Michel K, Roth S, Trautwein C, Gong W, Flemming P, Gressner A M
Department of Clinical Chemistry and Central Laboratory, Philipps University, Marburg, Germany.
Hepatology. 1998 Jun;27(6):1592-9. doi: 10.1002/hep.510270619.
Latent transforming growth factor beta binding protein (LTBP), a component of the extracellular matrix (ECM) of various tissues, is important for the secretion of TGF-beta and, furthermore, for the storage of TGF-beta in ECM. The proteolytic cleavage of LTBP is assumed to be the prerequisite for the activation of TGF-beta. We investigated the mRNA expression pattern of the three LTBP isoforms (LTBP-1, -2, -3) and the protein distribution of the components of the large latent TGF-beta complex, namely LTBP-1 and -2, latency-associated protein (LAP), and TGF-beta, in human liver using reverse-transcription polymerase chain reaction (RT-PCR) and immunohistochemical alkaline phosphatase anti-alkaline phosphatase (APAAP) staining. Parts of explanted livers diagnosed as hepatitis B, hepatitis C, primary biliary cirrhosis (PBC), and primary sclerosing cholangitis (PSC) and normal liver tissue were examined. LTBP transcripts were detected in the same manner in all liver specimens. Interestingly, we found a new splice variant of LTBP-1 (LTBP-1D), in which the sequence coding for the proteinase-sensitive hinge region is deleted. The corresponding parts of the human LTBP-2 and LTBP-3 cDNA coding for the hinge region were sequenced and show neither similar proteinase cleavage sites nor deleted cDNA sequences. The proposed proteinase cleavage site of mouse LTBP-3 seems not to be conserved in the human LTBP-3 gene. By immunohistochemistry, LTBP-1, -2, and LAP were detectable in normal and diseased livers and showed a different staining pattern for both LTBP isoforms. By contrast, TGF-beta showed a spotted staining pattern in diseased livers only, predominantly in the area of parenchymal cells that are close to fibrotic tissue. This strongly suggests the release of active TGF-beta from preexisting latent complexes. The LTBP-1D splice variant, which is probably less sensitive against proteolytic degradation and therefore may protect TGF-beta from activation, may have importance for modulating the biological activity of TGF-beta in normal and diseased liver.
潜伏转化生长因子β结合蛋白（LTBP）是各种组织细胞外基质（ECM）的一个组成部分，对TGF-β的分泌很重要，此外，对TGF-β在ECM中的储存也很重要。LTBP的蛋白水解切割被认为是TGF-β激活的先决条件。我们使用逆转录聚合酶链反应（RT-PCR）和免疫组织化学碱性磷酸酶抗碱性磷酸酶（APAAP）染色法，研究了三种LTBP亚型（LTBP-1、-2、-3）的mRNA表达模式以及大潜伏TGF-β复合物各成分，即LTBP-1和-2、潜伏相关蛋白（LAP）和TGF-β在人肝脏中的蛋白分布。对诊断为乙型肝炎、丙型肝炎、原发性胆汁性肝硬化（PBC）和原发性硬化性胆管炎（PSC）的部分切除肝脏以及正常肝脏组织进行了检查。在所有肝脏标本中均以相同方式检测到LTBP转录本。有趣的是，我们发现了LTBP-1的一种新的剪接变体（LTBP-1D），其中编码蛋白酶敏感铰链区的序列被删除。对人LTBP-2和LTBP-3 cDNA编码铰链区的相应部分进行了测序，结果显示既没有相似的蛋白酶切割位点，也没有缺失的cDNA序列。小鼠LTBP-3的推测蛋白酶切割位点似乎在人LTBP-3基因中不保守。通过免疫组织化学方法，在正常和患病肝脏中均可检测到LTBP-1、-2和LAP，并且两种LTBP亚型呈现出不同的染色模式。相比之下，TGF-β仅在患病肝脏中呈现斑点状染色模式，主要位于靠近纤维化组织的实质细胞区域。这强烈提示了活性TGF-β从预先存在的潜伏复合物中的释放。LTBP-1D剪接变体可能对蛋白水解降解不太敏感，因此可能保护TGF-β不被激活，这可能对调节正常和患病肝脏中TGF-β的生物学活性具有重要意义。
