Uchiyama H, Barut B A, Mohrbacher A F, Chauhan D, Anderson K C
Division of Tumor Immunology, Dana-Farber Cancer Institute, Boston, MA 02115.
Blood. 1993 Dec 15;82(12):3712-20.
Previous studies show that human myeloma-derived cell lines specifically adhere to fibronectin (FN) through very late antigen-4 (VLA-4; alpha 4 beta 1 integrin complex) and RGD-peptide mechanisms, which may contribute to the localization of tumor cells in bone marrow (BM). In these studies, we characterized the adhesion of myeloma-derived cell lines to both normal and myeloma BM stromal cells (BMSCs) and the effect of adhesion on DNA synthesis. Because interleukin-6 (IL-6) plays an important role in the pathogenesis of multiple myeloma, we also examined the effects of tumor cell adhesion on IL-6 secretion by BMSCs. In 51chromium binding assays, the U266, ARH-77, and IM-9 cell lines showed 52% +/- 12%, 55% +/- 6%, and 47% +/- 7% specific adherence, respectively, to normal BMSCs and 74% +/- 4%, 60% +/- 3%, and 61% +/- 6% specific adherence, respectively, to myeloma BMSCs. In contrast, only 12% to 13% specific binding of HS-Sultan cells to BMSCs was noted. The binding of myeloma cells to BMSCs was partially blocked with anti-beta 1 monoclonal antibody (MoAb), anti-beta 2 integrin MoAb, and excess RGD peptide, suggesting multiple mechanisms for the adhesion of myeloma cell lines to BMSCs. Binding of cell lines to FN or myeloma BMSCs did not affect cell line proliferation; however, adhesion of myeloma cell lines to normal BMSCs decreased DNA synthesis, ie, stimulation indices are 0.1 +/- 0.04, 0.2 +/- 0.1, 0.2 +/- 0.07, and 0.1 +/- 0.06 for the adherent non-IL-6-dependent U266, ARH-77, HS-Sultan, and IM-9 cells, respectively (n = 5, P < .01). In contrast, adherence of IL-6-dependent B9 cells increased their proliferation (stimulation index, 3.2 +/- 0.7). Significant (twofold to eightfold) increases in IL-6 secretion were evident in cell line-adherent (> or = 12 hours) normal and myeloma BMSC cultures. Paraformaldehyde fixation of BMSCs before adhesion completely abrogated IL-6 secretion, suggesting that IL-6 secretion was triggered in BMSCs rather than in cell lines. Partial blocking of cell line adhesion to BMSCs, using anti-beta 1 integrin and anti-beta 2 integrin MoAbs and RGD peptide, also partially blocked the triggering of IL-6 secretion by BMSCs. When cell lines were placed in Transwell inserts and then cultured with either normal or myeloma BMSCs, permitting juxtaposition without cell to cell contact between myeloma cell lines and BMSCs, no increase in IL-6 secretion was observed.(ABSTRACT TRUNCATED AT 400 WORDS)
先前的研究表明，人骨髓瘤来源的细胞系通过极晚期抗原-4（VLA-4；α4β1整合素复合物）和RGD肽机制特异性地黏附于纤连蛋白（FN），这可能有助于肿瘤细胞在骨髓（BM）中的定位。在这些研究中，我们对骨髓瘤来源的细胞系与正常和骨髓瘤骨髓基质细胞（BMSC）的黏附特性以及黏附对DNA合成的影响进行了表征。由于白细胞介素-6（IL-6）在多发性骨髓瘤的发病机制中起重要作用，我们还研究了肿瘤细胞黏附对BMSC分泌IL-6的影响。在51铬结合试验中，U266、ARH-77和IM-9细胞系对正常BMSC的特异性黏附分别为52%±12%、55%±6%和47%±7%，对骨髓瘤BMSC的特异性黏附分别为74%±4%、60%±3%和61%±6%。相比之下，HS-Sultan细胞与BMSC的特异性结合仅为12%至13%。骨髓瘤细胞与BMSC的结合被抗β1单克隆抗体（MoAb）、抗β2整合素MoAb和过量的RGD肽部分阻断，提示骨髓瘤细胞系与BMSC黏附存在多种机制。细胞系与FN或骨髓瘤BMSC的结合不影响细胞系增殖；然而，骨髓瘤细胞系与正常BMSC的黏附降低了DNA合成，即对于非IL-6依赖的黏附性U266、ARH-77、HS-Sultan和IM-9细胞，刺激指数分别为0.1±0.04、0.2±0.1、0.2±0.07和0.1±0.06（n = 5，P <.01）。相反，IL-6依赖的B9细胞的黏附增加了其增殖（刺激指数，3.2±0.7）。在细胞系黏附（≥12小时）的正常和骨髓瘤BMSC培养物中，IL-6分泌显著增加（两倍至八倍）。在黏附前用多聚甲醛固定BMSC可完全消除IL-6分泌，提示IL-6分泌是在BMSC中而非细胞系中触发的。使用抗β1整合素和抗β2整合素MoAb以及RGD肽部分阻断细胞系与BMSC的黏附，也部分阻断了BMSC触发IL-6分泌。当将细胞系置于Transwell小室中，然后与正常或骨髓瘤BMSC一起培养，使骨髓瘤细胞系与BMSC并列但不发生细胞间接触时，未观察到IL-6分泌增加。（摘要截短至400字）
