Functional skeletal muscle constructs from transdifferentiated human fibroblasts

Area of Science:

• Biotechnology
• Regenerative Medicine
• Cell Biology

Background:

• Transdifferentiation of human non-muscle cells into myogenic cells via MyoD is a promising strategy for generating human myogenic cells.
• The functional properties of engineered tissue constructs derived from these transdifferentiated cells remain largely uncharacterized.

Purpose of the Study:

• To investigate the functional properties of three-dimensional (3D) tissue constructs derived from transdifferentiated human dermal fibroblasts.
• To evaluate the impact of temporally controlled MyoD expression and specific signaling pathway modulators on myogenic differentiation and contractile force generation.

Main Methods:

• Engineered normal human dermal fibroblasts (NHDFs) with genes for human telomerase reverse transcriptase (hTERT) and doxycycline-inducible MyoD (iMyoD-hTERT-NHDFs).
• Constructed 3D tissue constructs using these engineered cells.
• Applied electrical stimuli to assess contractile forces upon MyoD induction.
• Manipulated doxycycline withdrawal timing and treated constructs with Wnt activator (CHIR99021) and Notch inhibitor (DAPT).

Main Results:

• Engineered 3D constructs generated detectable contractile forces upon MyoD expression via electrical stimulation.
• Withdrawing doxycycline during 3D culture significantly increased both twitch (3.05-fold) and tetanic (2.28-fold) forces.
• Treatment with CHIR99021 and DAPT further enhanced the contractile forces of the engineered muscle constructs.

Conclusions:

• Temporally controlled MyoD expression optimizes functional myogenic differentiation in transdifferentiated cells.
• Pharmacological modulation of Wnt and Notch signaling pathways can further improve the functional capacity of engineered skeletal muscle.
• These findings highlight the potential of patient-specific transdifferentiated cells for creating functional skeletal muscle constructs for applications like disease modeling.