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| Cloning of TLR4 Gene Fragment and Its Expression and Distribution Characteristic in Organs of Mongolian Gerbil (Meriones unguiculatus) |
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Abstract Toll-like receptors (TLRs) are highly conservative protein regulators participating in innate immune response. This test aimed to clone and analyse TLR4 gene fragment and distribution in organs of experimental animal Mongolian Gerbil (Meriones unguiculatus). Conservative region of TLR4 gene which was compared among rat (Rattus norvegicus), mouse (Mus musculus) and hamster (Mesocricetus auratus) was chosen to design the primers, and Mongolian Gerbil TLR4 was cloned. After sequencing, the sequences were further analyzed and compared homology with DNASTAR Meglign software between Mongolian Gerbil and other different species of animals. Semi-quantitative PCR and immunohistochemical assay were used to determine the expression levels of TLR4 mRNA and protein in heart, liver, spleen, lung and kidney of normal Mongolian Gerbil, respectively. Results demonstrated that TLR4 gene fragment of Mongolian Gerbil (GenBank accession No. KX137123) was successfully cloned and showed more than 87.4% homologous with other mouse genus, and the highest for 89.1% with hamster. Homology with human (Homo sapiens) and swine (Sus scrofa) showed 69.5% and 73.1%, respectively. These results showed that the kinship of TLR4 gene might be relatively closer between Mongolian Gerbil and hamster, and farther with human and swine. Semi-quantitative PCR showed that TLR4 differentially expressed in 5 different tissues, and the expression was the highest in lung but the lowest in liver. Immunohistochemical assay further confirmed the semi-quantitative PCR result in protein level. Positive reaction products mainly distributed in lung and spleen in Mongolian Gerbil. High abundance of TLR4 proteins distributed mainly in alveolar bronchial epithelial cells of lung. TLR4 was located mainly in marginal zone around splenic nodule and with little on nodule region of spleen. Sporadic TLR4 proteins could be found in kidney. But no observant reaction products was detected in heart and liver definitely. This study will be a pioneer test for further research on innate immunity of Mongolian Gerbil which performs excellent infection model for some specific pathogens.
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Received: 01 April 2016
Published: 06 August 2016
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