Estimate of cellular damage and application of several treatments in brain and lung diseases of neonate rat induced by hypoxia/hyperoxia

다국어 초록 (Multilingual Abstract)

A very preterm infant is a baby born before 37 weeks gestationor birth weights less than 1500g. Preterm baby organs, especially brain and lung, do not enough to allow normal postnatal survival and growth. Especially premature babies can not maintain saturation percentage of dissolved oxygen because their lung is immature that can not exchange of gases between inspired air and the blood. In the brain, low oxigen level leads high risk of death or major neurological and neuro developmental abnormalities resulting from injury in the basal ganglia/thalamus and cerebral cortex. Premature infants who deliver at 26？36 wk of gestation are born in the saccular phase of lung development and are particularly susceptible to bronchopulmonary dysplasia (BPD) and pulmonary fibrosis because premature infants have been received ventilatory support for oxygen supplementation for maintain saturation percentage of dissolved oxygen. For improvement of brian and lung diseases in premature baby, there are required new therapeutic targets and methods.
In this study was demonstrated brain and lung diseases animal model to improve the diseases that is caused hyper- and hypoxia in premature infant brain and lung is described.
In chapter 1, loss of mitochondrial permeability detection by flow cytometry was demonstrated, and examined the neuroprotective effect of permeability transition of mitochondrial membrane chemical on hypoxic-ischemic (HI) brain injury was also described in the newborn animal. HI brain injury was performed on 7-day old Sprague Dawley (SD) rats. The rat pups were sacrificed at 24h or 48h after hypoxia-ischemia under deep anesthesia, and the whole brain tissue was obtained. Thereafter, the brain was transected in the coronal plane and assayed by TTC stain, western blot, TUNEL and FACS.
In chapter 2, in vivo siRNA delivery was mentioned using PDMAEMA polymer and CTGF expression was silenced at the translational level using CTGF siRNA and whether CTGF mRNA knockdown could prevent the progression of bleomycin induced pulmonary fibrosis was also demonstrated. Four week old male SD rats were randomly exposed bleomycin or phosphate-buffered saline (PBS). siRNA-PDMAEMA complex was prepared with 100？l final volume by add deionized water and intratracheally administrated at 3h and 72h after the bleomycin treatment At P14, the harvested lungs were examined the level of CTGF, TGF-？, IL-6, and collagen content. For histological evaluation, lungs were removed and immersed in the same fixative overnight at room temperature. The fixed right lungs were embedded in paraffin. The sections were used with hematoxylin and eosin (H&E), Masson？s trichrome staining and immunostain.
In chapter 3, BPD rat animal models were established and then development therapy methods in that animals were applied using stem cell transplantation and growth factor. Wild-type SD rats were randomly exposed to 95%/90% oxygen or air from birth. In the transplantation groups, a single dose UCB-derived MSCs was administered at postnatal day (P) 5. In the GCSF injection groups, GCSF treatments of 20？g/kg were administered intraperitoneally on the fourth, fifth, and sixth postnatal days. At P14, the harvested lungs were examined for the myeoloperoxidase activity, the level of tumor necrosis factor (TNF)-？, interleukin (IL-6), and transforming growth factor (TGF-？) mRNA, ？-smooth muscle actin (SMA) protein, and collagen levels. Intratracheal transplantation of human UCB-derived MSCs is more effective than intraperitoneal transplantation in attenuating the hyperoxia-induced lung injury and GCSF treatment significantly attenuated hyperoxia-induced lung injury in neonatal rats. The result is that the intratracheal rather than the intraperitoneal transplantation of human UCB-derived MSCs significantly attenuated the hyperoxia-induced lung injury such as decreased alveolarization and increased fibrosis. This protective effect appears to be mediated primarily by their anti-inflammatory effects and less possibly by their differentiation into type II pneumocytes. GCSF treatment significantly attenuated hyperoxia-induced lung injuries such as decreased alveolarization by down-modulating the oxidative stress and inflammatory responses in neonatal rats. These findings support the potential use of GCSF as a new therapeutic agent for BPD.

목차 (Table of Contents)

• Contents
• Page
• Summary ⅰ
• List of Figures ⅵ
• List of Tables ⅸ
• Chapter 1. Evaluation of Mitochondrial Membrane Potential Change and Protection Effect of Cyclosporine A in Neonate Brain Damaged by Hypoxia-Ischemia
• Abstract 2
• Introduction 4
• Materials and Methods 7
• Results 15
• Discussion 26
• References 31
• Chapter 2. Attenuation of Bleomycin-Induced Pulmonary Fibrosis by CTGF siRNA in vivo Silencing
• Abstract 40
• Introduction 41
• Materials and Methods 43
• Results 52
• Discussion 64
• References 66
• Chapter 3. Development of Hyperoxia-Induced Lung Injury Therapy by Stem Cell Transplantation and Growth Factor in Neonatal Rats
• Abstract 71
• Introduction 73
• Materials and Methods 76
• Results 88
• Discussion 106
• References 114