Living Biointerfaces for the Maintenance of Mesenchymal Stem Cell Phenotypes

Abstract

Living interfaces are established as a novel class of active materials that aim to provide an alternative to traditional static cell culture methods by enabling users to accurately control cell behaviour in a precise, dynamic, and reliable system‐internal manner. To this day, the only reported biointerface has been a coculture between a biofilm of nonpathogenic genetically engineered bacteria and mammalian cells, where the recombinant proteins produced by the bacteria directly influence cell behaviour. In this work, a biointerface is presented between Lactococcus lactis (L. lactis) and human mesenchymal stem cells (hMSCs). L. lactis have been engineered to produce human C‐X‐C motif chemokine ligand 12, thrombopoietin, vascular cell adhesion protein 1, and the 7th–10th type III domains of human fibronectin, with the aim of recreating the native bone marrow conditions ex vivo. This active microenvironment has been shown to maintain key hMSC stemness markers, preventing their osteogenic and adipogenic differentiation, and maintaining high stem cell viability and physiological cell‐to‐substrate adhesion dynamics. This work presents proof of concept data that hMSC stemness can be regulated by living materials, using a system based on the symbiotic interaction between different engineered bacteria and mammalian cells.

Programmable Bacterial Biofilms as Engineered Living Materials

“…Researchers have also explored the engineering of other microbial biofilms for advanced applications, including the design of Lactococcus lactis to secrete biofilms fused with III 7–10 fragments of human fibronectin (FNIII 7–10 ) for mammalian cell adhesion supporting, the modification of surface layers (S-layer) in Caulobacter crescentus biofilms to generate 2D or 3D functional materials , and the engineering of Geobacter sulfurreducens conductive pili nanowires for novel biosensing applications . These examples underscore the universal applicability of the biofilm protein engineering strategy, yielding self-organizing living biofilm materials with diverse functionalities and application scenarios.…”

Engineered Living Materials For Sustainability

“…144 Hay et al engineered the nonpathogenic bacterium Lactobacillus lactis to display recombinant human fibronectin fragments (FNIII 7−10 ) on cell walls. 139,147,148 149 The developed biointerface resembling the bone marrow microenvironment prevented the differentiation of hMSCs and maintained their long-term stem cell phenotype. Thus, the differentiation or stemness of mammalian cells can be tuned ondemand by leveraging the tailor-made living bacterial interface.…”

“…Recently, the same group designed L. lactis to secrete several other biomolecules, including human C-X-C motif chemokine ligand 12 (CXCL12), thrombopoietin, and vascular cell adhesion protein 1, all of which are present in the native bone marrow . The developed biointerface resembling the bone marrow microenvironment prevented the differentiation of hMSCs and maintained their long-term stem cell phenotype.…”

Development of an optogenetic gene expression system inLactococcus lactisusing a split photoactivatable T7 RNA polymerase

“…Besides its widespread use in the food industry as a starter culture for the manufacturing of dairy products like buttermilk and cheese, it has gained traction as a live vector for mucosal vaccine delivery(1) and recombinant protein production without the disadvantages of E. coli such as the contamination of the recombinant proteins with lipopolysaccharides that can elicit immune response and inflammation, and the presence of robust protein secretion pathways in L. lactis compared to E. coli (2). L. lactis has been used recently in tissue engineering applications and in engineered living materials (ELMs) (3)(4)(5), a nascent field where a new class of materials based on an inert matrix and a living component, usually engineered using synthetic biology, combine together to produce "smart" functional materials with capabilities that surpass the current state-of-the-art. ELMs are able to respond to environmental physicochemical cues, namely electromagnetic and/or mechanical signals, chemical species and gradients, pH and others.…”