Open multi-culture platform for simple and flexible study of multi-cell type interactions

Abstract

The study of multi-cell-type (MCT) interactions has the potential to significantly impact our understanding of tissue and disease biology. Such studies require innovative culture tools for unraveling the contributions of each cell type. Micro- and macro-scale platforms for MCT culture each have different advantages and disadvantages owing to their widely different capabilities, availability, and ease-of-use. However, as evidenced in the literature, there are very few examples of MCT studies and culture platforms, suggesting both biological and technical barriers. We have developed an open multi-culture platform to promote more rapid progress by integrating advantages of both micro- and macro-scale culture devices. The proposed open multi-culture platform addresses technical barriers by allowing easy customization, independent control of basic physical culture parameters, and incorporation of multiple culture modalities (e.g., 2D, 3D, transwell, and spheroid). The design also permits the user to obtain independent endpoints for each culture region. We demonstrate use of the platform in two example studies where we evaluated how cell ratio and cell types influence the response of triple negative breast cancer cells to heat damage and Hedgehog signaling. We also show that the platform can improve soluble factor transport between cell types compared to compartmentalized macro- and micro-scale alternatives. Last, we examine current and future challenges of the platform. We envision simple, yet flexible and customizable, platforms such as this will be important for advancing in vitro study of tissue and tumor biology.

Mesenchymal Cells Support the Oncogenicity and Therapeutic Response of the Hedgehog Pathway in Triple-Negative Breast Cancer

“…A custom open-multi-culture platform was used for co-culture to maintain a 1:1 tumor and stromal ratio, while keeping local confluence of the stroma needed for growth arrest and strong signal response to SHH-ligand [41]. The microwell platform consisted of adjacent microwells contained within a parent well, as described before [25]. Results show that the tumor cell growth was driven in response to active Hh signaling in all fibroblasts examined (Figure 3A,B).…”

“…ALDH1, a well-established marker of breast cancer stem cells (CSCs) [24], was used to monitor changes in the CSCs proportion. The two-adjacent microwell culture device [25] (Figure 4A.1) was scaled-up by a factor of nine (Figure 4B.1) to obtain sufficient tumor cells for quantification of ALDH1 expression using image-based cytometry. Prior to tumor growth studies, Hh signaling activity was examined across culture modalities and compared to monocultures supplemented with SHH-ligand to determine whether GFP-tagged MDA-MB-231 secreted enough SHH-ligand to induce a robust Hh signaling activity in NIH3T3.…”

“…Fabrication of polystyrene-based microwells has been described in detail in several publications [25,26]. Tumor cells (5000 cells/microwell) and mesenchymal cell subtypes were seeded in adjacent compartments simultaneously.…”

“…In this study, we further confirmed a lack of sensitivity to Hh inhibition in TNBC cells and a paracrine interplay between the tumor and stroma in both in vitro and in vivo models. We adapted a previously developed microscale co-culture platform [25,26] to develop a paracrine model of Hh signaling in TNBC and examined the impact of mixed and adjacent culture modalities in the tumorigenicity and transcriptional activity of the Hh pathway. The culture model was composed of TNBC human cell lines and a subset of mesenchymal cells consisting of human mammary fibroblasts (HMFs), CAFs, tumor cells that undergo epithelial–mesenchymal transition (EMT), and adipose mesenchymal stem cells (ADMSCs).…”

Open microfluidic coculture reveals paracrine signaling from human kidney epithelial cells promotes kidney specificity of endothelial cells

“…Open microfluidic systems are characterized by the introduction of an air-liquid interface, where liquid flows in microscale channels devoid of at least one side-wall (2,5,6,13). Unlike typical microfluidic devices with enclosed culture chambers, open devices offer increased pipette accessibility at any point along the culture chamber and simpler fabrication processes, such as straightforward micro-milling and injection molding, allowing open microfluidic devices to become efficient tools with versatility and transferability across different research disciplines (11,18,30,31,40,49,58).…”