TY - JOUR

T1 - Papain-Based Solubilization of Decellularized Extracellular Matrix for the Preparation of Bioactive, Thermosensitive Pregels

AU - Almalla, Ahed

AU - Elomaa, Laura

AU - Bechtella, Leïla

AU - Daneshgar, Assal

AU - Yavvari, Prabhu

AU - Mahfouz, Zeinab

AU - Tang, Peter

AU - Koksch, Beate

AU - Sauer, Igor

AU - Pagel, Kevin

AU - Hillebrandt, Karl Herbert

AU - Weinhart, Marie

N1 - Funding Information: We acknowledge the financial support from the Federal Ministry of Research and Education (BMBF, FKZ 13N13523, M.W.) and the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) for SFB 1449 (Project ID 431232613, M.W., K.P., B.K.) and the Cluster of Excellence Matters of Activity, Image Space Material (M.W., I.M.S.) under Germany′s Excellence Strategy – EXC 2025. A.A. warmly thanks Helmholtz Graduate School Macromolecular Bioscience and Dahlem Research School of Freie Universität Berlin for their support. The authors are grateful to M.Sc. Peng Tang for taking the SEM images at the Core Facility of BioSupraMol supported by DFG. Some icons in the TOC Figure and A were used from flaticon.com or Biorender.com .

PY - 2023/12/11

Y1 - 2023/12/11

N2 - Solubilized, gel-forming decellularized extracellular matrix (dECM) is used in a wide range of basic and translational research and due to its inherent bioactivity can promote structural and functional tissue remodeling. The animal-derived protease pepsin has become the standard proteolytic enzyme for the solubilization of almost all types of collagen-based dECM. In this study, pepsin was compared with papain, α-amylase, and collagenase for their potential to solubilize porcine liver dECM. Maximum preservation of bioactive components and native dECM properties was used as a decisive criterion for further application of the enzymes, with emphasis on minimal destruction of the protein structure and maintained capacity for physical thermogelation at neutral pH. The solubilized dECM digests, and/or their physically gelled hydrogels were characterized for their rheological properties, gelation kinetics, GAG content, proteomic composition, and growth factor profile. This study highlights papain as a plant-derived enzyme that can serve as a cost-effective alternative to animal-derived pepsin for the efficient solubilization of dECM. The resulting homogeneous papain-digested dECM preserved its thermally triggered gelation properties similar to pepsin digests, and the corresponding dECM hydrogels demonstrated their enhanced bioadhesiveness in single-cell force spectroscopy experiments with fibroblasts. The viability and proliferation of human HepaRG cells on dECM gels were similar to those on pure rat tail collagen type I gels. Papain is not only highly effective and economically attractive for dECM solubilization but also particularly interesting when digesting human-tissue-derived dECM for regenerative applications, where animal-derived materials are to be avoided.

AB - Solubilized, gel-forming decellularized extracellular matrix (dECM) is used in a wide range of basic and translational research and due to its inherent bioactivity can promote structural and functional tissue remodeling. The animal-derived protease pepsin has become the standard proteolytic enzyme for the solubilization of almost all types of collagen-based dECM. In this study, pepsin was compared with papain, α-amylase, and collagenase for their potential to solubilize porcine liver dECM. Maximum preservation of bioactive components and native dECM properties was used as a decisive criterion for further application of the enzymes, with emphasis on minimal destruction of the protein structure and maintained capacity for physical thermogelation at neutral pH. The solubilized dECM digests, and/or their physically gelled hydrogels were characterized for their rheological properties, gelation kinetics, GAG content, proteomic composition, and growth factor profile. This study highlights papain as a plant-derived enzyme that can serve as a cost-effective alternative to animal-derived pepsin for the efficient solubilization of dECM. The resulting homogeneous papain-digested dECM preserved its thermally triggered gelation properties similar to pepsin digests, and the corresponding dECM hydrogels demonstrated their enhanced bioadhesiveness in single-cell force spectroscopy experiments with fibroblasts. The viability and proliferation of human HepaRG cells on dECM gels were similar to those on pure rat tail collagen type I gels. Papain is not only highly effective and economically attractive for dECM solubilization but also particularly interesting when digesting human-tissue-derived dECM for regenerative applications, where animal-derived materials are to be avoided.

UR - http://www.scopus.com/inward/record.url?scp=85177602068&partnerID=8YFLogxK

U2 - 10.1021/acs.biomac.3c00602

DO - 10.1021/acs.biomac.3c00602

M3 - Article

C2 - 38009757

AN - SCOPUS:85177602068

VL - 24

SP - 5620

EP - 5637

JO - BIOMACROMOLECULES

JF - BIOMACROMOLECULES

SN - 1525-7797

IS - 12

ER -