Changes in histology and fibrogenic cytokines in irradiated colorectum of two murine strains.

PURPOSE

A strain difference in the development of radiation-induced fibrosis of the colorectum was recently observed. C57B1/6 mice developed colorectal obstruction with significantly higher incidence compared to C3Hf/Kam mice after partial volume irradiation with 30 Gy. Previous reports have demonstrated differences in cytokine mRNA levels in fibrosis-prone and -resistant mice after lung irradiation. The aims of this study are to determine if there are strain differences in: 1) the histology of the lesion, 2) mRNA levels for transforming growth factor beta (TGFbeta) isoforms and tumor necrosis factor alpha (TNFalpha), and 3) immunohistochemical staining patterns using antibodies against the TGFbeta isoforms and latency-associated peptide (LAP).

METHODS AND MATERIALS

The colorectum of male C3Hf/Kam (C3H) and C57B1/6 (B6) mice were irradiated using a dose/length combination (30 Gy to 13.7 mm) that resulted in 10 or 100% incidence of obstruction by 6 months in each strain, respectively. Colorectal tissue was removed from 6 hours to 120 days after irradiation as well as from obstructed mice and prepared for histology, RNase protection assay, and immunofluorescence.

RESULTS

Distinct differences in the histological phenotype for the two strains were observed at times preceding obstruction. Samples from B6 mice showed increased hyperplastic crypts, colitis cystica profunda, and fibrosis within the lamina propria, compared to identically treated C3H mice. Fibrosis in the lamina propria of B6 mice appeared early, beginning at 75 days after irradiation, and was progressive, whereas fibrosis in C3H mice appeared simultaneous with obstruction and may have been a reaction to ulceration. No consistent strain difference in mRNA levels for TGFbeta1, 2, 3 or TNFalpha were observed, although mRNA levels of TGFbeta1 and TNFalpha were significantly elevated in both strains relative to nonirradiated controls. Immunofluorescent staining for TGFbeta1, 3 and LAP was observed in hyperplastic crypts and colitis cystica profunda adjacent to regions of fibrosis, histological changes that were present predominately in the B6 strain.

CONCLUSIONS

The response of the colorectum to irradiation involves changes in the expression of several different cytokines. However, the lack of a consistent strain difference in TGFbeta1, 2, 3 and TNFalpha mRNA levels, despite strain differences in both the incidence of colorectal obstruction and histological features preceding obstruction, suggests that mRNA changes in these fibrogenic cytokines are not the critical determinant of the strain difference and are not related to the process of radiation-induced colorectal fibrosis in these mouse strains. Strain-dependent differences were observed in the localization of active TGFbeta, but these differences were related to the histological changes specifically found in the irradiated colon of the B6 strain.