Interleukin-1 beta affects the biological properties of rat nucleus pulposus-derived mesenchymal stem cells

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Abstract

BACKGROUND:

Endogenous stem cells have no repair effects on the process of disc degeneration. Authors assumed that this maybe associate with abnormal effects of related etiological factor, resulting in an inhibitory effect on the function of nucleus pulposus-derived mesenchymal stem cells.

OBJECTIVE:

To investigate the effects of inflammatory cytokine interleukin-1β on biological characteristics of nucleus pulposus-derived mesenchymal stem cells of rats.

METHODS:

Lumbar spinal nucleus pulposus was obtained from 3-month-old male Sprague-Dawley rats. Nucleus pulposus-derived mesenchymal stem cells were isolated and cultured with collagenase and sequential trypsin digestion. The expression of CD24, CD34, CD45, CD90 and CD105 was detected using flow cytometry. Stem cell gene SOX2 and Nanog expression was measured using RT-PCR. Adipogenic, osteogenic and chondrogenic abilities of nucleus pulposus-derived mesenchymal stem cells were observed. The apoptotic rate of interleukin-1β-treated nucleus pulposus-derived mesenchymal stem cells was detected using flow cytometry. Fluorescent quantitative PCR was used to measure the expression of SOX9, proteoglycan, type II collagenase and caspase-3 gene after nucleus pulposus-derived mesenchymal stem cells were treated with interleukin-1β.

RESULTS AND CONCLUSION:

Cells isolated and cultured in vitro showed spindle-like fusiform. Nucleus pulposus-derived mesenchymal stem cells were positive for CD90 and CD105, negative for CD45, CD34 and CD24. Nucleus pulposus-derived mesenchymal stem cells expressed SOX2 and Nanog, and were able to differentiate into osteocytes and chondrocytes, but not adipocytes. By the cell counting kit-8, interleukin-1β inhibited the proliferation capacity of nucleus pulposus-derived mesenchymal stem cells, showing a dependent relationship with action time and action concentration. The optimum reaction time was 48 hours, and the optimal concentration was 50 μg/L. Flow cytometry detected that interleukin-1β induced a slight apoptosis of nucleus pulposus-derived mesenchymal stem cells. Fluorescent quantitative PCR results revealed that mRNA expression of SOX9, proteoglycan, and type II collagenase was decreased, but caspase-3 was increased. Results indicated that interleukin-1β inhibited the proliferation of nucleus pulposus-derived mesenchymal stem cells and induced the apoptosis of nucleus pulposus-derived mesenchymal stem cells, and was a reason that intervertebral own stem cells could not exert repair effects.

Subject headings:

intervertebral disk; mesenchymal stem cells; interleukin-1 beta; cell proliferation; apoptosis Funding: the Natural Science Foundation of Shanxi Province, No. 2011011042-3; the Undergraduate Innovation and Entrepreneurship Project of Taiyuan City, No. 120164064; the Excellent Innovation Project of Shanxi Province for Postgraduates, No. 20113069

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