Naumov George N, Bender Elise, Zurakowski David, Kang Soo-Young, Sampson David, Flynn Evelyn, Watnick Randolph S, Straume Oddbjorn, Akslen Lars A, Folkman Judah, Almog Nava
Department of Surgery, Children's Hospital, Harvard Medical School, Boston, MA, USA.
J Natl Cancer Inst. 2006 Mar 1;98(5):316-25. doi: 10.1093/jnci/djj068.
Microscopic human cancers can remain dormant for life. Tumor progression depends on sequential events, including a switch to the angiogenic phenotype, i.e., initial recruitment of new vessels. We previously demonstrated that human tumors contain tumor cell populations that are heterogeneous in angiogenic activity. Here, we separated angiogenic from nonangiogenic human tumor cell populations and compared their growth.
Severe combined immunodeficient (SCID) mice were inoculated with nonangiogenic human MDA-MB-436 breast adenocarcinoma, KHOS-24OS osteosarcoma, or T98G glioblastoma cells. Most of the resulting tumors remained microscopic (<1 mm diameter), but some eventually became angiogenic and enlarged and were used to isolate angiogenic tumor cells. Angiogenic and nonangiogenic tumor cells were inoculated into SCID mice, and time to the development of palpable tumors was determined. Cell proliferation was assayed in vitro by growth curves and in vivo by staining for proliferating cell nuclear antigen or Ki67. Microscopic tumors from both tumor cell populations were examined for histologic evidence of vascular development 14 days after inoculation in mice. Expression of the angiogenesis inhibitor thrombospondin-1 was examined by immunoblotting.
Nonangiogenic tumors of each tumor type developed palpable tumors after means of 119 days (range: 53-185 days) for breast cancer, 238 days (184-291 days) for osteosarcoma, and 226 days (150-301 days) for glioblastoma. Angiogenic cells developed palpable tumors within 20 days after inoculation. However, nonangiogenic and angiogenic cells of each tumor type had similar proliferation rates. Fourteen days after tumor cell inoculation, tumors from angiogenic cells showed evidence of functional vasculature. In contrast, nonangiogenic tumors remained microscopic in size with absent or nonfunctional vasculature. Thrombospondin-1 expression was statistically significantly lower (by five- to 23-fold, depending on tumor type) in angiogenic than nonangiogenic cells.
This model provides a conceptual framework and a reproducible in vivo system to study unresolved central questions in cancer biology regarding the initiation, reversibility, and molecular regulation of the timing of the angiogenic switch.
人类微观癌症可能终生保持休眠状态。肿瘤进展取决于一系列事件，包括转变为血管生成表型，即最初招募新血管。我们之前证明，人类肿瘤包含血管生成活性异质的肿瘤细胞群体。在此，我们将人类血管生成性肿瘤细胞群体与非血管生成性肿瘤细胞群体分离，并比较它们的生长情况。
将非血管生成性人类MDA-MB-436乳腺腺癌、KHOS-24OS骨肉瘤或T98G胶质母细胞瘤细胞接种到严重联合免疫缺陷（SCID）小鼠体内。大多数由此产生的肿瘤保持微观状态（直径<1毫米），但有些最终变为血管生成性并增大，被用于分离血管生成性肿瘤细胞。将血管生成性和非血管生成性肿瘤细胞接种到SCID小鼠体内，并确定可触及肿瘤出现的时间。通过生长曲线在体外测定细胞增殖，通过增殖细胞核抗原或Ki67染色在体内测定细胞增殖。在将肿瘤细胞接种到小鼠体内14天后，检查来自这两种肿瘤细胞群体的微观肿瘤有无血管发育的组织学证据。通过免疫印迹法检测血管生成抑制剂血小板反应蛋白-1的表达。
每种肿瘤类型的非血管生成性肿瘤在接种后出现可触及肿瘤的平均时间分别为：乳腺癌119天（范围：53 - 185天），骨肉瘤238天（184 - 291天），胶质母细胞瘤226天（150 - 301天）。血管生成性细胞在接种后20天内出现可触及肿瘤。然而，每种肿瘤类型的非血管生成性和血管生成性细胞具有相似的增殖率。肿瘤细胞接种14天后，来自血管生成性细胞的肿瘤显示出功能性脉管系统的证据。相比之下，非血管生成性肿瘤在大小上仍保持微观状态，脉管系统不存在或无功能。血管生成性细胞中血小板反应蛋白-1的表达在统计学上显著低于非血管生成性细胞（低5至23倍，取决于肿瘤类型）。
该模型提供了一个概念框架和一个可重复的体内系统，用于研究癌症生物学中关于血管生成开关启动、可逆性和时间分子调控的未解决核心问题。
