Stable cell fate changes in marrow cells induced by lung-derived microvesicles.
Stable cell fate changes in marrow cells induced by lung-derived microvesicles.
复制标题
DOI：
10.3402/jev.v1i0.18163
复制
发表时间：
2012
影响因子：
16
通讯作者：
Quesenberry PJ
中科院分区：
文献类型：
--
作者：
Aliotta JM;Pereira M;Li M;Amaral A;Sorokina A;Dooner MS;Sears EH;Brilliant K;Ramratnam B;Hixson DC;Quesenberry PJ
人们对细胞来源的微囊改变不同靶细胞命运的能力产生了兴趣。肺、肝、心和脑来源的囊泡可以改变小鼠骨髓细胞的遗传表型；然而，这种变化的稳定性和这些变化的机制尚不清楚。在本工作中，我们证明了肺源性微囊泡最初通过转录转移的基因和调控因子(S)改变了靶细胞的转录组和蛋白质组，但长期的表型改变仅仅是由于转录调控因子与靶细胞的转移。体内研究：将全骨髓细胞(WBM)与LDMV(均取自雄性C57BL/6小鼠)共培养或单独培养(对照组)。一周后，将培养的WBM移植到致死剂量照射的雌性C57BL/6小鼠体内。6周后处死受体小鼠，通过实时定量RT-PCR检测WBM、脾和肝脏中肺表面活性物质A、B、C和D、水通道蛋白-5和Clara细胞特异性蛋白的表达。肺组织进行免疫组织化学染色，以确定移植的骨髓来源(Y染色体+)II型肺泡细胞(前表面活性物质C+)的数量。移植LDMV联合培养的WBM的小鼠在骨髓、肝脏和脾细胞中表达肺上皮细胞基因，移植的骨髓来源的Y+/前表面活性C+细胞在肺中的数量是对照组的5倍以上。体外实验：将WBM(小鼠或大鼠)加入或不加入LDMV(小鼠或大鼠)培养1周，然后单独清洗和培养。每隔两周收集一次WBM，用于实时RT-PCR分析，使用物种特异性表面活性物质引物，并用于Western Blot分析。蛋白质组学和微RNA微阵列分析也在细胞上进行。LDMV共培养的WBM可维持肺上皮细胞基因和蛋白的表达达12周。在以后的时间点产生的表面活性物质仅针对培养中的骨髓细胞种类，表明从头开始转录。这些发现，除了LDMV共同培养的WBM的蛋白质和microRNA图谱发生变化外，还支持这些变化的稳定转录机制。这些数据表明，微泡对细胞命运的改变是强大和长期的，代表了细胞生物学的一个重要新方面。
Interest has been generated in the capacity of cellular-derived microvesicles to alter the fate of different target cells. Lung, liver, heart and brain-derived vesicles can alter the genetic phenotype of murine marrow cells; however, the stability of such changes and the mechanism of these changes remain unclear. In the present work, we show that lung-derived microvesicles (LDMV) alter the transcriptome and proteome of target marrow cells initially by mRNA and regulator(s) of transcription transfer, but that long term phenotype change is due solely to transfer of a transcriptional regulator with target cell. In vivo studies: Whole bone marrow cells (WBM) were co-cultured with LDMV (both isolated from male C57BL/6 mice) or cultured alone (control). One week later, cultured WBM was transplanted into lethally-irradiated female C57BL/6 mice. Recipient mice were sacrificed 6 weeks later and WBM, spleens and livers were examined for the presence of lung-specific gene expression, including surfactants A, B, C and D, aquaporin-5, and clara cell specific protein, via real-time RT-PCR. Immunohistochemistry was also performed on lungs to determine the number of transplanted marrow-derived (Y chromosome+) type II pneumocytes (prosurfactant C+). Mice transplanted with LDMV co-cultured WBM expressed pulmonary epithelial cell genes in the cells of their bone marrow, livers and spleens and over fivefold more transplanted marrow-derived Y+/prosurfactant C+cells could be found in their lungs (vs. control mice). In vitro studies: WBM (from mice or rats) was cultured with or without LDMV (from mice or rats) for 1 week then washed and cultured alone. WBM was harvested at 2-week intervals for real-time RT-PCR analysis, using species-specific surfactant primers, and for Western Blot analysis. Proteomic and microRNA microarray analyses were also performed on cells. LDMV co-cultured WBM maintained expression of pulmonary epithelial cell genes and proteins for up to 12 weeks in culture. Surfactant produced at later time points was specific only to the species of the marrow cell in culture indicating de novo mRNA transcription. These findings, in addition to the altered protein and microRNA profiles of LDMV co-cultured WBM, support a stable transcriptional mechanism for these changes. These data indicate that microvesicle alteration of cell fate is robust and long-term and represents an important new aspect of cellular biology.