Mechanism of Long Noncoding RNA HOTAIR in Nucleus Pulposus Cell Autophagy and Apoptosis in Intervertebral Disc Degeneration.

Sheng Song,Wei Cui,Xueguang Liu,Sakineh Kazemi Noureini,Zhenzhong Sun,Shujun Zhang

doi:10.1155/2022/8504601

Abstract

Objective Intervertebral disc degeneration (IDD) contributes to cervical and lumbar diseases. Long noncoding RNAs (lncRNAs) are implicated in IDD. This study explored the mechanism of lncRNA HOTAIR in IDD. Methods Normal and degenerative nucleus pulposus (NP) cells were isolated from NP tissues obtained in intervertebral disc surgery. Cell morphology was observed by immunocytochemistry staining and toluidine blue staining. NP cell markers were detected by RT-qPCR. Proliferation was detected by MTT assay. Autophagy-related proteins were detected by Western blot. Autophagosome was observed by monodansylcadaverine fluorescence staining. Apoptosis was detected by TUNEL staining and flow cytometry. si-HOTAIR and/or miR-148a inhibitor was introduced into degenerative NP cells. Binding relationships among HOTAIR, miR-148a, and PTEN were predicted and verified by dual-luciferase reporter assay and RNA pull-down. Finally, IDD rat models were established. Rat caudal intervertebral discs were assessed by HE staining. Expressions of HOTAIR, miR-148a, and PTEN were determined by RT-qPCR. Results HOTAIR was highly expressed in degenerative NP cells (p < 0.05). si-HOTAIR inhibited degenerative NP cell apoptosis and autophagy (p < 0.05). HOTAIR upregulated PTEN as a sponge of miR-148a. miR-148a was poorly expressed in degenerative NP cells. miR-148a deficiency partially reversed the inhibition of si-HOTAIR on degenerative NP cell autophagy and apoptosis (all p < 0.05). In vivo assay confirmed that si-HOTAIR impeded autophagy and apoptosis in intervertebral disc tissues, thus improving pathological injury in IDD rats (all p < 0.05). Conclusion LncRNA HOTAIR promoted NP cell autophagy and apoptosis via promoting PTEN expression as a ceRNA of miR-148a in IDD.

Highlights

Intervertebral discs (IVDs) are composed of nucleus pulposus (NP), annulus fibrosus (AF), and cartilage endplates, which experience a gradual degeneration under the influence of a variety of factors, such as aging and tissue damage caused by mechanical stress [1, 2]
To study the role of Long noncoding RNAs (lncRNAs) HOX transcript antisense intergenic RNA (HOTAIR) in NP cells, we first isolated and cultured normal and degenerative NP cells from NP tissue samples obtained from clinical disc surgery
Similar results were found in toluidine blue staining, which showed blue color in normal NP cell cytoplasm and light blue color in degenerative NP cell cytoplasm (Figure 1(b)), together with reduced aggrecan generation compared with normal NP cells

Summary

Introduction

Intervertebral discs (IVDs) are composed of nucleus pulposus (NP), annulus fibrosus (AF), and cartilage endplates, which experience a gradual degeneration under the influence of a variety of factors, such as aging and tissue damage caused by mechanical stress [1, 2]. Long noncoding RNAs (lncRNAs) are characterized by limited protein-coding ability and are involved in multiple biological processes, such as transcription, protein activity, and aging-related degenerative musculoskeletal diseases, including IDD [7]. Growing studies have proposed that lncRNAs interact with microRNAs (miRNAs), thereby exerting effects on cell autophagy and apoptosis via a lncRNA-miRNA-mRNA competing endogenous RNA (ceRNA) network [12, 13]. LncRNA FAM83H-AS1 can promote the growth of NP cells and maintain the tissue homeostasis of IVDs by inhibiting miR-22-3p [14]. LncRNA H19 can promote autophagy and apoptosis of NP cells, aggravating IDD via the miR-139/CXCR4/NF-κB axis [15]. LncRNA CDKN2B-AS1 can inhibit ox-LDL-induced proliferation of vascular smooth muscle cells and promote apoptosis through the ceRNA network of CDKN2B-AS1/miR-126-5p/ PTPN7 [16]

Methods

Results

Conclusion