Date of Award


Degree Type


Degree Name

Doctor of Philosophy


Fibrosis is an essential feature of tissue repair and is characterized by the deposition of collagen. Fibrosis in the myocardium is prevalent among patients with heart disease and can significantly impair cardiac function. The evaluation and treatment of myocardial scarring is however limited by a lack of quantitative and practical methods for studying fibrosis. My thesis was that both the amount and the relative maturity of fibrotic collagen could be quantified directly from histologic sections by taking advantage of the birefringent properties of collagen and assessing these with computer-based image analysis. A system, using polarized light microscopy and video microscopy, was therefore developed.;To quantify fibrosis, the high brightness intensity of collagen stained with picrosirius red was exploited. Using color filtering and image processing, a digital video image of myocardium was generated in which only collagen was visible. The collagen was quantified as the area-fraction of visible pixels. This video approach was compared to that of hydroxyproline analysis using samples from 14 autopsy hearts. A strong correlation (r = 0.98) between the two techniques was demonstrated. Also, in 22 endomyocardial biopsy specimens, the collagen content determined by the video technique was shown to correlate well with the collagen content determined by stereology (r = 0.95).;To quantify the relative maturity of fibrosis, the increasing birefringence of maturing fibrotic collagen was utilized. The median grey-level of all pixels depicting collagen served as the index of collagen brightness and reflected fiber birefringence. I measured the brightness of collagen in the scar that formed after superficial injury to the rat gracilis muscle. In this model, the diffuse arrangement of the collagen fibers was similar to that in interstitial myocardial fibrosis. Collagen brightness increased progressively over a 63-day period. I also measured brightness in the scar that formed after myocardial infarction in dogs and found that collagen brightness in 6-week infarcts was greater than that in 3-week infarcts (p {dollar}<{dollar} 0.01).;The clinical utility of the system was confirmed by two studies in which fibrosis in the human transplanted heart was studied. In the first study, I examined the relationship between the length of time the heart was not perfused during transport from donor to recipient (graft ischemic time), and the amount of fibrosis present one week after transplantation. Collagen content was measured in endomyocardial biopsy samples from 36 transplant recipients and found to be linearly related to graft ischemic time (r = 0.60). In the second study, I examined the relationship between allograft rejection and the amount and activity of fibrosis in five transplant patients. Serial biopsy samples, taken over a period of up to 1.5 years after transplantation, were evaluated. The collagen content of samples taken one year after surgery did not correlate with the frequency or severity of rejection. However, the activity of fibrosis was greatest in the early weeks ({dollar}<{dollar}12) after transplantation which was time when most of the rejection episodes had occurred.;In conclusion, both the content and the activity of myocardial fibrosis can be quantified using the polarization-video microscopy technique.



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