Zarnegar R, Muga S, Rahija R, Michalopoulos G
Department of Pathology, Duke University Medical Center, Durham, NC 27710.
Proc Natl Acad Sci U S A. 1990 Feb;87(3):1252-6. doi: 10.1073/pnas.87.3.1252.
Hepatopoietin-A (HPTA) is a heparin-binding polypeptide growth factor which consists of a heavy and a light polypeptide chain with molecular weights of 70,000 and 35,000, respectively. It stimulates DNA synthesis in primary cultures of normal rat hepatocytes in serum-free medium. The complete purification and characterization of HPTA from rabbit serum were reported by us elsewhere. Recently we have determined the amino-terminal amino acid sequence of the rabbit HPTA light chain up to 24 residues and have shown that the sequence is not homologous with other known sequences. [N.B. Human hepatocyte growth factor, recently sequenced by two other groups, is the same molecular species as HPTA.] In the present paper we report the production of a neutralizing polyclonal antiserum raised in chicken against purified rabbit HPTA. This antiserum does not inhibit the mitogenic effect of other potent inducers of hepatocyte DNA synthesis (epidermal growth factor or acidic fibroblast growth factor), nor does it interact with these growth factors in an enzyme-linked immunosorbent assay (ELISA). The antibody recognizes HPTA, as was determined by Western immunoblotting. Since the tissue origin of HPTA is not known, this anti-HPTA antiserum was used to investigate the tissue distribution of HPTA in rabbits by immunohistostaining methods. Acinar cells of the pancreas, neurons of the brain, C cells of the thyroid, ductal cells of the salivary glands, and Brunners glands of the duodenum stained with anti-HPTA antibody. Liver, spleen, thymus, and kidney do not seem to contain appreciable amounts of HPTA. We confirmed these findings by extracting and purifying active HPTA from the stained tissues listed above. The anti-HPTA antibody recognizes HPTA purified from different tissues, as was determined by ELISA, Western immunoblotting, and immunoneutralization experiments.
肝生成素 - A（HPTA）是一种肝素结合多肽生长因子，它由一条重链和一条轻链多肽组成，分子量分别为70,000和35,000。它能在无血清培养基中刺激正常大鼠肝细胞原代培养物中的DNA合成。我们已在其他地方报道了从兔血清中对HPTA进行的完全纯化和特性鉴定。最近，我们已确定兔HPTA轻链的氨基末端氨基酸序列，长达24个残基，并表明该序列与其他已知序列不同源。[注意：最近另外两个研究小组测定了序列的人肝细胞生长因子与HPTA是同一分子种类。]在本文中，我们报道了用纯化的兔HPTA在鸡体内产生的一种中和多克隆抗血清。这种抗血清不抑制其他肝细胞DNA合成有效诱导剂（表皮生长因子或酸性成纤维细胞生长因子）的促有丝分裂作用，在酶联免疫吸附测定（ELISA）中也不与这些生长因子相互作用。通过蛋白质免疫印迹法确定，该抗体能识别HPTA。由于HPTA的组织来源尚不清楚，因此用这种抗HPTA抗血清通过免疫组织化学染色方法研究兔体内HPTA的组织分布。胰腺的腺泡细胞、脑的神经元、甲状腺的C细胞、唾液腺的导管细胞和十二指肠的布伦纳腺均被抗HPTA抗体染色。肝、脾、胸腺和肾似乎不含大量的HPTA。我们通过从上述染色组织中提取和纯化活性HPTA证实了这些发现。通过ELISA、蛋白质免疫印迹法和免疫中和实验确定，抗HPTA抗体能识别从不同组织中纯化的HPTA。
