The cell’s resistance to cell death by adhesion loss to extracellular matrix (anoikis), contributes to tumor progression and metastasis. Various adhesion molecules are involved in the anoikis resistance, including the syndecan-4 (SDC4), a heparan sulfate proteoglycan (HSPG) present on the cell surface. Changes in the expression of SDC4 have been observed in tumor and transformed cells, indicating its involvement in cancer. In previous works, we demonstrated that acquisition of anoikis resistance resistance by blocking adhesion to the substrate up-regulates syndecan-4 expression in endothelial cells. This study investigates the role of SDC4 in the transformed phenotype of anoikis resistant endothelial cells. Anoikis-resistant endothelial cells (Adh1-EC) were transfected with micro RNA interference (miR RNAi) targeted against syndecan-4. The effect of SDC4 silencing was analyzed by real-time PCR, western blotting and immunofluorescence. Transfection with miRNA-SDC4 resulted in a sequence-specific decrease in syndecan-4 mRNA and protein levels. Furthermore, we observed a reduction in the number of heparan and chondroitin sulfate chains in the cell extract and culture medium. The SDC4 silencing led to downregulation of proliferative and invasive capacity and angiogenic abilities of anoikis-resistant endothelial cells. Compared with the parental cells (Adh1-EC), SDC4 silenced cells (SDC4 miR-Syn-4-1-Adh1-EC e miR-Syn-4-2-Adh1-EC) exhibited an increase in adhesion to collagen and laminin and also in the apoptosis rate. Moreover, transfection with miRNA-SDC4 caused a decrease in the number of cells in the S phase of the cell cycle. This is accompanied by an increase in the heparan sulfate synthesis after 12 h of simulation with fetal calf serum (FCS). SDC4 silencing cells are more dependent of growth factors present in the FCS to synthesize heparan sulfate than parental cells. Similar data were obtained for the wild-type cell line (EC). Our results indicated that downregulation of SDC4 expression reverses the transformed phenotype of anoikis resistant endothelial cells. These and other findings suggest that syndecan-4 is suitable for pharmacological intervention, making it an attractive target for cancer therapy.

细胞对细胞外基质（失巢凋亡）的粘附丧失而对细胞死亡的抵抗力有助于肿瘤进展和转移。各种粘附分子参与抗失巢凋亡，包括 Syndecan-4 (SDC4)，一种存在于细胞表面的硫酸乙酰肝素蛋白聚糖 (HSPG)。在肿瘤和转化细胞中观察到 SDC4 表达的变化，表明其与癌症有关。在以前的工作中，我们证明了通过阻断与底物的粘附来获得失巢凋亡抗性可上调内皮细胞中的 syndecan-4 表达。本研究调查了 SDC4 在抗失巢凋亡内皮细胞的转化表型中的作用。抗失巢凋亡的内皮细胞 (Adh1-EC) 用靶向 syndecan-4 的微 RNA 干扰 (miR RNAi) 转染。通过实时 PCR、蛋白质印迹和免疫荧光分析 SDC4 沉默的影响。用 miRNA-SDC4 转染导致 syndecan-4 mRNA 和蛋白质水平的序列特异性降低。此外，我们观察到细胞提取物和培养基中乙酰肝素和硫酸软骨素链的数量减少。SDC4 沉默导致抗失巢凋亡内皮细胞的增殖和侵袭能力以及血管生成能力的下调。与亲本细胞 (Adh1-EC) 相比，SDC4 沉默细胞 (SDC4 miR-Syn-4-1-Adh1-EC e miR-Syn-4-2-Adh1-EC) 表现出对胶原蛋白和层粘连蛋白的粘附增加，并且也体现在细胞凋亡率上。此外，转染 miRNA-SDC4 导致细胞周期 S 期的细胞数量减少。这伴随着用胎牛血清 (FCS) 模拟 12 小时后硫酸乙酰肝素合成增加。SDC4 沉默细胞比亲代细胞更依赖 FCS 中存在的生长因子来合成硫酸乙酰肝素。野生型细胞系 (EC) 获得了类似的数据。我们的结果表明，SDC4 表达的下调逆转了抗失巢凋亡内皮细胞的转化表型。这些和其他发现表明，syndecan-4 适用于药物干预，使其成为癌症治疗的有吸引力的靶点。我们的结果表明，SDC4 表达的下调逆转了抗失巢凋亡内皮细胞的转化表型。这些和其他发现表明，syndecan-4 适用于药物干预，使其成为癌症治疗的有吸引力的靶点。我们的结果表明，SDC4 表达的下调逆转了抗失巢凋亡内皮细胞的转化表型。这些和其他发现表明，syndecan-4 适用于药物干预，使其成为癌症治疗的有吸引力的靶点。