[Effect of intrauterine lipopolysaccharide infusion on Toll-like receptor 4 signaling transduction pathway in lungs of perinatal rats].
OBJECTIVE
To investigate the influence of intrauterine infection caused by lipopolysaccharide on Toll-like receptor 4 (TLR4) signaling pathway in fetal and neonatal rat lungs in order to explore immunomodulating activity of innate immunity responding to intrauterine infection and its effect on lung development.


METHODS
On day 17 of pregnancy, 30 pregnant Sprague-Dawley (SD) rats were randomly divided into two groups: LPS group and saline group. For LPS group, LPS (10 microl, 40 microg/ml) was intrauterine injected between every two embryonic sacs of the pregnant rats, while the rats in the control group were injected with the same volume of pyrogen-free saline. Lung tissues of fetal rats and corresponding placental tissues were collected on the embryonic day 18 (E18), E20, and E22. Neonatal lung tissues were also harvested on postnatal day 1 (P1), P3, and P7. Lung sections and placental tissues were stained with hematoxylin and eosin for histological examination. Reverse transcription quantitative polymerase chain reaction (RT-PCR) analysis was performed to test mRNA expression for TLR4, myeloid differentiation 88 (MyD88) and IL-1beta, while immunohistochemistry was used to evaluate TLR4 and MyD88 expression in lung tissues. All data were analyzed with one-way analysis of variance (ANOVA) and q test.


RESULTS
(1) Placental hematoxylin-eosin staining showed a great number of neutrophils infiltration, obvious interstitial hyperplasia and narrow capillaries in placental tissues in the LPS group which indicated that intrauterine infection occurred. However, there were no obvious inflammatory cells in the control group. (2) On E18, E20 and E22, the lung of LPS group showed no obvious pathological changes, and there were no apparent neutrophils infiltrated in alveoli, then some structural changes appeared. On P7, we found that the number of alveoli decreased, space of alveoli was larger than ever, septa thickened, but without significant constructive disorder. (3) In the LPS group, the TLR4, MyD88 and IL-1beta mRNA levels increased compared with control group, higher than control group at E20 and E22 (P < 0.05), and peaked at E22. Then the expression levels of these substances decreased slowly. (4) The result of immunohistochemistry showed that in lung tissues of the two groups at E18, there was no remarkable positive staining of TLR4 and MyD88, which mainly expressed in cytoplasm of bronchiole and alveolar epithelial cells, then positive cells increased slowly.


CONCLUSIONS
(1) For perinatal rat lungs, intrauterine LPS infusion can induce an increased expression levels of TLR4 and MyD88 to a certain extent, which then returned to normal level gradually. At the same time, lung tissues showed a mild pathological change and inflammatory reaction. We propose that innate immune system of fetal lungs controls the magnitude of the LPS-induced cytokine response during the perinatal period. (2) The findings confirmed that LPS-activated signaling transduction pathway was the MyD88-dependent pathway.
Yu
Xiao
X
Department of Pediatrics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Xiao
Cai-ju
CJ
Dai
Hong-mei
HM
Wang
Wei
W
Meng
Jing
J
Zhang
Xiao-ting
XT
Ning
Qin
Q
Luo
Xiao-ping
XP
D000818:Animals; D000831:Animals, Newborn; D005260:Female; D007239:Infections / Q000378:metabolism; D053583:Interleukin-1beta / Q000378:metabolism; D008070:Lipopolysaccharides / Q000008:administration & dosage / Q000494:pharmacology*; D008168:Lung / Q000254:growth & development / Q000378:metabolism*; D053594:Myeloid Differentiation Factor 88 / Q000378:metabolism; D011247:Pregnancy; D011251:Pregnancy Complications, Infectious / Q000378:metabolism*; D011297:Prenatal Exposure Delayed Effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects*; D051197:Toll-Like Receptor 4 / Q000378:metabolism*
D053583:Interleukin-1beta; D008070:Lipopolysaccharides; C507781:Myd88 protein, rat; D053594:Myeloid Differentiation Factor 88; C495347:Tlr4 protein, rat; D051197:Toll-Like Receptor 4