Thompson L, Chang B, Barsky S H
Department of Pathology, UCLA School of Medicine 90024, USA.
Am J Surg Pathol. 1996 Mar;20(3):277-85. doi: 10.1097/00000478-199603000-00003.
Malignant mixed tumors (carcinosarcomas) are examples of unusual neoplasms whose occurrences have been observed in increasingly diverse sites but whose pathogenesis remains a complete mystery. Two antithetical hypotheses that have been advanced to explain the histogenesis of these tumors include the convergence hypothesis, which proposes an origin from two or more stem cells (multiclonal hypothesis), and the divergence hypothesis, which proposes an origin from a single totipotential stem cell that differentiates into separate epithelial and mesenchymal directions (monoclonal hypothesis). To test these hypotheses, a novel strategy for the determination of clonality from as few as 100 tumor cells obtained by enzymatic digestion of either fresh or formalin-fixed, paraffin-embedded tissues and cell sorting was used that exhibited the polymerase chain reaction (PCR) in amplifying a 511-bp region located within the first intron of the human hypoxanthine phosphoribosyl transferase gene, a site that contains inactive X chromosomal obligately methylated HpaII/MspI sites and single-base allelic polymorphisms in 5% females. Carcinoma cells gated on the basis of fluorescein isothiocyanate (FITC)-anti-cytokeratin and sarcoma cells gated on the basis of FITC-antivimentin or FITC-anti-desmin were sorted to homogeneity on FACSTAR and then subjected to genomic DNA extraction and Hpa II digestion before PCR amplification and subsequent analysis of the product on denaturing gradient gel electrophoresis. The comigrations of the single homoduplexes generated from both the carcinoma cells and sarcoma cells in six different malignant mixed tumors obtained from four different organs indicated clonal identity and monoclonality in all cases. These findings of monoclonality were confirmed independently by two other methods of clonality determination. The findings of a monoclonal origin of carcinosarcomas support the single totipotential stem-cell-divergence hypothesis.
恶性混合瘤（癌肉瘤）是一类罕见肿瘤，其发病部位日益多样，但其发病机制仍是个谜。关于这些肿瘤的组织发生，有两种对立的假说：趋同假说，认为起源于两个或更多干细胞（多克隆假说）；分歧假说，认为起源于单个全能干细胞，该干细胞分化为不同的上皮和间叶方向（单克隆假说）。为验证这些假说，采用了一种新策略，即通过酶消化新鲜或福尔马林固定、石蜡包埋组织获得肿瘤细胞，经细胞分选后，从少至100个肿瘤细胞中确定克隆性。该策略利用聚合酶链反应（PCR）扩增位于人次黄嘌呤磷酸核糖转移酶基因第一内含子内的一个511bp区域，该位点在5%的女性中含有失活的X染色体特异性甲基化HpaII/MspI位点和单碱基等位基因多态性。基于异硫氰酸荧光素（FITC）-抗细胞角蛋白分选的癌细胞和基于FITC-抗波形蛋白或FITC-抗结蛋白分选的肉瘤细胞在FACSTAR上被分选至均一性，然后进行基因组DNA提取和Hpa II消化，再进行PCR扩增，随后对产物进行变性梯度凝胶电泳分析。在从四个不同器官获得的六个不同恶性混合瘤中，癌细胞和肉瘤细胞产生的单个同源双链体的共迁移表明在所有病例中均存在克隆同一性和单克隆性。单克隆性的这些发现通过另外两种克隆性测定方法得到了独立证实。癌肉瘤单克隆起源的发现支持了单个全能干细胞分歧假说。
