Research On The Regulation Of Fast-growing Antler Extract On Chondrocytes

Objective: Study on the regulatory effect of velvet antler extract on chondrocytes and related biological mechanisms.Methods: Preparation of fast-growing velvet antler extract by homogenization,low-temperature extraction and freeze-drying.Proteome research on apical growth tissues of velvet antler in primary,rapid growth and ossification stages using isotope labeling and mass spectrometry identification techniques.Combined with bioinformatics analysis methods,screen differential analysis of proteomic databases and analyze related functions.The CCK-8 method and Alixin blue staining method were used to study the role of velvet antler extract in the rapid growth period on chondrocytes.Using high-throughput RNA-seq technology to establish a transcriptome database of primary chondrocytes after deer antler extract(DAE),combined with bioinformatics analysis methods,differentially expressed gene analysis and related functional annotation of the transcriptome database were performed.Results:(1)The average yield of fast-growing deer antler extract(DAE)reached 2%,and the protein content in DAE reached 70%.(2)An in-depth analysis was performed on the active proteins whose expression was significantly up-regulated in the rapid growth phase of antler antler compared to the nascent phase.They mainly included related active proteins involved in blood circulation and chondrocyte growth and development;An in-depth analysis of differential proteins that significantly increase the expression of velvet antler in fast-growing phase compared with ossifying phase,found that these differential proteins are mainly involved in the circulatory system and oxygen transport,nerve growth and regeneration,cartilage and bone tissue development,and ATP synthesis Related proteins.(3)The results of CCK-8 experiments show that DAE can promote chondrocyte proliferation in a dose-dependent manner.The effects are most obvious at drugconcentrations of 0.64 mg / ml and 1.28 mg / ml,and the effects are basically the same.Alixin blue staining test verified the promotion effect of DAE on chondrocyte proliferation,and the optimal concentration was consistent with the results of CCK-8 experiment.(4)A transcriptome database of mouse primary chondrocytes with DAE effect was constructed.Through bioinformatics analysis,a total of 639 differentially expressed genes were identified.The DAE treatment group significantly increased 365 genes and significantly decreased 274 genes.(5)Screening and analysis of differentially expressed genes,screen out differentially expressed genes(Dbf4,Usp3,E2 f,etc.)that promote cell proliferation from genes whose expression is significantly up-regulated,participate in oxidative stress(MT2,MT1,SOD3,etc.),and inflammatory responses(Ptpn2,Relb,S1pr3,etc.)and immune response genes(Lbp,C1 ra,C1sa,etc.)differentially expressed genes.From genes whose expression levels were significantly down-regulated,we screened genes related to the regulation of chondrocyte differentiation(Lect1,Tgf?3,Sfrp1,etc.).(6)The results of Q-PCR experiments are consistent with the results of transcriptome sequencing,which proves that the transcriptome sequencing results are authentic and reliable.Conclusion: The fast-growing antler apical growth tissue contains a large number of related active protein components involved in blood circulation,oxygen transport,nerve growth,chondrocyte proliferation,and ATP synthesis.The high expression of these proteins makes the fast-growing velvet antler have the highest quality.The fast-growing velvet antler extract can promote the proliferation of primary chondrocytes in a dose-dependent manner.The mechanism of action is to significantly increase the proliferative activity of chondrocytes,while inhibiting the proliferation and differentiation of chondrocytes.DAE can also improve the anti-oxidation and anti-inflammatory properties of chondrocytes.And immune response.