Journal Club: In vitro culturing of rat bone marrow stem cells to make hair-cell like cells.

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Today's journal article

Peyvandi AA, Davoudi S, Bazgir N, Janahmadi M, Norioun H, Khoshsirat S, Niknazar S. A time course analysis of the electrophysiological and gene expression properties during differentiation of hair cell-like cells in culture. 

Why I picked this article

Auditory hair cells are lost in many forms of sensorineural hearing loss. Currently, it is not possible to restore lost hair cells in humans. Regenerative therapy, where cochlear hair cells can be regenerated or transplanted to restore the cochlear function, remains one of the central goals in hearing research. 

To achieve such a therapy, scientists will need to be able to find a way to either create hair cells in a petri dish and transplant them into the cochlea or find some way for cells to become new hair cells in the cochlea. For the former approach, one of the potential cells to make hair cells from is mesenchymal stem cells (MSCs). MSCs can be cultured to become different cell types, including neurons and cells that have proteins like myosin VIIa found in hair cells. 

This research tracks, over time, how bone marrow–derived mesenchymal stromal cells (BMSCs) acquire hair cell-like features in a dish. 

Some of the research findings

Starting cells:
  • Male Wistar rats, 4-6 week-old.
  • BMSCs were collected from the femurs and tibias' bone marrow.
  • Cultured cells were triangular in shape with spindles. 
Culturing to produce hair-cell-like cells: 
  • Induction medium included B27, EGF, FGF, and IGF-1. 
  • Cells were sampled at day 17 and days 21–26 for molecular and electrophysiological readouts.
  • Immunocytochemistry, RNA sequencing and electrophysiological recordings were made. 
Changes in protein and RNA profiles of cells over time: 
  • Immunostaining: myosin VIIA (MYO7A) and SOX2 were observed in some cells in culture at days 21–26. 
  • This may indicate an emergence of a hair-cell like cells in late culture. 
  • Sox2-positive cells and myosin IIA-positive cells seem to be different, as expected. 
  • RNA sequencing data showed that eight hair cell/neuronal-related transcripts were elevated during differentiation (Wnt7a, Mgat5b, Myo7a, Pou4f3, Sox2, Atoh1, Map2k3, and Actin).
  • Ontology analysis from RNA sequencing pointed towards increased hair cell genes and genes related to synapses and ion channels. 
  • Electrophysiology (whole-cell patch clamp) of the hair-cell-like cells showed hyperpolarised membrane potential near −58.96 ± 1.10 mV at days 21-26. 
    • the average membrane resistance was 171.66 ± 29.12 MΩ
    • time constant 10.73 ± 0.45 ms
    • capacitance 0.0625 ± 0.0087 pF
    • Researchers interpret that these parameters are consistent with compact, high-resistance cells typical of differentiating sensory-like phenotypes.

Haruna's takeaway

Creating a particular desired cell type in culture is very interesting work. It also makes us realise how much we don't know about that particular cell type; where is the threshold of "critical features" that hair cell-like cells must have, before they can be used for regenerative therapy, or have some functional restoration in the inner ear? I wouldn't think it needs to be "perfect" hair cells. 

Some research in the retina showed that the most successful transplantation is when cells are "precursor" cells, or a developing but immature version of the cell. If it's the same in the cochlea, the good timing to transplant cells would be when stem cells started becoming hair-cell-like, but still in an immature form, so that they can finish development once inside the cochlear environment. 

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This is Haruna's 95/100 of the 100-day challenge to post a science blog article every day! I love inner ear biology & cochlear physiology.