Abstract 3726: Right Ventricular Microcirculatory Changes in Pulmonary Hypertension-Associated Right Heart Failure Imaged by Three-Dimensional Confocal Microscopy

Abstract

Background: Right ventricular (RV) ischemia occurs in pulmonary hypertension (PH) patients despite normal coronary angiograms. O 2 supply-demand balance in the pressure-overloaded RV is determined by the interplay between reduced coronary perfusion (due to systemic hypotension and compression of coronary vessels) and a wall stress-related increase in myocardial O 2 consumption. The role of capillary rarefaction, which is involved in pressure overload induced left heart failure, has not been investigated in PH-associated RV failure. Methods: Rats received a single dose of the VEGF receptor blocker SU5416 and were exposed to 4 wks of normobaric hypoxia and 2 wks of sea level room air. This regimen induces angioproliferative changes in the pulmonary circulation, severe PH and RV failure. At 6 wks, the rats received intravital injections of Texas-Red conjugated tomato lectin, which is an N-acetylglucosamine specific marker of blood vessels in rodents. After circulation of the lectin for 5 min, papaverin was injected to promote maximal vasodilation of all perfused blood vessels. Animals were exsanguinated and perfusion-fixed with paraformaldehyde. After overnight fixation, 300 μm thick transverse sections of the myocardium were bleached in a methanol/DMSO mixture and nuclei were stained using DAPI. Prior to 3-dimensional imaging under a confocal microscope, further tissue clearing was achieved using organic solvents. Results: Z-stacks of 2-dimensional images were made in whole-mount cardiac specimens, using a step size of 0.32 μm. 3-dimensional reconstruction allowed quantification of the total capillary volume, which was reduced in the PH model compared to controls (2.9±1% vs. 5.7±1% of tissue volume, p<0.05). The reconstructions showed a remarkable increased morphological heterogeneity of the capillaries in the failing RV. In contrast, images of the LV microcirculation in the SU5416 treated animals were not different from those of controls. Conclusions: RV failure in experimental PH is associated with capillary rarefaction. 3-dimensional imaging of the myocardial microcirculation can be achieved using intravital injections of tomato lectin and subsequent confocal imaging.