Assay of premorbid murine jejunal fibrosis based on mechanical changes after X irradiation and hyperthermia

Abstract

Preparative surgery immobilized 15 mm of functional jejunum against the peritoneal surface of the ventral abdominal wall in C3H/HeJ mice. The surgery allowed subsequent treatments with single fractions of 44 degrees C hyperthermia and X irradiation to be selective to this portion of small intestine. With each doubling of time since treatment, 1 through 70 weeks, a sample of mice was killed and specimens of their intestines were excised and radially stretched in a tensile-testing apparatus that measured tension as a continuous function of circumference. Preconditioning with repeated cycles of stretch and relaxation before specimens were irreversibly stretched enabled measurement of the limit collagen placed on the extensibility of the intestinal wall by physiologic forces and the stiffness of the intestinal collagen once that limit was exceeded. Both kinds of measurements made possible dose-response characterization of radiation fibrosis for treatments that killed no mice. Response increased linearly with X-ray dose above a threshold. After X rays alone the threshold remained constant at 9.7 +/- 0.6 Gy for the assays at 1 through 8 weeks and subsequently decreased to about 6 Gy by 35 weeks. With adjuvant hyperthermia of 15 min at 44 degrees C beginning 10 min after X irradiation, the threshold of approximately 5 Gy at 2-4 weeks decreased to about 2 Gy by 17 weeks; the thermal enhancement ratio as calculated from slope-ratio analysis of the dose-response curves was 1.50 +/- 0.08 at 2-4 weeks post-treatment and 1.96 +/- 0.05 at 17-70 weeks post-treatment. Up to 20 min at 44 degrees C by itself was without effect. From comparisons of these data with results of crypt microcolony assays, it was concluded that intestinal fibrosis was both a chronic sequela of acute mucosal injury and a late effect of X irradiation. Adjuvant hyperthermia both hastened the expression of the late effect and increased its severity beyond that predicted from the acute injury.