Effects of 6-mercaptopurine in pressure overload induced right heart failure

Figures

• 

  Fig 1. Study design. Male Wistar rats were randomized to sham or pulmonary trunk banding (PTB) operation. The PTB rats were subsequently randomized to either placebo or 6-mercaptopurine treatment. Vehicle or 6-mercaptopurine were given via drinking water from one day before the surgery or two weeks after. Two weeks after the procedure, an echocardiography was performed on all the rats. Seven weeks after the surgery, cardiac function was evaluated by echocardiography, MRI, and invasive pressure-volume measurements and the rats were euthanized.

• 

  Fig 2. Effects of pulmonary trunk banding and 6-mercaptopurine treatment on right ventricular function at the end of the study. PTB: Pulmonary trunk banding; prev: Prevention group receiving 6-mercaptopurine treatment from one day before surgery; rev: Reversal group receiving 6-mercaptopurine treatment from two weeks after surgery. (A) Arterial elastance. (B) Right ventricular (RV) systolic pressure. (C) RV end-systolic (red dots) and end-diastolic volumes (black dots). (D) Cardiac index. (E) RV ejection fraction. (F) Tricuspid annular plane systolic excursion. (G) End-diastolic elastance. (H) End-systolic elastance. (I) Ventriculo-arterial coupling. Results are expressed as scatterplots with mean ± SEM. *P<0.05; ***P<0.001; ****P<0.0001.

• 

  Fig 3. Effects of pulmonary trunk banding and 6-mercaptopurine treatment on anatomy and histology. PTB: Pulmonary trunk banding; prev: Prevention group receiving 6-mercaptopurine treatment from one day before surgery; rev: Reversal group receiving 6-mercaptopurine treatment from two weeks after surgery. (A) Right ventricular (RV) cardiomyocyte cross sectional area. (B) RV fibrosis. (C) RV weight/tibia length. Histological sections stained with hematoxylin and eosine from (D) Sham and (E) Pulmonary trunk banding (PTB) control group and sections stained with picrosirius red from (F) Sham and (G) PTB-control. Results are expressed as scatterplots with mean ± SEM and are analysed by comparing PTB-control vs sham, PTB-control vs PTB-prevention, and PTB-control vs PTB-reversal.**P<0.01; ***P<0.001; ****P<0.0001.

• 

  Fig 4. Effects of pulmonary trunk banding and 6-mercaptopurine treatment on fibrosis, hypertrophy, and heart failure markers. Gene expression levels quantified by real-time polymerase chain reaction (real-time PCR) and normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH). Results presented by box plots. PTB: Pulmonary trunk banding; rev: Reversal group receiving 6-mercaptopurine treatment from two weeks after surgery. (A) mRNA expression of collagen 1. (B) mRNA expression of collagen 3a. (C) Collagen 1/collagen 3a ratio. (D) mRNA expression of osteopontin-1. (E) mRNA expression of connective tissue growth factor (CTGF). (F) mRNA expression of lysyl oxidase (LOX). (G) mRNA expression of fibronectin 1 (FN1). (H) mRNA expression of brain natriuretic peptide (BNP). (I) mRNA expression of myosin heavy chain-β (MHC-β).^P = 0.058; *P<0.05; ***P<0.001; ****P<0.0001.

• 

  Fig 5. Effects of pulmonary trunk banding and 6-mercaptopurine treatment on inflammation and apoptosis in the right ventricle. Gene expression levels of interleukin 6 (IL-6) and monocyte chemotactic protein 1 (MCP-1) quantified by real-time polymerase chain reaction (real-time PCR) and normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH). Results presented by box plots. Western blot analyses of CD45 and cleaved caspase-3 normalized to GAPDH. Results expressed as scatterplots with mean ± SEM. PTB: Pulmonary trunk banding; rev: Reversal group receiving 6-mercaptopurine treatment from two weeks after surgery.(A) mRNA expression of IL-6. (B) mRNA expression of MCP-1. (C) Protein expression of CD45 with representative lanes from a single blot. (D) Protein expression of cleaved caspase-3 with representative lanes from a single blot.

• 

  Fig 6. Effects of pulmonary trunk banding and 6-mercaptopurine treatment on Nur77 expression in the right ventricle. Gene expression levels of Nur77 quantified by real-time polymerase chain reaction (real-time PCR) and normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH) presented by box plots. Western blot analyses of total Nur77 normalized to GAPDH and Nur77 expression in isolated cytoplasmic and nuclear fraction normalized to GAPDH and proliferating cell nuclear antigen (PCNA), respectively. Results are expressed as scatterplots with mean ± SEM and with representative lanes from a single blot. PTB: Pulmonary trunk banding; rev: Reversal group receiving 6-mercaptopurine treatment from two weeks after surgery.(A) mRNA expression of Nur77. (B) Protein expression of total Nur77. (C) Representative images of immunofluorescence staining for Nur77 (green), DAPI (blue), and Wheat Germ Agglutinin (WGA; red). (D) Protein expression of Nur77 from cytoplasmic fraction. (E) Protein expression of Nur77 from nuclear fraction.