Nijmegen Breakage Syndrome fibroblasts and iPSCs: cellular models for uncovering disease-associated signaling pathways and establishing a screening platform for anti-oxidants

Barbara Mlody,James Adjaye,Soraia Martins,Wasco Wruck,Karl Sperling

doi:10.1038/s41598-017-07905-2

Abstract

Nijmegen Breakage Syndrome (NBS) is associated with cancer predisposition, premature aging, immune deficiency, microcephaly and is caused by mutations in the gene coding for NIBRIN (NBN) which is involved in DNA damage repair. Dermal-derived fibroblasts from NBS patients were reprogrammed into induced pluripotent stem cells (iPSCs) in order to bypass premature senescence. The influence of antioxidants on intracellular levels of ROS and DNA damage were screened and it was found that EDHB-an activator of the hypoxia pathway, decreased DNA damage in the presence of high oxidative stress. Furthermore, NBS fibroblasts but not NBS-iPSCs were found to be more susceptible to the induction of DNA damage than their healthy counterparts. Global transcriptome analysis comparing NBS to healthy fibroblasts and NBS-iPSCs to embryonic stem cells revealed regulation of P53 in NBS fibroblasts and NBS-iPSCs. Cell cycle related genes were down-regulated in NBS fibroblasts. Furthermore, oxidative phosphorylation was down-regulated and glycolysis up-regulated specifically in NBS-iPSCs compared to embryonic stem cells. Our study demonstrates the utility of NBS-iPSCs as a screening platform for anti-oxidants capable of suppressing DNA damage and a cellular model for studying NBN de-regulation in cancer and microcephaly.

Highlights

Nijmegen Breakage Syndrome (NBS) is a rare autosomal recessive genetic disorder, first described 1981 in Nijmegen, the Netherlands[1]
In line with the known predisposition of NBS patients to cancer, we found Pathways in cancer as the most enriched pathway in NBS-fibroblasts compared to healthy fibroblasts as well as in NBS-induced pluripotent stem cells (iPSCs) compared to embryonic stem cells via DAVID analysis[34] (Fig. 3a,b)
The DNA damage sensing NBN is an adapter protein which can bind to a variety of other DNA signaling and repair proteins ATM, which is a kinase that amplifies and transduces the DNA damage signal[53]

Summary

Introduction

Nijmegen Breakage Syndrome (NBS) is a rare autosomal recessive genetic disorder, first described 1981 in Nijmegen, the Netherlands[1]. Characteristics of NBS include genomic instability (resulting in early onset of malignancies), premature aging, microcephaly and other growth retardations, immune deficiency, impaired puberty and infertility in females. The consequence of these manifestations is a severe decrease in average life span, caused by cancer or infection of the respiratory and urinary tracts[2]. Several cases of NBS with a variety of mutations in NBN exist but over 90% of the patients carry a 5 base pair deletion (657del5) within the NBN exon 66 This hypomorphic mutation leads to a truncated 26 kD amino-terminal protein and a 70 kD carboxy-terminal protein due to alternative translation from a cryptic start site upstream of the deletion[7]. Cells counteract ROS by antioxidant production and enzymatic removal but ROS have cellular signaling functions which must be maintained in a controlled balance[11].One strategy to minimize endogenous ROS levels is to regulate mitochondrial respiration, which plays a special role in stem cells

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