A microfluidic interface for the culture and sampling of adiponectin from primary adipocytes

Abstract

Secreted from adipose tissue, adiponectin is a vital endocrine hormone that acts in glucose metabolism, thereby establishing its crucial role in diabetes, obesity, and other metabolic disease states. Insulin exposure to primary adipocytes cultured in static conditions has been shown to stimulate adiponectin secretion. However, conventional, static methodology for culturing and stimulating adipocytes falls short of truly mimicking physiological environments. Along with decreases in experimental costs and sample volume, and increased temporal resolution, microfluidic platforms permit small-volume flowing cell culture systems, which more accurately represent the constant flow conditions through vasculature in vivo. Here, we have integrated a customized primary tissue culture reservoir into a passively operated microfluidic device made of polydimethylsiloxane (PDMS). Fabrication of the reservoir was accomplished through unique PDMS “landscaping” above sampling channels, with a design strategy targeted to primary adipocytes to overcome issues of positive cell buoyancy. This reservoir allowed three-dimensional culture of primary murine adipocytes, accurate control over stimulants via constant perfusion, and sampling of adipokine secretion during various treatments. As the first report of primary adipocyte culture and sampling within microfluidic systems, this work sets the stage for future studies in adipokine secretion dynamics.

Macro-to-micro interfacing to microfluidic channels using 3D-printed templates: application to time-resolved secretion sampling of endocrine tissue

“…5,12 In this work, we used similar passive control and added the capability of temporally-resolved secretion sampling. Again, ease-of-use and device disposability are major advantages compared to other approaches that use syringe pumps or electroosmotic flow control.…”

“…5,6 We previously addressed this problem using hand-fabricated templates to create a moat where dispersed primary adipocytes could be anchored down in collagen, adjacent to sampling channels. 5 In this work, adipose tissue explants were used, rather than dispersed adipocytes. This permitted the use of similar, albeit larger, interface designs compared to islet sampling devices.…”

“…11 We have previously shown that rigid polymer templates are functional as moulds for fluidic reservoirs in passive, monolithic PDMS microdevices designed for endocrine tissue sampling. 5,12 This work has highlighted the potential gains in device function that are available by extending patterning into the PDMS substrate above the channels, beyond the typical manual punching or drilling of reservoirs. Particularly for on-chip tissue culture, this bulk PDMS sculpting approach permits unique macro-to-micro interfacing that can be customized to the application or cell type.…”

“…12 In another study, the challenge of culturing buoyant primary adipocytes on-chip was addressed using custom fluidic moats for 3D culture in collagen. 5 However, these polymer templates were hand-constructed, increasing fabrication time as well as batch-to-batch variability. The templates required manual alignment with the photo-patterned wafer prior to PDMS curing, and failures were often encountered during transfer to an oven for curing, when templates would shift out of alignment with channels or even topple over into the uncured PDMS.…”

Monitoring cell secretions on microfluidic chips using solid-phase extraction with mass spectrometry

“…Perfusate can also be analyzed by other analytical techniques to measure cell secretions and other effects of the cells on the chemical environment. Manually sampling perfusate from chips for detection of peptide or metabolite release from cells has been reported [17–20]; however, manual sampling of perfusate and off-line assay fails to fully to take advantage of the integration and automation potential of microfluidic chips.…”