Abstract
One of the main problems of the decellularization technique is the subjectivity of the final evaluation of its efficacy in individual organs. This problem can result in restricted cell repopulation reproducibility and worse responses to transplant tissues. Our proposal is to analyze the optical profiles produced by hearts during perfusion decellularization, as an additional method for evaluating the decellularization process of each individual organ. An apparatus comprised of a structured LED source and photo detector on an adjustable base was developed to capture the relationship between transmitted light during the perfusion of murine hearts, and residual DNA content. Voltage-time graphic records were used to identify a nonlinear mathematical model to discriminate between decellularizations with remaining DNA above (Incomplete Decellularization) and below (Complete Decellularization) the standardized limits. The results indicate that temporal optical evaluation of the process enables inefficient cell removal to be predicted in the initial stages, regardless of the apparent transparency of the organ. Our open system also creates new possibilities to add distinct photo detectors, such as for specific wavelengths, image acquisition, and physical-chemical evaluation of the scaffold, in order to collect different kinds of information, from dozens of studies. These data, when compiled and submitted to machine learning techniques, have the potential to initiate an exponential advance in tissue bioengineering research.
Highlights
Studies have demonstrated that 40–50% of transplanted organs are lost within ten to twelve years, and a high proportion of the remainder suffer from side effects related to immune suppression[1,2]
The decellularization process must be capable of preserving the original extracellular matrix (ECM), which is essential for maintaining the three-dimensional geometry of the organ, and on which the recipient multipotent stem cells will be cultured, through cell repopulation[13]
Despite standardization of the decellularization protocol, simultaneous optical monitoring has demonstrated that there is an optical profile for each individual heart decellularization, which can be correlated with residual DNA concentrations
Summary
Studies have demonstrated that 40–50% of transplanted organs are lost within ten to twelve years, and a high proportion of the remainder suffer from side effects related to immune suppression[1,2]. Efficient removal of the donor’s original cells, as well as any cellular material, including DNA, is required to minimize or even avoid triggering such reactions[8] This concept guided the development of the decellularization technique by conferring low immunogenicity and producing scaffolds with unique structure and properties[9,10,11,12]. It is widely accepted that a successful decellularization can be confirmed by histological analysis with DAPI or Hematoxylin and Eosin (H&E), an observed lack of visible nuclear material, and through spectrophotometric quantification of DNA, collagen and glycosaminoglycans[8,11,22,23,24,25] These criteria are associated with destructive methods at the end point of the decellularization process, which creates an obstacle to assess the organs individually before recellularization without causing them damage. This will enable inefficient decellularizations to be identified, even where there is apparent organ transparency, providing an exponential advance in tissue bioengineering research
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