Establishment and Validation of a Rat Model of Pulmonary Arterial Hypertension Associated with Pulmonary Fibrosis

This research investigates the pathogenesis of group three pulmonary hypertension, focusing on molecular mechanisms in case associated with pulmonary fibrosis. In contrast to pulmonary arterial hypertension, the mechanisms underlying interstitial lung disease associated pulmonary hypertension are still poorly understood. Recent developments in this field include efficacy in reversing vascular remodeling. The discovery of cancer-like endothelia phenotypes and metabolic findings, linking amino acids and the lipid dysregulation to the pathogenesis of pulmonary hypertension. Bulk and the single cell I-A sequencing metabolimics and functional assays, such as annoy case resistance testing are pivotal technologies for mapping pulmonary hypertension, heterogeneity, and identifying therapeutic targets.

[Instructor] To begin, intubate the trachea of the rat using a PE-50 tube, inserted to a depth of three centimeters, which is the distance from the glottis to the end of the tube. Using a one milliliter syringe with a 26 gauge needle, draw 0.2 milliliters of bleomycin and inject it into the airway through a cannula as a single dose. Place the rat in a supine position to ensure an even distribution of bleomycin. Gently hold and slowly rotate the rat, alternating sides at an angle of 30 degrees to 45 degrees. Maintain each rotation for 10 to 15 seconds and repeat three to four times. Open the PFT software and check that the device is recognized. Now turn on the main control instrument switch and ensure each signal amplifier switch is off. To set the airflow, turn on the suction airflow regulating nut and set inspiration to five. Then turn on the expiratory airflow regulating nut and set slow expiration to minus four. Set the pressure value to 60. Calibrate the instrument in the order of FRC flow, flow, high flow, and lung pressure with an error of less than 5%. After anesthetizing the rat, cut the skin along the center of the neck. Separate the muscles layer by layer to expose the upper portion of the cervical trachea. Make a horizontal incision between the tracheal cartilage rings. Quickly insert the metal cannula into the tracheal incision, ensuring correct positioning, and secure with four zero silk thread sutures. Place the rat in the PFT animal compartment and connect the tracheal cannula to the airflow channel interface outside the instrument. Click the start button and examine the forced vital capacity or FVC and dynamic lung compliance. For the detection of right ventricular systolic pressure, or RVSP, fix the abdomen of the anesthetized rat upward on the experimental bench. Make an incision about four centimeters long on the right side of the neck, using surgical scissors. Separate the external jugular vein gently with microsurgical forceps, ensuring a length of about one centimeter is isolated. Insert two surgical lines at the distal and proximal ends of the separated vein. Ligate the distal end of the external jugular vein, using four zero silk thread. Gently lift the proximal surgical line and place it on the distal vein wall. Make a small incision of about 0.3 millimeters with small scissors. After inserting the PE-50 tube into the external jugular vein, ligate the PE-50 tube and vein together using four zero silk thread to prevent blood leakage. Observe the venous pressure waveform on the recorder. Push the catheter slowly into the right atrium, advancing it into the right ventricle to display the right ventricle pressure waveform. When the pressure is stable, record the right ventricular pressure for one minute and save the data for analysis. After thoracotomy, remove the intact heart of the rat using tweezers. Cut off the oracle and the connective tissue near the heart with scissors. Cut the free wall of the right ventricle along the pulmonary artery. This tissue is the right ventricle and the remaining tissues are the left ventricle plus the intraventricular septum. After separation, drain the surface of the right ventricle and the left ventricle plus the intraventricular septum, using filter paper. Weigh the right ventricle and left ventricle plus ventricular septum separately with an analytical balance before calculating their weight ratio. This figure illustrates the hemodynamic changes, pulmonary vascular remodeling, and impaired lung function observed in the bleomycin-induced PF PH rat model. Right ventricular systolic pressure and right ventricle to left ventricle plus septum ratio were significantly increased in the model group compared to the control group. Histological staining revealed more blue stained areas around pulmonary arteries in the model group, indicating greater collagen accumulation. Forced vital capacity and dynamic lung compliance were significantly reduced in the model group compared to the control group. Ratios of pulmonary artery acceleration time to ejection time, tricuspid annular plane systolic excursion, right ventricular fractional area change, and cardiac output were significantly reduced in the model group. Hydroxyproline concentration was significantly higher in the model group, indicating increased collagen content in lung tissue. Liver and kidney weights were not significantly different between the control and model groups.