Isolation and Culture of Primary Human Gingival Epithelial Cells using Y-27632

Zhiwei Xie; Jizhou Shi; Min Zong; Qiuping Xu; Chang Liu; Jie Wen; Qun Zhang; Panpan Liu; Guanyi Liu; Jing Guo; Xunwei Wu

doi:10.3791/62978

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Method Article

Isolation and Culture of Primary Human Gingival Epithelial Cells using Y-27632

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DOI:

10.3791/62978

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November 6th, 2021

Zhiwei Xie¹^,²^,³ , Jizhou Shi⁴ , Min Zong³ , Qiuping Xu¹ , Chang Liu¹ , Jie Wen¹ , Qun Zhang¹ , Panpan Liu¹ , Guanyi Liu¹ , Jing Guo²^,⁵ , Xunwei Wu¹

¹Department of Tissue Engineering and Regeneration, School and Hospital of Stomatology, Cheeloo College of Medicine, Shandong University & Shandong Key Laboratory of Oral Tissue Regeneration & Shandong Engineering Laboratory for Dental Materials and Oral Tissue Regeneration, ²Department of Orthodontics, School and Hospital of Stomatology, Cheeloo College of Medicine, Shandong University & Shandong Key Laboratory of Oral Tissue Regeneration & Shandong Engineering Laboratory for Dental Materials and Oral Tissue Regeneration, ³Department of Stomatology, Shengli Oilfield Central Hospital, ⁴Department of Pediatric Surgery, Shengli Oilfield Central Hospital, ⁵Ningbo Stomatology Hospital of Savaid Stomatology School, Hangzhou Medical College

Summary

Here we present a modified method for the isolation and culture of human gingival epithelial cells by adding the Rock inhibitor, Y-27632, to the traditional method. This method is easier, less time-consuming, enhances stem cell properties, and produces larger numbers of high-potential epithelial cells both for the laboratory and for clinical applications.

Abstract

The gingival tissue is the first structure that protects periodontal tissues and plays meaningful roles in many oral functions. The gingival epithelium is an important structure of gingival tissue, especially in the repair and regeneration of periodontal tissue. Studying the functions of gingival epithelial cells has crucial scientific value, such as repairing oral defects and detecting the compatibility of biomaterials. As human gingival epithelial cells are highly differentiated keratinized cells, their lifespan is short, and they are difficult to passage. So far, there are only two ways to isolate and culture gingival epithelial cells, a direct explant method and an enzymatic method. However, the time required to obtain epithelial cells using the direct explant method is longer, and the cell survival rate of the enzymatic method is lower. Clinically, the acquisition of gingival tissue is limited, so a stable, efficient, and simple in vitro isolation and culture system is needed. We improved the traditional enzymatic method by adding Y-27632, a Rho-associated kinase (ROCK) inhibitor, which can selectively promote the growth of epithelial cells. Our modified enzymatic method simplifies the steps of the traditional enzymatic method and increases the efficiency of culturing epithelial cells, which has significant advantages over the direct explant method and the enzymatic method.

Introduction

The human gingiva, the first line defense structure that protects periodontal tissue, is not only a physical and chemical barrier¹, but also secretes different classes of inflammatory mediators that participate in immune responses and constitute an immune barrier²^,³. The gingival epithelium plays an important role in the repair and regeneration of periodontal tissue. Therefore, studying the defense and immunity of the gingival epithelium is of great significance for understanding the occurrence, diagnosis, and treatment of periodontitis. The isolation and culture of gingival epithelial ....