Study Of MicroRNA Expression Of Osteogegesis And Angiogenesis During Canine Mandibular Distraction Osteogenesis

Objective: To establish surgical model of canine mandibular distraction osteogenesis and to obtain the differentially expression of microRNA expression profile of bone tissue from distraction gap,as well as bone tissue from mandibles of the dog embryo,normal new born dogs fracture,adult dogs fracture and adult dogs through high throughput sequencing technique.To predict microRNA target genes and to find the signal pathways associated with osteogenesis and angiogenesis by KEGG enrichment analysis.Method: The experiment were divided into 11 groups: distraction group without consolidation(DOI),distraction group with 2-week-consolidation(DO2),the fracture group for 2 weeks(corresponding to 1 week latency period and 1week distraction period)(MF1),the fracture group for 4 weeks(corresponding to1 week latency period,1 week distraction period and 2 weeks consolidation period)(MF2),30 days embryo group(E1),35 days embryo group(E2),50 days embryo group(E3),new born group(NB),the puppy 2-week group(P2),thepuppy 4-week group(P4)and the control group(AC).Each group includes 3dogs.DO1 and DO2 groups were selected three healthy adult dogs,in the right mandibular distraction osteogenesis.After 7 days latency period,the distraction was performed at a rate of 1mm/day for 7 days.Then the groups were to be executed and the right mandible were collected immediately after the completion of distraction or after 2 weeks of consolidation period respectively.Then the animals were sacrificed and the right mandibles was collected.MF1 and MF2 groups were selected three healthy adult dogs separately,the right mandibulars were cut off.The fracture space was 7mm.E1,E2,E3 groups were selected 9 pregnant dogs about 30 days,35 days and 50 days.Then those embryos were taken into the experiment.We also selected three new born dogs,three 2 weeks old dogs,three 4 weeks old dogs and three healthy adult dogs into NB group,P2 group,P4 group and AC group respectively.These groups were subjected to high-throughput microRNA sequencing.Results:1)General observation: All groups had successfully undergone operation and the subsequent follow-up observation,without death,infection and serious complications.The distractor and titanium plate were fixed firmly and no loosening in DOI and DO2 groups.Most new born tissues in distraction areas,florid and soft,were fibrous tissues and cartilage without obvious callus.2)Adequate total RNAs was extracted from all 11 samples.High-throughput microRNA sequencing of each group was completed successfully.A total of 354 microRNAs were expressed in the 11 samples.3)Compared with the DO group,E group,P group and AC group,we found 27 differentially expressed microRNAs,of which 18 were up-regulatedand 9 were down-regulated.PTEN was predicted as the target gene of mi R-494,which may be one of the reason of rapid osteogenesis in distraction osteogenesis.Conclusion: MiR-494 expression has significant difference in DO group,E group,P group and AC group,suggesting that distraction osteogenesis,embryo development and rapid growth may have similar regulation.MiR-494 could through the negative control target gene PTEN,thus activating PI3K-Akt signaling pathway to promote angiogenesis,vasculogenesis and osteogenesis.