Latent transforming growth factor β1 activation in situ: quantitative and functional evidence after low‐dose γ‐irradiation 1

Abstract

The biological activity of transforming growth factor β1 (TGF‐β) is controlled by its secretion as a latent complex in which it is noncovalently associated with latency‐associated peptide (LAP). Activation is the extracellular process in which TGF‐β is released from LAP, and is considered to be a primary regulatory control. We recently reported rapid and persistent changes in TGF‐β immunoreactivity in conjunction with extracellular matrix remodeling in γ‐irradiated mouse mammary gland. Our hypothesis is that these specific changes in immunoreactivity are indicative of latent TGF‐β activation. In the present study, we determined the radiation dose response and tested whether a functional relationship exists between radiation‐induced TGF‐β and collagen type III remodeling. After radiation exposures as low as 0.1 Gy, we detected increased TGF‐β immunoreactivity in the mammary epithelium concomitant with decreased LAP immunostaining, which are events consistent with activation. Quantitative image analysis demonstrated a significant ( P = 0.0005) response at 0.1 Gy without an apparent threshold and a linear dose response to 5 Gy. However, in the adipose stroma, loss of LAP demonstrated a qualitative threshold at 0.5 Gy. Loss of LAP paralleled induction of collagen III immunoreactivity in this tissue compartment. We tested whether TGF‐β mediates collagen III expression by treating animals with TGF‐β panspecific monoclonal antibody, 1D11.16, administered i.p. shortly before irradiation. Radiation‐induced collagen III staining in the adipose stroma was blocked in an antibody dose‐dependent manner, which persisted through 7 days postirradiation. RN‐ase protection assay revealed that radiation‐induced elevation of total gland collagen III mRNA was also blocked by neutralizing antibody treatment. These data provide functional confirmation of the hypothesis that radiation exposure leads to latent TGF‐β activation, support our interpretation of the reciprocal shift in immunoreactivity as evidence of activation, and implicate TGF‐β as a mediator of tissue response to ionizing radiation. The sensitivity of activation to low radiation doses points to a potential role for TGF‐β in orchestrating tissue response to oxidative stress. As such, radiation may be useful as a probe to delineate the consequences of latent TGF‐β activation in situ.—Ehrhart, E. J., Segarini, P., Tsang, M. L.‐S., Carroll, A. G., Barcellos‐Hoff, M. H. Latent transforming growth factor‐β1 activation in situ: quantitative and functional evidence after low‐dose γ‐irradiation. FASEB J. 11, 991–1002 (1997)