2026-11-092026-11-09https://scholars.csmu.edu.tw/handle/123456789/37921Cell behavior is tightly controlled by coordination of signals elicited from growth factors, cell-cell and cell-extracellular matrix (ECM) interactions. Dynamic and reciprocal interplay between cells and their microenvironment occurs in order for cells to adjust themselves and the external milieu to maintain proper phenotype. In mammary epithelia, cell differentiation requires at least two types of signal, the lactogenic hormone (prolactin, IGF-I and hydrocortisone) and the basement membrane (BM). Crosstalk between ECM- and growth factor-mediated pathways occurs whereby cell adhesion to ECM affects growth factor signal propagation. Our previous work has shown that cell-ECM interactions affect insulin-like growth factor-I (IGF-I) signaling in mammary cells. IGF-I signaling downstream of IGF-I receptor (IGFIR) propagates better in cells cultured on BM than those cultured on plastic, suggesting that a novel ECM-dependent control point in IGF-I signaling is at the level of tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1). Thus, BM regulates IGF-I-triggered mammary functions by controlling the permissiveness of IGF-I signaling. In this study, we will further investigate into the mechanism for ECM-mediated regulation of IGF-I signaling in mammary epithelial cells. Our working hypothesis is that ECM via its cognate receptor and the downstream signaling events alters gene expression, in turn, affecting IGF-I signaling. Three different strategies are used to solve this problem. Firstly we will try to approach this question from the point of view of IGF-I signal transduction, especially focusing on those possibilities that cause a block in tyrosine phosphorylation of IRS-1, including the aberrant association of protein tyrosine phosphatase (PTP) or inhibitory molecules with IGFIR/IRS-1, or improper serine phosphorylation of IRS-1. BM, its cognate receptor(s) and the downstream signaling pathways are absolutely the determinants for modulation of IGF-I signaling in mammary cells. Therefore, seeking out the differences in signaling pathways triggered by cell adhesion to plastic or BM may help to spot the molecules involved in the modulation of IGF signaling. One dramatic effect of BM on cells is that it signals cells to form 3D spheroid morphology. Whether this cell shape change is prerequisite for IGF-I signaling will also be examined. Lastly, identification of the proteins whose expression is differentially regulated by cell adhesion to ECM may lead to a revelation of the mechanism as we speculate that ECM controls IGF-I signaling through reprogramming of gene expression. TGF-b and LAR have been shown to be upregulated in cells cultured on plastic and BM, respectively. We will thus start with the inspection of their involvement in the modulation of IGF-I signaling. Examination of the involvement of other genes identified by microarray analysis will be performed afterwards. Following the delineation of the mechanism for ECM-mediated control on IGF-I signaling, we wish to apply this information in cancer studies. Whether the dependency of IGF signaling on BM is missing and what causes this deregulaton in cancer cells will be examined. We hope these studies would help for deciphering the control of normal development of mammary gland and furthermore, for designing strategies to tackle progressive diseases such as breast cancer.細胞的生理反應是經由統整週遭環境中種種訊息所控制，而這些訊息是來自生長因子、相鄰的細胞以及細胞外基質(extracellular matrix)。細胞與其環境會相互影響並作持續調整以維持細胞特有的表型。對乳腺上皮細胞而言，細胞分化需要至少兩種訊號；一是來自激素，包括了泌乳激素、類胰島素生長因子(IGF-I)和氫皮質酮，另一種則是細胞與基底膜(basement membrane)的接觸。我們發現在乳腺上皮細胞，細胞外基質會影響IGF-I訊息傳遞。IGF-I受體的酪胺酸磷酸化並不受細胞外基質的影響 而自IGF-I受體下游的訊息傳遞則需要細胞與基底膜(basement membrane)的接觸。這表示細胞外基質調節IGF-I訊息傳遞在於控制insulin receptor substrate-1 (IRS-1)的酪胺酸磷酸化。因此我們認為乳腺上皮細胞之所以同時需要基底膜和恰當的生長因子來推動其功能是因為當細胞黏附於基底膜上時，它所產生的訊號可以促進生長因子的訊號傳遞。 在這個研究計劃中，我們將更進一步探討基底膜調節IGF-I訊息傳遞所扮演的角色。我們的假設是基底膜經由其細胞膜上的受體激活特定的訊息路徑改變基因的表現，再藉此影響IGF-I訊息傳遞。三種不同的策略將被用來解決此問題。首先我們單純以訊息傳遞的角度來看，會造成IRS-1酪胺酸磷酸化被抑制但受體磷酸化卻不受影響的原因有酪胺酸去磷酸脢的參與促進去磷酸化;一些抑制分子阻斷IRS-1結合到IGF-I受體上。另外當IRS-1產生絲胺酸磷酸化時會抑制其酪胺酸磷酸化。我們將探討當細胞培養在一般的塑膠盤上其IGF-I訊息傳遞受阻是否肇因於此。另外，從另一個的角度來看，基底膜的存在與否決定訊息傳遞是否可以進行，我們推測基底膜所誘發的特殊訊息路徑可能是調控IGF-I訊息傳遞的主要因子。因此，我們將找出那些是基底膜所誘發的特殊訊息並檢測其對IGF-I訊息傳遞的影響。此外細胞培養在基底膜上會形成腺體狀的構造，是否細胞的型態會影響IGF-I訊息傳遞也將被探討。以前的結果顯示，基底膜和第一型膠原蛋白對IRS-1酪胺酸磷酸化所產生的影響需等到細胞貼附兩天後才會最明顯。因為需要如此長的時間，因此我們推測細胞外基質調節IGF-I的訊息傳遞是藉由一些新合成的分子來達成。找出此種分子並檢測其對IGF-I訊息傳遞的影響則是我們的第三種策略。因細胞培養在一般的塑膠盤上具較高的TGF-b，而細胞培養在基底膜上則是LAR的表現較高。所以我們將先檢測此二分子對IGF-I訊息傳遞的影響。在釐清細胞外基質調節IGF-I訊息傳遞機制之後，我們希望將此資訊運用在癌症的研究上。癌細胞常能逃脫細胞內的調控以提高其生長，移動及存活。我們接著將探討在乳癌細胞中IGF-I訊息傳遞是否也受細胞外基質的影響。我們希望藉此研究能更加了解細胞外間質對正常乳腺細胞的發育所扮演的角色，並藉此提供治療癌症(乳癌)的方法。基底膜類胰島素生長因子訊息傳遞basement membraneIGFsignal transductionProject