransferase in gastric carcinoma and normal gastric mucosa tissues

Background Orotate phosphoribosyltransferase (OPRT; EC 2.4.2.10), a key enzyme that catalyzes one of the primary steps in the phosphorylation of fluoropyrimidine, was recently recognized as an important enzyme that determines the anticancer effects of the dihydropyrimidine dehydrogenase-inhibitory fluoropyrimidine, S-1. Methods Levels of OPRT were examined in 97 gastric carcinoma tissues and 65 normal gastric mucosa tissues obtained from patients with gastric carcinoma. The relation between OPRT levels and clinicopathological variables was evaluated, and correlations of OPRT with thymidylate synthase and dihydropyrimidine dehydrogenase levels in gastric carcinoma tissues were evaluated. Results Although OPRT levels were high in well-differentiated and localized carcinomas, they were not correlated with other clinicopathological variables or with the pathological stage of gastric carcinoma. Levels of OPRT were significantly higher in gastric carcinoma tissue than in normal gastric mucosa. OPRT levels were not correlated with levels of either thymidylate synthase or dihydropyrimidine dehydrogenase. In samples of gastric carcinoma tissues and normal gastric mucosa tissues obtained simultaneously from 24 patients, no correlation was found between OPRT levels in gastric carcinoma and levels in normal gastric mucosa. Conclusion These results suggest that the OPRT level is significantly higher in gastric carcinoma tissue than in normal gastric mucosa and that the OPRT level in gastric carcinoma is a novel variable that is independent of the levels of other previously known enzymes related to 5-fluorouracil (FU) metabolism.     

大鼠胰腺导管上皮细胞体外诱导分化

目的 探讨上皮细胞生长因子(EGF)、尼克酰胺(NIC)、肝细胞生长因子(HGF)、葡萄糖(Glu)、角脘蛋白生长因子(KGF)和β细胞素在大鼠胰腺导管上皮细胞体外诱导分化的作用.方法 采用健康SD大鼠胰腺导管上皮细胞,分于70个培养孔中,加入含有多种营养因子(不含血清)的PRMI 1640培养液中培养.每个孔中6种诱导剂均进行随机组合.在胰岛样细胞团(ICCs)形成前后的不同天数进行细胞计数、电镜观察、胰岛素测定以及免疫细胞化学和荧光分析.结果 胰腺导管上皮细胞培养7d后,单个贴壁上皮细胞分裂增殖,呈鹅卵石样细胞片,单层覆盖;加入诱导和促生长因子后,单层覆盖上皮细胞中逐渐产生一定数量类似胰岛样球状细胞团,直径为80～130μm.结论 6种促生长和诱导因子中,Glu、NIC、KGF、EGF作用较强;在所设定的浓度范围内,其作用均随着浓度的增加而增强.     

复方叶酸B_(12)注射液导数光谱测定法

复方叶酸B_(12)注射液在江苏省药品质量标准中只测定叶酸含量,用还原-重氮化-偶合-比色法,操作步骤多,易造成误差。本文采用紫外分光光度法直接测定叶酸含量,并用一阶导     

7-ribosyl-3-deazaguanine—Mechanism of antibacterial action

The mechanism by which 7-ribosyl-3-deazaguanine $\text{[7}\text{R}\text{3}\text{DG}\text{, 6-}\text{amino}\text{-3-β-}\text{d}\text{-}\text{ribofuranosylimidazo}\text{[4,5-c]}\text{pyridin}\text{-4(5H)-}\text{one}\text{]}$ exerts its antibacterial effect was examined. Escherichia coli was found to contain an enzyme that exhibited the properties of a nucleoside phosphorylase and that converted 7R3DG to 3-deazaguanine (3DG, 6-aminoimidazo[4,5-c]pyridin-4(5H)-one], but no mammalian system that was examined (Erilch ascites, rat liver and human liver) was able to convert 7R3DG to 3DG. The 3DG arising from the phosphorolysis of 7R3DG was converted to 3-deaza-GMP [3DGMP, 6-amino-l-β-D-riboluranosylimidazo [4,5-c]pyridin-4(5H)-one-5′-phosphate] by the guanine phosphoribosyltransferase present in E. coli. A strain of E. coli, resistant to 7R3DG, was found to lack this enzyme and, therefore, was unable to convert 3DG to 3DGMP.     

Inhibition of orotate phosphoribosyltransferase and orotidine-5'-phosphate decarboxylase of human erythrocytes by purine and pyrimidine nucleotides.

Orotidine-5′-phosphate decarboxylase of human hemolysates exhibits triphasic kinetics with K_m values of 33, 1.7 and 0.082 μM. Inhibition of this enzyme at low OMP concentrations (<3 μM) by several naturally occurring purine and pyrimidine nucleotides was investigated. No significant inhibition was observed with IMP, GMP, TMP, ADP, and TTP at 5 mM. Inhibition constants for CMP, AMP, and dAMP were 31 μM, 0.11 mM and 0.21 mM, respectively. The results are discussed in relation to inhibition by nucleotides of orotate phosphoribosyltransferase, previously measured with a method which depends on orotidine-5′-phosphate decarboxylase activity.     

Partial hypoxanthine-guanine phosphoribosyltransferase deficiency presenting as acute renal failure

Hyperuricemia and secondary urate nephropathy are uncommon in the paediatric setting outside of tumour lysis syndrome. We describe the case of a 12-year-old boy who presented at 3 years of age with acute renal failure. The cause of this remained unknown until the development of uric acid renal calculi 9 years later. This, and the availability of the previously unknown family history, provided the subsequent diagnosis of partial hypoxanthine-guanine phosphoribosyltransferase (HPRT) deficiency. Detailed family history is important for early detection of this heterogeneous group of disorders. Early treatment may minimise long-term renal morbidity and mortality from renal insufficiency.     

Abnormal Property of Human Mutant Hypoxanthine-Guanine Phosphoribosyltransferase: Insensitivity of Fibroblast Enzyme to Stabilization against Freezing and Thawing by 5-Phosphoribosyl-1-Pyrophosphate

Abstract. Freezing and thawing of dilute normal human fibroblast suspensions causes partial inactivation of hypoxanthine-guanine phosphoribosyltransferase (HGPRT) and adenine phosphoribosyltransferase (APRT). Phosphoribosyl-pyrophosphate (PRPP) stabilizes both phosphoribosyltrans-ferases against this inactivation. Mutant HGPRT enzymes from a patient with the Lesch-Nyhan syndrome and from a gouty patient with partial HGPRT deficiency were similarly inactivated by freezing and thawing, but only the former mutant enzyme could be stabilized by PRPP. The insensitivity of the mutant HGPRT from the patient with partial enzyme deficiency to PRPP stabilization indicates a structural enzyme alteration. The different sensitivity of the two HGPRT mutants to PRPP stabilization reflects the heterogeneity of HGPRT mutations in man.     

Relationships between the expression of thymidylate synthase,dihydropyrimidine dehydrogenase,and orotate phosphoribosyltransferase and cell proliferative activity and 5-fluorouracil sensitivity in colorectal carcinoma

Background: The site of action of the 5-fluorouracil (5-FU) antitumor effect has been explicated in recent years. Many studies have investigated enzymes involved in 5-FU metabolism in attempts to predict this effect, and a correlation of enzyme activity with the 5-FU drug sensitivity test has been reported. The aim of this study was to identify the biochemical response determinants of 5-FU. Additionally, we aimed to clarify the association between cell proliferative activity and the response to 5-FU of colorectal cancer. Methods: Our research subjects were 54 patients with colorectal carcinoma who had undergone operations between August 1999 and July 2001 in our department. Assays of thymidylate synthase (TS), dihydropyrimidine dehydrogenase (DPD), and orotate phosphoribosyltransferase (OPRT) activities in colorectal carcinoma tissue and assays of 5-FU sensitivity by the collagen gel droplet embedded culture drug sensitivity test (CD-DST) were conducted to investigate the relationships between each enzyme activity and 5-FU sensitivity. In addition, the proliferative activity of cancer cells was evaluated with Ki-67 antibody, and the relationship of this activity to each enzyme activity and 5-FU sensitivity were investigated. Results: 5-FU sensitivity was high in the low-TS-activity group and in the high-OPRT-activity group. Cancers with high cell proliferative activity showed good sensitivity to 5-FU, and TS and OPRT activities were high in such cancers. Conclusion: The results suggest that OPRT activity can predict sensitivity to 5-FU, and high OPRT activity may cause good 5-FU sensitivity in cancers with high cell proliferative activity. Received: April 23, 2002 / Accepted: January 20, 2003 Correspondence to:R. Fujii