Chinese Journal of Tissue Engineering Research ›› 2021, Vol. 25 ›› Issue (8): 1172-1176.doi: 10.3969/j.issn.2095-4344.3036

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Morphology and activity of human nucleus pulposus cells under different hydrostatic pressures

Liu Zhichao1, 2, Zhang Fan3, Sun Qi3, Kang Xiaole3, Yuan Qiaomei3, Liu Genzhe4, Chen Jiang3, 5   

  1. 1Department of Orthopedics, Southern Branch of Guanganmen Hospital, China Academy of Chinese Medical Sciences, Beijing 102600, China; 2Beijing University of Chinese Medicine, Beijing 100029, China; 3Department of Orthopedics, Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing 100700, China; 4Department of Orthopedics, Beijing Traditional Chinese Medicine Hospital, Capital Medical University, Beijing 100010, China; 5Hunan University of Chinese Medicine, Hunan Province, China
  • Received:2020-06-06 Revised:2020-06-12 Accepted:2020-07-11 Online:2021-03-18 Published:2020-12-10
  • Contact: Chen Jiang, MD, Associate chief physician, Department of Orthopedics, Dongzhimen Hospital, Beijing University of Chinese Medicine, Beijing 100700, China; Hunan University of Chinese Medicine, Changsha 410208, Hunan Province, China
  • About author:Liu Zhichao, Master, Department of Orthopedics, Southern Branch of Guanganmen Hospital, China Academy of Chinese Medical Sciences, Beijing 102600, China; Beijing University of Chinese Medicine, Beijing 100029, China
  • Supported by:
    the National Natural Science Foundation of China, No. 81603638; General Program of China Postdoctoral Science Foundation, No. 2019M662791; Fundamental Research Funds for the Central Universities, No. 2018-JYB-XJQ010; 2017 Basic Scientific Research Business Expenses Project of Beijing University of Chinese Medicine, No. 2017-JYB-JS-085; 2019 Basic Scientific Research Business Expenses Project of Beijing University of Chinese Medicine, No. 2019-JYB-JS-042; Beijing Youth Talent Project of Dongzhimen Hospital of Beijing University of Chinese Medicine, No. DZMYS-201702 

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Abstract:

BACKGROUND: Compressive stress can change the morphology and activity of cells, but whether the morphology and activity of nucleus pulposus cells change under hydrostatic pressure still needs further study.

OBJECTIVE: To study the morphology and activity of human nucleus pulposus cells in vitro. 
METHODS: The human nucleus pulposus cells were separated, cultured and passed on for three generations, and pressurized for 2, 4 and 6 hours under the hydrostatic pressure of 0.3, 1, and 3 MPa. Then, the morphological changes and growth of the cells before and after pressurization were observed by inverted phase contrast microscope. Transmission electron microscope was used to observe the ultrastructural changes and differences of the cells. Cell counting kit-8 was used to detect the proliferation activity, morphology and activity of the cells under different hydrostatic pressures. 
RESULTS AND CONCLUSION: (1) Cell culture and passage: The growth curves of the first, second and third generations of human nucleus pulposus cells were S-shaped, and the cells proliferated fastest in a straight line from 3 to 7 days. The protuberances of the 5th and 6th generation cells were long shuttle shaped, grew slowly and degenerated. (2) Cell morphology: the human nucleus pulposus cells were shrunk under hydrostatic pressures of 0.3, 1, 3 MPa. At 0.3 and 1 MPa, the cells became slightly smaller and the morphology was basically complete; at 3 MPa, the cells were most obviously shrunk and the morphology was incomplete. The results showed that when the human nucleus pulposus cells were pressurized for 2, 4 and 6 hours under 0.3, 1 and 3 MPa hydrostatic pressures, the change of cell morphology was the most obvious under 3 MPa hydrostatic pressure, but there was no obvious change under the same hydrostatic pressure for different time. (3) Cell viability: Under 0.3 MPa hydrostatic pressure, the proliferation rate of human nucleus pulposus cells first increased and then decreased with the increase of time, and the cell proliferation rate reached the peak at 4 hours. Under 1 and 3MPa hydrostatic pressures, the proliferation rate of the cells gradually decreased with the increase of time, and the cell proliferation rate under 1 MPa hydrostatic pressure was significantly higher than that under 3 MPa hydrostatic pressure at the same action time (P < 0.05). These findings indicate that proper hydrostatic pressure stimulation helps to promote the proliferation of human nucleus pulposus cells, and long-term improperly high hydrostatic pressure stimulation can reduce the proliferation rate of human nucleus pulposus cells, leading to the occurrence of intervertebral disc degeneration.

Key words: hydrostatic pressure, intervertebral disc, nucleus pulposus cells, morphology, cell viability, growth kinetics

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