Dental pulp stem cells therapy overcome photoreceptor cell death and protects the retina in a rat model of sodium iodate-induced retinal degeneration

https://doi.org/10.1016/j.jphotobiol.2019.111561Get rights and content

Highlights

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    Protection of ERG waveforms from sodium iodate insult in rat model by transplanted DPSCs.

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    Preservation of retinal morphology in rat model by transplanted DPSCs

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    Integration of transplanted DPSCs in the photoreceptor layer of treated rat model

Abstract

Blindness and vision loss contribute to irreversible retinal degeneration, and cellular therapy for retinal cell replacement has the potential to treat individuals who have lost light sensitive photoreceptors in the retina. Retinal cells are well characterized in function, and are a subject of interest in cellular replacement therapy of photoreceptors and the retinal pigment epithelium. However, retinal cell transplantation is limited by various factors, including the choice of potential stem cell source that can show variability in plasticity as well as host tissue integration. Dental pulp is one such source that contains an abundance of stem cells. In this study we used dental pulp-derived mesenchymal stem cells (DPSCs) to mitigate sodium iodate (NaIO3) insult in a rat model of retinal degeneration. Sprague-Dawley rats were first given an intravitreal injection of 3 × 105 DPSCs as well as a single systemic administration of NaIO3 (40 mg/kg). Electroretinography (ERG) was performed for the next two months and was followed-up by histological analysis. The ERG recordings showed protection of DPSC-treated retinas within 4 weeks, which was statistically significant (* P ≤ .05) compared to the control. Retinal thickness of the control was also found to be thinner (*** P ≤ .001). The DPSCs were found integrated in the photoreceptor layer through immunohistochemical staining. Our findings showed that DPSCs have the potential to moderate retinal degeneration. In conclusion, DPSCs are a potential source of stem cells in the field of eye stem cell therapy due to its protective effects against retinal degeneration.

Introduction

Retinal degenerative diseases are caused by many genetic and environmental factors, and can lead to incurable blindness even in developed countries. This complete impairment of vision tends to be the major outcome when left untreated due to the loss of photoreceptors [1]. The pathological pathways of these diseases differ on onset, but the ensuing degeneration share similar mechanisms [1]. The apoptosis that occurs in the degenerating retina can involve for example, the outer nuclear layer (ONL), inner nuclear layer (INL), and photoreceptor layer [2]. Some retinal cells are capable of evading apoptosis, and this may be utilized in visual restoration with treatments such as implants [3]. Other than implants, transplanting functional retinal cells may be key to restoring vision.

In animal models, replacing dysfunctional photoreceptors was shown to have an influence on retinal function and morphology. This replacement and protection approach in preclinical research are being done with various cell types, and is a more feasible, short-term aim due to the slow-progressing nature of most clinical conditions [4]. It is postulated that this protection and maintenance of retinal function is from the release of growth factors by donor cells. Apart from retinal neurons that make up most of the retina, these methods can also improve retinal pigment epithelium dystrophy [5]. Currently, age-related macular degeneration (AMD) and retinitis pigmentosa (RP) are the main research focus in the application of a suitable stem cell therapy [4,[6], [7], [8]].

Dental pulp stem cells (DPSCs) are a type of stem cell that have been recently hypothesized to have regenerative and protective properties in eye diseases model [9]. The use of these neural crest-derived cells in neural regenerative medicine has gained widespread popularity because of its relative ease of isolation from adult teeth, which makes it easily accessible from patients of all ages [10]. They also surmised that the neural crest properties of DPSCs make these cells more suitable than other mesenchymal stem cells, for example bone marrow derived mesenchymal stem cells in treating central nervous system (CNS) injury [9]. However, there are contradicting evidence regarding the differentiation of DPSCs into neurons in vitro that either supports it [11,12] or are against it [9,13].

Interestingly, it has been emphasized that DPSCs function in neural support by way of paracrine secretions [14], which was shown in a study on spinal cord injury treatment in rodent models to improve loco motor function [15]. The paracrine mechanism was found to involve the expression of neurotrophic factors (NTFs) such as nerve growth factor (NGF), brain derived-neurotrophin factor (BDNF), and neurotrophin-3 (NT-3). In our study, the main objective is to utilize DPSCs as a stem cell source to both protect and replace the degenerating retinal cells in a Sprague Dawley rat model of retinal degeneration.

Section snippets

Dental pulp stem cells culture and expansion conditions

Dental pulp stem cells were obtained from a stem cell bank, CryoCord (Cyberjaya, Selangor, Malaysia) at passage 2, and stored in liquid nitrogen until further use. In brief, the 1 ml ampoule of DPSCs was thawed in a water bath at 37 °C for 1 min (min) and transferred to a 15 ml centrifuge tube containing 9 ml of complete culture medium. The medium consisted of DMEM-F12 (Thermo Fisher Scientific, Massachusetts, USA) supplemented with 10% fetal bovine serum (Thermo Fisher Scientific,

Immunophenotyping of dental pulp stem cells

Dental pulp stem cells exhibit fibroblast-like morphology in standard culture conditions (Fig. 1). The DPSC immunophenotype consisted of highly expressed CD90 (99.3%), CD73 (99.7%), CD44 (99.3%), and a slightly lower expression of CD166 (60.9%). On the other hand, there were no expressions of endothelial and hematopoietic cell markers – CD19, HLA-DR, CD34, and CD45 (Fig. 2).

Electroretinographic and histological analysis of DPSC-treated retinas

Electroretinography was performed throughout the study. The scotopic response at 3.0 cd.s/m2 was detected and recorded for

Discussion

This study provides the first evidence showing human DPSC-mediated rescue of the retina in a rodent model of retinal degeneration. Stem cells are unspecialized cells that have the potential to differentiate into various cell lineages. DPSC, also known as postnatal DPSC, is a type of MSC that was first isolated by Gronthos et al. (2002) [16]. These cells are known to have high proliferative and clonogenicity levels, and are able to generate highly calcified colonies [16]. In standard culture

Conclusion

Stem cell therapy has a high potential in enhancing the life quality of patients experiencing ocular dysfunctions. In this study, mesenchymal stem cells taken from dental pulp (DPSCs) have been shown to possess therapeutic capabilities in protecting the retina from retinal degeneration. DPSCs were able to protect the visual functions and, to a certain extent, the retinal morphology of NaIO3-induced rats within two months. The cells were also found to have localized in the photoreceptor layer.

Author contributions

Suresh Kumar Subbiah and Pooi Ling Mok conceived the experimental study design; Hiba Amer Alsaeedi and Avin Ee-Hwan Koh equally contributed in carrying out the research study. Munirah binti Abd Rashid, Chenshen Lam, and Mohd Hairul Nizam Harun, aided in the electroretinographic study; Muhamad Fakhri bin Mohd Saleh aided in the histological study; Hiba Amer Alsaeedi, Avin Ee-Hwan Koh, and Seoh Wei Teh composed the manuscript and prepared the figures; Kong Yong Then, Suresh Kumar Subbiah,

Declaration of Competing Interest

The authors declare no competing or financial interests. The funding sponsors played no role in the study design; in the data collection and analysis; in the manuscript writing; and in the decision to publish the results.

Acknowledgements

This research was supported by the Research and Management Centre of Universiti Putra Malaysia through the Putra Graduate Initiative (IPS) grant (Grant No.: 9544300) and Geran Putra Berimpak (Grant No.: 9674600).

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