On-demand dissolution of modular, synthetic extracellular matrix reveals local epithelial-stromal communication networks
Methods to parse paracrine epithelial-stromal communication networks are a vital need in drug development, as disruption of these networks underlies diseases ranging from cancer to endometriosis. Here, we describe a modular, synthetic, and dissolvable extracellular matrix (MSD-ECM) hydrogel that fos...
| Main Authors: | , , , , , , , , , , , , |
|---|---|
| Other Authors: | |
| Format: | Article |
| Published: |
Elsevier BV
2018
|
| Online Access: | http://hdl.handle.net/1721.1/117633 https://orcid.org/0000-0002-6673-087X https://orcid.org/0000-0001-8272-6419 https://orcid.org/0000-0001-6005-6901 https://orcid.org/0000-0001-6718-2265 https://orcid.org/0000-0002-5506-236X https://orcid.org/0000-0002-8106-0640 https://orcid.org/0000-0001-6865-4084 https://orcid.org/0000-0001-8373-1463 https://orcid.org/0000-0002-0050-989X https://orcid.org/0000-0002-1545-1651 https://orcid.org/0000-0002-1801-5548 |
| _version_ | 1826207582036951040 |
|---|---|
| author | Ahrens, Caroline Chopko Valdez Macias, Jorge Luis Cook, Christi Dionne Wang, Alex J-S Brown, Alexander Thomas Kumar, Manu Prajapati Stockdale, Linda Rothenberg, Daniel Abram Renggli-Frey, Kasper Gordon, Elizabeth A Lauffenburger, Douglas A Griffith, Linda G White, Forest M. |
| author2 | Massachusetts Institute of Technology. Biotechnology Process Engineering Center |
| author_facet | Massachusetts Institute of Technology. Biotechnology Process Engineering Center Ahrens, Caroline Chopko Valdez Macias, Jorge Luis Cook, Christi Dionne Wang, Alex J-S Brown, Alexander Thomas Kumar, Manu Prajapati Stockdale, Linda Rothenberg, Daniel Abram Renggli-Frey, Kasper Gordon, Elizabeth A Lauffenburger, Douglas A Griffith, Linda G White, Forest M. |
| author_sort | Ahrens, Caroline Chopko |
| collection | MIT |
| description | Methods to parse paracrine epithelial-stromal communication networks are a vital need in drug development, as disruption of these networks underlies diseases ranging from cancer to endometriosis. Here, we describe a modular, synthetic, and dissolvable extracellular matrix (MSD-ECM) hydrogel that fosters functional 3D epithelial-stromal co-culture, and that can be dissolved on-demand to recover cells and paracrine signaling proteins intact for subsequent analysis. Specifically, synthetic polymer hydrogels, modified with cell-interacting adhesion motifs and crosslinked with peptides that include a substrate for cell-mediated proteolytic remodeling, can be rapidly dissolved by an engineered version of the microbial transpeptidase Sortase A (SrtA) if the crosslinking peptide includes a SrtA substrate motif and a soluble second substrate. SrtA-mediated dissolution affected only 1 of 31 cytokines and growth factors assayed, whereas standard protease degradation methods destroyed about half of these same molecules. Using co-encapsulated endometrial epithelial and stromal cells as one model system, we show that the dynamic cytokine and growth factor response of co-cultures to an inflammatory cue is richer and more nuanced when measured from SrtA-dissolved gel microenvironments than from the culture supernate. This system employs accessible, reproducible reagents and facile protocols; hence, has potential as a tool in identifying and validating therapeutic targets in complex diseases. |
| first_indexed | 2024-09-23T13:51:48Z |
| format | Article |
| id | mit-1721.1/117633 |
| institution | Massachusetts Institute of Technology |
| last_indexed | 2024-09-23T13:51:48Z |
| publishDate | 2018 |
| publisher | Elsevier BV |
| record_format | dspace |
| spelling | mit-1721.1/1176332022-10-01T17:37:41Z On-demand dissolution of modular, synthetic extracellular matrix reveals local epithelial-stromal communication networks Ahrens, Caroline Chopko Valdez Macias, Jorge Luis Cook, Christi Dionne Wang, Alex J-S Brown, Alexander Thomas Kumar, Manu Prajapati Stockdale, Linda Rothenberg, Daniel Abram Renggli-Frey, Kasper Gordon, Elizabeth A Lauffenburger, Douglas A Griffith, Linda G White, Forest M. Massachusetts Institute of Technology. Biotechnology Process Engineering Center Massachusetts Institute of Technology. Department of Biological Engineering Massachusetts Institute of Technology. Department of Biology Massachusetts Institute of Technology. Department of Chemical Engineering Massachusetts Institute of Technology. Department of Mechanical Engineering Valdez Macias, Jorge Luis Cook, Christi Dionne Wang, Alex J-S Brown, Alexander Thomas Kumar, Manu Prajapati Stockdale, Linda Rothenberg, Daniel Abram Renggli-Frey, Kasper Gordon, Elizabeth A Lauffenburger, Douglas A White, Forest M Griffith, Linda G Methods to parse paracrine epithelial-stromal communication networks are a vital need in drug development, as disruption of these networks underlies diseases ranging from cancer to endometriosis. Here, we describe a modular, synthetic, and dissolvable extracellular matrix (MSD-ECM) hydrogel that fosters functional 3D epithelial-stromal co-culture, and that can be dissolved on-demand to recover cells and paracrine signaling proteins intact for subsequent analysis. Specifically, synthetic polymer hydrogels, modified with cell-interacting adhesion motifs and crosslinked with peptides that include a substrate for cell-mediated proteolytic remodeling, can be rapidly dissolved by an engineered version of the microbial transpeptidase Sortase A (SrtA) if the crosslinking peptide includes a SrtA substrate motif and a soluble second substrate. SrtA-mediated dissolution affected only 1 of 31 cytokines and growth factors assayed, whereas standard protease degradation methods destroyed about half of these same molecules. Using co-encapsulated endometrial epithelial and stromal cells as one model system, we show that the dynamic cytokine and growth factor response of co-cultures to an inflammatory cue is richer and more nuanced when measured from SrtA-dissolved gel microenvironments than from the culture supernate. This system employs accessible, reproducible reagents and facile protocols; hence, has potential as a tool in identifying and validating therapeutic targets in complex diseases. National Institutes of Health (U.S.) (R01EB010246) National Institutes of Health (U.S.) (UH2TR000496) Institute for Collaborative Biotechnologies (W911NF-09-0001) National Institutes of Health (U.S.) (T32GM008334) United States. Defense Advanced Research Projects Agency. Microphysiological Systems Program (W911NF-12-2-0039) John and Karinne Begg Fund Begg New Horizon Fund for Undergraduate Research at MIT Massachusetts Institute of Technology. Biophysical Instrumentation Facility Manton Foundation Ludwig Postdoctoral Fellowship for Cancer Research Swiss National Science Foundation (Postdoctoral Fellowship) 2018-09-05T12:35:06Z 2018-09-05T12:35:06Z 2017-03 2017-03 2018-08-30T17:16:56Z Article http://purl.org/eprint/type/JournalArticle 01429612 http://hdl.handle.net/1721.1/117633 Valdez, Jorge, Christi D. Cook, Caroline Chopko Ahrens, Alex J. Wang, Alexander Brown, Manu Kumar, Linda Stockdale, et al. “On-Demand Dissolution of Modular, Synthetic Extracellular Matrix Reveals Local Epithelial-Stromal Communication Networks.” Biomaterials 130 (June 2017): 90–103. https://orcid.org/0000-0002-6673-087X https://orcid.org/0000-0001-8272-6419 https://orcid.org/0000-0001-6005-6901 https://orcid.org/0000-0001-6718-2265 https://orcid.org/0000-0002-5506-236X https://orcid.org/0000-0002-8106-0640 https://orcid.org/0000-0001-6865-4084 https://orcid.org/0000-0001-8373-1463 https://orcid.org/0000-0002-0050-989X https://orcid.org/0000-0002-1545-1651 https://orcid.org/0000-0002-1801-5548 http://dx.doi.org/10.1016/J.BIOMATERIALS.2017.03.030 Biomaterials Creative Commons Attribution-NonCommercial-NoDerivs License http://creativecommons.org/licenses/by-nc-nd/4.0/ application/pdf Elsevier BV Elsevier |
| spellingShingle | Ahrens, Caroline Chopko Valdez Macias, Jorge Luis Cook, Christi Dionne Wang, Alex J-S Brown, Alexander Thomas Kumar, Manu Prajapati Stockdale, Linda Rothenberg, Daniel Abram Renggli-Frey, Kasper Gordon, Elizabeth A Lauffenburger, Douglas A Griffith, Linda G White, Forest M. On-demand dissolution of modular, synthetic extracellular matrix reveals local epithelial-stromal communication networks |
| title | On-demand dissolution of modular, synthetic extracellular matrix reveals local epithelial-stromal communication networks |
| title_full | On-demand dissolution of modular, synthetic extracellular matrix reveals local epithelial-stromal communication networks |
| title_fullStr | On-demand dissolution of modular, synthetic extracellular matrix reveals local epithelial-stromal communication networks |
| title_full_unstemmed | On-demand dissolution of modular, synthetic extracellular matrix reveals local epithelial-stromal communication networks |
| title_short | On-demand dissolution of modular, synthetic extracellular matrix reveals local epithelial-stromal communication networks |
| title_sort | on demand dissolution of modular synthetic extracellular matrix reveals local epithelial stromal communication networks |
| url | http://hdl.handle.net/1721.1/117633 https://orcid.org/0000-0002-6673-087X https://orcid.org/0000-0001-8272-6419 https://orcid.org/0000-0001-6005-6901 https://orcid.org/0000-0001-6718-2265 https://orcid.org/0000-0002-5506-236X https://orcid.org/0000-0002-8106-0640 https://orcid.org/0000-0001-6865-4084 https://orcid.org/0000-0001-8373-1463 https://orcid.org/0000-0002-0050-989X https://orcid.org/0000-0002-1545-1651 https://orcid.org/0000-0002-1801-5548 |
| work_keys_str_mv | AT ahrenscarolinechopko ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks AT valdezmaciasjorgeluis ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks AT cookchristidionne ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks AT wangalexjs ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks AT brownalexanderthomas ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks AT kumarmanuprajapati ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks AT stockdalelinda ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks AT rothenbergdanielabram ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks AT rengglifreykasper ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks AT gordonelizabetha ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks AT lauffenburgerdouglasa ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks AT griffithlindag ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks AT whiteforestm ondemanddissolutionofmodularsyntheticextracellularmatrixrevealslocalepithelialstromalcommunicationnetworks |