Effect of cell therapy with stem cells from amniotic fluid (AF-MSC) and growth factor (TGF) factors in the treatment of secondary pulmonary hypoplasia in fetal sheep with tracheal occlusion

• Grant number: 14/14551-1
• Support Opportunities: Scholarships abroad - Research
• Start date: February 02, 2015
• End date: February 01, 2016
• Field of knowledge: Health Sciences - Medicine - Maternal and Child Health
• Principal Investigator: Walusa Assad Gonçalves Ferri
• Grantee: Walusa Assad Gonçalves Ferri
• Host Investigator: Jose Luis Peiro Ibanez
• Host Institution: Faculdade de Medicina de Ribeirão Preto (FMRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
• Institution abroad: Vall d'Hebron Research Institute (VHIR), Spain

Abstract

Congenital diaphragmatic hernia (CDH) affects 1:2500 live births and constitutes 8% of congenital anomalies and mortality (32-62%) remains high due to hypoplasia and pulmonary hypertension (PH). In studies of HDC, sheep have been used successfully as animal models and some results from this model demonstrated that the tracheal occlusion leads to lung growth. Tracheal occlusion in utero prevents the outflow of amniotic fluid produced by the lung and causes acceleration of lung maturation and growth, reducing the effects of hypoplasia and HP. Researchers have studied the role of stem cells in (AF-MSCs) amniotic fluid and observed that feature role in lung morphogenesis, stimulating growth factor Hepatocyte Growth Factor (HGF), considered as an initiator of normal lung organogenesis. Studies with Transforming Growth Factor (TGF) compared with administration of TGF tracheal occlusion, and concluded that there was little invasive, improved pulmonary hypoplasia and promoted differentiation of type II pneumocytes and, suggesting a potential role in the treatment of CHD. Analyze the VEGF is useful is stimulating angiogenesis factor, acts in mitosis and migration of endothelial cells in angiogenesis and neovascularization. Anyway, the HDC has a high mortality and searches for understanding the pathophysiology and future treatment strategies collaborate to improve the treatment of CDH. OBJECTIVE Evaluate the cellular, molecular, and functional changes that occur in hypoplastic fetal lung after intrapulmonary fluid retention, caused by intra-tracheal early tracheal occlusion, intra-tracheal administration of mesenchymal stem cells from amniotic fluid (AF-MSCs) and/or administration of growth factor (TGF) factors. Material and methods: Pregnant sheeps (n = 20) are also divided into 5 groups (n = 4) experimental: 1 - Congenital diaphragmatic hernia (CDH) created at 65 days of gestation. 2 - HDC created at 65 days of gestation + early tracheal occlusion (TO) created at 85 days of gestation. 3 - HDC created at 65 days of gestation + TO created at 85 days of gestation Introduction + stem cells from amniotic fluid (AF-MSC) at tracheal occlusion. 4 - HDC created at 65 days of gestation + TO created at 85 days of gestation + Introduction of growth factor (TGF) factors at tracheal occlusion. 5 - HDC created at 65 days of gestation + TO created at 85 days of gestation + AF-MSC + TGF at tracheal occlusion. The diaphragmatic defect will be done through surgical intervention (gestational age 65 days). The cesarean occur with gestational age 135 days, followed by cardiopulmonary resuscitation, ventilation for 6 hours with tidal volume 4-6 ml/kg, euthanasia and sample processing. The sample size was estimated considering that all the sheep can get 1-2 fetuses and that the average rate of abortion after fetal surgery is 30%. Analysis: Morphometric: fetal position, fetal body weight, Lung weight, lung to body weight ratio, diaphragmatic area, area of diaphragmatic hernia, hernia area ratio: diaphragm area. Histology: Pneumocytes type I, capillary network, alveoli, degree of pulmonary hypoplasia, collagen I and III, elastin, and neovascularization of the lung tissue. Cell Imuno-histoquimica/biologia: Platelet endothelial cell adhesion molecule-1, vascular-endothelial level-growth-factor-A (VEGF) and its receptors, angiopoietins 1 and 2, metalloproteinases, and cyclo-2 oxigenasa. Functional Dynamic and Static compliance, inspiratory resistance, oxygenation rate, tidal volume, minute volume, mean pressure in the airways, capnography, C20/C, pulmonary artery pressure, cardiac contractility and ventricular ejection fraction. (AU)