title: A hyaluronic acid-based hydrogel enabling CD44-mediated chondrocyte binding and gapmer oligonucleotide release for modulation of gene expression in osteoarthritis.
abstract: Hyaluronic acid (HA) is an attractive biomaterial for osteoarthritis (OA) treatment due to inherent functional and compatibility properties as an endogenous knee joint component. In this work, we describe a HA-based hydrogel with the dual functionality of increased CD44-dependent chondrocyte binding and controlled release of gapmer antisense oligonucleotides for unassisted cellular entry and subsequent gene silencing activity. A Schiff base-mediated gelation method was used to produce a panel of hydrogels varying in the aldehyde-modified HA (900kDa) to chitosan ratios (3:7, 5:5 and 7:3) for identifying designs displaying optimal engagement of OA patient-derived CD44-expressing chondrocytes. Correlation was found between cell binding and CD44 expression, with maximal binding exhibited at a HA/chitosan ratio of 7:3, that was 181% higher than CD44-negative MCF-7 cell control cells. Transfection agent-free uptake into OA chondrocytes of fluorescent 13-mer DNA oligonucleotides with a flanked locked nucleic acid (LNA) gapmer design, in contrast to naked siRNA, was demonstrated by confocal and flow cytometric analysis. A sustained and complete release over 5days was found with the 7:3 hydrogel, in contrast, the 5:5 and 3:7 hydrogel released 60% and 43% of loaded gapmers, respectively over the same period. A COX-2-specific gapmer designed with maximal chondrocyte gene silencing (~70% silencing efficiency at 500nM compared with a mismatch gapmer sequence) resulted in effective COX-2 silencing over 14days in hydrogels seeded with OA chondrocytes, with significant difference exhibited between day 3 and 10. This work introduces a novel HA-based CD44-mediated cellular binding and gapmer controlled release platform to modulate cellular gene expression.
authors.affiliation: The Interdisciplinary Nanoscience Center (iNANO), Department of Molecular Biology and Genetics, Aarhus University, 8000 Aarhus C, Denmark.
authors.forename: Yunpeng
authors.initials: Y
authors.lastname: Cai
authors.affiliation: The Interdisciplinary Nanoscience Center (iNANO), Department of Molecular Biology and Genetics, Aarhus University, 8000 Aarhus C, Denmark; Department of Health Sciences, University of Jaén, Jaén E-23071, Spain.
authors.forename: Elena
authors.initials: E
authors.lastname: López-Ruiz
authors.affiliation: Nucleic Acid Center, Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, 5230 Odense M, Denmark.
authors.forename: Jesper
authors.initials: J
authors.lastname: Wengel
authors.affiliation: Department of Orthopaedics, University Medical Center Utrecht, 3584 CX, Utrecht, Netherlands.
authors.forename: Laura B
authors.initials: LB
authors.lastname: Creemers
authors.affiliation: The Interdisciplinary Nanoscience Center (iNANO), Department of Molecular Biology and Genetics, Aarhus University, 8000 Aarhus C, Denmark. Electronic address: kenh@inano.au.dk.
authors.forename: Kenneth A
authors.initials: KA
authors.lastname: Howard
mesh_terms: D000447:Aldehydes / Q000737:chemistry; D048271:Chitosan / Q000737:chemistry; D019902:Chondrocytes / Q000378:metabolism*; D051546:Cyclooxygenase 2 / Q000235:genetics*; D015870:Gene Expression; D020868:Gene Silencing; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism*; D006820:Hyaluronic Acid / Q000737:chemistry; D020100:Hydrogels / Q000008:administration & dosage* / Q000737:chemistry; D061986:MCF-7 Cells; D009841:Oligonucleotides / Q000008:administration & dosage* / Q000737:chemistry; D010003:Osteoarthritis / Q000235:genetics
chemical_list: D000447:Aldehydes; C497483:CD44 protein, human; D018960:Hyaluronan Receptors; D020100:Hydrogels; D009841:Oligonucleotides; C477371:locked nucleic acid; D006820:Hyaluronic Acid; D048271:Chitosan; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human
keywords: Antisense; CD44 receptor; Gapmer; Hyaluronic acid; Hydrogel; Osteoarthritis