μOrgano: A Lego®-Like Plug & Play System for Modular Multi-Organ-Chips

Abstract

Human organ-on-a-chip systems for drug screening have evolved as feasible alternatives to animal models, which are unreliable, expensive, and at times erroneous. While chips featuring single organs can be of great use for both pharmaceutical testing and basic organ-level studies, the huge potential of the organ-on-a-chip technology is revealed by connecting multiple organs on one chip to create a single integrated system for sophisticated fundamental biological studies and devising therapies for disease. Furthermore, since most organ-on-a-chip systems require special protocols with organ-specific media for the differentiation and maturation of the tissues, multi-organ systems will need to be temporally customizable and flexible in terms of the time point of connection of the individual organ units. We present a customizable Lego®-like plug & play system, μOrgano, which enables initial individual culture of single organ-on-a-chip systems and subsequent connection to create integrated multi-organ microphysiological systems. As a proof of concept, the μOrgano system was used to connect multiple heart chips in series with excellent cell viability and spontaneously physiological beat rates.

Fig 1. Challenges and solution for multi-organ-systems.

a) General requirements for multi-organ-chips: i) initial separate loading of the respective cells; ii) individual culture for differentiation, formation, equilibration, and maturation of the tissues; and, iii) combined culture for drug screening purposes. b) Underlying concept of the μOrgano system: Schematics depicting the basic μOrgano components: the master-organ-chip and exemplary plug & play connectors. Conceptual idea of the usage principle of the μOrgano system for the connection of two MPSs in series via a simple linear channel connector with a close-up of the connected system highlighting the resulting media flow.

Fig 2. Fabrication of μOrgano building blocks.

Schematic protocol for the fabrication of connectors (and MPSs) with precise in- and outlet positions via multi step UV-lithography: i) microscopic channel structures are patterned in photoresist using UV lithography; ii) macroscopic in- and outlets are patterned as pillars on top of the microscopic channel structures using a second UV lithography step; iii) microfluidic PDMS devices are fabricated with predefined in- and outlets via exclusion molding; iv) PDMS connectors are cut and bonded to pre-cut microscope slides; and, v) glass capillaries are inserted and bonded into the in- and outlets of the connectors.

Fig 3. Characterization of μOrgano building blocks.

a) Transition time of the interface of a liquid advancing through a system of two MPSs and a linear connector. The time necessary to advance from the cell chamber in MPS 1 to the cell chamber in MPS 2 is plotted versus the inner diameters of the glass capillaries in the respective systems. Insets show pictures of the respective glass capillaries (scale bars = 2 mm). b) Scatter plot of the transition times for ten independent systems connected by the same type of connectors featuring 50 μm ID capillaries. c) Time series of microscopy images from a channel section in the proximity of the inlet of the second MPS initially filled with clear water. The continuous transition occurring after connection to a MPS filled with coloured water using a food dye reveals the bubble less connection ability of the system (scale bar = 100 μm). d) Time series of pictures showing two MPSs connected by a linear connector whereby MPS 1 is prefilled with red dyed water, and MPS 2 and the connector with blue dyed water. Pumping red dyed water into MPS 1 leads to the replacement of the blue dyed water in both the connector and MPS 2 without the occurrence of leakage. e) Volume flown through MPS 2 (left; in flow direction) and MPS 3 (right) plotted as percentage of the total volume after connection to MPS 1 via a bifurcation connector.

Fig 4. Proof of concept of the μOrgano system.

a) General procedure for biological experiments with the μOrgano system. b) Combined culture of two devices with 3T3 fibroblasts: Live (green) /dead (red) staining in both devices after 1 day of individual and 2 days of combined culture show that viability can be maintained. c) In-series culture of two heart-on-a-chip devices: tracings of the beating motion of cardiac tissue formed by hiPSC-cardiomyocytes—i) optical microscopy image—in two connected MPSs (ii) MPS 1; iii) MPS 2). The analysis using computational motion tracking reveals that a physiological phenotype is retained and individual cardiac devices beat with distinct frequencies. (scale bars = 200 μm).