Metalloproteinase activity is the sole factor responsible for the growth-promoting effect of conditioned medium in Trichoplusia ni insect cell cultures

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• 作者：Ulrika Eriksson ; Jenny Hassel ; Elke Lü ; llau and Lena H4 ; ggströ ; m
• 关键词：Trichoplusia ni ; Serum-free medium ; Conditioned medium ; Gel filtration fractions ; Metalloproteinase activity ; Autocrine regulation of proliferation
• 刊名：Journal of Biotechnology
• 出版年：2005
• 出版时间：22 September 2005
• 年：2005
• 卷：119
• 期：1
• 页码：76-86
• 全文大小：205 K

文摘

Conditioned medium (CM) taken from a serum-free culture of Trichoplusia ni (BTI-Tn-5B1-4, High Five) cells on days 2 and 3, shortened the lagphase and increased the maximum cell density when added to T. ni cultures with low-inoculum cell density. Gel filtration fractions of CM, eluting at around 45 kDa, stimulated cell proliferation even better than CM. A protein in the gel filtration fraction was identified by N-terminal amino acid sequencing as a proteinase, related to a snake venom metalloproteinase. Casein zymography showed, multiple metalloproteinase bands between 48 and 25 kDa, as well as precursor forms above 48 kDa. Metalloproteinase bands below the main band at 48 kDa were autocatalytic degradation products. Metalloproteinase activity was the sole factor responsible for the growth stimulating effect of CM as shown by using the specific metalloproteinase inhibitor dl-thiorphan. Metalloproteinases have recently been shown to release growth factors from sequestering extracellular proteins. We propose that the metalloproteinase is involved in autocrine regulation of T. ni proliferation in serum-free media. In addition, a gel filtration fraction of CM, eluting at about 10 kDa, inhibited cell growth. Apart from a lysozyme precursor protein and a cyclophilin-like protein, a kazal-type proteinase inhibitor could be identified in this fraction.