Proteomic Identification of Insulin-like Growth Factor-binding Protein-6 Induced by Sublethal H2O2 Stress from Human Diploid Fibroblasts

Abstract

Fibroblasts are the most ubiquitous cell types within our body. They produce various factors to maintain the texture and structure of a particular organ or tissue. To identify protein factors secreted by fibroblasts and alteration of these protein factors upon oxidative stress, HCA 3 human skin diploid fibroblasts were exposed to a sublethal dose of H 2 O 2 , which induces a prematurely senescent phenotype. Conditioned media from prematurely senescent cells versus control cells were analyzed for proteins using an LC-MS/MS-based proteomic technique. Collagen ␣1(VI), collagen ␣2(I), fibronectin, lumican, and matrix metalloproteinase 2 were among the proteins consistently detected from control and H

Promotion of Cancer Cell Migration

“…It is noteworthy that IGFBP-6 expression is increased by cellular stressors, including H 2 O 2 -induced oxidative stress (47). p38 activation has apparently contradictory effects.…”

TCDD induces the expression of insulin-like growth factor binding protein 4 in 5L rat hepatoma cells: A cautionary tale of the use of this cell line in studies on dioxin toxicity

“…However, several observations (Park et al, 2003;Guo et al, 2007) indicated that IGFBP-6 induction by TCDD is not mediated in the classical AhR and XRE-mediated way. In fact, evidence has been presented that IGFBP-6 induction is a general response to cell degeneration or injury in vitro and in vivo and unrelated to the mechanism of induction of AhR gene battery members (Xie et al, 2005).…”

Genomic and proteomic profiling of oxidative stress response in human diploid fibroblasts

“…The cell lysates used here do not contain secreted proteins, although microarray can detect genes encoding secreted proteins, for example MMP3 and IGFBP6. IGFBP6 was found elevated in H 2 O 2 treated HDFs while profiling the secreted proteins (Xie et al 2005). In addition to the absence of secreted proteins for proteomic detection, we used human 5 K cDNA gene chip for the microarray analyses in this study.…”

“…Each sample was diluted by and dialyzed against 0.01 N NH 4 HCO 3 , passed through a 0.45-μm filter to remove insoluble cell debris, and concentrated down using a speed vacuum concentrator. After protein concentration determination by the Bradford method (Bio-Rad, Hercules, CA), protein mixtures from whole cell lysates were digested overnight with trypsin at a 50:1 ratio (Xie et al 2005). A microbore HPLC system (Paradigm MS4, Michrom, Auburn, CA) was used with two separation columns: a reverse phase (RP) column and a strong cation exchange (SCX) column (Whatman, Clifton, NJ).…”