| Objective: To observe the difference of Brn-4 mRNA expression inhippocampus between the fimbria/fornix-transected rats and normal ones,and to investigate the molecular biology mechanism of neural regenerationand reparation after fimbria/fornix-transection. Methods: (1) RT-PCR:Forty-two SD rats were randomly divided into 7 groups, 6 rats in each group.One group served as normal control and the others served as fimbria/fornixtransected 1st, 3rd, 7th, 14th, 21st and 28th day group, respectively. Thenhippocampi were isolated and total RNA was extracted. Semi-quantitativereverse transcriptase-polymerase chain reaction (RT-PCR) method was usedin detection of the expression of Brn-4 mRNA in hippocampus. The relativeexpression level of Brn-4 mRNA was indicated by the ratio of the opticaldensity value of Brn-4 to that of GAPDH. The Stata7．0 software was used inone-factor analysis of variance and comparison between every two groups.(2) In Situ Hybridization: Six SD rats' right fimbrias were transected. Onthe 14th day after transection, the rats were perfused and fixed, then cryostatsections of hippocampus were prepared. Digoxigenin-labeled Brn-4 RNAprobe was prepared and used in hybridization to observe expression changeof Brn-4 mRNA in hippocampus after operation. Three sections were takenfrom each rat in anterior, middle and posterior location, respectively. Thenumber and the mean optical density value of Brn-4 mRNA positive cells intransected and normal sides was measured, respectively. Paired t test analysiswas used in statistic analysis of the number and the mean optical densityvalue of Brn-4 mRNA positive cells with Stata7．0 software. Results: (1) RT-PCR: In normal group, The relative expression level of Brn-4 mRNAwas 0．138±0．06．It started to increase on day3(0．256±0．54) aftertransection, and the peak appeared on day 14(0．719±0．124) , then decreasedslowly to pre-transection level on day28(0．189±0．059) . (2) In SituHybridization: Brn-4 mRNA was expressed in pyramidal layer ofhippocampus and granular layer of dentate gyrus in both sides. The numberof Brn-4 mRNA positive cells has no discrepancy between two sides. But themean optical density value of Brn-4 mRNA positive cells in transectedside(0．27±0．06) was higher than that in normal side(0．14±0．02) , t=11．4709,P<0．01．Brn-4 mRNA positive cells was also seen in hilus and subgranularlayer of dentate gyrus. In transected side, the number of Brn-4 mRNApositive cells was 40．5±9．37, the mean optical density value was 0．28±0．05．In normal side, there was only few Brn-4 mRNA positive cells (6．5±2．07) , and the mean optical density value was 0．09±0．02．For the twoobjects, P values were both less than 0．01．Conclusion: The expressionlevel of Brn-4 mRNA increases after fimbria/fornix transection. Theexpression of Brn-4 mRNA is changed with time, that is, it increases at firstand then decreases to normal level, and reaches peak on day 14; Brn-4mRNA is expressed in pyramidal layer of hippocampus and granular layer ofdentate gyrus in both sides. The expression level of Brn-4 mRNA issignificantly higher in transected side than that in normal side; Brn-4 mRNAis also expressed in hilus and subgranular layer of dentate gyrus in both sides.The number and the mean optical density value of Brn-4 mRNA positivecells were both significantly higher in transected side than these in normalside. The results suggest that the process in which the expression of Brn-4mRNA increases after fimbria/fornix transection mights be related to theneural stem cells differentiating into neurons in hippocampus.
|
PDF Full Text Request