Strong Components of Epigenetic Memory in Cultured Human Fibroblasts Related to Site of Origin and Donor Age

Abstract

Differentiating pluripotent cells from fibroblast progenitors is a potentially transformative tool in personalized medicine. We previously identified relatively greater success culturing dura-derived fibroblasts than scalp-derived fibroblasts from postmortem tissue. We hypothesized that these differences in culture success were related to epigenetic differences between the cultured fibroblasts by sampling location, and therefore generated genome-wide DNA methylation and transcriptome data on 11 intrinsically matched pairs of dural and scalp fibroblasts from donors across the lifespan (infant to 85 years). While these cultured fibroblasts were several generations removed from the primary tissue and morphologically indistinguishable, we found widespread epigenetic differences by sampling location at the single CpG (N = 101,989), region (N = 697), “block” (N = 243), and global spatial scales suggesting a strong epigenetic memory of original fibroblast location. Furthermore, many of these epigenetic differences manifested in the transcriptome, particularly at the region-level. We further identified 7,265 CpGs and 11 regions showing significant epigenetic memory related to the age of the donor, as well as an overall increased epigenetic variability, preferentially in scalp-derived fibroblasts—83% of loci were more variable in scalp, hypothesized to result from cumulative exposure to environmental stimuli in the primary tissue. By integrating publicly available DNA methylation datasets on individual cell populations in blood and brain, we identified significantly increased inter-individual variability in our scalp- and other skin-derived fibroblasts on a similar scale as epigenetic differences between different lineages of blood cells. Lastly, these epigenetic differences did not appear to be driven by somatic mutation—while we identified 64 probable de-novo variants across the 11 subjects, there was no association between mutation burden and age of the donor (p = 0.71). These results depict a strong component of epigenetic memory in cell culture from primary tissue, even after several generations of daughter cells, related to cell state and donor age.

Origin and functional heterogeneity of fibroblasts

“…Distinct DNA methylation patterns are associated with various anatomical sources of human fibroblasts that impact transcriptional activity . Variability in the epigenetic landscape of dermal fibroblasts may be modulated by environmental stressors . The pathologic role of epigenetic modification of fibroblast genomes during fibrosis is evident in the kidney .…”

“…The functional heterogeneity and unique plasticity of fibroblast origin may operate through epigenetic memory. 105 Although a variety of epigenetic regulators exists, DNA methylation has been well studied in the context of fibroblasts biology. Distinct DNA methylation patterns are associated with various anatomical sources of human fibroblasts that impact transcriptional activity.105 Variability in the epigenetic landscape of dermal fibroblasts may be modulated by environmental stressors.…”

“…Distinct DNA methylation patterns are associated with various anatomical sources of human fibroblasts that impact transcriptional activity.105 Variability in the epigenetic landscape of dermal fibroblasts may be modulated by environmental stressors. 105 The pathologic role of epigenetic modification of fibroblast genomes during fibrosis is evident in the kidney.106-108 Hypermethylation of the Rasal1 gene promotor sustains the activated phenotype of fibroblasts, driving renal fibrogenesis in vivo. 106 Differential DNA methylation patterns are also reported in lung fibroblasts in the context of chronic obstructive pulmonary disease (COPD)-associated lung fibrosis109 and idiopathic pulmonary fibrosis,110 in synovial fibroblasts ( Table 2) in rheumatoid arthritis,111 and in dermal fibroblasts in systemic sclerosis.…”

Pluripotent stem cells in neuropsychiatric disorders

“…Although induced neuron technology may better preserve the epigenetic landscape of the patient donor cell, epigenetic modifications vary across different cell types and therefore changes in skin fibroblasts may not capture important neuron-specific disease-related epigenetic differences. 94 The emerging field of transgenerational epigenetic inheritance provides evidence that environmental influences, including nutrition or stress, can modify epigenetic marks inherited across the germline. 95 , 96 Thus, such marks may persist through the reprogramming process and be of use in the study of mental illness with environmental contributions.…”

The silver lining of induced pluripotent stem cell variation

“…It is surprising that this point was not addressed by Yoshida and colleagues while describing the cell lines used and the potential causes for variation between them. Another factor related to aberrant DMRs is the age of the donor, and it would be interesting to stratify the different iPSC lines by age, as other studies have done (14).…”