Comparison of Two Different Methods of Establishment of Mouse Asthma Model / 医学研究杂志

Objective To compare the advantages and disadvantages of the two methods.Methods Thirty BALB/c mice were randomly divided into 3 groups:normal control group,OVA group and OVA/RSV group.With ovalbumin and hydroxide suspension sensitized,OVA nouse asthma model group and OVA/RSV group were established respectively by continuous atomization of ovalbumin and repeated nasal drops of RSV.The control group was established by sensitization and atomization excitation of sterile injection water.24 hours after the last stimulation,the lung function was measured.The cell sorting and counting in bronchoalveolar lavage fluid were performed.The pathological changes of lung tissue were observed by HE staining.Results Compared with the control group,the model groups have obvious symptoms of asthma,airway wall thickening,lumen stenosis and a large number of inflammatory cell infiltration.And pathological damage of lung tissue in OVA/RSV group was significantly increased,compared with OVA group.The enhanced expiratory pause (Penh)values of OVA/RSV group were significantly higher than those in group OVA (P ＜ 0.05),and the number of neutrophils,monocytes,lymphocytes and eosinophils in BALF were significantly higher than those in OVA group (P ＜ 0.05).Conclusion The establishment of asthma mouse model by OVA sensitization and RSV induction is a better way to simulate the human asthma pathogenesis.

High-bone-turnover Osteoporosis and Aortic Calcification in Opg Knockout Mice / 生物化学与生物物理进展

Bone turnover is regulated by local concentrations of cytokines such as osteoprotegerin (OPG) and receptor activator of nuclear factor kappaB ligand (RANKL). To explore the in vivo biological function of Opg and the mechanism of osteoporosis due to deficiency of Opg, Opg knockout mice have been generated through homologous recombination. Opg-/- mice exhibit a sharply decrease in bone density and strength as expected. The number of osteoclasts in Opg-/- mice significantly increases. Morphologically, osteoclasts appear more cuboidal in shape in Opg-/- mice than those of wt mice, suggesting that active osteoclastogenesis occurs in the absence of Opg. In consistent with this finding, an increase of osteoblast activity was also observed with accelerated mineral accumulation rate by histomorphometric measurement and elevated serum alkaline phosphatase activity (ALP) in Opg-/- mice. Interestingly, more than 50% of 2-month-old Opg-/- mice manifest medial calcification of aorta with comparable serum concentrations of calcium and phosphorus to wt mice. In conclusion, Opg-/- mice have a high-bone-rurnover type osteoporosis. The aortic calcification in Opg-/- mice is not due to abnormality of calcium and phosphorus metabolism. The mechanism underlying aortic calcification in Opg-/- mice needs to be further investigated.

Generation of Adiponectin Gene Knock-out and LacZ Gene Knock-in Mouse Model / 生物化学与生物物理进展

Adiponectin is an adipocyte-derived secretory protein. It was found to be associated with insulin resistance, inflammation and arteriosclerosis. To further study the biological function and expression of adiponection in vivo, adipoenctin gene knock-out and LacZ gene knock-in mouse model was constructed. Gene targeting strategy was designed to replace part of exon 2 and exon 3 of adiponectin gene with full length LacZ gene in frame with remaining upstream ATG and signal peptide sequence of exon 2. The targeting vector (Adipo-LacZ-XpPNT) was constructed and verified by restriction enzyme digestion and sequencing. CJ7 ES cells were transfected with targeting vector linearized by NotⅠ digestion, selected in the medium containing both G418 and ganciclovoir. Resistant clones were screened by PCR and further confirmed by Southern blot for correct homologous recombinants. Chimera mice were obtained by routing microinjection of homologous recombined ES cells into blastocysts. After mating, mice heterozygous and further homozygous for adiponectin knockout and LacZ gene knock-in were established. Expression of both endogenous adiponectin and exogenous LacZ gene in mouse tissues and sera were detected by RT-PCR, Northern-blot, Western blot and ELISA. The results show that adiponectin was disrupted at both mRNA and protein levels. LacZ gene is expressed exclusively in adipose tissue of mutant mice. Its expression profile is identical to endogenous adiponection. Unexpectedly, LacZ activity could not be detected in both adipose tissue and serum although LacZ protein can be detected in adipose tissue but not in serum of mutant mice. In conclusion, mice homozygous for adiponectin knockout and LacZ gene knock-in have been successfully constructed. Mutant mice display LacZ expression profile identical to endogenous adiponectin albeit neither LacZ activity nor protein can be detected in serum of mutant mice.