A patient-derived cellular model for Huntington’s disease reveals phenotypes at clinically relevant CAG lengths

Abstract

The huntingtin protein participates in several cellular processes that are disrupted when the polyglutamine tract is expanded beyond a threshold of 37 CAG DNA repeats in Huntington’s disease (HD). Cellular biology approaches to understand these functional disruptions in HD have primarily focused on cell lines with synthetically long CAG length alleles that clinically represent outliers in this disease and a more severe form of HD that lacks age onset. Patient-derived fibroblasts are limited to a finite number of passages before succumbing to cellular senescence. We used human telomerase reverse transcriptase (hTERT) to immortalize fibroblasts taken from individuals of varying age, sex, disease onset, and CAG repeat length, which we have termed TruHD cells. TruHD cells display classic HD phenotypes of altered morphology, size and growth rate, increased sensitivity to oxidative stress, aberrant adenosine diphosphate/adenosine triphosphate (ADP/ATP) ratios, and hypophosphorylated huntingtin protein. We additionally observed dysregulated reactive oxygen species (ROS)-dependent huntingtin localization to nuclear speckles in HD cells. We report the generation and characterization of a human, clinically relevant cellular model for investigating disease mechanisms in HD at the single-cell level, which, unlike transformed cell lines, maintains functions critical for huntingtin transcriptional regulation and genomic integrity.

Image-based profiling for drug discovery: due for a machine-learning upgrade?

“…Nevertheless, many reproducible image-based phenotypes have been discovered, often inadvertently, as scientists stained and visually examined cells, typically using common markers such as organelle dyes. For example, unusual mitochondrial structure was identified in fibroblasts and lymphocytes from patients with bipolar disorder 52 and in fibroblasts from patients with Leigh syndrome 53 , and normal human fibroblasts can be differentiated from Huntington disease fibroblasts using only tubulin staining 54 . Image-based profiling offers a way to scale-up and systematize this kind of serendipitous discovery.…”

Juvenile Huntington’s Disease Skin Fibroblasts Respond with Elevated Parkin Level and Increased Proteasome Activity as a Potential Mechanism to Counterbalance the Pathological Consequences of Mutant Huntingtin Protein

“…However, these models mainly include studies on post-mortem tissues, in vitro neuronal cells, non-neuron relevant cellular models, and non-human, particularly mouse and rat primary cells. HD is a neurodegenerative disease; however, alteration of cellular homeostasis is observed in peripheral tissues as well [6,7,8,9,10,63,64]. Thus, studying peripheral tissues in HD might add additional details for the development of biomarkers of the disease or model system for drug testing.…”

“…However, the huntingtin protein is also expressed in peripheral tissues [5,6]. Skin primary fibroblasts of adult onset HD patients are an attractive model for studying the disease due to the expanded polyglutamine stretch in the huntingtin protein in these fibroblasts [7]. Several studies describe the alteration of mitochondrial bioenergetics, increased oxidative stress, and changes in gene expression profile in skin fibroblasts derived from adult HD patients [8,9,10,11].…”

“…Therefore, we used skin primary fibroblasts derived from juvenile HD patients. Skin fibroblasts and cells directly converted from fibroblasts to neurons are currently being used to study HD biology [7,39]. We analyzed two juvenile primary HD fibroblast cell lines.…”

High-mobility group box 1 links sensing of reactive oxygen species by huntingtin to its nuclear entry

“…Tissue culture T-antigen immortalized, WT mouse striatal STHdh Q7/Q7 cells (kind gift from M. E. MacDonald, MGH) were cultured in Dulbecco's modified Eagle's medium with 10% fetal bovine serum (FBS) at 33°C with 5% CO 2 . hTERT-immortalized WT TruHD-Q21Q18 and HD mutant TruHD-Q43Q17 patient-derived fibroblast cells (36) were cultured in minimal essential medium with 15% FBS and 1% GlutaMax at 37°C with 5% CO 2 . All media and supplements were from Life Technologies unless stated otherwise.…”