Page 137 - Assessing right ventricular function and the pulmonary circulation in pulmonary hypertension Onno Anthonius Spruijt

Data Analysis
PET image data were sorted into 0.5mm sinogram bins and 33 time frames and images were reconstructed using filtered back projection with CT‐based attenuation correction. The frame durations used for the study were 12×5s, 4×15s, 6×30s, 11×300s. The reconstructed CT and PET images were analyzed using the 3‐dimensional visualization toolbox in the Inveon Research Workplace software. (Siemens Healthcare Molecular Imaging).
The whole lung tissue time‐activity curves (TACs) were calculated from lung PET images co‐ registered with ROI on CT lung images, covering the lung volume with clearly visible boundaries adjusted by CT density thresholding. Cumulative images over 0 to 60 min were used for kinetic analysis of tracer uptake. The image derived input functions (IDIFs) were determined individually by sampled arterial plasma time activity curves or the PET images derived activity curve from the LV blood pool.
Decay corrected whole lung TACs were normalized for injected activity (kBq) and body weight to obtain standardized uptake value (SUV). The dynamic image data was fitted to a 2‐compartment 4k model using the Matlab‐based in house kinetic analysis software package (CLICKFIT). In this model, K1 represents the rate constant for the transport of 18FLT from blood into tissue, k2 represents the rate constant of the transport of 18FLT from tissue back to blood, k3 reflects the rate constant of phosphorylation of 18FLT, and k4 describes the rate of 18FLT dephosphorylation [29].
Bio‐distribution
The plasma and tissue samples (lung, RV, LV, septum, liver, and kidney) harvested following PET scanning were weighed and measured for radioactivity in a gamma counter. All data were corrected for radioactivity decay. The amount of radioactivity was expressed as percentage of injected dose per gram of tissue or blood (%ID/g).
Histology Examination
Transverse lung sections were processed for elastic Van Gieson (EVG) and hematoxylin and eosin (H&E) staining. Peripheral vessels less than 100 μm in lung were counted blindly under microscope (40X) and pulmonary vascular remodelling was expressed as the proportion of vessels with double elastic lamina to total vessels counted. Lung sections were also processed for Ki‐67 (1:50, Thermo Scientific), CD68 (1:50; Serotec), TK1 (1:50; Abcam) and ENT1 (1:100; Abcam) immunostaining. Ki‐67
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