Abstract
The proper functioning of many organs and tissues containing smooth muscles greatly depends on the intricate organization of the smooth muscle cells oriented in appropriate directions. Consequently controlling the cellular orientation in three-dimensional (3D) cellular constructs is an important issue in engineering tissues of smooth muscles. However, the ability to precisely control the cellular orientation at the microscale cannot be achieved by various commonly used 3D tissue engineering building blocks such as spheroids. This paper presents the formation of coiled spring-shaped 3D cellular constructs containing circumferentially oriented smooth muscle-like cells differentiated from dedifferentiated fat (DFAT) cells. By using the cell fiber technology, DFAT cells suspended in a mixture of extracellular proteins possessing an optimized stiffness were encapsulated in the core region of alginate shell microfibers and uniformly aligned to the longitudinal direction. Upon differentiation induction to the smooth muscle lineage, DFAT cell fibers self-assembled to coiled spring structures where the cells became circumferentially oriented. By changing the initial core-shell microfiber diameter, we demonstrated that the spring pitch and diameter could be controlled. 21 days after differentiation induction, the cell fibers contained high percentages of ASMA-positive and calponin-positive cells. Our technology to create these smooth muscle-like spring constructs enabled precise control of cellular alignment and orientation in 3D. These constructs can further serve as tissue engineering building blocks for larger organs and cellular implants used in clinical treatments.
Highlights
Smooth muscle cells are indispensable parts of many organs of the gastrointestinal, cardiovascular, urinary, and reproductive systems [1,2]
dedifferentiated fat (DFAT) cell fibers were formed by the cell fiber technology using the hydrogel core-shell microfiber fabrication device (Fig. 2A)
This microfluidic device generated two co-axial laminar flow streams consisting of a core stream of DFAT cells suspended in extracellular matrix (ECM) protein and a shell stream of sodium alginate that solidified upon contacting the sheath stream of CaCl2
Summary
Smooth muscle cells are indispensable parts of many organs of the gastrointestinal, cardiovascular, urinary, and reproductive systems [1,2]. After the cells aligned to form into fiber structures, they are subsequently induced to differentiate into the smooth muscle cell lineage During this differentiation process as cells’ inherent traction forces [15,16] increase, DFAT cell fibers self-assemble into smooth muscle cell-like uniformly coiled spring structures with circumferentially oriented cells that serve as useful models for circumferential smooth muscles. This simple method takes advantage of cells’ natural tendency to align and self-assemble into micro-scale 3D spring constructs without laborious pre-patterning of proteins or subsequent assembling manipulations that may bring undesired chemical and mechanical stresses to the cells. We investigate the requirements for extracellular matrix (ECM) proteins to maintain fiber structure, characterize various parameters of the spring structures, and demonstrate expression of smooth muscle specific markers in the fibers
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