ABSTRACT
This study aimed to use the sika deer as a model to study the influence of IGF-1 on the expression of Col Ⅰ in antler chondrocytes.The chondrocytes were separated from sika deer antlers,cultured and were treated with recombinant human IGF-1 protein (rIGF-1),both rIGF-1 and PQ401,and transfected with IGF-1 over-expression plasmid or IGF-1 siRNA,respectively.The expression of Col Ⅰ which,a well-known marker for chondrocytes dedifferentiation,was detected by real-time PCR.The results showed that administration of rIGF-1 to antler chondroctyes resulted in an obvious decrease of Col Ⅰ mRNA levels,while PQ401 pretreatment could dramatically attenuate the effects of rIGF-1 on the expression of Col Ⅰ mRNA.After transfection with IGF-1 over-expression plasmid,the expression of Col Ⅰ mRNA was obviously reduced in antler chondrocytes compared with control.Conversely,knockdown IGF1 with specific siRNA could increase the expression of Col Ⅰ in antler chondrocytes.These results indicate that IGF-1 may play an important role in process of antler chondrocyte dedifferentiation.
ABSTRACT
Objective:To construct the pleiotrophin (PTN )overexpression vector,and to explore the effect of PTN on the decidualization of uterine stromal cells in the mice.Methods:The specific primers containing restriction enzyme cutting sites were designed according to the PTN gene sequences published in GenBank for PCR amplification.The amplified fragment of PTN was recovered from the agarose gel and cloned into the pGEM-T vector.The pGEMT-PTN was cut by double enzyme digestion and ligated into pcDNA3.1 (+)to construct the PTN overexpression plasmid.After transfection with PTN overexpression plasmid,the expression levels of PTN mRNA in the uterine stromal cells and the expression levels of decidualization markers Prl8a2 and Prl3c1 were detected by qRT-PCR method.The uterine stromal cells transfected with pcDNA3.1 (+)empty vector were used as control group. Results:The results of identification by double enzyme digestion indicated that the bands of PTN overexpression plasmid were consistent with those of the target gene,and the clone sequencing results suggested that it had 100% homology with mouse PTN gene sequence published in GenBank.Compared with control group, the expression levels of PTN,Prl8a2 and Prl3c1 mRNA in mouse uterine stromal cells in PTN overexpression group were significantly increased (P < 0.05).Conclusion:PTN overexpression could increase the expression levels of decidualization markers in mouse uterine stromal cells,indicating that PTN might play an enhancement effect during uterine decidualization in the mice.