Bulbul M A, Pavelic K, Slocum H K, Frankfurt O S, Rustum Y M, Huben R P, Bernacki R J
J Urol. 1986 Aug;136(2):512-6. doi: 10.1016/s0022-5347(17)44933-0.
Surgical specimens of fifty urologic tumors, 19 renal, 19 bladder, nine prostate and three testicular, were disaggregated into a cell suspension by a two step mechanical and enzymatic method. Viability, cytology and flow cytometry (FCM) for DNA ploidy were subsequently determined. Growth of urological tumors on extracellular matrix (ECM) was carried out as follows: 2.5 to 5 X 10(5) cells were placed in plastic T25 flasks in RPMI 1640 + 10 per cent fetal bovine serum (FBS). 1.0 to 5 X 10(4) cells were plated in wells coated with ECM derived from bovine corneal endothelial cells with RPMI 1640 + 10 per cent FBS. Cultures were incubated for seven to 10 days at 37 degrees C. Only one (renal) out of 28 tumors grew on plastic. Forty out of 50 tumors (80 per cent) established primary cultures on ECM as determined by cell counting, protein determination, and/or [3H]thymidine incorporation. Previous experience with 106 urologic tumors grown on double layer agar demonstrated an overall success rate of 48 per cent. On ECM renal tumors showed 95 per cent growth success, prostate 89 per cent, bladder 63 per cent and testicular 67 per cent. Unlike viability by trypan blue exclusion, tumor DNA ploidy and percentage of malignant cells plated on ECM had no effect on growth success. The malignant nature of the cultured cells was confirmed by cytology. Twelve high grade and metastatic tumors caused degradation of the ECM. DNA ploidy was similar in four and different in six tumors before and after culture. Five tumors underwent in vitro drug testing on ECM with significant growth inhibition observed in three cases. The extracellular matrix seems to be a promising model for growing urologic tumors with excellent potential for drug testing in vitro.
采用两步机械和酶法将50例泌尿系统肿瘤的手术标本（19例肾癌、19例膀胱癌、9例前列腺癌和3例睾丸癌）解离成细胞悬液。随后测定细胞活力、细胞形态学及DNA倍体的流式细胞术（FCM）。泌尿系统肿瘤在细胞外基质（ECM）上的生长实验如下：将2.5至5×10⁵个细胞置于含RPMI 1640 + 10%胎牛血清（FBS）的塑料T25培养瓶中。将1.0至5×10⁴个细胞接种于涂有牛角膜内皮细胞来源的ECM的孔中，培养基为含10% FBS的RPMI 1640。培养物在37℃孵育7至10天。28例肿瘤中只有1例（肾癌）在塑料培养瓶上生长。通过细胞计数、蛋白质测定和/或[³H]胸腺嘧啶核苷掺入法确定，50例肿瘤中有40例（80%）在ECM上建立了原代培养。之前对106例在双层琼脂上生长的泌尿系统肿瘤的经验表明总体成功率为48%。在ECM上，肾癌的生长成功率为95%，前列腺癌为89%，膀胱癌为63%，睾丸癌为67%。与台盼蓝排斥法测定的活力不同，肿瘤DNA倍体及接种于ECM上的恶性细胞百分比对生长成功率没有影响。通过细胞形态学证实了培养细胞的恶性性质。12例高级别转移性肿瘤导致ECM降解。4例肿瘤培养前后DNA倍体相似，6例不同。5例肿瘤在ECM上进行了体外药物测试，3例观察到明显的生长抑制。细胞外基质似乎是一个有前景的用于培养泌尿系统肿瘤的模型，在体外药物测试方面具有极好的潜力。
