Bioinformatics analysis of surfactant-associated protein a in cotton rat and its relationship with lung injury / 第二军医大学学报

ABSTRACT

Objective To analyze the pulmonary surfactant-associated protein A (SP-A) gene sequence and its bioinforma tic characteristics and to observe SP-A mRNA and protein levels in lung injury model of cotton rats, so as to explore the correlation between lung injury and SP-A expression. Methods A total of 32 cotton rats were randomly divided into control (normal saline, NS) group and three lipopolysaccharide (LPS)-induced lung injury groups (the cotton rats were injected intraperitoneally with 2 mg/kg LPS for 24, 48 and 96 hours, respectively). The total RNA was extracted from lung tissue and SP-A gene was amplified by RT-PCR. Then bioinformatic analysis was done for SP-A characteristics. Histopathological changes of lung tissue were observed at different time points by hematoxylin-eosin staining; the mRNA level of SP-A was detected by real time-PCR and the protein level of SP-A was detected by Western blotting analysis. Results The coding region of cotton rat SP-A gene had 744 bp and encoded 248 amino acids, containing six cysteine conserved sites, four crhelices and two predicted N-glycosylation sites. Meanwhile, cotton rat SP-A shared high level of homology in the nucleotide sequences (75. 4%-90. 1%) and in deduced amino acid sequences (70. 6%-87. 1%) with other species. Moreover, histopathological changes of lung tissues in three LPS groups were more notable and severe than those in the control group, and the changes were related to LPS treatment time. Compared with the control group, the mRNA and protein levels of SP-A in lung tissues were significantly increased in three LPS groups, with rapid increase starting from 24 h after treatment (P<0. 001, P<0. 01), with continuous increase at 48 h (P<0. 001 and P<0. 01), and slight decrease and still keeping at a high level at 96 h (P<0. 05, P<0. 01). Conclusion SP-A gene of cotton rat is highly conservative; the mRNA and protein levels of SP-A are closely associated with the severity of lung injury.