BACKGROUND: A middle cerebral artery occlusion and reperfusion(MCAO/R) model in rats with suture has been widely used in the researches of acute focal ischemic cerebral infarction, while the model in rabbits by the same method is relatively rare. Magnetic resonance diffusion weighted imaging (MR DWI) has been paid close attention recently for its sharp sensitivity of cerebral ischemia.OBJECTIVE: To establish rabbit models of MCAO/R by intraluminal thread, and study the characteristics of MR DWI after cerebral ischemia and reperfusion.DESIGN: Random controlled animal experiment.SETTING: Institute of Cerebrovascular Diseases, Affiliated Hospital of Qingdao University Medical College.MATERIALS: The experiment was accomplished at the Key Laboratory of Brain Diseases Prevention and Cure of Shandong Province from March to June in 2005. A total of 103 adult healthy New Zealand rabbits of either sex, 10-12 weeks old and 1.8-3.3 kg weight were provided by the Experimental Animal Center of Shandong Agricultural Academy (SCX20040013).They were bred at quiet, sanitary and dry conditions.METHODS: Animal groups: 103 rabbits were divided randomly into group A (n=53) and group B (n=50). The rabbits in group A were treated with suture of 0.51-0.55 mm as the diameter of thread, while group B was reassigned into B1 (0.46-0.50 mm), B2 (0.51-0.55 mm) and B3 (0.56-0.60 mm).The successful MCAO/R models in 57 cases were randomly divided into permanent ischemia group (n=30, ischemia 1, 3, 6, 12, 24 andl 48 hours, 5ones at each time point) and ischemic reperfusion group (n=27, reperfusion 0, 2 and 5 hours, 5 ones at each time point; reperfusion 11, 23 and 47hours, 4 ones at each time point). Another 10 rabbits receiving sham operations were regarded as contrasts for permanent ischemia group and ischemia reperfusion group, with 5 ones in each.MAIN OUTCOME MEASURES: The changes of hyperintensity area on DWI and apparent diffusion coefficient (ADC) were measured in permanent ischemia group and ischemic reperfusion group.RESULTS: The data of 57 successful model rabbits were involved in the result analysis.①The successful rate in group A (26 cases, 49.1%) was significantly lower than that in group B (31 cases, 62.0%).②In ischemia group:The hyperintensity area on DWI with declined ADC appeared at ischemia 1 hour. The hyperintensity areas on DWI at different times increased gradually from ischemia 1 hour and unchanged within 24 hours. The mean ADC at different times declined at first and then gradually increased.③In reperfusion group: Comparing with ischemia 1 hour, the hyperintensity area on DWI reduced while ADC increased at reperfusion 2 hours and 5 hours, and enlarged with ADC high at reperfusion 11 hours, then continued to enlarge with ADC reduced significantly at 23 hours and 47 hours.CONCLUSION: The diameter of thread tip and the inserting distance of thread are main factors for establishing successful MCAO/R models. The hyperintensity area on DWI and the decreasing ADC after acute cerebral ischemia can be improved by early reperfusion, but the secondary decreasing ADC may be induced by continuously reperfusion.

The cloning and identification of frc gene from Oxalobacter formigenes in the intestines of Chinese people were conducted. The genomic DNA of Oxalobacter formigenes was extracted. frc gene fragment was amplified by polymerase chain reaction (PCR) and linked with pEGFP-C1. The recombinant plasmid was designated pEGFP-frc and was identified by restriction-enzyme digestion and sequencing. Human embryo kidney 293 cells were transfected with pEGFP-frc, then RT-PCR and Western blotting were performed to detect the expression of frc gene. The length of frc gene was found to be 1287 bp, and the homology of nucleotides and amino-acid residue with the sequence in GenBank was 95.88% and 99.07%. Bright green fluorescent light could be observed in 293 cells transfected with the pEGFP-frc. frc mRNA and fusion protein FCoAT-EGFP were detected in the cells. It is concluded that frc gene cloned from the Oxalobacter formigenes in the intestines of Chinese people can be expressed in eucaryotic 293 cells and keep its enzyme activity.

The cloning and identification of frc gene from Oxalobacter formigenes in the intestines of Chinese people were conducted. The genomic DNA of Oxalobacter formigenes was extracted. frc gene fragment was amplified by polymerase chain reaction (PCR) and linked with pEGFP-C1. The recombinant plasmid was designated pEGFP-frc and was identified by restriction-enzyme digestion and sequencing. Human embryo kidney 293 cells were transfected with pEGFP-frc, then RT-PCR and Western blotting were performed to detect the expression offrc gene. The length of frc gene was found to be 1287 bp, and the homology of nucleotides and amino-acid residue with the sequence in GenBank was 95.88% and 99.07%. Bright green fluorescent light could be observed in 293 cells transfected with the pEGFP-frc. frc mRNA and fusion protein FCoAT-EGFP were detected in the cells. It is concluded that frc gene cloned from the Oxalobacterformigenes in the intestines of Chinese people can be expressed in eucaryotic 293 cells and keep its enzyme activity.