Research review: DNA polymerases as molecular markers of the regenerating capacity of hepatocytes

Hepatocyte regeneration has been widely investigated, with the mitotic index and the incorporation of [³H]thymidine being used as regeneration markers. We focused on the induction of DNA replication enzymes, particularly DNA polymerases (pol) α, δ, and ε. Using rat models, we have shown that the activity of pol α in crude liver extract well represents the regenerating capacity of hepatocytes. Using pol α as an indicator, we analyzed liver regeneration in rat models under various conditions: obstructive jaundice, external or internal biliary drainage, and the obstruction of portal vein branches. It has been revealed that the ligation of the common bile duct alone induces a certain amount of hepatocyte proliferation. It was striking that external biliary drainage suppressed regeneration capacity in cholestatic rat liver after partial hepatectomy. The strong regeneration in nonligated lobes induced by portal branch ligation was similar to the liver regeneration seen after partial hepatectomy with respect to the induction of DNA polymerases. Taken together, the aspects of DNA replication, particularly the induction of DNA polymerases, may contribute to shedding new light on the regeneration of human liver. This work was supported in part by a Grant-in-Aid for General Scientific Research and for Cancer Research from the Ministry of Education, Science and Culture, Japan, and by grants from the Uehara Memorial Foundation     

Molecular oncogene markers and their significance in cutaneous malignant melanoma

Background: Oncogenes and other molecular tumor markers that predict tumor aggressiveness may allow individualization and optimization of surgical therapy of intermediate-thickness malignant melanoma. We examined the expression of selected markers, including the HLA-DR antigen, the heat shock protein-70 (HSP-70), and the c-myc oncogene in primary melanoma and regional nodes and related these findings to metastatic potential and survival. Methods: Forty patients with primary melanoma (1.5–4.0 mm) were studied, all of whom had prophylactic lymph node dissection and were followed for 18 months to 7 years. The primary tissue and nodes were examined using immunohistochemical techniques for the presence of HLA-DR antigen and HSP-70 protein and the expression of the c-myc oncogene. Results: Of 40 patients, there were 23 with lesions 1 to 2.9 mm thick and 17 with lesions 3 to 4 mm thick. Nodal metastases were present in 25 of the 40 patients who had elective node dissection. HLA-DR antibody stained the primary tumor in 10 patients (25%), but there was no correlation with survival in this group. HLA-DR antibody stained the stroma and cellular infiltrates surrounding the primary tumor in 28 of 40 patients; in this group there was a correlation of HLA-DR staining of the peritumoral stroma with improved survival overall. HLA-DR staining of the peritumoral stroma also influenced survival when patients were stratified by tumor thickness groups 1 to 2.9 mm and 3 to 4 mm and presence of nodal metastases. HSP-70 was demonstrated in the primary tumor in 25% of patients, who were also shown to have significantly improved survival when compared with those whose primary tumor did not stain with HSP-70. C-myc was expressed in the primary tumor in 25%, but showed no correlation with survival. None of these proteins correlated with or predicted the presence of nodal metastases. Conclusion: We conclude that the use of specific molecular-oncogene markers in intermediate-thickness primary melanoma may identify patients at high risk for conventional treatment failure and reduced survival who may profit from more aggressive surgery, adjuvant therapy, or both.Presented at the 48th Annual Cancer Symposium of The Society of Surgical Oncology, Boston, Massachusetts, March 23–26, 1995.     

胃肠癌患者血清及癌组织癌胚抗原的检测意义

应用放射免疫测定法,对20例胃肠癌患者进行了血清癌胚抗原(CEA)测定,并应用免疫组化法对其中16例癌组织中的CEA进行了检测.检测结果,血清CEA阳性6例,癌组织CEA阳性9例.低分化癌的血清CEA显著高于高分化癌(P<0.05).血清CEA与癌灶部位、癌组织CEA与分化程度及血清CEA均无关(P>0.05).提示血清CEA的含量与癌组织的分化程度有关,血清CEA与癌组织内的CEA无平行关系.作者并对其机制及检测CEA的意义进行讨论.     

SA、F/TPSA及PSAD在前列腺癌诊断中的作用

目的 :探讨血清前列腺特异抗原 (PSA) ,血清总PSA及游离PSA比值 (F/TPSA)及前列腺特异性抗原密度(PSAD)在前列腺癌诊断中的作用。方法 :对 5 1例前列腺癌患者及 14 5例良性前列腺增生症患者PSA、F/TPSA及PSAD值的差异进行分析、比较。结果 :前列腺癌组血清PSA及PSAD高于良性前列腺增生组 ;而F/TPSA值低于良性前列腺增生组 ,差异均有显著性。结论 :PSA >4ng·ml-1作为筛选前列腺癌的临界值存在一定缺陷 ;当PSA <10ng·ml-1,F/T值有助于鉴别前列腺癌和良性前列腺增生 ;而PSAD对于筛选前列腺活检病例亦有一定价值。     

逆转录-聚合酶链反应检测胃癌淋巴结微转移

目的 检测胃癌常规病理检查阴性的淋巴结微转移的发生及与其它临床参考指标的关系。方法 利用逆转录－聚合酶链反应（RT－PCR）方法检测68枚胃癌胃周淋巴结癌胚抗原（CEA）mRNA基因表达，同时比较RT－PCR与免疫组化（IHC）方法的检测敏感性。结果 CEAmRNA RT－PCR是一种很敏感的方法，可以检测1／10^6个转移的癌细胞；检测19例胃癌患者取材的68枚胃周淋巴结，IHC阳性率28％（19／68），RT－PCR阳性率57％（39－68），两组之间差异有非常显著性意义（P＜0.01）；RT－PCR阳性率与胃部临床参考指标密切相关，且随着病期进展而增大。结论 CEA mRNA RT-PCR是比免疫组化更敏感的方法，可以预测胃癌淋巴结微转移，能够有效地避免已有微小转移的患者被漏诊。     

Ca^＋＋对链霉素抗原—抗体反应的抑制作用

为探讨临床中使用钙制剂抢救链霉素过敏休克病人有良好效果的原因,由被动血凝试验、被动皮肤过敏试验和ELISA法证实,Ca~( )可以抑制链霉素抗原-抗体的反应。这种抑制作用是通过Ca~( )链霉素抗原结合,封闭链霉素抗原决定簇来加以实现的。     

Antibody reactivity to a synthetic peptide (P62) of the Epstein-Barr nuclear antigen in sera of patients with X-linked lymphoproliferative syndrome

An enzyme-linked immunosorbent assay (ELISA) was used to measured IgG antiboody titers againt a synthetic peptide whose sequence was derived from the glycine-alanine repeating region of Epstein-Barr virus nuclear associated antigen 1 (EBNA-1). Antibody titers were determined in sera from 15 normal subjects, sera from 21 normal male siblings of X-linked lymphoproliferative syndrome (XLP) patients, from 20 XLP patients comprising a total of 42 samples, and ten samples before and ten samples after gamma-globulin therapy in ten patients with XLP. Data analysis demonstrated that while there are differences between the ELISA and ACIF, they appear to measure a similar response as demonstrated by their correlation coefficient (0.77) and the GMT to EBNA observed by both methods. No cross-reactivity of cytomegalovirus antibodies to the EBNA-1 peptide was observed by immunobv using adsorption against AD-169 infected MRC-5 cells.. However, non-specific binding was observed if samples were not pre-incubated in a 10% goat serum PBS-Tween 20 solution. This pre-treatment removed the non-specific binding that falsely elevated GMT in approximately 15% of both normal and XLP samples in ELISA. The ELISA system appears to be a sensitive, reproducible and objective test that may be useful for assessing the antibody responses of patients to the EBNA-1 protein.     

简单灵敏的免疫复合物抗原特异性固相酶联检测法

本文将聚乙二醇(PEG)比浊法和固相酶联免疫法(ELISA)结合,建立了—较灵敏的免疫复合物(IC)直接固相吸附抗原特异性检测法。利用牛清蛋白(BSA)为已知抗原组份的IC模型,分别对IC直接固相吸附的条件和影响因素、方法的灵敏度、重复性等进行了研究。结果发现IC在解离状态下直接固相吸附后的抗原特异性检测灵敏度明显高于未解离者。该法具有简单易行,灵敏度较高、适于临床测定大量血清样品等优点。     

癌胚抗原DNA疫苗对大肠癌特异性细胞免疫的激活效果

目的检测以质粒pIRES为载体构建的带有全序列癌胚抗原(CEA)基因的核苷酸疫苗对机体特异性抗肿瘤免疫反应的激活效果。方法将CEA基因片段连接于真核表达质粒pIRES中,用肌肉注射方法接种核酸疫苗;检测CEA在小鼠肌肉组织中的表达情况及其对小鼠脾细胞CEA特异性细胞免疫反应的激活效果。结果小鼠经肌肉注射质粒后,免疫组化证实该核酸疫苗可在体内有效表达CEA;分子免疫检测显示注射后小鼠特异性淋巴细胞增值反应明显并且伴有自然杀伤细胞NK活性显著增高。结论实验所构建的核酸疫苗pIRESCEA可在体外及小鼠体内高效表达并表现出良好的细胞免疫原性。     

三种实验性IgA肾病模型的比较

目的探讨建立一种理想的IgA肾病（IgAN）动物模型方法。方法分别采用葡聚糖G200、大肠杆菌外膜蛋白和金葡菌的细胞膜20肽抗原决定簇诱导小鼠IgA肾病模型。用分子生物学和病理学方法对3组IgAN模型小鼠进行鉴定和比较。结果（1）葡聚糖组尿蛋白增高，伴有血尿；免疫荧光显示部分肾小球大量IgA沉积；光镜下肾小球系膜细胞增多，肝和脾可见弥漫性的粉染物质沉积；电镜下肾小球系膜区少量低电子密度的致密沉积物，肝和脾可见淀粉丝样物质沉积。（2）大肠杆菌外膜蛋白组尿蛋白增高，伴有血尿；免疫荧光显示肾小球有少量IgA沉积；光镜下肾小球系膜细胞轻度增多，间质炎细胞浸润明显；电镜下肾小球系膜区无电子致密沉积物。（3）金葡菌细胞膜20肽抗原决定簇组尿蛋白增高，伴有血尿；免疫荧光显示多数肾小球均可见大量IgA沉积；光镜下肾小球系膜细胞增多，伴系膜基质轻度增生；电镜下肾小球系膜区和基底膜的内皮细胞下可见高电子密度的致密沉积物。结论金葡菌细胞膜20肽抗原决定簇组诱导的IgAN模型从临床表现和病理学变化与人IgAN极其相似，是3种IgAN模型中最理想的IgAN模型。