Recombinant human growth hormone promotes osteogenic differentiation of human dental pulp stem cells

doi:10.12307/2024.103

Abstract

Abstract: BACKGROUND: Previous studies have shown that human dental pulp stem cells have good osteogenic differentiation potential and are potential seed cells in bone tissue engineering, and the effect of recombinant human growth hormone on the proliferative osteogenic differentiation of human dental pulp stem cells is still unclear.
OBJECTIVE: To explore the effect of recombinant human growth hormone on the proliferation and osteogenic differentiation of human dental pulp stem cells.
METHODS: Human dental pulp stem cells were isolated and cultured by tissue block culture method. After screening according to the drug concentration gradient, recombinant human growth hormone containing 10, 100, 250, 500, 1 000 μg/L was selected as the experimental group, and 0 μg/L without recombinant human growth hormone was selected as the control group. CCK-8 detection reagents were used on days 1, 3, 5, and 7 after the drug intervention to detect the proliferation of human dental pulp stem cells. Different concentrations (10, 100, 250, 500, and 1 000 μg/L) of recombinant human growth hormone were added to the osteogenesis induction solution to intervene in human dental pulp stem cells. Alkaline phosphatase activity was detected by alkaline phosphatase staining and semi-quantitative analysis on day 7 of mineralization induction. The mRNA expression levels of osteogenic gene type I collagen, osteocalcin and Runt-related transcription factor 2 were detected by fluorescence quantitative RT-qPCR. Alizarin red staining was performed on day 14 of mineralization induction to detect osteogenic mineralized nodules.
RESULTS AND CONCLUSION: (1) CCK-8 assay results showed that from the third day of intervention, the 100, 250, 500, 1 000 μg/L recombinant human growth hormone group could promote the proliferation of human dental pulp stem cells compared with the control group (P < 0.01). (2) The alkaline phosphatase activity of human dental pulp stem cells in the 100, 250, and 500 μg/L recombinant human growth hormone group was significantly increased compared with the control group (P < 0.01). The number of alizarin-stained mineralized nodules in human dental pulp stem cells in the 100, 250 μg/L recombinant human growth hormone group was significantly increased compared with the control group (P < 0.01). Compared with the control group, the mRNA expression of type I collagen and osteocalcin increased in the 250 μg/L recombinant human growth hormone group (P < 0.05, P < 0.01). mRNA expression of Runt-associated transcription factor 2 increased in the 100 and 250 μg/L recombinant human growth hormone groups (P < 0.01). (3) According to the above results, recombinant human growth hormone at a concentration of 250 μg/L is a more suitable concentration to promote the proliferation and osteogenic differentiation of human dental pulp stem cells.

Key words: mesenchymal stem cell, human dental pulp stem cell, growth hormone, recombinant human growth hormone, cell proliferation, osteogenesis, differentiation

CLC Number:

Sun Jing, Liao Jian, Sun Jiangling, Cheng Ping, Feng Hongchao. Recombinant human growth hormone promotes osteogenic differentiation of human dental pulp stem cells[J]. Chinese Journal of Tissue Engineering Research, 2024, 28(1): 56-61.

Figures/Tables 8

References

[1] AL MARUF DSA, GHOSH YA, XIN H, et al. Hydrogel: A Potential Material for Bone Tissue Engineering Repairing the Segmental Mandibular Defect. Polymers (Basel). 2022;14(19):4186.
[2] DEC P, MODRZEJEWSKI A, PAWLIK A. Existing and Novel Biomaterials for Bone Tissue Engineering. Int J Mol Sci. 2022;24(1):529.
[3] GRABER E, REITER EO, ROGOL AD. Human Growth and Growth Hormone: From Antiquity to the Recominant Age to the Future. Front Endocrinol (Lausanne). 2021;12:709936.
[4] BAMBA V, KANAKATTI SHANKAR R. Approach to the Patient: Safety of Growth Hormone Replacement in Children and Adolescents. J Clin Endocrinol Metab. 2022;107(3):847-861.
[5] 朱怀安,尹苗,陈婉丽,等.基因重组人生长激素对骨髓基质细胞增殖与成骨性分化的影响[J].临床口腔医学杂志,2018,34(3):131-134.
[6] WANG JR, AHMED SF, GADEGAARD N, et al. Nanotopology potentiates growth hormone signalling and osteogenesis of mesenchymal stem cells. Growth Horm IGF Res. 2014;24(6):245-250.
[7] TAIHI I, PILON C, COHEN J, et al. Efficient isolation of human gingival stem cells in a new serum-free medium supplemented with platelet lysate and growth hormone for osteogenic differentiation enhancement. Stem Cell Res Ther. 2022;13(1):125.
[8] MOHANRAM Y, ZHANG J, TSIRIDIS E, et al. Comparing bone tissue engineering efficacy of HDPSCs, HBMSCs on 3D biomimetic ABM-P-15 scaffolds in vitro and in vivo. Cytotechnology. 2020;72(5):715-730.
[9] ZHANG S, ZHANG X, LI Y, et al. Clinical Reference Strategy for the Selection of Treatment Materials for Maxillofacial Bone Transplantation: A Systematic Review and Network Meta-Analysis. Tissue Eng Regen Med. 2022;19(3): 437-450.
[10] JANEBODIN K, HORST OV, IERONIMAKIS N, et al. Isolation and characterization of neural crest-derived stem cells from dental pulp of neonatal mice. PLoS One. 2011;6(11):e27526.
[11] AL-MASWARY AA, O’REILLY M, HOLMES AP, et al. Exploring the neurogenic differentiation of human dental pulp stem cells. PLoS One. 2022;17(11): e0277134.
[12] HATORI A, FUJII Y, KAWASE-KOGA Y, et al. VCAM-1 and GFPT-2: Predictive markers of osteoblast differentiation in human dental pulp stem cells. Bone. 2023;166:116575.
[13] ZHANG Z, OH M, SASAKI JI, et al. Inverse and reciprocal regulation of p53/p21 and Bmi-1 modulates vasculogenic differentiation of dental pulp stem cells. Cell Death Dis. 2021;12(7):644.
[14] PERIĆ KAČAREVIĆ Ž, RIDER P, ALKILDANI S, et al. An introduction to bone tissue engineering. Int J Artif Organs. 2020;43(2):69-86.
[15] CAO S, HAN J, SHARMA N, et al. In Vitro Mechanical and Biological Properties of 3D Printed Polymer Composite and β-Tricalcium Phosphate Scaffold on Human Dental Pulp Stem Cells. Materials (Basel). 2020;13(14):3057.
[16] DI TINCO R, SERGI R, BERTANI G, et al. Effects of a Novel Bioactive Glass Composition on Biological Properties of Human Dental Pulp Stem Cells. Materials (Basel). 2020;13(18):4049.
[17] GU Y, ZHUANG R, XIE X, et al. Osteogenic stimulation of human dental pulp stem cells with self-setting biphasic calcium phosphate cement. J Biomed Mater Res B Appl Biomater. 2020;108(4):1669-1678.
[18] YAMADA Y, NAKAMURA-YAMADA S, KUSANO K, et al. Clinical Potential and Current Progress of Dental Pulp Stem Cells for Various Systemic Diseases in Regenerative Medicine: A Concise Review. Int J Mol Sci. 2019;20(5):1132.
[19] FAGEEH HN. Preliminary Evaluation of Proliferation, Wound Healing Properties, Osteogenic and Chondrogenic Potential of Dental Pulp Stem Cells Obtained from Healthy and Periodontitis Affected Teeth. Cells. 2021; 10(8):2118.
[20] KARAMZADEH R, ESLAMINEJAD MB, AFLATOONIAN R. Isolation, characterization and comparative differentiation of human dental pulp stem cells derived from permanent teeth by using two different methods. J Vis Exp. 2012;(69):4372.
[21] YU Z, GAUTHIER P, TRAN QT, et al. Differential Properties of Human ALP+ Periodontal Ligament Stem Cells vs Their ALP- Counterparts. J Stem Cell Res Ther. 2015;5(7):292.
[22] 杨俊辉,罗金莉,袁小平.人生长激素对人牙周膜干细胞增殖及成骨分化的影响[J].中国组织工程研究,2021,25(25):3956-3961.
[23] KOFFI KA, DOUBLIER S, RICORT JM, et al. The Role of GH/IGF Axis in Dento-Alveolar Complex from Development to Aging and Therapeutics: A Narrative Review. Cells. 2021;10(5):1181.
[24] LINDSEY RC, MOHAN S. Skeletal effects of growth hormone and insulin-like growth factor-I therapy. Mol Cell Endocrinol. 2016;432:44-55.
[25] CRIPPA GE, BELOTI MM, CARDOSO CR, et al. Effect of growth hormone on in vitro osteogenesis and gene expression of human osteoblastic cells is donor-age-dependent. J Cell Biochem. 2008;104(2):369-376.
[26] FENG J, MENG Z. Insulin growth factor-1 promotes the proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells through the Wnt/β-catenin pathway. Exp Ther Med. 2021;22(2):891.
[27] YUAN Y, DUAN R, WU B, et al. Gene expression profiles and bioinformatics analysis of insulin-like growth factor-1 promotion of osteogenic differentiation. Mol Genet Genomic Med. 2019;7(10):e00921.
[28] BIAN M, YU Y, LI Y, et al. Upregulating the Expression of LncRNA ANRIL Promotes Osteogenesis via the miR-7-5p/IGF-1R Axis in the Inflamed Periodontal Ligament Stem Cells. Front Cell Dev Biol. 2021;9:604400.
[29] 周思佳,姜文学,尤佳.骨缺损修复材料:现状与需求和未来[J].中国组织工程研究,2018,22(14):2251-2258.
[30] TONIETTO L, VASQUEZ AF, DOS SANTOS LA, et al. Histological and structural evaluation of growth hormone and PLGA incorporation in macroporous scaffold of α-tricalcium phosphate cement. J Biomater Appl. 2019;33(6): 866-875.