| Objective:To observe the effect of acupotomy therapy on matrix aggrecan(AGG),type II collagen(Col-Ⅱ)and matrix metalloproteinase 13(MMP-13)in cartilage endplate of rabbits with cervical spondylosis,and to explore the regulatory effect of acupotomy on cartilage endplate matrix of rabbits with cervical spondylosis,in order to provide laboratory basis for acupotomy treatment of cervical spondylosis.METHODS:1.Forty New Zealand rabbits were randomly divided into blank group,model group,acupotomy group and electro-acupuncture group(10 in each group).2.The experimental animals that need to be modeled were placed in a self-made wooden modeling box to keep the head flexion 45 °for 5 hours each time,once a day for 12 weeks.Before and after modeling,two rabbits in each group were randomly selected from blank group,acupotomy group,electroacupuncture group and model group to take lateral X-ray films.Before shooting,rabbits were tied to a suitable rectangular plank with tape,and each rabbit was photographed in the same position twice to verify the success of the model.3.One week after successful modeling,electroacupuncture intervention was performed in the electroacupuncture group.The neck hair of rabbits with cervical spondylosis was removed,disinfected with iodophor,acupuncture was performed at "Dazu"(BL 11)acupoint,"Bailao"(EX-HN 15)acupoint and "Tianzhu"(BL 10)acupoint,and electroacupuncture was received on the ipsilateral Tianzhu(BL 10)acupoint and Dazhu(BL 11)acupoint for 20 minutes each time,3 times a week and once every other day for a total of 3 weeks.One week after the model was successfully made,the acupotomy group was treated with acupotomy.First,the experimental rabbits were pacified,and after calming,the neck hair of the rabbits with cervical spondylosis was removed.Then,according to the four-step needle method,(1)Fixed point:the acupotomy operator touched the soft tissue on both sides of the cervical spinous process of the cervical spondylosis model rabbit,according to the neck muscle stiffness and the position of the cord.The acupotomy entry point was selected within the range of C2-C7 spinous process point and peri-spinous process 1-1.5cm.Each time,two points on both sides of the spinous process and the spinous process point in the lesion site were selected as the acupotomy entry point,and each time the acupotomy entry point was selected alternately.The needle entry point to be treated with the acupotomy was marked with a marker pen and disinfected with iodophor.(2)orientation: the acupotomy operator wear aseptic gloves,the acupotomy edge line was parallel to the median line of the rabbit model of cervical spondylosis,and the acupotomy body was perpendicular to the skin of the posterior part of the cervical vertebra.(3)compression: the tissue at the needle entry site was pressurized and separated with the thumb of the left hand.(4)puncture: the acupotomy operator held the acupotomy in his right hand,and quickly inserted the acupotomy into the subcutaneous tissue of the needle point along the edgeof the left fingernail to reach the bone surface of the cervical vertebrae.The diseased tissue was cut longitudinally for 3 times within the range of 0.3cm,then the needle was stripped horizontally for 2 times,and then the needle point was pressurized to stop bleeding.Acupotomy was treated once a week for 3 weeks.The model group was not treated.4.One week after the end of the intervention,all rabbits were killed by air embolization.The c4-5 intervertebral disc was quickly removed and the cartilage endplate was separated under a surgical microscope.The cartilage endplate was placed in a plastic test tube,sealed and frozen at-80 ℃.5.The m RNA and protein expression levels of cartilage endplates aggrecan(AGG),type II collagen(Col-Ⅱ)and matrix metalloproteinase 13(MMP-13)were detected by Real-time PCR and Western blot.RESULTS:1.Real-time PCR:Compared with the blank group,the expression of aggrecan m RNA in the model group was significantly decreased(p<0.01),the expression of type II collagen m RNA was significantly decreased(p<0.01),and the expression of matrix metalloproteinase 13 m RNA was significantly increased(p<0.01).The expression of type II collagen m RNA in the electro-acupuncture group decreased(p<0.05).Compared with the model group,the expression of aggrecan m RNA in the electroacupuncture group increased(p<0.05)and the expression of matrix metalloproteinase13 m RNA decreased(p<0.05).the expression of aggrecan m RNA in the acupotomy group increased significantly(p<0.01),type II collagen m RNA expression increased(p<0.05),and matrix metalloproteinase 13 m RNA expression decreased(p<0.05).2.Western blot:Compared with the blank group,the aggrecan protein expression of the model group decreased significantly(p<0.01),the type II collagen protein expression decreased(p<0.05),and the matrixmetalloproteinase 13 protein expression increased significantly(p<0.01).Compared with the model group,the aggrecan protein expression of the electro-acupuncture group increased(p<0.05)and the matrix metalloproteinase 13 protein expression decreased(p<0.05).The aggrecan protein expression of the acupotomy group increased significantly(p<0.01)and the type II collagen protein expression increased(p<0.05),matrix metalloproteinase 13 protein expression decreased significantly(p<0.01).Conclusion:Acupotomy therapy for cervical spondylosis can regulate the expression of aggrecan,type II collagen,matrix metalloproteinase 13 m RNA and protein,repair the degenerated cartilage endplate.which may be one of the effect mechanisms of acupotomy therapy for cervical spondylosis. |
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