Comparative characterization of decellularized renal scaffolds for tissue engineering

Abstract

Native extracellular matrix (ECM) provides scaffolds for tissue engineering with natural architecture and biochemical composition. Maintaining the native ECM in decellularized tissues provides cues for cells, which promote their tissue specific arrangement and function. Several approaches have been used to decellularize ECM from the kidney in order to reestablish renal tissue but their comparability is hampered because methods for decellularization and assessment of ECM vary widely. Therefore, we applied a standardized immersion protocol to decellularize porcine kidney tissue with three detergents Triton X-100, SDS and sodium deoxycholate (SDC) at variable temperatures. For comparative analysis decellularization efficacies, structural preservation, composition and cell attachment and viability were analyzed. Structural ECM-conservation is strongly dependent on decellularization temperature, while preservation of glycosaminoglycans (GAG), collagens and cytokines was affected by the detergents used. GAG and collagens were best maintained by 1% SDS at 4 °C, whereas cytokines were best maintained in 1% SDC at 4 °C. Viability and attachment of human induced pluripotent stem cell derived renal precursor cells were best in SDC-ECM and thus not associated with the degree of GAG and collagen maintenance but the cytokine preservation. Based on structural and functional characteristics, we developed a scoring system that allows intra- and inter-study comparison of decellularization strategies. Application of the scoring system to our experimental data showed that decellularization with 1% SDS at 4 °C provided the highest structural and composition scores, while 1% SDC at 4 °C had lower structural and composition but a significantly better cell performance score. Inclusion of multiple published studies in the scoring matrix for comparison identified the highest structural and composition scores when decellularization was performed with SDS at low concentration, for a short period of time and at low temperature. Furthermore, the scoring system indicated that cell attachment and viability cannot be concluded from any other parameter and should therefore always be included in evaluation of decellularization strategies.

A Non-woven Path: Electrospun Poly(lactic acid) Scaffolds for Kidney Tissue Engineering

“…However, cells within the Kidney do not exist within the two dimensional axis of cell culture plastic, with greater amounts of research underlining the importance of a 3D structure [17,44]. 3D kidney tissue engineering has focused on the use of decellularised tissue with promising results [25,45]; this method is not without its shortcomings, with issues surrounding decontamination [46] and a lack of a standardized approach to decellularisation leading to a variation between scaffolds [21,26,27]. Decellularised tissue brings favourable physical and chemical characteristics to help support cells and control physiology [47], but as cell produce their own ECM a synthetic scaffold can provide an excellent foundation to build upon [48,49].…”

“…This acellular extracellular matrix (ECM) is then recellularized and in some cases has been documented to produce rudimentary urine [25]. However, the decellularisation of tissue is not a simple process and the material left behind is often poorly characterised and mechanically weak [21,26,27], added to this the formidable task of recellularization [28] yielding enough uncertainty that other avenues should be pursued alongside.…”

Charting the unexplored extracellular matrix in cancer

“…Recently, researchers have also begun investigating the way that decellularization approaches affect the integrity of the ECM including the preservation of glycosaminoglycans (GAG), collagens and cytokines (Fischer et al . ). A clear benefit of tissue decellularization is that it improves the optical clarity of large volumes of tissue for studying the ECM using conventional approaches such as confocal and multiphoton microscopy as well as emerging techniques such as light‐sheet microscopy.…”

Molecular mobility and activity in an intravital imaging setting – implications for cancer progression and targeting

“…Furthermore, new optical clearing techniques for large mammalian systems, such as Clarity, Scale, CUBIC (clear, unobstructed brain/body imaging cocktails and computational analysis), 3DISCO (3-dimensional imaging of solvent-cleared organs), SeeDB (see deep brain), ISDoT (in situ decellularization of tissues) or PACT (photoacoustic computed tomography), have improved the imaging resolution at depth by removing the cellular components of tissues that cause its opacity (Chung and Deisseroth, 2013;Fischer et al, 2017;Mayorca-Guiliani et al, 2017;Richardson and Lichtman, 2015;Tainaka et al, 2014;Tomer et al, 2014;Yu et al, 2017). Future combination of intravital studies with post-intravital optical clearing and subsequent high-resolution imaging, similar to CLEM, may provide new insight into disease aetiology and treatment.…”