Creating living cellular machines

Abstract

Development of increasingly complex integrated cellular systems will be a major challenge for the next decade and beyond, as we apply the knowledge gained from the sub-disciplines of regenerative medicine, synthetic biology, micro-fabrication and nanotechnology, systems biology, and developmental biology. In this prospective, we describe the current state-of-the-art in the assembly of source cells, derived from pluripotent cells, into populations of a single cell type to produce the components or building blocks of higher order systems and finally, combining multiple cell types, possibly in combination with scaffolds possessing specific physical or chemical properties, to produce higher level functionality. We also introduce the issue, questions and ample research opportunities to be explored by others in the field. As these "living machines" increase in capabilities, exhibit emergent behavior and potentially reveal the ability for self-assembly, self-repair, and even self-replication, questions arise regarding the ethical implications of this work. Future prospects as well as ways of addressing these complex ethical questions will be discussed.

Figure 1

The fusion of different disciplines and specialties needed to develop living machines.

Figure 2

Synthesis and examples of the different disciplines: (a) gene networks in bacteria can be programmed to result in different patterns, (b) the “Vacanti mouse” as an example of tissue engineering and control of biological phenotype, (c) systems biology depicted in terms of increasing complexity of mechanistic, biological, and systems understanding, (d) developmental biology examples such as growth of a zebrafish embryo (left) and examples of phenotypic control by altering the electrical polarization of adult stem cells to generate a 2nd head on a planarian, or to develop a second working eye induced on tadpole gut, (e) stem cell differentiation to produce neurons, muscle or endothelial cells, (f) systems design and top-down decomposition of complex systems such as an airliner and design specification of the sub components, and (f) examples in microfluidics, lab on chip, and 3-D fabrication using stereolithographic printing of cells and polymers for tissue engineering and 3-D soft systems

Figure 3

Conceptual schematic of the interdisciplinary foundations of the New Biology with implications for many solutions to societal problems. Reproduced from A New Biology for the 21^st Century.

Figure 4

Conceptual schematic of a Biological Machine with cells, scaffolds and physical or chemical cues to result in machines that exhibit specific functionalities.

Figure 5

Increasing levels of complexity to go from cells, modules, and machines.

Figure 6

Formation of muscle strips anchored to flexible posts. A) Sequential images of myoblasts in a collagen/Matrigel solution after seeding into a rectangular well with two compliant posts. B) Schematic of the resulting muscle strip formed around the two compliant posts. C) Fluorescent image showing cell membranes (green) and nuclei (red). D) Muscle strip as in (C) stained for actin showing cell alignment at 3 days post seeding. E) Striated actin (red) and multi-nucleated (green) cells. Adapted from (Sakar et al., 2012).

Figure 7

Neuronal circuits on a chip. (a) Network of individual neurons patterned by laser, (b) 30 μm lines and 80 μm square nodes at 21 days in culture. (c) Neuropil structure separated by 500 μm with 3 μm wide lines. (d) Cross pattern of 80 μm nodes and 30 μm lines, stained for neurons (green), astroglia (red), and nuclei (blue). Adapted from .

Figure 8

Vascular networks formed in microfluidic platforms. Left: A) Microfluidic system. B) Schematic showing four parallel channels of the device with two outside gel regions (LO, RO), two media channels (LI, LO) and a central gel region. C) – F) Different seeding conditions for forming vascular networks. Reproduced from . Right top: A) – C) Vascular networks formed in the central gel region as in Left (A). Right bottom: A) Confocal image of a perfusible vascular network grown in a microfluidic gel system. B) Slices of the network in (A) showing lumens and the 3D nature of the formed vessels. Reproduced from .

Figure 9

Autonomous bio-hybrid muscle actuators capable of (a) swimming in fluid – bioengineering of a artificial jelly fish like structure capable of swimming in fluid autonomously or being pulsed by an external electric field (Adapted from Nawroth, et al. 2012), (b) walking in fluid - Biological Biomorph cantilever structure actuation with the beating of primary cardiac cells resulting in a net motion with maximum velocity 236 μm/s, average displacement 354 μm/stroke and average beating frequency ~1.5 Hz Adapted from .

Figure 10

Body-on-a-chip. Reproduced from . Conceptual image of how the various existing organs-on-a-chip might be assembled to simulate the entire physiological system of a human for the purpose of drug screening. A) Lung. Reproduced from . B) Blood brain barrier. Reproduced from . C) Heart tissue. Reproduced from . D) Liver. Reproduced from . E) Intestinal villi. Reproduced from . F) Muscle. Reproduced from . G) Blood vessels. Reproduced from .