Effect of electroacupuncture at "Jiaji" (EX-B2) on senescence of nucleus pulposus cells of degenerated lumbar intervertebral disc in rabbits.

Abstract

Objectives: To observe the effect of electroacupuncture (EA) at "Jiaji"(EX-B2) on body mass, motor function, expression of caveolin-1 (Cav-1) in nucleus pulposus cells and annulus fibrosus tissue, telomerase activi-ty, relative telomere length and different cell cycle ratio of nucleus pulposus cells in rabbits with intervertebral disc degeneration(IVDD), so as to investigate its mechanism underlying delaying senescence of the degenerated lumbar intervertebral disc nucleus pulposus cells.

Methods: Twenty-five male New Zealand rabbits with mature bones were divided into control, sham operation, model, EA, and acupuncture groups, with 5 rabbits in each group. The IVDD model was established by inserting kirschner wires to the vertebral bone surface between the lumbar (L)4 and L5 vertebrae, followed by applying continuous axial pressure for 28 d. EA (2 Hz/15 Hz, 1-2 mA) or acupuncture (only insertion of acupuncture needles into bilateral EX-B2, but without electrical stimulation) was applied to bilateral EX-B2 for 20 min, once daily, 6 times a week for 4 weeks. The hindlimb locomotor function (locomotor score) was assessed by using Faden's and colleagues' methods. The general conditions of rabbits in each group were observed, and their body weight changes were measured every week. Nucleus pulposus cells were isolated using enzyme digestion method. After the treatment, the Cav-1 positive cell counts in nucleus pulposus cells and annulus fibrosus tissues were detected by immunohistochemistry, and the telomerase activity of nucleus pulposus cells was detected by PCR-ELISA. The relative telomere length of nucleus pulposus cells was measured by real-time quantitative polymerase chain reaction (real-time qPCR), and the cell cycle of nucleus pulposus was detected by flow cytometry.

Results: Compared with the sham operation group, the body mass from 4 to 11 week, locomotor score at 4, 7 and 11 week, telomerase activity, relative telomere length and the proportion of cells in G2/M phase of nucleus pulposus cells were significantly decreased (P<0.01), while Cav-1 positive cell counts of nucleus pulposus and annulus fibrosus tissue, and the proportion of nucleus pulposus cells in the G0/G1 phase considerably increased (P<0.01) in the model group. Relevant to the model group, the EA group rather than the acupuncture group had an increase in the body mass from 8 to 11 week, locomotor score at 11 week, telomerase activity, relative telomere length of nucleus pulposus cells, and the proportion of nucleus pulposus cells in G2/M phase (P<0.01), and a decrease in the Cav-1 positive cell counts of nucleus pulposus and annulus fibrosus tissue and the proportion of cells in G0/G1 phase (P<0.01). No significant differences were found between the model and acupuncture groups in all the indexes mentioned above.

Conclusions: EA at EX-B2 has a bene-ficial effect in improving motor function in rabbits with IVDD, which may be related to its functions in reducing the expression of Cav-1 in nucleus pulposus cells and annulus fibrosus, improving cycle arrest, enhancing the telomerase activity and the relative telomere length of nucleus pulposus cells, delaying the senescence of nucleus pulposus cells of the degenerated lumbar intervertebral discs.