three-dimensional-culture-vascularized-thermogenic-adipose-tissue

Three-Dimensional Culture of Vascularized Thermogenic Adipose Tissue from Microvascular Fragments

Engineering thermogenic adipose tissue (e.g., beige or brown adipose tissues) has been investigated as a potential therapy for metabolic diseases or for ...

Injectable hydrogels represent a valuable tool for the delivery of therapeutic molecules aimed to restore the functionality of damaged tissues. In this study, we report the design of a nanocomposite DNA-based hydrogel crosslinked with oxidized alginate (OA) via the formation of reversible imine linkages. The formulated hydrogel functioned as an injectable carrier for the sustained delivery of a small molecule drug, simvastatin. The degree of oxidation of alginate and the concentration of silicate-based nanoparticles (nSi) were varied to modulate the rheological properties of the hydrogels. Specifically, the formulations consisting of OA with higher degree of oxidation displayed the highest value of storage moduli, yield stress, yield strain, and rapid recovery after removal of cyclic stress. The hydrogel formulations exhibited self-healing and shear-thinning properties due to the reversible nature of the covalent imine bonds formed between the aldehyde groups of OA and the amine groups present in the DNA nucleotides. Moreover, the incorporation of charged nSi further enhanced the shear strength of the formulated hydrogels by establishing electrostatic interactions with the phosphate groups of the DNA network. The optimized hydrogel was able to promote the sustained release of simvastatin for more than a week. The bioactivity of the released drug was confirmed by testing its ability to induce osteogenic differentiation and migration of human adipose-derived stem cells in vitro. Overall, the results obtained from this study demonstrate that DNA could be used as a natural biopolymer to fabricate self-healing injectable hydrogels with sustained release properties for minimally invasive therapeutic approaches. STATEMENT OF SIGNIFICANCE: Dynamic covalent chemistry, especially Schiff base reactions have emerged as a promising route for the formation of injectable hydrogels. Our study demonstrated the development of a DNA-based self-healing hydrogel formed via Schiff base reaction occurring at physiological conditions. The hydrogels functioned as sustained delivery vehicles for the hydrophobic drug simvastatin, which requires a polymeric carrier for controlled delivery of therapeutic concentrations of the drug without exhibiting cytotoxic effects. Presently available hydrogel-based drug delivery systems encounter major challenges for the delivery of hydrophobic drugs due to the hydrophilic nature of the base matrix. Our strategy presents a platform technology for the design of minimally invasive approaches for the sustained delivery of hydrophobic drugs similar to simvastatin.

Self-healing DNA-based injectable hydrogels with reversible covalent linkages for controlled drug delivery.

The development of ectopic adipose tissue in skeletal muscle is associated with several skeletal muscle and metabolic pathologies, including Type II Diabetes Mellitus. The adipogenic differentiation of muscle precursor cells (MPCs) has been postulated to occur in skeletal muscle in vivo in a three-dimensional (3-D) configuration; therefore, it is appropriate to investigate this phenomenon using 3-D matrices in vitro. The capacity for MPC adipogenic differentiation in a 3-D environment was investigated in fibrin hydrogels by treating MPCs derived from healthy or diabetic animals with adipogenic induction medias that differed in their ability to increase lipid accumulation and activate the expression of genes associated with adipogenic differentiation (peroxisome proliferator-activated receptor gamma (PPARG), adiponectin (ADIPOQ), and fatty acid synthase (FAS)). The capacity for adipogenic differentiation was diminished, but not prevented, if myogenic differentiation preceded MPC exposure to adipogenic induction conditions. Conversely, adipogenic differentiation was greater in hydrogels containing MPCs from diabetic rats as compared to those derived from lean rats, as evidenced by an increase in lipid accumulation and adipogenic gene expression. Collectively, the data herein support a role for the MPCs in adipogenesis in a 3-D environment and that they may contribute to the ectopic accumulation of adipose tissue. The observation that the potential for adipogenic differentiation is maintained even after a period of myogenic differentiation alludes to the possibility that adipogenesis may occur during different phases of muscle development. Finally, the increase in adipogenic differentiation in hydrogels containing MPCs derived from diabetic animals provides strong evidence that a pathological environment in vivo increases their capacity for adipogenesis.

Divergent effects of myogenic differentiation and diabetes on the capacity for muscle precursor cell adipogenic differentiation in a fibrin matrix.

Skeletal muscle is a tissue that is directly involved in the progression and persistence of type 2 diabetes (T2D), a disease that is becoming increasingly common. Gaining better insight into the mechanisms that are affecting skeletal muscle dysfunction in the context of T2D has the potential to lead to novel treatments for a large number of patients. Through its ability to emulate skeletal muscle architecture while also incorporating aspects of disease, tissue-engineered skeletal muscle (TE-SkM) has the potential to provide a means for rapid high-throughput discovery of therapies to treat skeletal muscle dysfunction, to include that which occurs with T2D. Muscle precursor cells isolated from lean or obese male Zucker diabetic fatty rats were used to generate TE-SkM constructs. Some constructs were treated with adipogenic induction media to accentuate the presence of adipocytes that is a characteristic feature of T2D skeletal muscle. The maturity (compaction and creatine kinase activity), mechanical integrity (Young's modulus), organization (myotube orientation), and metabolic capacity (insulin-stimulated glucose uptake) were all reduced by diabetes. Treating constructs with adipogenic induction media increased the quantity of lipid within the diabetic TE-SkM constructs, and caused changes in construct compaction, cell orientation, and insulin-stimulated glucose uptake in both lean and diabetic samples. Collectively, the findings herein suggest that the recapitulation of structural and metabolic aspects of T2D can be accomplished by engineering skeletal muscle .

Diabetic Conditions Confer Metabolic and Structural Modifications to Tissue-Engineered Skeletal Muscle.

Cryogels are a particular type of hydrogels that possess great potential in both fields of drug delivery and tissue engineering. Based on these premises, the goal of this work was to develop a cytocompatible polymeric cryogel, which could be used as a spongy scaffold to promote the delivery of biomolecules. Precisely, the novel formulation was fabricated by combining dextran methacrylate (DEX-MA) and polyethylene glycol dimethacrylate (PEG-DMA) through radical polymerization at a temperature of -15 °C. The swelling, porosity, mechanical properties, and the drug release profile of vitamin B12 from the optimized cryogel were evaluated and compared to hydrogels fabricated at room temperature. The use of the cryo-gelation technique enabled the formation of scaffolds with improved swelling, increased interconnected porosity, and higher mechanical resistance than conventional hydrogels. The cryogels proved to be non-toxic and suitable carriers for the delivery of water-soluble biomolecules. Overall, the novel cytocompatible cryogel formulation could be used for biomedical applications that require the need of a macroporous scaffold for localized delivery of bioactive molecules.

Dextran-polyethylene glycol cryogels as spongy scaffolds for drug delivery.

Hydrogels can be fabricated and designed to exert direct control over stem cells' adhesion and differentiation. In this study, we have investigated the use of polydopamine (pDA)-treatment as a binding platform for bioactive compounds to create a versatile gelatin-alginate (Gel-Alg) hydrogel for tissue engineering applications. Precisely, pDA was used to modify the surface properties of the hydrogel and better control the adhesion and osteogenic differentiation of human adipose-derived stem cells (hASCs). pDA enabled the adsorption of different types of bioactive molecules, including a model osteoinductive drug (dexamethasone) as well as a model pro-angiogenic peptide (QK). The pDA treatment efficiently retained the drug and the peptide compared to the untreated hydrogel and proved to be effective in controlling the morphology, cell area, and osteogenic differentiation of hASCs. Overall, the findings of this study confirm the efficacy of pDA treatment as a valuable strategy to modulate the biological properties of biocompatible Gel-Alg hydrogels and further extend their value in regenerative medicine.

Investigation of human adipose-derived stem-cell behavior using a cell-instructive polydopamine-coated gelatin-alginate hydrogel.

This work investigates a sequential strategy to develop DNA-based hydrogel scaffolds with interpenetrating polymeric network. The scaffolds were formed via a two-step procedure. First, a covalently cross-linked DNA-based cryogel was formed by the chemical reaction between DNA strands and a bifunctional cross-linker, polyethylene glycol diepoxide at subzero temperatures. In the second step, alginate chains were absorbed into the preformed macroporous DNA cryogel network, followed by ionic cross-linking with divalent calcium ions. The individual and synergistic effects of covalent and ionic cross-linkings on mechanical and physical properties of the IPN cryogel were tested. The IPN cryogels were able to sustain large deformations higher than 95% of strain under compressive forces without exhibiting any failure. Addition of a physically cross-linked alginate network to the covalently linked DNA cryogel significantly enhanced its toughness and energy dissipation compared to the covalent network alone. The formulated hydrogels also exhibited excellent biocompatibility with human stem cells. Overall, this DNA-based IPN cryogel has the potential to be used as a biomaterial scaffold for a diverse range of tissue engineering applications.

Article	Year
Self-healing DNA-based injectable hydrogels with reversible covalent linkages for controlled drug delivery. Acta biomaterialia\| PubMed ID: 31972367	2020
Divergent effects of myogenic differentiation and diabetes on the capacity for muscle precursor cell adipogenic differentiation in a fibrin matrix. Biochemical and biophysical research communications\| PubMed ID: 32192775	2020
Diabetic Conditions Confer Metabolic and Structural Modifications to Tissue-Engineered Skeletal Muscle. Tissue engineering. Part A\| PubMed ID: 32878567	2021
Dextran-polyethylene glycol cryogels as spongy scaffolds for drug delivery. International journal of biological macromolecules\| PubMed ID: 33161086	2021
Investigation of human adipose-derived stem-cell behavior using a cell-instructive polydopamine-coated gelatin-alginate hydrogel. Journal of biomedical materials research. Part A\| PubMed ID: 34189837	2021
Fabricating Tough Interpenetrating Network Cryogels with DNA as the Primary Network for Biomedical Applications. ACS macro letters\| PubMed ID: 35638638	2020

Settimio Pacelli

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