Background: MicroRNA-449b (miR-449b) is a ~22-base pair long nucleotide, that post-transcriptionally affects processes like proliferation, differentiation and apoptosis by directly targeting the HDAC and DNMTs family genes that regulate DNA methylation. 
 
Methods: This study aimed to investigate the effect of miR-449b mimic, inhibitor and scrambled sequence on buffalo fibroblast cells containing the human insulin (hINS) gene. A transgenic buffalo cell line containing human insulin gene (hINS) was treated with 40 nM miR-449b mimic and inhibitor by transfection method (lipofectamine-3000). 
 
Result: miR-449b mimic treated cells showed a significant decrease (P&lt;0.05) in the expression of epigenetic-related genes and found a significant reduction (P&lt;0.05) in the expression of HDAC1, DNMT3A and DNMT3B genes. In contrast, DNMT1 showed no significant change (P&lt;0.05) in their expression level as compared to control whereas miR-449b inhibitor-treated cells found significantly increased (P&lt;0.05) expression levels of HDAC1, DNMT 3A, DNMT 3B and DNMT1 genes as compared to control. In apoptosis-related genes, miR-449b mimic significantly increased (P&lt;0.05) the expression of BCL-XL and MCL-1, while miR-449b inhibitor significantly increased (P&lt;0.05) the expression of MCL-1 gene whereas there was no significant change (P&lt;0.05) in the expression of BCL-XL. These findings indicate that treatment of miR-449b mimic on transgenic cells reduces the DNA methylation and apoptosis levels. This reduced methylation level helps in the nuclear reprogramming of transgenic cells. Thus, these cells can be a better choice as donor somatic cells for efficient handmade transgenic cloned embryo production.

Microrna-449b Reduces Methylation and Apoptosis-related Gene Expression in Fibroblast Cells Containing Human Insulin Gene