ABSTRACT
Objective To screen for microRNA (miRNA) involved in fetal rat lung development.Methods Fetal lungs were collected at their 16 d,19 d and 21 d of gestational age,and were observed after HE staining.Differentially expressed miRNA (fold change＞ 1.0) were screened by miRCURYTM locked nucleic acids chip.Some differentially expressed miRNA were selected for further analysis to investigate their change trends in 16 d,19 d and 21 d of gestational age.Results (1) Under the observation after HE staining,in gestational age 16 d group,original bronchus was dendritic distributed,with thick interstitial,rare capillary and no alveolar structure existed; in gestational age 19 d group,primary alveolar was seen,interstitial became thinner,and more capillaries were found; in gestational age 21 d group,more alveolar septa were identified and pulmonary acinus cavity was extremely expanded.(2) Two hundred and two differentially expressed miRNA were found.Among them,many miRNA were firstly reported in rat fetal lung development,suchas miRNA-3560 (8.4211415,4.8889050),miRNA-126 * (7.5239524,1.5118160),miRNA-186* (0.980 325 0,0.688 447 5),miRNA-466c* (0.977 220 0,0.877 227 0),miRNA-195 (13.549 629 0,0.985 488 8),miRNA-34a (12.426 133 0,0.604 066 2) and miRNA-466b-1 *(0.993 153 1,1.732 802 3).(3)The expression of miRNA-466c * and miRNA 186 * decreased as the gestational age increased from 16 d to 21 d,while expression of miRNA-195,miRNA-3560,miRNA-466b-1 *,miRNA-126 * and miRNA-let-7b increased; miRNA-34a expression increased during 16 d to 19 d.miRNA 17-92 family expression decreased,while expression of most let-7 family members (except let-7i and let-7e) increased from 16 d to 21 d of gestational age.Conclusions These miRNA might play an important role in the physiological mechanisms of fetal lungs development.
ABSTRACT
Objective To investigate the role of prenatal single-dose administration of dexamethasone and ambroxol on the expression of Toll-like receptor 4 (TLR4) of fetal and neonatal rats. Methods Fifty-four pregnant rats were randomly divided into three groups with eighteen rats in each group:rats treated with 0.2 mg/kg dexamethasone (group 1),0.2 mg/kg dexamethasone and 100 mg/kg ambroxol (group 2),or saline(controls) on the 17th day of gestation.The lung tissues of the offsprings were harvest independently on the 19th day of gestation,the postnatal 3 days and 7 days.The expressions of TLR4 in fetal/neonatal rat lungs of each pregnant rat were analyzed by reverse transcription-polymerase chain reaction(RT-PCR),immunohistochemistry stain,and Western blot. ANOVA and two independent samples t-test were applied. Results On the 19th day of pregnancy,TLR4 mRNA expression was up-regulated in lungs of the two treatment groups compared with controls(controls:0.26 ± 0.18,group 1:0.39 ± 0.21,t =5.866,P＜ 0.05 ; control:0.27 ± 0.22,group 2:0.46 ± 0.13,t =9.572,P＜ 0.01 ).TLR4 mRNA expression was up-regulated in group 2 compared with controls on the postnatal 3 days and 7 days(postnatal 3 d:0.59 ± 0.23 and 0.47 ±0.24,t=2.295,P＜0.05;postnatal 7 d:0.52±0.12 and 0.35±0.17,t=4.219,P＜0.05),while no significant difference was found in group 1 compared with the controls(postnatal 3 d:0.45±0.22 and 0.44±0.14,t=0.128,P＞0.05; postnatal 7 d:0.40±0.16 and 0.36 ±0.12,t=1.365,P＞0.05).Results of the immunohistochemistry demonstrated that on the 19th day of pregnancy,the protein expression of TLR4 was significantly increased in the two treatment groups (controls:0.20 ± 0.29,group 1:0.35±0.32,t=7.179,P＜0.05 ;controls:0.20±0.29,group 2:0.39±0.25,t=10.764,P＜0.01).The protein expression of TLR4 was significantly increased in group 2 on the postnatal 3 days and 7 days(postnatal 3 d:0.55±0.32 and 0.37±0.18,t=7.121,P＜0.05;postnatal 7 d:0.41±0.29and 0.25±0.24,t=6.355,P＜0.05),while no notable difference was found between group 1 and the control (postnatal 3 d:0.40±0.21 and 0.37±0.18,t=0.683,P＞0.05 ;postnatal 7 d:0.28±0.31 and 0.25±0.24,t=0.462,P＞0.05).Results of the Western blot demonstrated that on the 19th day of pregnancy,the protein expression of TLR4 was significantly increased in the two treatment groups (controls:0.15 ± 0.12,group 1:0.27± 0.20,t =7.835,P＜0.05; controls:0.16 ± 0.18,group 2:0.34±0.16,t=10.470,P＜0.01).The protein expression of TLR4 was significantly increased in lungs of the combination administration group on the postnatal 3 days and 7 days(postnatal 3 d:group 2:0.37±0.20 and 0.25±0.22,t=6.379,P＜0.05; postnatal 7 d:0.35±0.15 and 0.24±0.13,t=5.152,P＜0.05),while no notable difference could be found between group 1 and the control (postnatal3 d:0.32±0.26 and 0.25±0.16,t=1.167,P＞0.05; postnatal 7 d:0.29±0.19 and 0.24±0.10,t =1.248,P ＞ 0.05 ). Conclusions Prenatal single-dose administration of dexamethasone may up-regulate the expression of TLR4 in the rat fetal lung.The up-regulation of TLR4 might be one of the critical factors for glucocorticoid-induced maturity of fetal lung.Prenatal single-dose administration of dexamethasone and ambroxol may have effects on the regulation of TLR4 not only in fetal rats,but also in neonatal rats.