RESUMEN

Aligned collagen I (COL1) fibers guide tumor cell motility, influence endothelial cell morphology, control stem cell differentiation, and are a hallmark of cardiac and musculoskeletal tissues. To study cell response to aligned microenvironments in vitro, several protocols have been developed to generate COL1 matrices with defined fiber alignment, including magnetic, mechanical, cell-based, and microfluidic methods. Of these, microfluidic approaches offer advanced capabilities such as accurate control over fluid flows and the cellular microenvironment. However, the microfluidic approaches to generate aligned COL1 matrices for advanced in vitro culture platforms have been limited to thin "mats" (<40 µm in thickness) of COL1 fibers that extend over distances less than 500 µm and are not conducive to 3D cell culture applications. Here, we present a protocol to fabricate 3D COL1 matrices (130-250 µm in thickness) with millimeter-scale regions of defined fiber alignment in a microfluidic device. This platform provides advanced cell culture capabilities to model structured tissue microenvironments by providing direct access to the micro-engineered matrix for cell culture.

Asunto(s)

Colágeno , Hidrogeles , Técnicas de Cultivo de Célula/métodos , Microambiente Celular , Colágeno Tipo I

RESUMEN

Randomly oriented type I collagen (COL1) fibers in the extracellular matrix are reorganized by biophysical forces into aligned domains extending several millimeters and with varying degrees of fiber alignment. These aligned fibers can transmit traction forces, guide tumor cell migration, facilitate angiogenesis, and influence tissue morphogenesis. To create aligned COL1 domains in microfluidic cell culture models, shear flows have been used to align thin COL1 matrices (<50µm in height) in a microchannel. However, there has been limited investigation into the role of shear flows in aligning 3D hydrogels (>130µm). Here, we show that pure shear flows do not induce fiber alignment in 3D atelo COL1 hydrogels, but the simple addition of local extensional flow promotes alignment that is maintained across several millimeters, with a degree of alignment directly related to the extensional strain rate. We further advance experimental capabilities by addressing the practical challenge of accessing a 3D hydrogel formed within a microchannel by introducing a magnetically coupled modular platform that can be released to expose the microengineered hydrogel. We demonstrate the platform's capability to pattern cells and fabricate multi-layered COL1 matrices using layer-by-layer fabrication and specialized modules. Our approach provides an easy-to-use fabrication method to achieve advanced hydrogel microengineering capabilities that combine fiber alignment with biofabrication capabilities.

Asunto(s)

Colágeno , Hidrogeles , Técnicas de Cultivo de Célula , Matriz Extracelular , Hidrogeles/farmacología

RESUMEN

Intervertebral disc (IVD) degeneration is a major cause of low back pain and represents a massive socioeconomic burden. Current conservative and surgical treatments fail to restore native tissue architecture and functionality. Tissue engineering strategies, especially those based on 3D bioprinting and electrospinning, have emerged as possible alternatives by producing cell-seeded scaffolds that replicate the structure of the IVD extracellular matrix. In this review, we provide an overview of recent advancements and limitations of 3D bioprinting and electrospinning for the treatment of IVD degeneration, focusing on future areas of research that may contribute to their clinical translation.