Photoconversion and chromatographic microfluidic system reveals differential cellular phenotypes of adhesion velocity versus persistence in shear flow.

An integrated photoconversion and cell sorting parallel-plate chromatography channel enabling the measurement of instantaneous and average velocities of cells mediating adhesion in flow fields was engineered to study the mechanisms underlying adhesion to selectins by metastatic cancer cells. Through the facile enrichment of cells into subfractions of differing adhesive behaviors and a fluorescent velocity probe amenable to off-chip analysis, underlying, causal molecular profiles implicated in differing adhesive phenotypes of metastatic cancer cells could be interrogated. This analytical method revealed selectin-mediated rolling adhesion to be strongly associated with expression of selectin ligands, correlations that vary with ligand type and rolling velocity magnitude. Discrete selectin ligand expression profiles were also found to underlie persistent versus non-persistent adhesion on selectins, suggestive of divergent regulatory mechanisms. This integrated cell sorting and photoconversion microfluidic platform thus enables in vitro analysis and comparisons of adhesive phenotypes as they relate to mechanisms of cancer cell metastasis in the context of selectin mediated adhesion, revealing new insights into potential cancer dissemination pathways.

Lymph Node Subcapsular Sinus Microenvironment-On-A-Chip Modeling Shear Flow Relevant to Lymphatic Metastasis and Immune Cell Homing.

A lymph node sinus-on-a-chip adhesion microfluidic platform that recapitulates the hydrodynamic microenvironment of the lymph node subcapsular sinus was engineered. This device was used to interrogate the effects of lymph node remodeling on cellular adhesion in fluid flow relevant to lymphatic metastasis. Wall shear stress levels analytically estimated and modeled after quiescent and diseased/inflamed lymph nodes were experimentally recapitulated using a flow-based microfluidic perfusion system to assess the effects of physiological flow fields on human metastatic cancer cell adhesion. Results suggest that both altered fluid flow profiles and presentation of adhesive ligands, which are predicted to manifest within the lymph node subcapsular sinus as a result of inflammation-induced remodeling, and the presence of lymph-borne monocytic cells may synergistically contribute to the dynamic extent of cell adhesion in flow relevant to lymph node invasion by cancer and monocytic immune cells during lymphatic metastasis.

Analyzing Mechanisms of Metastatic Cancer Cell Adhesive Phenotype Leveraging Preparative Adhesion Chromatography Microfluidic.

An integrated, parallel-plate microfluidic device is engineered to interrogate and fractionate cells based on their adhesivity to a substrate surface functionalized with adhesive ligand in a tightly controlled flow environment to elucidate associated cell-intrinsic pathways. Wall shear stress levels and endothelial presentation of E-selectin are modeled after the inflamed vasculature microenvironment in order to simulate in vitro conditions under which in vivo hematogenous metastasis occurs. Based on elution time from the flow channel, the collection of separate fractions of cells-noninteracting and interacting-at high yields and viabilities enables multiple postperfusion analyses, including flow cytometry, in vivo metastasis modeling, and transcriptomic analysis. This platform enables the interrogation of flow-regulated cell molecular profiles, such as (co)expression levels of natively expressed selectin ligands sLex , CD44, and carcinoembryonic antigen, and cancer stem cell marker CD24. This additionally reveals E-selectin adhesivity exhibited by metastatic human colon carcinoma cells to be a transient phenotype. Facile and rapid, this methodology for unbiased, label free sorting of large populations of cells based on their adhesion in flow represents a method of studying flow-regulated adhesion in vitro for the identification of molecular drug targets for development as antimetastatic cancer therapeutics.