SLIT2 Overexpression in Periodontitis Intensifies Inflammation and Alveolar Bone Loss, Possibly via the Activation of MAPK Pathway

Liping Wang; Jing Zheng; Janak L Pathak; Yunxin Chen; Dongliang Liang; Luxi Yang; Haobo Sun; Mei Zhong; Lihong Wu; Li Li; Shuhua Deng; Lingyun Zheng; Yongyong Yan; Dan Hou; Lijing Wang; Linhu Ge

doi:10.3389/fcell.2020.00593

SLIT2 Overexpression in Periodontitis Intensifies Inflammation and Alveolar Bone Loss, Possibly via the Activation of MAPK Pathway

Front Cell Dev Biol. 2020 Jul 14:8:593. doi: 10.3389/fcell.2020.00593. eCollection 2020.

Authors

Liping Wang¹, Jing Zheng¹, Janak L Pathak¹, Yunxin Chen¹, Dongliang Liang¹, Luxi Yang¹, Haobo Sun¹, Mei Zhong¹, Lihong Wu¹, Li Li², Shuhua Deng², Lingyun Zheng², Yongyong Yan¹, Dan Hou¹, Lijing Wang^{1

2}, Linhu Ge¹

Affiliations

¹ Guangzhou Key Laboratory of Basic and Applied Research in Oral Regenerative Medicine, Affiliated Stomatology Hospital of Guangzhou Medical University, Institute of Oral Disease, Guangzhou Medical University, Guangzhou, China.
² Vascular Biology Research Institute, School of Life Sciences and Biopharmaceutics, Guangdong Pharmaceutical University, Guangzhou, China.

Abstract

SLIT2, a member of neuronal guidance cues, has been reported to regulate inflammation and cancer progression. Periodontitis is an oral inflammatory disease that degenerates periodontal tissue, alveolar bone and tooth. This study aims to explore the expression pattern of SLIT2 in periodontitis and its role in disease progression and bone loss. Gingival tissue of 20 periodontitis patients and 20 healthy-controls was obtained. Ligature-induced periodontitis (LIP) mice-model was developed in Slit2-Tg and wild-type mice. The effect of SLIT2 on inflammation, immune cell infiltration, M1 macrophage polarization, and alveolar bone loss in periodontitis was analyzed extensively. In periodontitis-affected gingival-tissue, SLIT2 expression was 4.4-fold higher compared to healthy-volunteers. LIP enhanced SLIT2 expression in mice periodontitis-affected periodontal tissue (PAPT) and blood circulation of wild-type mice by 4. 6-, and 5.0-fold, respectively. In Slit2-Tg-mice PAPT, SLIT2 expression was 1.8-fold higher compared to wild-type mice. Micro-CT and histomorphometric analysis revealed a 1.3-fold higher cement-enamel-junction to the alveolar-bone-crest (CEJ-ABC) distance and alveolar bone loss in LIP Slit2-Tg-mice compare to LIP wild-type mice. Results from RNA-sequencing, RT-qPCR, and ELISA showed a higher expression of Cxcr2, Il-18, TNFα, IL-6, and IL-1β in Slit2-Tg-mice PAPT compared to wild-type-mice. Slit2-Tg-mice PAPT showed a higher number of osteoclasts, M1 macrophages, and the upregulation of Robo1 expression. Slit2-Tg-mice PAPT showed upregulation of M1 macrophage marker CD16/32 and osteoclastogenic markers Acp5, Ctsk, and Nfatc1, but osteogenic markers (Alp, Bglap) remained unchanged. Immunohistochemistry unveiled the higher vasculature and infiltration of leucocytes and macrophages in Slit2-Tg-mice PAPT. RNA-sequencing, GO-pathway enrichment analysis, and western blot analysis revealed the activation of the MAPK signaling pathway in Slit2-Tg mice PAPT. In conclusion, SLIT2 overexpression in periodontitis intensifies inflammation, immune cells infiltration, M1 macrophage polarization, osteoclastogenesis, and alveolar bone loss, possibly via activation of MAPK signaling, suggesting the role of SLIT2 on exacerbation of periodontitis and alveolar bone loss.

Keywords: MAPK signaling; SLIT2; alveolar bone loss; osteoclasts; periodontitis.