SEQUENTIAL DEGRADATION OF HEPARAN-SULFATE IN THE SUBENDOTHELIAL EXTRACELLULAR-MATRIX BY HIGHLY METASTATIC LYMPHOMA-CELLS
SEQUENTIAL DEGRADATION OF HEPARAN-SULFATE IN THE SUBENDOTHELIAL EXTRACELLULAR-MATRIX BY HIGHLY METASTATIC LYMPHOMA-CELLS
复制标题
DOI：
10.1002/ijc.2910350411
复制
发表时间：
1985-01-01
影响因子：
6.4
通讯作者：
VLODAVSKY, I
中科院分区：
文献类型：
--
作者：
BARNER, M;KRAMER, MD;VLODAVSKY, I
甲基胆蒽诱导的T淋巴瘤的高转移性变体（ESb）比其非转移性亲本亚系（Eb）更高程度地阐述硫酸乙酰肝素（HS）降解糖苷内切酶（乙酰肝素酶）。而无血清培养基中含有胰蛋白酶样丝氨酸蛋白酶的亚系条件，乙酰肝素酶活性检测仅在ESB条件培养基（CM）。将ESb CM与天然产生的硫酸盐标记的内皮下细胞外基质（ECM）或与可溶性高分子量标记的蛋白聚糖一起孵育，所述蛋白聚糖首先通过与Eb CM或与部分纯化的ESb蛋白酶一起孵育从ECM释放。在Sepharose 6 B上通过凝胶过滤分析硫酸盐标记的降解产物。ECM结合HS降解的最佳pH为6.2，而可溶性蛋白聚糖降解的最佳pH为5.2。肝素酶介导的ECM结合和可溶性HS的降解被肝素抑制。添加胰蛋白酶，纤溶酶或在较低程度上，纯化的ESb蛋白酶，刺激之间的S-和20倍的ESb CM介导的ECM结合HS的降解，但对乙酰肝素酶介导的可溶性蛋白多糖的降解没有影响。这种刺激在肝素或蛋白酶抑制剂的存在下被抑制。这些结果表明，蛋白酶和肝素酶都参与了ESB介导的ECM结合的HS的降解，并且1种酶为下一种酶产生更易接近的底物。这种连续切割是多分子结构（如内皮下ECM）降解的特征，因此在其分离组分的研究中无法检测到。
A highly metastatic variant (ESb) of a methylcholanthrene-induced T lymphoma elaborates a heparan sulfate (HS) degrading endoglycosidase (heparanase) to a much higher extent than its non-metastatic parental subline (Eb). Whereas a serum-free medium conditioned by either subline contained a trypsin-like serine protease, heparanase activity was detected only in the ESb-conditioned medium (CM). ESb CM was incubated with a naturally produced, sulfate-labeled subendothelial extracellular matrix (ECM) or with a soluble, high-MW labeled proteoglycan first released from the ECM by incubation with Eb CM or with the partially purified ESb protease. Sulfate labeled degradation products were analyzed by gel filtration on Sepharose 6B. The optimal pH for degradation of ECM-bound HS was 6.2 as compared to pH 5.2 for degradation of the soluble proteoglycan. Heparanase-mediated degradation of both ECM-bound and soluble HS was inhibited by heparin. Addition of either trypsin, plasmin or to a lower extent, the purified ESb protease, stimulated between S- and 20-fold the ESb CM-mediated degradation of ECM-bound HS but had no effect on heparanase-mediated degradation of the soluble proteoglycan. This stimulation was inhibited in the presence of heparin or protease inhibitors. These results indicate that both a protease and heparinase are involved in the ESb-mediated degradation of ECM-bound HS and that 1 enzyme produces a more accessible substrate for the next enzyme. This sequential cleavage is characteristic of degradation of a multimolecular structure such as the subendothelial ECM and hence cannot be detected in studies with its isolated constituents.