Ultrastructure Change Of Skeletal Muscle Cell In Pelvic Floor Induced By Stress

Objective:Apply loading force on the pelvic floor skeletal muscle cells with Flexercell4000cell loading system to clarify the relationship between loading time, loading tension and cell ultrastructure injury, to clarify the change cell apoptosis rate, to discuss alpha actin gene expression level changes through loading force on the pelvic floor skeletal muscle.Methods:First take levator muscle tissue from the patients with abdominal perineal joint colorectal cancer radical operation and carry on the in vitro primary culture, purification and identification. Take two to five generation stability batches of skeletal muscle cells, for subsequent strength provides the material basis.And then apply loading force on the pelvic floor skeletal muscle cells with Flexercell4000cell loading system. Set strength3%and6%respectively, time6h and24h respectively, not load force of static cells as control. Finally collected cells, and with the help of transmission electron microscopy (tem), make a qualitative description of morphological and ultrastructure changes of skeletal muscle cells before and after the strength;Apply flow cytometry technology to detect the change of cells apoptosis rate in skeletal muscle cells before and after the strength; By real-time quantitative PCR (RT-PCR) method, determine alpha actin gene expression level changes through loading force on the pelvic floor skeletal muscle.Results:1. By transmission electron microscope, under the condition of strength, pelvic floor skeletal muscle cells morphology and its internal ultrastructure show series of adaptive change or traumatic change.Pelvic floor skeletal muscle cells were found that under the condition of strength by3%for6hours,compared to the control group,cell form is not obvious difference, organelles shape is normal, cells is in neat rows and it is not seen obvious nucleus changed; With the increase of strength intensity of the extension of time and strength, cells form are gradually broken, arranged disorder, the membrane structure is not clear or even disappear, mitochondria swelling, ridge become shorter or even disappear, cell nucleus invaginate and pycnosis, heterochromatin increased.2. Flow cytometry instrument testing results showed that under the condition of strength intensity, skeletal muscle cell apoptosis rate increased. Under the condition of fixed strength time,apoptosis rate in6%strength group increased higher than in3%strength group.They were statistically significant (F=44.742,P<0.05).With the increase of strength intensity, apoptosis rate in pelvic floor skeletal muscle increase. Under the condition of fixed strength intensity,apoptosis rate in strength group for24h increased higher than in strength group for6h.They were statistically significant (F=15.998,P<0.05). With the increase of strength time, apoptosis rate in pelvic floor skeletal muscle increase. Strength time and the strength intensity existed the interaction (F=3.575, P＜0.05), and under the condition of6%strength group for24h, the cell apoptosis rate increased most obviously.This experiment showed that under the effect of mechanical stretching force, apoptosis rate in pelvic floor skeletal muscle change with the strength of the strength of time and strength.3. RT-PCR results show that under the condition of strength intensity, skeletal muscle cell alpha actin mRNA expression levels decreased. Under the condition of fixed strength time, cell alpha actin mRNA expression levels in6%strength group decreased more obviously than in3%strength group.They were statistically significant (F=90.743,P<0.05). With the increase of strength intensity, alpha actin mRNA expression levels in pelvic floor skeletal muscle decrease. Under the condition of fixed strength intensity, cell alpha actin mRNA expression levels in strength group for24h decreased more obviously than in strength group for6h.They were statistically significant (F=23.151,P<0.05).With the increase of strength time, the alpha actin mRNA expression levels in pelvic floor skeletal muscle decrease. Strength time and the strength intensity existed the interaction (F=12.908, P<0.05), and under the condition of6%strength group for24h, the alpha actin mRNA expression levels increased most obviously. This experiment found that with loading force time and strength on the pelvic floor skeletal muscle cells increasing, alpha actin (alpha actin) gene expression level showed a trend of decline.Conclusion:This experiment found that under mechanical stretch after stimulation, pelvic floor anal muscle cells morphology, ultrastructure and skeleton protein gene expression level were in a series of changes.This cause the damage of cell function, and then it may cause pelvic floor tissues relaxation,finally leading to pop.