Construction of millimeter-scale vascularized engineered myocardial tissue using a mixed gel.

Engineering myocardium has shown great clinal potential for repairing permanent myocardial injury. However, the lack of perfusing blood vessels and difficulties in preparing a thick-engineered myocardium result in its limited clinical use. We prepared a mixed gel containing fibrin (5 mg/ml) and collagen I (0.2 mg/ml) and verified that human umbilical vein endothelial cells (HUVECs) and human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) could form microvascular lumens and myocardial cell clusters by harnessing the low-hardness and hyperelastic characteristics of fibrin. hiPSC-CMs and HUVECs in the mixed gel formed self-organized cell clusters, which were then cultured in different media using a three-phase approach. The successfully constructed vascularized engineered myocardial tissue had a spherical structure and final diameter of 1-2 mm. The tissue exhibited autonomous beats that occurred at a frequency similar to a normal human heart rate. The internal microvascular lumen could be maintained for 6 weeks and showed good results during preliminary surface re-vascularization in vitro and vascular remodeling in vivo. In summary, we propose a simple method for constructing vascularized engineered myocardial tissue, through phased cultivation that does not rely on high-end manufacturing equipment and cutting-edge preparation techniques. The constructed tissue has potential value for clinical use after preliminary evaluation.

Layer-by-Layer Assembly of a Polysaccharide "Armor" on the Cell Surface Enabling the Prophylaxis of Virus Infection.

Airborne pathogens, such as the world-spreading severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), cause global epidemics via transmission through the respiratory pathway. It is of great urgency to develop adequate interventions that can protect individuals against future pandemics. This study presents a nasal spray that forms a polysaccharide "armor" on the cell surface through the layer-by-layer self-assembly (LBL) method to minimize the risk of virus infection. The nasal spray has two separate components: chitosan and alginate. Harnessing the electrostatic interaction, inhaling the two polysaccharides alternatively enables the assembly of a barrier that reduces virus uptake into the cells. The results showed that this approach has no obvious cellular injury and endows cells with the ability to resist the infection of adenovirus and SARS-CoV-2 pseudovirus. Such a method can be a potential preventive strategy for protecting the respiratory tract against multiple viruses, especially the upcoming SARS-CoV-2 variants.

Programmable dual responsive system reconstructing nerve interaction with small-diameter tissue-engineered vascular grafts and inhibiting intimal hyperplasia in diabetes.

Small-diameter tissue-engineered vascular grafts (sdTEVGs) with hyperglycemia resistance have not been constructed. The intimal hyperplasia caused by hyperglycemia remains problem to hinder the patency of sdTEVGs. Here, inspired by bionic regulation of nerve on vascular, we found the released neural exosomes could inhibit the abnormal phenotype transformation of vascular smooth muscle cells (VSMCs). The transformation was a prime culprit causing the intimal hyperplasia of sdTEVGs. To address this concern, sdTEVGs were modified with an on-demand programmable dual-responsive system of ultrathin hydrogels. An external primary Reactive Oxygen Species (ROS)-responsive Netrin-1 system was initially triggered by local inflammation to induce nerve remolding of the sdTEVGs overcoming the difficulty of nerve regeneration under hyperglycemia. Then, the internal secondary ATP-responsive DENND1A (guanine nucleotide exchange factor) system was turned on by the neurotransmitter ATP from the immigrated nerve fibers to stimulate effective release of neural exosomes. The results showed nerve fibers grow into the sdTEVGs in diabetic rats 30 days after transplantation. At day 90, the abnormal VSMCs phenotype was not detected in the sdTEVGs, which maintained long-time patency without intima hyperplasia. Our study provides new insights to construct vascular grafts resisting hyperglycemia damage.

Engineering hiPSC-CM and hiPSC-EC laden 3D nanofibrous splenic hydrogel for improving cardiac function through revascularization and remuscularization in infarcted heart.

Cell therapy has been a promising strategy for cardiac repair after myocardial infarction (MI), but a poor ischemic environment and low cell delivery efficiency remain significant challenges. The spleen serves as a hematopoietic stem cell niche and secretes cardioprotective factors after MI, but it is unclear whether it could be used for human pluripotent stem cell (hiPSC) cultivation and provide a proper microenvironment for cell grafts against the ischemic environment. Herein, we developed a splenic extracellular matrix derived thermoresponsive hydrogel (SpGel). Proteomics analysis indicated that SpGel is enriched with proteins known to modulate the Wnt signaling pathway, cell-substrate adhesion, cardiac muscle contraction and oxidation-reduction processes. In vitro studies demonstrated that hiPSCs could be efficiently induced into endothelial cells (iECs) and cardiomyocytes (iCMs) with enhanced function on SpGel. The cytoprotective effect of SpGel on iECs/iCMs against oxidative stress damage was also proven. Furthermore, in vivo studies revealed that iEC/iCM-laden SpGel improved cardiac function and inhibited cardiac fibrosis of infarcted hearts by improving cell survival, revascularization and remuscularization. In conclusion, we successfully established a novel platform for the efficient generation and delivery of autologous cell grafts, which could be a promising clinical therapeutic strategy for cardiac repair and regeneration after MI.

Precisely and Efficiently Enzyme Response Microspheres with Immune Removal Escape Loaded with MCC950 Ameliorate Cardiac Dysfunction in Acute Myocardial Infarction.

Although the percutaneous coronary intervention (PCI)treatment can improve the survival rate of acute myocardial infarction (AMI) patients, the early granulocytes response within 6 hours can induce second injuries during the reperfusion process. The new drug delivery system MMP9 hydrolytic microspheres (NMM) with negatively charged surface was designed out and MCC950 (MCC) was loaded into NMM (NMM-M), MCC is the inhibitor of nucleotide binding oligomerization domain (NOD)-like receptor, pyrin containing domain 3 (NLRP3)-inflammasome which is the key promoter of granulocytes-induced injury. NMM-M could effectively escape the phagocytosis of immune phagocytes in the blood, and target the ischemic region based on the electrostatic attraction and the attraction of enzyme to substrate, and sudden release the loaded MCC within 2 hours. The released MCC can inhibit the NLRP3-inflammasome activity, and then further inhibit the secretion of inflammatory factors in granulocytes which are the main factors of early inflammatory damage, and improving cardiac function, realizing the goal of pre-treatment. Therefore, NMM may be a new delivery system, which can provide the accurately, sufficient and rapidly drug deliver, and MCC may be a novel candidate drug in AMI treatment, which may be hopeful in the future.

Surface-Engineered Monocyte Inhibits Atherosclerotic Plaque Destabilization via Graphene Quantum Dot-Mediated MicroRNA Delivery.

Rupture-prone atherosclerotic plaque is the cause of the high mortality and morbidity rates that accompany atherosclerosis-associated diseases. MicroRNAs can regulate the expression of a variety of atherosclerotic inflammation-related genes in macrophages. There are currently no definitive methods for delivering microRNAs into the interior of plaque. Monocytes typically possess a pathological feature that allows them to be recruited to atherosclerotic plaque resulting in rupture-prone; however, whether monocytes can be modified to be gene carriers remains unclear. In this study, a novel monocyte surface-engineered gene-delivery system based on graphene quantum dots (GQDs) is developed. Briefly, GQDs-microRNA223 linked by disulfide bonds are grafted onto the monocyte membrane via a carefully designed C18-peptide (C18P) containing a hydrophobic end to afford the designed monocyte-C18P-GQDs-miR223 architecture. The system can reach and enter the interior of the plaque and release the GQDs-miRNA via C18P digestion. The released GQDs-miRNA are taken up by the macrophages in atherosclerotic plaques, and the disulfide linkages between the GQDs and the miRNA are cleaved through γ-interferon-inducible lysosomal thiol reductase (GILT) in the lysosome. Under the protection of GQDs, miRNA cargos are transfected into the cytosol and subsequently undergo nuclear translocation, allowing a significantly reduced plaque burden by regulating inflammatory response in vivo.

Engineered-Macrophage Induced Endothelialization and Neutralization via Graphene Quantum Dot-Mediated MicroRNA Delivery to Construct Small-Diameter Tissue-Engineered Vascular Grafts.

Rapid endothelialization of tissue-engineered blood vessels (TEBVs) is an essential strategy to inhibit thrombosis, chronic inflammation and intimal hyperplasia after transplantation into the body. Monocytes will be recruited to the transplantation site and converted to macrophages after TEBV implantation. Macrophages play an important role in angiogenesis; however, whether engineered macrophages can be utilized to promote rapid endothelialization of TEBVs remains unclear. Thus, a cell bioreactor that can engineer macrophages via graphene quantum dot (GQD)-mediated microRNA (miR) delivery was built in the TEBV. Briefly, GQD-miR-150 linked by disulfide bonds was adopted to functionalize both the inner and outer TEBVs. The GQD-miR-150 conjugation as an intracellular gene delivery system was taken up by macrophages. Under the protection of GQDs, miR-150 was transfected into the cytosol, allowing continuous secretion of vascular endothelial growth factor (VEGF) via upregulation of HIF-1α protein expression, and promoted the migration of endothelial cells (ECs) in vitro. An in vivo study showed a rapid endothelialization of the inner TEBVs after transplantation for 7 days, especially a holonomic endothelial layer after 30 days. For the outer TEBVs, neovascularization (vasa vasorum) accompanied by nerve growth was observed around the adventitia on day 90. In conclusion, the designed cell bioreactor consisting of GQD-miR-engineered macrophages can effectively promote endothelialization and neuralization in vivo for TEBVs.

Construction of a small-caliber tissue-engineered blood vessel using icariin-loaded ß-cyclodextrin sulfate for in situ anticoagulation and endothelialization.

The rapid endothelialization of tissue-engineered blood vessels (TEBVs) can effectively prevent thrombosis and inhibit intimal hyperplasia. The traditional Chinese medicine ingredient icariin is highly promising for the treatment of cardiovascular diseases. ß-cyclodextrin sulfate is a type of hollow molecule that has good biocompatibility and anticoagulation properties and exhibits a sustained release of icariin. We studied whether icariin-loaded ß-cyclodextrin sulfate can promote the endothelialization of TEBVs. The experimental results showed that icariin could significantly promote the proliferation and migration of endothelial progenitor cells; at the same time, icariin could promote the migration of rat vascular endothelial cells (RAVECs). Subsequently, we used an electrostatic force to modify the surface of the TEBVs with icariin-loaded ß-cyclodextrin sulfate, and these vessels were implanted into the rat common carotid artery. After 3 months, micro-CT results showed that the TEBVs modified using icariin-loaded ß-cyclodextrin sulfate had a greater patency rate. Scanning electron microscopy (SEM) and CD31 immunofluorescence results showed a better degree of endothelialization. Taken together, icariin-loaded ß-cyclodextrin sulfate can achieve anticoagulation and rapid endothelialization of TEBVs to ensure their long-term patency.