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Global Translational Medicine Advancements in cardiac regenerative therapy

signaling pathways, and epigenetic modifications that 2.2. iPSC differentiation into cardiac subtypes
ultimately lead to the formation of functional CMs with the Differentiating iPSCs into specific cardiac subtypes: atrial,
ability to contract, propagate action potentials (APs), and ventricular, Purkinje fibers, nodal cells, and endothelial
respond to physiological stimuli. Differentiation protocols cells involves manipulating distinct signaling pathways and
also enhance the expression of cardiac-specific markers environmental conditions, which leads to the development
and ion channels, improving their electrophysiological of cells with specialized structural and functional
properties. 15,16 This process also promotes the development properties. Below is an elaboration of how differentiation
of sarcomeres, contractile apparatus, and mitochondrial occurs for each subtype:
density, leading to enhanced contractility and calcium
handling. 17,18 In addition, differentiation promotes the For atrial CM (iPSC-aCMs), a key factor is the
formation of functional gap junctions, enabling proper addition of retinoic acid (RA), which is pivotal for
cell-cell coupling and synchronous beating. 19,20 Moreover, atrial differentiation. RA helps to guide progenitor
differentiated iPSC-CMs exhibit an improved response cells toward the atrial phenotype by influencing atrial-
to beta-adrenergic stimulation and increased sensitivity specific gene expression, such as atrial natriuretic
33
to pharmacological agents. 21,22 This process also reduces peptide. Compared to ventricular CMs (vCMs), aCM’s
the activation of pluripotency markers and tumorigenic exhibit functional differences, including a shorter AP
potential. 23,24 duration and faster beating rates. iPSC-aCMs also show
distinctive triangular-shaped APs, reflecting faster
The maintenance and differentiation of cardiac repolarization and higher beating frequencies, which
stem cells from in vitro iPSCs rely on the recreation are essential for their role in controlling the atrial
of the heart’s intrinsic signaling microenvironment, contraction rhythm. 34,35
achieving through the targeted use of growth factors
(GFs), neurotransmitters, epigenetic modulators, and In attaining vCMs, differentiation involves the
extracellular matrix proteins, which necessitates the inhibition of RA and the enhancement of Wnt signaling.
selective activation or suppression of distinct signaling The absence of RA guides cells away from an atrial fate and
25
cascades. Various differentiation protocols have been toward the ventricular lineage. The iPSC-vCMs display
developed, including modulation of Wnt/β-catenin longer APs with a prominent plateau phase, reflecting the
signaling, ACTIVIN/NODAL activation, BMP signaling sustained contraction necessary for ventricular pumping
activation, and the application of small molecules that as its functional differences. These cells also have a slower
trigger mesodermal specification. This is followed by the beating frequency and demonstrate different calcium
inhibition of Wnt signaling to form cardiac mesoderm and, transient kinetics, which are crucial for the forceful and
consequently, cardiac-specific progenitors, culminating sustained contractions of the ventricles. 36,37
in the expression of cardiac markers, including structure Nodal (Pacemaker) cells are induced by high levels
genes which encode sarcomeric proteins characteristic of of cyclic adenosine monophosphate (cAMP) and the
terminally differentiated CMs. 26-29 transcription factor T-box 3 (TBX3), which suppresses

Inadequate differentiation of iPSCs into CMs typical chamber-specific genes, thus enabling pacemaker
can result in immature or partially differentiated characteristics. Bone morphogenetic protein (BMP)
cells, leading to reduced contractility, abnormal pathway is also involved in their specification. Nodal cells
electrophysiology, increased susceptibility to disease exhibit automaticity, meaning they generate spontaneous
modeling, limited response to pharmacological agents, APs to regulate the heartbeat. This pacemaker activity is
reduced engraftment and survival, and tumorigenic a defining feature, where nodal cells exhibit slower AP
potential. These immature CMs may exhibit decreased upstrokes and lack the fast sodium current seen in working
contractile force and velocity (less effective for CMs. 38,39
contractility to pump blood), aberrant electrical activity Differentiation into Purkinje fibers is less commonly
(leading to arrhythmia), and decreased response to drugs studied in iPSC research. However, these cells can be
targeting mature CM functions. Furthermore, they may derived by modulating Notch signaling and transforming
be more prone to disease-related phenotypes, such as GF-beta (TGF-β) pathways, which influence the
hypertrophy or fibrosis, and have lower survival rates specialized conduction system. Purkinje fibers are
and engraftment efficiency when transplanted into the specialized for fast transmission of electrical signals in
heart. The use of iPSC-CMs with immature properties the heart. They have unique AP profiles with a quick rise
may also result in the development of teratomas or other velocity, essential for coordinating timely ventricular
kinds of tumor. 30-32 contraction. Their primary function is fast conduction

Volume 4 Issue 1 (2025) 3 doi: 10.36922/gtm.5745