Isolation, karyotype, and clonal growth of heterogeneous subpopulations of human malignant gliomas.
Isolation, karyotype, and clonal growth of heterogeneous subpopulations of human malignant gliomas.
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发表时间：
1981-06
期刊：
影响因子：
11.2
通讯作者：
J. Shapiro;W. Yung;W. Shapiro
中科院分区：
文献类型：
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作者：
J. Shapiro;W. Yung;W. Shapiro
人类恶性胶质瘤的组织学多形性及其对治疗的可变反应表明，这类肿瘤不是同质的，而是由异质的细胞群组成的。为了寻找这些肿瘤的异质性，我们制定了一项通过核型鉴定、分离和克隆肿瘤亚群的方案。新鲜切除的肿瘤被机械分离成单细胞，在悬浮和短期单层培养中生长。在切除后的前6至72小时收集染色体准备。制备的核型包括在每个肿瘤中发现的“参考组”染色体。分离的单细胞也被稀释，进行单层培养。附着的细胞被标记并分离为克隆，其核型与参考组中的克隆进行比较，以确定哪些克隆是肿瘤的细胞代表。从同一肿瘤中建立的外植体培养在单层培养中生长并克隆；他们的核型同样与参考组的核型进行了比较。我们分析了8个胶质瘤。每个肿瘤都有染色体数目从次二倍体到高倍体不等的中期，但亚群的频率分布在不同的肿瘤中有所不同。将肿瘤中的细胞亚群定义为在参考集中至少产生5个核型相同的细胞，我们发现每个肿瘤包含3到21个亚群。除了一个肿瘤外，所有肿瘤都产生了100个或更多的克隆，在7个肿瘤中，超过50%的克隆可以扩增进行核型分析；这些克隆中有7.6 - 25%的人的核型在其肿瘤的参考组中可识别。克隆形态与其核型或生长动力学之间的关系不大。虽然外植体培养生长良好，但在参考集合中无法鉴定出来自此类培养的克隆。该方案允许研究克隆系可识别为患者肿瘤的细胞代表，而不是在组织培养中产生的变异。这些研究证实，人类恶性胶质瘤是由核型异质性细胞亚群组成的。初步研究表明，它们的表型行为可能具有相似的异质性。
Abstract The histological pleomorphism of human malignant gliomas as well as their variable response to therapy suggests that such tumors are not homogeneous but are composed of a heterogeneous population of cells. To search for heterogeneity in these tumors, we developed a protocol to identify by karyotype, isolate, and clone the subpopulations of the tumor. Freshly resected tumors were mechanically dissociated into single cells that were grown in suspension and short-term monolayer cultures. Chromosomal preparations were collected over the first 6 to 72 hr postresection. The karyotypes prepared comprised the “reference set” of chromosomes found in each tumor. The dissociated single cells were also dilution plated for monolayer culture. Cells that attached were marked and isolated as clones whose karyotypes were compared to those in the reference set in order to identify those clones that were cellular representatives of the tumor. Explant cultures established from the same tumor were grown in monolayer culture and cloned; their karyotypes were likewise compared to those in the reference set. Eight gliomas were analyzed. Each had metaphases that ranged in chromosome number from hypodiploid to hyperploid, but the frequency distribution of the subpopulations differed among the tumors. Defining a cellular subpopulation in the tumor as one yielding at least five karyotypically identical cells in the reference set, we found that each tumor contained from three to 21 subpopulations. All but one tumor produced 100 or more clones, and in seven tumors, more than 50% of the clones could be expanded for karyotyping; 7.6 to 25% of these clones had karyotypes identifiable in the reference set of their tumor. There was little relationship between the morphology of the clones and their karyotypes or growth kinetics. While explant cultures grew well, no clone derived from such cultures could be identified in the reference set. The protocol permits studies of clonal lines identifiable as cellular representatives of a patient9s tumor and not variants generated in tissue culture. These studies confirmed that human malignant gliomas are comprised of karyotypically heterogeneous cellular subpopulations. Preliminary work suggests that their phenotypic behavior may be similarly heterogeneous.