Effects of arachidonic acid and prostaglandins on proliferation, differentiation, and fusion of bovine myoblasts

Abstract Text:

Arachidonic acid (AA) is a major lipid component of the plasma membrane and the precursor of prostaglandins (PG) in skeletal muscle. The objective of this study was to determine the effects of AA and its major PG derivatives PGE₂, PGF_2α, and PGI₂ on the proliferation, differentiation, and fusion of bovine myoblasts. Satellite cells were isolated from 6 Angus or Angus crossbred steers (experimental unit) and were expanded as myoblasts in growth medium for a week before being used in the following tests. In the proliferation test, myoblasts were cultured in growth medium with 10 μM AA, 1 μM PGE₂, 1 μM PGF_2α, 1 μM PGI₂, or vehicle control for 24 h. Proliferating cells were identified by EdU labeling. This test revealed that AA, PGE₂, PGF_2α, and PGI₂ each increased the number of proliferating myoblasts by 13%, 24%, 16%, and 16%, respectively, compared to the control (P<0.05). In the differentiation and fusion test, myoblasts were induced to differentiate and fuse into myotubes in the presence of the aforementioned treatments or control, for 24, 48, and 72 h. The differentiation status of myoblasts was assessed by reverse transcription-quantitative PCR of myogenin (MYOG), myosin heavy chain 3 (MYH3), and muscle creatine kinase (CKM) mRNAs, which are markers of differentiated myoblasts. The fusion level of myoblasts was estimated by calculating the percentage of nuclei located in myotubes, i.e., fusion index. Compared to the control, AA increased MYOG mRNA expression at 24 and 48 h, MYH3 mRNA expression at 24 and 48 h, and CKM mRNA expression at 24, 48, and 72 h of differentiation (P<0.05); PGE₂ increased MYOG mRNA expression at 24 and 72 h, and MYH3 mRNA expression at 72 h of differentiation (P<0.05); PGF_2α increased CKM mRNA expression at 72 h of differentiation (P<0.05); and PGI₂ had no effect on mRNA expression of any of the three markers at any of the three times of differentiation. Compared to the control, PGE₂ increased the fusion index by 14% (P<0.05) but the remaining treatments had no effect on this index. In conclusion, this study demonstrates that AA, PGE₂, PGF_2α, and PGI₂ stimulate the proliferation, that AA and PGE₂ stimulate the differentiation, and that PGE₂ stimulates the fusion of bovine myoblasts in vitro.

Keywords:

cattle, myoblasts, prostaglandins