Carbohydrate sulfotransferase 3 (CHST3) overexpression promotes cartilage endplate‐derived stem cells (CESCs) to regulate molecular mechanisms related to repair of intervertebral disc degeneration by rat nucleus pulposus

Abstract

To investigate the regulatory effect of carbohydrate sulfotransferase 3 (CHST3) in cartilage endplate‐derived stem cells (CESCs) on the molecular mechanism of intervertebral disc degeneration after nucleus pulposus repair in rats. We performed GO and KEGG analysis of GSE15227 database to select the differential genes CHST3 and CSPG4 in grade Ⅱ, Ⅲ and Ⅳ intervertebral disc degeneration, IHC and WB to detect the protein profile of CHST3 and CSPG4, Co‐IP for the interaction between CHST3 and CSPG4. Then, immunofluorescence was applied to measure the level of CD90 and CD105, and flow cytometry indicated the level of CD73, CD90 and CD105 in CESCs. Next, Alizarin red staining, Alcian blue staining and TEM were performed to evaluate the effects of CESCs into osteoblasts and chondroblasts, respectively, CCK8 for the cell proliferation of osteoblasts and chondroblasts after induction for different times; cell cycle of osteoblasts or chondroblasts was measured by flow cytometry after induction, and WB for the measurement of specific biomarkers of OC and RUNX in osteoblasts and aggrecan, collagen II in chondroblasts. Finally, colony formation was applied to measure the cell proliferation of CESCs transfected with ov‐CHST3 or sh‐CHST3 when cocultured with bone marrow cells, WB for the protein expression of CHST3, CSPG4 and ELAVL1 in CSECs, transwell assay for the migration of CESCs to bone marrow cells, TEM image for the cellular characteristics of bone marrow cells, and WB for the protein profile of VCAN, VASP, NCAN and OFD1 in bone marrow cells. CHST3 and CSPG4 were differentially expressed and interacted in grade Ⅱ, Ⅲ and Ⅳ intervertebral disc degeneration; CD73, CD90 and CD105 were lowly expressed in CESCs, osteogenic or chondroblastic induction changed the characteristics, proliferation, cell cycle and specific biomarkers of osteoblasts and chondroblasts after 14 or 21 days,; CHST3 affected the cell proliferation, protein profile, migration and cellular features of cocultured CESCs or bone marrow cells. CHST3 overexpression promoted CESCs to regulate bone marrow cells through interaction with CSPG4 to repair the grade Ⅱ, Ⅲ and Ⅳ intervertebral disc degeneration.

Cartilage Endplate-Derived Stem Cells for Regeneration of Intervertebral Disc Degeneration: An Analytic Study

“…54,57,61,63,[68][69][70]76 Eight studies elaborated on the biological potential of CESCs in intervertebral disc regeneration. 55,67,[71][72][73][74][75]77…”

“…77 Similarly, Guan et al identified that overexpression of carbohydrate sulfotransferase 3 (CHST3) enhances the proliferation and migration capabilities of CESCs. 72 Insights gleaned from these studies offer a promising avenue for identifying potential molecular targets, which could inform the development of novel, more effective treatments for IDD. These advances in understanding the molecular dynamics of CESCs contribute significantly to the evolving landscape of IDD research, potentially leading to breakthroughs in regenerative medicine and therapeutics.…”

“…10,80 Moreover, several studies have reported the direct isolation of CESCs using standard cell culture techniques without specific interventions. [71][72][73][74][75] Despite these varied techniques, a consensus on the optimal method for CESCs isolation remains elusive, particularly in the absence of comparative studies. Therefore, further investigation is required to develop strategies for the simple and economical isolation of pure CESCs.…”

The role of microenvironment in stem cell-based regeneration of intervertebral disc

“…In addition to providing topographic support and mechanical stimulus for cells, IVD grafts could also be loaded with bioactive agents ( Guan et al, 2021 ; Chen and Jiang, 2022 ). In order to achieve antioxidant properties in NP, Cheng et al fabricated an injectable thermosensitive hydrogel scaffold incorporated with ferulic acid as a controlled release system.…”

Mechanism of the Mitogen-Activated Protein Kinases/Mammalian Target of Rapamycin Pathway in the Process of Cartilage Endplate Stem Cell Degeneration Induced by Tension Load

“…[5][6][7] Moreover, studies have shown that IVDD is not only caused by cell aging but also mainly because of the aging of adult stem cells that fail to timely replenish the aging and damaged cells in the tissues, suggesting that the CESCs in IVD may play an important role in IVDD. 8,9 Tension load (TL) contributes a lot to the initiation and progression of IVDD. 10,11 Therefore, the in-depth study of the specific mechanism of TL-induced CESCs degeneration is of great significance for the early prevention and treatment of IVDD.…”