Lung Tissues Of Neonatal Rats Research Articles

AMPK/MTOR/TP53 Signaling Pathway Regulation by Calcitonin Gene-Related Peptide Reduces Oxygen-Induced Lung Damage in Neonatal Rats through Autophagy Promotion.

Our previous studies indicated that calcitonin gene-related peptide (CGRP) alleviates hyperoxia-induced lung injury and suggested the possible involvement of autophagy in this process. Herein, we aimed to further explore the potential involvement of tumor protein p53 (TP53) and autophagy in the mode of action of CGRP against hyperoxia-induced lung injury in vitro and in vivo. The study conducted tests on type II alveolar epithelial cells (AECII) and rats that were subjected to hyperoxia treatment or combined treatment of hyperoxia with CGRP, CGRP inhibitor, rapamycin (an autophagy agonist), 3-methyladenine (3-MA, an autophagy inhibitor), TP53 silencing/inhibitor (pifithrin-α), or expression vector/activator (PRIMA-1 (2,2-bis(hydroxymethyl)-3-quinuclidinone)) and their corresponding controls. We found that oxidative stress, apoptosis, and autophagy were all increased by hyperoxia treatment in vitro. However, treating AECII cells with CGRP reversed hyperoxia-induced oxidative stress and apoptosis but further promoted autophagy. In addition, the combined treatment with rapamycin or TP53 silencing with CGRP promoted the effect of CGRP, while contrary results were obtained with combined therapy with 3-MA or TP53 overexpression. In vivo, the number of hyperoxia-induced autophagosomes was promoted in the lung tissue of neonatal rats. Furthermore, hyperoxia increased the expression levels of AMP-activated protein kinase (AMPK) alpha 1 (also known as protein kinase AMP-activated catalytic subunit alpha 1 (PRKAA1)) but inhibited TP53 and mechanistic target of rapamycin (MTOR); these expression trends were regulated by CGRP treatment. In conclusion, we showed that CGRP can attenuate hyperoxia-induced lung injury in neonatal rats by enhancing autophagy and regulating the TP53/AMPK/MTOR crosstalk axis.

Effects of celecoxib on AQP-1, NF-κB and apoptosis in lung tissue of neonatal rats with hyperoxia-induced lung injury.

To analyze the effects of AQP-1, NF-κB and apoptosis in lung tissue of neonatal rats with hyperoxia-induced lung injury (HILI).162 neonatal SD rats were divided into control group (COG), model group (MOG) and celecoxib group (CEG), 54 rats in each group. Each group was subdivided into 1st, 5th and 10th-day groups with 18 rats in each group. The neonatal rat model of HILI was established. Nine rats in each group were randomly selected on the 3rd, 6th and 12th day respectively. The level of oxidative stress (OS) in bronchoalveolar lavage fluid (BALF), the pathological changes of lung tissue, the ratio of lung wet weight to dry weight (W/D), the expression of inflammatory factors in lung tissue, the l AQP-1 and NF-κB level and apoptosis in lung tissue were tested. In comparison with COG, the level of MDA in BALF, the ratio of lung W/D, the expression level of IL-6, TNF- α, NF- κB and the number of apoptotic cells in lung tissue were significantly increased, while the level of SOD in BALF and AQP-1 in lung tissue notably decreased in the MOG (P<0.05). The level of malondialdehyde (MDA) in BALF, the ratio of lung W/D, IL-6, TNF-α, NF-κB and plenty of apoptotic cells in lung tissue in CEG were notably decreased than MOG(P<0.05), while the level of SOD in BALF and AQP-1 in lung tissue were raised in CEG. In the COG, the lung tissue structure was complete, the arrangement was neat, the alveolar cavity was clear, and there was no inflammatory infiltration. With the extension of time, the inflammatory infiltration of lung tissue in the MOG gradually increased, the plenty of red blood cells gradually increased, and the size of the alveolar cavity varied; compared with the MOG, the inflammatory infiltration in the CEG decreased and the plenty of red blood cells decreased. Celecoxib can improve oxidative injury, inhibit inflammation, up-regulate AQP-1 and down-regulate NF-κB, inhibit apoptosis, and have a certain protective effect on HILI in neonatal rats.

MicroRNA-214 promotes alveolarization in neonatal rat models of bronchopulmonary dysplasia via the PlGF-dependent STAT3 pathway

BackgroundRecently, the role of several microRNAs (miRNAs or miRs) in pulmonary diseases has been described. The molecular mechanisms by which miR-214 is possibly implicated in bronchopulmonary dysplasia (BPD) have not yet been addressed. Hence, this study aimed to investigate a putative role of miR-214 in alveolarization among preterm neonates with BPD.MethodsMicroarray-based gene expression profiling data from BPD was employed to identify differentially expressed genes. A BPD neonatal rat model was induced by hyperoxia. Pulmonary epithelial cells were isolated from rats and exposed to hyperoxia to establish cell injury models. Gain- and loss-of-function experiments were performed in BPD neonatal rats and hyperoxic pulmonary epithelial cells. MiR-214 and PlGF expression in BPD neonatal rats, and eNOS, Bcl-2, c-myc, Survivin, α-SMA and E-cadherin expression in hyperoxic pulmonary epithelial cells were measured using RT-qPCR and Western blot analysis. The interaction between PlGF and miR-214 was identified using dual luciferase reporter gene and RIP assays. IL-1β, TNF-a, IL-6, ICAM-1 and Flt-1 expression in the rat models was measured using ELISA.ResultsThe lung tissues of neonatal rats with BPD showed decreased miR-214 expression with elevated PlGF expression. PlGF was found to be a target of miR-214, whereby miR-214 downregulated PlGF to inactivate the STAT3 pathway. miR-214 overexpression or PlGF silencing decreased the apoptosis of hyperoxic pulmonary epithelial cells in vitro and restored alveolarization in BPD neonatal rats.ConclusionOverall, the results demonstrated that miR-214 could facilitate alveolarization in preterm neonates with BPD by suppressing the PlGF-dependent STAT3 pathway.

Hyperoxia reduces STX17 expression and inhibits the autophagic flux in alveolar typeII epithelial cells in newborn rats.

Supplemental oxygen therapy can be life-saving for premature infants. Our previous study revealed a defect in the autophagic flux in the lung tissues of neonatal rats with hyperoxia-induced bronchopulmonary dysplasia (BPD), but the underlying mechanism remains unknown. Moreover, there are few innovative treatments that can completely alter the course of BPD. The present study examined the expression of Syntaxin 17 (STX17), a protein necessary for autophago-some-lysosome binding, in alveolar type II (AT-II) epithelial cells of neonatal rats with BPD. Neonatal Sprague-Dawley rats were randomly exposed to elevated O2 levels [fraction of inspired oxygen (FiO2), 0.8; model group] or normal room air (FiO2, 0.21; control group), and the expression levels of STX17, autophagy-related [Microtubule-associated protein 1A/1B-light chain 3B (LC3B)-II, p62, lysosomal-associated membrane protein 1)] and apoptosis-related (cleaved caspase3) mRNA and proteins were examined in lung tissues. Moreover, the expression levels of the aforementioned proteins were measured in isolated primary AT-II cells cultured in vitro under hyperoxic conditions in the presence or absence of pharmacological modulators of autophagy. Transmission electron microscopy identified that AT-II cell apoptosis and autophagosome aggregation were elevated in the lungs of BPD rats compared with control rats on postnatal day 7. STX17 mRNA and protein expression levels were decreased in lung tissue and isolated AT-II cells as early as postnatal day 3 in BPD rats, while the expression levels of LC3B-II, p62 and cleaved caspase3 were increased, reaching a peak on postnatal day 7. This early reduction in STX17 expression, followed by increased expression in autophagy- and apoptosis-related proteins, was also observed in isolated AT-II cells exposed to hyperoxia in vitro. However, treatment with the autophagy inducers rapamycin or LiCl eliminated the hyperoxia-induced reduction in STX17, partially restored the autophagy flux and increased the survival of AT-II cells exposed to hyperoxia. Collectively, these results indicated that STX17 expression in AT-II cells was reduced in the early stages of BPD in neonatal rats and may be related to the subsequent hyperoxia-induced block in autophagic flux.

Cell Division Cycle 2 Protects Neonatal Rats Against Hyperoxia-Induced Bronchopulmonary Dysplasia.

PurposeHyperoxia-induced bronchopulmonary dysplasia (BPD) is a lung disease in preterm infants. We aimed to explore the role of cell division cycle 2 (CDC2) on histopathologic changes of lung tissues, as well as the viability, apoptosis, and inflammation of lung cells in rats with hyperoxia-induced BPD.Materials and MethodsHyperoxia-induced BPD in neonatal rats and hyperoxia-induced A549 cells were constructed. The mRNA expression of CDC2 was detected by qRT-PCR. The fibrosis score of lung tissues was evaluated by hematoxylin-eosin staining. The viability and apoptosis of A549 cells were detected by cell counting kit-8 assay and flow cytometry. The protein expressions of bcl-2, bax, and caspase-3 were measured by western blot. The levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-6, and IL-1β in A549 cells were detected by enzyme-linked immunosorbent assay. The pcDNA3.1-CDC2 was injected into rats to determine the role of CDC2 in hyperoxia-induced BPD in vivo.ResultsThe expression of CDC2 was decreased in lung tissues of neonatal rats with hyperoxia-induced BPD and hyperoxia-induced A549 cells. The fibrosis score was increased in the lung tissues of neonatal rats with hyperoxia-induced BPD. Overexpression of CDC2 increased the viability and protein expression of bcl-2; and inhibited the apoptosis, inflammation, and protein expression of bax and caspase-3 in hyperoxia-induced A549 cells. Up-regulation of CDC2 alleviated the histopathologic changes in lung tissues of neonatal rats with hyperoxia-induced BPD.ConclusionOverexpression of CDC2 promoted the viability and inhibited the apoptosis and inflammation of hyperoxia-induced cells, and alleviated the histopathologic changes of lung tissues in neonatal rats with hyperoxia-induced BPD.

The expression and effect of long non-coding RNA H19 in BPD neonatal rats

Objective To investigate the expression of long non-coding RNA H19(LncRNA H19)and its regulation of histone methyltransferase 2(enhancer of zeste homolog 2, EZH2)in the lung tissue of neonatal rats with bronchopulmonary dysplasia(BPD), and to lay a foundation for elucidating the pathogenesis of BPD lung epithelium-interstitial transformation(EMT). Methods The BPD model of SD neonatal rats was induced by hyperoxia(inhalation oxygen concentration was 85%)(n=50), and oxygen inhalation concentration of the control group was 21%(n=50). The two groups were collected at 1d, 3d, 7d, 14d and 21d after birth in lung tissue.Immunohistochemistry, Western blot, real-time quantitative PCR and other techniques were used to detect the intracellular localization, and the expression level of EZH2 protein and the mRNA expression level of H19 and EZH2. Results Immunohistochemistry showed that EZH2 protein was located in the nucleus and cytoplasm of alveolar epithelial cells, and the expression of EZH2 protein in the model group was significantly enhanced compared with the control group.Similarly, the results of Western blot demonstrated that the expression of EZH2 protein in the model group increased from 1d(control group: 0.196±0.030, model group 0.650±0.149)to 21d(control group 0.934±0.215, model group 1.785±0.298)rather than the control group(P<0.05). Compared with the control group, the mRNA expression level of H19 in the model group increased from 7d(control group 2.591±0.211, model group 3.558±0.093, P<0.05)and the expression level of EZH2 mRNA started to increase from 3d(control group 1.246±0.015, model group 2.148±0.215, P<0.05). Moreover, the differences between the two groups were obvious with the time of hyperoxia exposure. Conclusion In the development of BPD, the expression levels of H19 and EZH2 protein in lung tissue is up-regulated, and the peak of H19 expression precedes EZH2, which suggest that H19 might be involved in the pathogenesis of pulmonary dysplasia induced by EZH2-mediated EMT. Key words: Long non-coding RNA H19; Enhancer of zeste homolog 2; Bronchopulmonary dysplasia; Newborn rat

Protective effects of propofol on experimental neonatal acute lung injury.

The present study aimed to investigate the effects of propofol on neonatal acute lung injury (ALI) in a rat model and to examine the molecular mechanisms underlying propofol function. A rat model of ALI was established by intraperitoneal injection of lipopolysaccharides (LPS). The neonatal rats were treated with various concentrations of propofol and a lung injury score was assessed. The protein expression levels of pro‑inflammatory cytokines was detected using ELISA. In the present study, oxidative stress was determined by measuring the level of malondialdehyde (MDA) and the activity of superoxide dismutase (SOD) in lung tissues. Reverse transcription quantitative‑polymerase chain reaction and western blot analysis were used to examine the mRNA and protein expression levels of the factors downstream to LPS signaling pathway. Treatment with propofol significantly alleviated LPS‑induced lung injury in neonatal rats as suggested by the decreased lung injury score, increased partial pressure of oxygen and decreased lung wet‑dry weight ratio. LPS promoted the upregulation of tumor necrosis factor α (TNF‑α), interleukin (IL)‑6 and IL‑1β in lung tissues and bronchoalveolar lavage fluid from neonatal rats exhibiting ALI. Notably, treatment with propofol decreased the expression levels of these factors. Additionally, LPS caused an increase in the levels of MDA, and a decrease in SOD activity, and treatment with propofol suppressed these effects in a dose‑dependent manner. Furthermore, LPS induced the upregulation of phosphorylated (p‑)p38, nuclear factor κ‑light‑chain‑enhancer of activated B cells (NF‑κB), p‑p65, NLR family pyrin domain containing 3 (NLRP3), apoptosis‑associated speck‑like protein containing CARD and caspase‑1 in lung tissues of neonatal rats, and treatment with propofol was able to downregulate these factors in a dose‑dependent manner. Propofol alleviated lung injury in neonatal rats with LPS‑induced ALI by preventing inflammation and oxidative stress via the regulation of the activity of the p38 mitogen‑activated protein kinase/NF‑κB signaling pathway and the expression levels of the NLRP3 inflammasome.

Effects of MiR-26a on respiratory distress syndrome in neonatal rats via the wnt/β-catenin signaling pathway.

Micro ribonucleic acids (miRNAs) are crucial to post-transcriptional regulation of the gene expression. Whether miR-26a affects respiratory distress syndrome (RDS) in neonatal rats through the Wnt/β-catenin signaling pathway was investigated in this study. The neonatal rat model of RDS was established, and the expressions of miR-26a and glycogen synthase kinase-3β (GSK-3β) in RDS in neonatal rats and their correlation were analyzed. The cascade relationship between miR-26a and the Wnt/β-catenin signaling pathway and the influence of miR-26a on the expression of inflammatory cytokines were subsequently verified. Finally, the influences of miR-26a on the expressions of important markers, receptor for advanced glycation endproducts (RAGE), high mobility group box 1 (HMGB1), and plasminogen activator inhibitor-1 (PAI-1), through the Wnt/β-catenin signaling pathway were analyzed. Compared with those in normal tissues, the expression of miR-26a in lung tissues of neonatal rats with RDS was significantly decreased (p<0.05), while the expression of GSK-3β messenger RNAs (mRNAs) was notably increased (p<0.01), and the GSK-3β expression was negatively correlated with the miR-26a expression (r=-0.6693, p=0.0064). In addition, miR-26a mimics significantly inhibited the GSK-3β protein expression and activated the Wnt/β-catenin signaling pathway. Moreover, miR-26a could reduce the expressions of tumor necrosis factor-alpha (TNF-α), interleukin-1β (IL-1β), and IL-6, as well as RAGE, HMGB1, and PAI-1. MiR-26a can affect inflammatory responses and markers through the Wnt/β-catenin signaling pathway in neonatal rats with RDS.

Effects of CASZI on bronchopulmonary development of neonatal rats.

The effect of CASZI on bronchopulmonary development in neonatal rats was investigated. Forty SD rats were randomly divided into the control group (n=20) and the experimental group (n=20) using a random number table. The experimental group was treated with hyperoxia to establish the bronchopulmonary dysplasia (BPD) model, while the control group received no treatment. At 14 days of experiment, rats in each group were treated, and lung tissues were taken for hematoxylin and eosin (H&E) staining. Radial alveolar count (RAC) and morphological changes in lung tissues were observed under a microscope, and the thickness of respiratory membrane was detected via IPP6 image. Reverse transcription-polymerase chain reaction (RT-PCR) and western blotting were used to detect the expression levels of CASZI, Rho kinase, RhoA and ET-A receptor in lung tissues. There were no obviously pathological changes in lung tissues of neonatal rats in the control group, while BPD could be seen in lung tissue structures of neonatal rats in the experimental group. RAC in experimental group was significantly lower than that in the control group (P<0.05), and the thickness of respiratory membrane was significantly higher than that in the control group (P<0.05). CASZI mRNA expression level in the experimental group was significantly lower than that in control group (P<0.05), and Rho kinase, RhoA and ET-A receptor mRNA expression levels were significantly higher than those in the control group (P<0.05). CASZI protein expression level in the experimental group was significantly lower than that in the control group (P<0.05), and the Rho kinase, RhoA and ET-A receptor protein expression levels were significantly higher than those in the control group (P<0.05). In conclusion, CASZI expression was downregulated during the development of BPD. CASZI downregulation may have an effect on the pulmonary microvascular regeneration through RhoA/ROCK signaling pathway, thus participating in the pathogenesis of BPD.

Genome-Wide Analysis of DNA Methylation in Hyperoxia-Exposed Newborn Rat Lung.

Oxygen therapy is often required to treat newborn infants with respiratory disorders. Prolonged exposure of neonatal rats to hyperoxia reduced alveolar septation, increased terminal air space size, and increased lung fibrosis; these conditions are very similar to those of human bronchopulmonary dysplasia. Epigenetic regulation of gene expression plays a crucial role in bronchopulmonary dysplasia development. We reared Sprague-Dawley rat pups in either room air (RA, n=24) or an atmosphere containing 85% O2 (n=26) from Postnatal Days 1 to 14. Methylated DNA immunoprecipitation (MeDIP) was used to analyze genome-wide DNA methylation in lung tissues of neonatal rats. Hyperoxia-exposed rats exhibited larger air spaces and thinner septa than RA-exposed rats did on Postnatal Day 14. The rats exposed to hyperoxia exhibited significantly higher mean linear intercepts than did the rats exposed to RA. We applied MeDIP next-generation sequencing for profiling changes in DNA methylation in the rat lungs exposed to hyperoxia and RA. We performed bioinformatics and pathway analyses on the raw sequencing data to identify differentially methylated candidate genes. Our in vivo model revealed that neonatal hyperoxia exposure arrested alveolarization on Postnatal Day 14. We found that the ErbB, actin cytoskeleton, and focal adhesion signaling pathways are epigenetically modulated by exposure to hyperoxia. We demonstrated that hyperoxia exposure contribute in delaying lung development through an epigenetic mechanism by disrupting the expression of genes in lungs that might be involved in alveolarization. These data indicate that aberrant DNA methylation and deregulation of the actin cytoskeleton and focal adhesion pathways of lung tissues may be involved in the pathophysiology of hyperoxia-induced arrested alveolarization.

Protective effect of heat shock protein 70 on lungs in newborn rats with hypoxic pulmonary hypertension

Objective To investigate the protective effect of adenovirus mediated heat shock protein 70 (HSP70) on lungs in neonatal rats with hypoxic pulmonary hypertension(HPH). Methods One hundred and twenty-eight 7-10 d healthy Wistar neonatal rats were randomly divided into HPH model group and control group.HPH group was divided into saline group, empty virus group, and HSP70 group according to the transfection solution.HPH model was established in the hypoxia cabin of 80 mL/L nitrogen oxygen mixed gas after transfection.The mean pulmonary artery pressure(mPAP) was measured after 3, 7, 10 and 14 days of hypoxia in each group.The mRNA and protein expression of HSP70, hypoxia inducible factor-1 alpha(HIF-1α), endothelin-1(ET-1) and inducible nitric oxide synthase(iNOS) in the lung tissues of neonatal rats were detected by using reverse transcription-PCR and Western blot respectively. Results (1)The mPAP level was significantly higher in saline group(M, Q: 12.00, 2.50; 15.00, 2.00; 18.00, 1.75; 20.00, 2.25) than that in control group(M, Q: 9.50, 4.75; 10.50, 1.00; 13.00, 1.00; 15.50, 3.25), and the differences were significant(z=-3.28, -3.40, -3.34, -3.06, all P 0.05). (2)The expressions of HSP70 mRNA among the groups were statistically significant(F=6.321, 9.669, 6.333, all P<0.01), and the expressions of HSP70 protein also had significant difference(F=16.463, 3.637, 17.749, all P<0.01). (3)The level of HIF-1α mRNA in saline group was significantly higher than that of the control group, and the differences were statistically significant (q=4.312, 9.106, 6.151, all P<0.01); and the level of HIF-1α mRNA in empty virus group was also significantly higher than that in the control group, and the differences were statistically significant ( q=3.982, 9.235, 5.352, all P<0.01) in 3, 7, and 10 days; hypoxia in HSP70 group was lower than that of the empty virus group in 3, 7 days, and the differences were statistically significant (q=6.083, 11.031, all P<0.05). The level of ET-1 mRNA in saline group was significantly higher than that in the control group(q=5.112, 10.086, 6.264, all P<0.01), in empty virus group was significantly higher than that in the control group, and the differences were statistically significant (q=4.182, 12.238, 5.864, all P<0.01) in 3, 7, and 10 days, but in HSP70 group it was lower than that in the empty virus group in 3, 7, and 10 days, and the differences were statistically significant (q=6.912, 10.235, 7.021, all P<0.05). The level of iNOS mRNA in saline group was significantly higher than that of the control group, and the differences were statistically signi-ficant (q=4.998, 8.056, 5.369, all P<0.01), in empty virus group was significantly higher than that in the control group, and the differences were statistically significant (q=4.778, 10.138, 5.154, all P<0.01) in 3, 7, and 10 days, but in HSP70 group it was lower than that in the empty virus group in 3, 7, and 10 days, and the differences were statistically significant (q=7.819, 9.838, 6.156, all P<0.05). The level of HIF-1α protein in saline group was significantly higher than that of the control group in 3, 7, and 10 days, and the differences were statistically significant (q=3.146, 3.012, 4.106, all P<0.05), in empty virus group was significantly higher than that of the control group in 10 days, and the difference was statistically significant (q=3.468, P<0.05); but in HSP70 group it was lower than that in the empty virus group in 3, 7, and 10 days, and the differences were statistically significant (q=3.876, 4.108, 4.021, all P<0.05). The level of ET-1 protein of HSP70 group was lower than that of the saline group, the differences were statistically significant(q=3.367, 2.983, 3.246, all P<0.05), in HSP70 group was lower than that of the empty virus, and the differences were statistically significant (q=3.268, 2.678, 3.567, all P<0.05). The level of iNOS protein in saline group was significantly higher than that in the control group in 3, 7, and 10 days, and the diffe-rences were statistically significant (q=3.360, 3.567, 3.567, all P<0.05), but in HSP70 group it was lower than that in the empty virus group, and the differences were statistically significant (q=3.126, 3.908, 3.087, all P<0.05). Conclusion Adenovirus mediated HSP70 can improve the HSP70 expression in HPH, down-regulate the expression of HIF-1α, ET-1, iNOS, and reduce pulmonary arterial pressure. Key words: Heat shock protein 70; Adenovirus; Hypoxia inducible factor -1α; Hypoxic pulmonary hypertension; Neonatal rat

Effects of prolonged oxygen exposure at different concentrations on expressions of vascular endothelial growth factor and its receptors mRNA in pulmonary tissues of neonatal rat

Objective To study the effects of prolonged oxygen exposure at different concentrations on expressions of vascular endothelial growth factor (VEGF) and its receptors (VEGF receptor-1 ,VEGFR1 and VEGF receptor-2, VEGFR2)mRNA in lungs of neonatal rat. Methods Thirty-two SD rat pups were randomly divided into control and experimental group. Rats in the experimental group were further divided into three subgroups: 30% O2, 50% O2 and 75% O2 group. Each group had 8 rats. The rats were ,sacrificed at postnatal day 21 and their right lower lobes were collected. Expressions of VEGF, VEGFR1 and VEGFR2 mRNA were detected by RT-PCR. Results There was no effect on VEGF, VEGFR1 or VEGFR2 mRNA expression in 30% O2 group. The expressions of VEGF mRNA in the 75% O2 group was 0. 48 times of the control (P<0. 05). The expressions of VEGFR1 mRNA(P<0.01) and VEGFR2 mRNA (P<0.01) in 50% O2 and 75% O2 group were lower than those of control. Condusions Prolonged exposure to moderate or high oxygen concentration may reduce the expression of VEGF and its receptors' mRNA in the lung tissues of neonatal rats. Key words: Vascular endothelial growth factor As Vascular endothelial growth factor receptor-1; Vascular endothelial growth factor receptor-2; Oxygen inhalation therapy; Oxygen; Rats

Dynamic changes of renin-angiotensin system, collagen I and intercellular adhesion molecule I in lung tissue of neonatal rats with hyperoxia-induced lung fibrosis

Objective To observe the dynamic changes of renin-angiotensinsystem,collagen I(CoI) and intercellular adhesion molecule 1(ICAM-1) in hmg tissue of neonatal rats with lung fibrosis induced by hyperoxia.Methods One hundred and twenty term neonatal Wistar rats were randomly assigned into model group and air control group.The model group was continuously exposed to hyperoxia (FiO2=0.9)for 21 days,and the air control group was exposed to room air(FiO2=0.21).On the Ist,3rd,7th,14th and 21st day of exposure,the subjects were sacrificed.And then,the protein levels of CoI,ICAM-1 were measured by enzyme-linked immunosorbentassay and angiotensinII(AngII) by radio-immunity technique,the activity of angiotensin-convertion enzyme(ACE) were measured by biochemistry analysator,and the mRNA expression of ACE,AngII,CoI was measured by RT-polymerase chain reaction.The changes of lung histomorphology were observed.Results The ACE,AngII,CoI and ICAM-1 protein levels of model group were (249.20±20.19)μg/g,(838.22±197.75)μg/g,(539.85±81.03) μg/g and (577.06±61.17)μg/g on the 14th day respectively and in.eased significantly as compared to the air control group (P＜0.05),and the ACE,AngII,CoI mRNA expression of modal group also increased signficantly on the 14th day (P＜0.05).The ACE,AngII,CoI and ICAM-1 protein levels of model group were(249.20±20.19)μg/g,(838.22±197.75)μ/g,(539.85±81.03)μg/g and (577.06±61.17)μg/g on the 21st day and increased to the peak respectively(P＜0.05),and the ACE,AngII and CoI mRNA expression had the same changes as well.The histopathological examination demonstrated different degrees of fibrosis in the model group.Conclusion The ACE,AngII,CoI and ICAM-1 protein levels and their mRNA expression were increased significantly on the 14th and 21st day in lung tissue of neonatal rats with lung fibrosis induced by hyperoxia.The renin-angiotensin system played a role in lung fibrosis induced by hyperoxia. Key words: Rat; Hyperoxia; Fibrosis; Renin-angiotensin system

Changes of MMP-8 and TIMP-2 expression in the lung tissue of neonatal rats with hyperoxia induced lung injury

目的 探讨高氧致新生大鼠肺损伤肺组织基质金属蛋白酶-8(MMP-8)和基质金属蛋白酶抑制物-2(TIMP-2)表达的动态变化及其对慢性肺疾病(CLD)纤维化形成的作用.方法 足月新生大鼠,于生后12 h内分别持续吸入0.90～0.95的高氧和空气,于1、3、7、14、21 d,取肺组织行Masson染色,图像定量分析,计算胶原纤维阳性面积百分比,以评估胶原纤维沉积程度;应用RT-PCR和免疫组化技术检测肺组织中MMP-8、TIMP-2mRNA和蛋白表达的动态变化.结果 高氧暴露7 d开始高氧组的胶原纤维在肺间质沉积,胶原纤维阳性面积高于空气组.高氧组MMP-8mRNA和蛋白表达在7 d后低于空气组,两组TIMP-2的表达在各时间点上无差异.结论 随着高氧暴露时间的延长,新生大鼠肺组织中MMP-8表达降低,而TIMP-2表达不变,MMP-8/TIMP-2的平衡状态遭到破坏,胶原降解减少,从而促进了胶原在肺间质的沉积。

Effect of in vivo hyperoxia on the glutathione system in neonatal rat lung.

Relative resistance to oxygen toxicity in newborn animals of some species has been associated with a rapid increase in antioxidants in lung tissue homogenate. This study investigated the effect of hyperoxia on the glutathione system antioxidants in lung tissue of neonatal rats exposed to hyperoxia for 15 days. Neonatal rats were exposed to either 100% oxygen or air for 0, 3, 6, 9, 12, and 15 days prior to sacrifice for determination of glutathione and the glutathione system antioxidant enzymes in whole lung homogenate. There were significantly higher levels of total glutathione at 3, 6, and 9 days and of reduced glutathione at 3 and 6 days in oxygen-exposed animals compared to air-exposed controls. These differences were no longer present after 12 or 15 days of exposure to hyperoxia. Glutathione peroxidase and glucose-6-phosphate dehydrogenase remained higher in lung tissue from oxygen-exposed animals from 6 through 12 days of hyperoxia. The failure to maintain sustained high levels of total glutathione during hyperoxia might suggest that glutathione depletion is a factor in the timing of death from oxygen toxicity in these animals. The absence of a sustained increase in oxidized glutathione disulfide is more consistent with other explanations for this transient increase in total glutathione.

The influence of maternal nicotine exposure on neonatal lung carbohydrate metabolism

The influence of maternal nicotine exposure (1 mg/kg body mass/day) during pregnancy and lactation on energy metabolism of lung tissue of neonatal rats were investigated. The glucose turnover of the lung tissue of the neonatal rats exposed to nicotine via the placenta and mother's milk was 86,4% higher than that of the controls. Glycolysis was however suppressed by 22,7% (P<0,01). The adenine nucleotide pool (ATP + ADP + AMP) was 32,8% higher for the lungs of the 3 week old neonates exposed to nicotine than that of the control rat lung. After 4 weeks of nicotine withdrawal glycolysis of those animals exposed to nicotine were still inhibited to the same extent than during exposure. The adenine nucleotide pool was 69,95% higher than that of the controls. It is proposed that the inhibition of glycolysis was due to the high ATP/ADP ratio of the lungs of the nicotine exposed rats.

Effect of maternal nicotine exposure on growth in vivo of lung tissue of neonatal rats.

The effect of maternal nicotine exposure on lung growth in vivo in neonatal rats was investigated. Nicotine (0.25 and 1.0 mg/kg/day) administered subcutaneously to the pregnant animal from day 7 of gestation until weaning resulted in smaller neonatal lungs that were about 15% smaller on postnatal day 8. On day 21 no difference in lung mass occurs. Maternal nicotine exposure also causes enhanced lung cellular multiplication as judged by the calculated daily increase in DNA of 0.19 mg/g for control lung and 0.31 mg/g for experimental lung. Comparison of the protein/DNA ratio 5.75 +/- 0.22 of the control and the 3.59 +/- 0.21 of nicotine-exposed lungs showed that the cells of the latter was smaller. The lower lung mass was attributed to the smaller cell size. It is proposed that nicotine's marked inhibitory (42%) effect on glycolysis probably results in type I cell injury and consequently enhanced cell proliferation.