June 2023
Volume 64, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2023
Pharmacologic reprogramming human adipose derived stem cells (ADSCs) into retinal ganglion cells
Author Affiliations & Notes
  • Jingna He
    Department of Ophthalmology and Visual Sciences, The Chinese University of Hong Kong, Kowloon, Hong Kong, China
  • WAI KIT CHU
    Department of Ophthalmology and Visual Sciences, The Chinese University of Hong Kong, Kowloon, Hong Kong, China
  • Kam-lung, Kelvin Chong
    Department of Ophthalmology and Visual Sciences, The Chinese University of Hong Kong, Kowloon, Hong Kong, China
  • Poemen Pui Man Chan
    Department of Ophthalmology and Visual Sciences, The Chinese University of Hong Kong, Kowloon, Hong Kong, China
  • Calvin C P Pang
    Department of Ophthalmology and Visual Sciences, The Chinese University of Hong Kong, Kowloon, Hong Kong, China
  • Clement C. Tham
    Department of Ophthalmology and Visual Sciences, The Chinese University of Hong Kong, Kowloon, Hong Kong, China
  • Footnotes
    Commercial Relationships   Jingna He None; WAI KIT CHU None; Kam-lung, Kelvin Chong None; Poemen Pui Man Chan None; Calvin Pang None; Clement Tham None
  • Footnotes
    Support  United College Early Career Research Excellence Award (2019 to W.K.C); The Chinese University of Hong Kong Direct Grant (2020.067 and 2021.046 to W.K.C).
Investigative Ophthalmology & Visual Science June 2023, Vol.64, 1559. doi:
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      Jingna He, WAI KIT CHU, Kam-lung, Kelvin Chong, Poemen Pui Man Chan, Calvin C P Pang, Clement C. Tham; Pharmacologic reprogramming human adipose derived stem cells (ADSCs) into retinal ganglion cells. Invest. Ophthalmol. Vis. Sci. 2023;64(8):1559.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Loss of retinal ganglion cells (RGCs) is an important hallmark feature in glaucoma. Stem cell therapy offers an opportunity to regenerate RGCs. In this study, human adipose derived stem cells (ADSCs) were developed as a new cell source for reprogramming into RGCs. ADSCs are multipotent cells that can be easily extracted from adipose tissue, are capable of expansion in vitro, and have the capacity to differentiate into multiple cell lineages, which have great potentials for use in regenerative medicine. Instead of using transgenic reprogramming factors, we aim to differentiate ADSCs into RGC-like cells with small molecules which can be easily mass produced and are more suitable for clinical therapy.

Methods : ADSCs were purified from periorbital (eyelid, orbital) and subcutaneous (abdominal) adipose tissues by digestion with 0.075% type 4 collagenase in DMEM-F12 medium for 2 hours. ADSCs were treated with a set of small molecules (valproic acid, GSK3 inhibitor CHIR99021, TGFβ receptor inhibitor RepSox, Forskolin, Wnt/β-catenin pathway inhibitor IWR-1 and Notch signalling inhibitor DAPT). On day 6 after treatment, RNA was extracted for real-time PCR. Three RGC markers were detected, TUBB3, RBPMS and THY1. Mean ± standard deviation of three independent experiments was analyzed by using t-test. P<0.05 was considered statistically significant.

Results : Real-time PCR showed the mRNA levels of TUBB3, RBPMS and THY1 in eyelid and abdominal ADSCs were significantly increased after small molecules treatment (Fig 1). The fold change of TUBB3 mRNA levels was induced 8.33 ± 2.10 and 5.39 ± 2.07 times in eyelid and abdominal ADSCs respectively (P<0.05, Fig 1A). The mRNA levels of RBPMS and THY1 were also induced 2.71 ± 0.36 and 3.05 ± 1.92 fold in eyelid ADSCs, 3.41 ± 1.25 and 3.47 ± 1.09 fold in abdominal ADSCs (P<0.05, Fig 1B and 1C). In orbital ADSCs, the induction of TUBB3 mRNA level was insignificant, while RPBMS and THY1 mRNA levels significantly increased after treatment (P<0.05, Fig 1). The mRNA level of THY1 and RPBMS in orbital ADSCs was 5.00 ± 2.24 and 3.10 ± 1.74 times higher after treatment (P<0.05, Fig 1B and 1C).

Conclusions : We have established a protocol to differentiate ADSCs into RGC-like cells by using small molecules.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

 

The mRNA level of TUBB3 (A), RBPMS (B) and THY1 (C) in ADSCs before and after treatment. * indicates P value < 0.05.

The mRNA level of TUBB3 (A), RBPMS (B) and THY1 (C) in ADSCs before and after treatment. * indicates P value < 0.05.

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