Heparin modification improves the re-endothelialization and angiogenesis of decellularized kidney scaffolds through antithrombosis and anti-inflammation in vivo

Abstract

Background: Constructing tissue-engineered kidneys using decellularized kidney scaffolds (DKS) has attracted widespread attention as it is expected to be the key to solving the shortage of donor kidneys.However, thrombosis and the host inflammatory response are unfavorable factors that hider the reendothelialization and vascularization of the decellularized scaffolds.Methods: Heparin was immobilized into the DKS using the method of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide/N-hydroxysuccinimide (EDC/NHS) activation. Fourier-transform infrared (FTIR) spectra were used to verify the heparinization of DKS. Human umbilical vein endothelial cells (HUVECs) were seeded and cultured in the DKS, then the sliced scaffolds were transplanted subcutaneously into nude mouse. Scanning electron microscopy and a series of histochemical stains including hematoxylin and eosin (H&E), elastic Verhöeff-Van Gieson (EVG), Sirius red, Masson's trichrome, and toluidine blue (TB) staining were used for morphological characterization. The qRT-PCR analysis, immunohistochemistry (IHC), and immunofluorescence (IF) staining were used to determine the expression of related molecular markers. Results:The rat DKS completely retained the extracellular matrix and heparinized modification. The H&E staining results showed there were more HUVECs covering the internal surfaces of tubular structures in the HEP-DKS group compared with the DKS group. The IF analysis results revealed that CD31, Ki67, and CD206 had higher positive rates in HUVECs in the HEP-DKS group compared to the DKS group. Both groups of scaffolds showed blood vessel formation via H&E staining, and there were more blood vessels in the HEP-DKS group compared with the native DKS group (P<0.05). The qRT-PCR results showed that the levels of IL-1β, IL-6, and TNF-α in the HEP-DKS group were significantly lower than those of the native DKS group, while the expression level of IL-10 was significantly higher than that in the native DKS group (P<0.05).Conclusions: Heparin modification improves the re-endothelialization and vascular regeneration of the DKS through anticoagulation in vitro and in vivo. The anti-inflammatory effect of heparin on the transplanted host was initially confirmed, and it is considered that this effect may play a non-negligible role in promoting DKS re-endothelialization and angiogenesis. Heparinized DKS is therefore a promising candidate for kidney tissue engineering.

In-Vitro Endothelialization Assessment of Heparinized Bovine Pericardial Scaffold for Cardiovascular Application

“…As mentioned earlier, endothelial cells were confirmed to attach to a heparin-modified material surface [ 11 , 12 , 13 , 17 ], which was also found in our current result. The results also showed that the endothelialization effect on the heparinized scaffolds was not comparable to the un-modified one, which might raise a contradiction on the initial purpose of employing heparin on the bovine pericardial scaffolds.…”

“…Endothelial cells were shown not to express direct receptors for heparin. The positive effects of the heparin immobilized scaffold on endothelial cells were described via “bridging” molecules, e.g., chitosan, gelatin [ 11 , 17 ], or binding and stabilizing cell growth factors, e.g., VEGF [ 12 , 13 ]. Therefore, the decrease in cell adherence or proliferation on HepBPS could be explainable.…”

“…For early endothelialization, biochemical modification of the surfaces and immobilization of biomolecules to the surfaces have been useful. The present publications have provided evidence for the attachment of endothelial cells on heparin-modified material surfaces [ 11 , 12 , 13 ]. At the same time, pericardial scaffolds were also proven to support endothelial cell adherence and proliferation [ 14 ].…”

Crosslinking of dialdehyde heparin: a new strategy for improving the anticoagulant properties of porcine acellular dermal matrix

“…2b shows the FTIR spectrum of LMWH and OL, it was found that both of them had characteristic absorption peaks at 892 cm −1 and 936 cm −1 . 23,24 The C–O–S stretching vibration absorption peak of the sulfuric acid group on amino hexose appeared around 830 cm −1 , the strong absorption peak of O–S stretching vibration appeared around 1240 cm −1 , and the C O stretching vibration peak of the sulfuric acid group in the carboxyl group appeared around 1621 cm −1 was also found in both, all of these indicated that the main structure of the OL obtained after oxidation did not change significantly. It has been known that the reason for the conspicuous anticoagulant activity and cytocompatibility of LMWH is the presence of a reactive pentose structure in its molecular chain, 25 they can bind to antithrombin III (AT III), change the conformation of the antithrombin molecules, and inhibit the activity of coagulation factors IIa, Xa and IXa, exhibiting strong anticoagulant properties.…”

Heparin-Loaded Alginate Hydrogels: Characterization and Molecular Mechanisms of Their Angiogenic and Anti-Microbial Potential

“…It binds to many growth factors such as Transforming Growth Factor (TGF), VEGF and Fibroblast Growth Factor (FGF) [ 14 , 18 ]. It is widely used due to its anti-inflammatory and other pharmacological properties [ 19 ]. It binds to a wide number of proteins and helps in regulating their function [ 20 ].…”