David A. Tulis
Thank you kindly for your note and comments. Yes, indeed, the duration of our study was 7 days, and therefore we cannot ascertain whether the changes observed at this time point (ie, cellular hypertrophy and/or nuclear polyploidy) will eventually lead to enhanced cellular hyperplasia as the cell cycle procedes which might be observed at later time points. If indeed we have cellular hyperplasia in the larger ileal artery and cellular hypertrophy in the second-order vessel (as observed), then this regional heterogeneity in vascular responses might be explained by differential GF expression. The ileal artery demonstrated significant medial expression of PDGF-A after 24 hours, with subsequent endothelial expression at 3 and 7 days. In addition, we found significant endothelial expression of PDGF-B after 7 days in the ileal artery (data not illustrated). This GF induction was not observed in the smaller second-order branch; therefore, the highly replicative state of cells in the ileal artery might be directly due to an exaggerated GF expression which was not observed in the smaller branch vessel.
In terms of connective tissue results, we used a modification of Masson's Trichrome Stain to stain all connective tissue; therefore, we cannot distinguish between the different components found in the ECM such as elastin and various forms of collagen. The percent of medial wall occupied by ECM components was between 29% and 34% in the control vessels, and between 34% and 41% in the high flow vessels. As the vessel walls hypertrophied in response to elevated flow, a concomitant increase in ECM components was observed in an effort to maintain a normal vessel wall environment.
I hope these comments adequately address your questions. If you have any additional questions or comments, please do not hesitate to contact me.
Very truly yours,
David A. Tulis