Decellularisation of tissue matrices for bladder implantation
Inventors
Korossis, Sotirios • Ingham, Eileen • Bolland, Fiona • Southgate, Jennifer
Assignees
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
Publication Number
US-8828448-B2
Publication Date
2014-09-09
Expiration Date
Abstract
The invention provides an improved method of producing a natural, acellular matrix scaffold for subsequent use in tissue-engineered replacement of tissues such as the bladder. Decellularization is carried out on an expanded or distended bladder and the product retains the strength and compliance of natural material. The invention also provides use of the matrix scaffolds as wound healing material and to investigate tissue structure and function in vitro.
Core Innovation
The invention relates to a method of decellularisation of an intact bladder. The method immerses the bladder in a buffer solution at a mild alkaline pH that includes a proteolytic inhibitor, while maintaining the structural character of the bladder material.
The method expands the bladder by introducing a sufficient volume of the same buffer solution into an interior cavity of the bladder. This stretching and thinning of the bladder wall enables subsequent decellularisation, and the bladder is maintained in an expanded state rather than relaxed during processing.
Decellularisation is continued by replacing and introducing fresh solutions both around the exterior surface and into the bladder interior. This repeated interior and exterior solution replacement maintains expansion during decellularisation and maintains the histoarchitecture of the bladder material, with complete cell removal while retaining native bladder strength and compliance.
Claims Coverage
The partial content provides one independent claim directed to an intact-bladder decellularisation method, with multiple dependent claims refining the buffer/inhibitor composition and optional downstream treatments. The independent claim includes three main inventive features, and the dependent claims add further inventive feature refinements for specific inhibitor selection, nucleases, and immunoacceptability enhancement.
Intact-bladder decellularisation in a mild alkaline buffer with proteolytic inhibitor
The method includes immersing the intact bladder in a buffer solution at a mild alkaline pH that includes a proteolytic inhibitor.
Bladder expansion by interior infusion of the same mild alkaline buffer
The method expands the bladder by introducing a sufficient volume of the same buffer solution into an interior cavity to stretch and thin the bladder wall.
Continuing decellularisation with interior and exterior solution replacement to maintain expansion and histoarchitecture
The method continues decellularisation by replacing and introducing fresh solutions around the exterior surface and into the bladder interior to maintain expansion during decellularisation and to maintain the histoarchitecture.
Proteolytic inhibitor selection using EDTA
The proteolytic inhibitor is EDTA, with amounts defined as 1 mM to 100 mM or 0.01% to 1.0% w/v.
Proteolytic inhibitor selection using aprotinin
The proteolytic inhibitor is Aprotinin, with amounts defined as 1KIU to 100KIU.
Nuclease treatment with DNase Type I, DNase Type II, and RNase
The method includes treatment with one or more enzymes comprising DNase Type I, DNase Type II, and RNase.
Optional cryoprotectant or storage medium for later use
The method optionally places the biological bladder material in a cryoprotectant medium or storage medium for later use.
Enzymic digestion to remove α-Gal epitopes
The method uses an enzymic agent specifically an α-galactosidase or glycosidase digestion for removal of α-gal epitopes.
Fluid-loss control by clamping during solution replacement
The method includes clamping the bladder before and after each fluid replacement step to prevent fluid loss and maintain bladder expansion.
Overall, the claims coverage is centered on maintaining an intact bladder in an expanded condition during decellularisation using a mild alkaline buffer with a proteolytic inhibitor, followed by interior and exterior solution replacement to preserve histoarchitecture. Dependent claim refinements add specified proteolytic inhibitors, nucleases, optional cryoprotectant or storage handling, enzymic α-gal epitope removal, and clamping to prevent fluid loss while maintaining expansion.
Stated Advantages
Enables permeation and complete cell removal while retaining native bladder strength, compliance, and histoarchitecture.
Maintains bladder histoarchitecture during decellularisation by maintaining expansion.
Provides biocompatible acellular bladder matrix with retained biomechanical properties versus fresh bladder and comparisons to SIS/PERMACOL/pericardium.
Demonstrates minimal foreign-body response in a mouse subcutaneous implant model.
Supports shelf-life with mechanical integrity up to about 30 months.
Documented Applications
Bladder reconstruction and tissue repair/wound healing using the acellular bladder matrix.
Cosmetic/augmentation applications.
Tissue-engineering scaffold.
An in vitro model of bladder function.
Interested in licensing this patent?