Skin tissue is regenerated by the combinational function of skin cells, extracellular matrix (ECM), and bioactive molecules. As an artificial ECM, supramolecular hydrogels exhibited outstanding capability to mimic the physical properties of ECM. However, the lack of biochemical function in supramolecular hydrogels has limited further tissue engineering applications. Here, they developed self-assembling supramolecular drug delivery hydrogels to mimic the skin tissue regeneration process. The supramolecular hydrogels were prepared to encapsulate fibroblasts by the host–guest interaction of cyclodextrin-modified gelatin (GE-CD) and adamantane-modified hyaluronate (Ad-HA) in conjugation with human growth hormone (hGH) for accelerated skin tissue regeneration. In vitro, GE-CD/Ad-HA-hGH hydrogels showed highly facilitated cell growth by the controlled hGH delivery. After a subcutaneous injection into the back of mice, IVIS imaging of bioengineered fibroblasts to express red fluorescence protein (RFP) revealed prolonged cell survival and proliferation in the supramolecular hydrogels for more than 21 days. They could also observe the improved skin tissue regeneration by the facilitated fibroblast proliferation with angiogenesis. Taken together, they could confirm the feasibility of biomimetic supramolecular drug delivery GE-CD/Ad-HA-hGH hydrogels for various tissue engineering applications.
Tissue regeneration is based on the signal transduction between cells and extracellular matrix (ECM) via growth factors and hormones.Tissue engineering scaffolds mimicking cellular mechanisms in the body have been widely investigated for tissue regeneration. In particular, fibroblasts encapsulated in ECM play an important role in skin regeneration, activating other cells with relevant protein production. The composition and structure of biomolecules determine the physical and biochemical properties of ECM, which affect the fibroblast cellular behaviors. Collagen (Col) and hyaluronate (HA) are the primary biopolymer backbones to form skin tissue ECM with different physicochemical characteristics. The RGD peptide in Col regulates cellular behavior and morphology. Proteins, including growth hormones (GHs) and factors, stimulate cell proliferation and tissue regeneration. Accordingly, tissue engineering scaffolds mimicking not only structural characteristics but also biochemical components such as RGD and proteins of ECM are strongly needed for accelerated skin tissue regeneration.
Schematic illustration of the biomimetic supramolecular GE-CD/Ad-HA-hGH hydrogel for accelerated skin tissue regeneration.
Supramolecular host-guest assembled hydrogels have been suggested as a cell delivery scaffold for tissue regeneration. Non-toxic and reversible strong interactions between host and guest molecules enable high viability of encapsulated cells and minimally invasive transplantation of hydrogels via injection. However, the structure and biochemical cues of skin tissue ECM were not fully mimicked in supramolecular hydrogels.
Characterization of Ad-HA-hGH. (a) HPLC with 1H NMR (inset images) of Ad-HA-hGH (upper) and Ad-HA (bottom), (b) circular dichroism, and (c) ELISA/ Bradford assay. (d) Proliferation rate of Nb2-11 cells stimulated by hGH and Ad-HA-hGH. (e) Serum stability of hGH and Ad-HA-hGH (n = 4). (f) In vitro release of hGH and Ad-HA-hGH from the GE-CD/Ad-HA hydrogels (n = 4). The remaining hGH in the hydrogels was quantified after degradation by adding excess CD solution. (g) Fluorescence images of FITC-modified hGH, Ad-HA, and Ad-HA-hGH taken up into fibroblast cells (scale bars: 200 μm). (h) Relative quantification of the hGH, Ad-HA, and Ad-HA-hGH taken up by the analysis of fluorescence intensity (n = 4).
The complicated structure and low solubility of Col prohibit the conjugation of host–guest molecules to the polymer backbone and the development of supramolecularly crosslinked hydrogels. Meanwhile, the delivery of proteins to cells in the hydrogels has suffered from the burst release, low stability, and low cellular uptake. To overcome these problems, proteins are tethered to the hydrogel scaffolds, loaded into nanoparticles,or micelles,for the sustained release of proteins from the hydrogels and the increased stability under physiological conditions. However, there are strong unmet needs for the triggered and controlled protein delivery for tissue engineering applications.
(a) Confocal Z-stack fluorescent images of live/dead assay of stained fibroblasts in supramolecular hydrogels on day 4 and 10 (scale: 600 × 600 × 300 μm3). (b) Quantitative analysis of cell viability in confocal fluorescent images (n = 5). (c) Proliferation rate of fibroblasts encapsulated in supramolecular hydrogels by CCK-8 analysis for 10 days (n = 4).
Here, they developed biomimetic supramolecular hydrogels using gelatin (GE) and HA with hGH conjugation for skin tissue regeneration. GE is a hydrolyzed form of Col with increased solubility, showing similar structural and biological properties. Supramolecular hydrogels were prepared by the host–guest interaction between β-cyclodextrin-modified GE (GE-CD) and adamantane-modified HA (Ad-HA). HGH conjugated to Ad-HA (Ad-HA-hGH) was continuously released from GE-CD/Ad-HA-hGH hydrogels with improved cellular uptake. Supramolecular GE-CD/Ad-HA-hGH hydrogels were characterized and assessed for facilitated cell proliferation and growth by the controlled release of hGH. After that, they assessed the feasibility of biomimetic supramolecular hydrogels for skin tissue regeneration in mice with sufficient mechanical strength to retain fibroblast cells and biochemical environment with RGD and hGH.
(a) Confocal Z-stack (600 × 600 × 300 μm3) and cross-section (middle and bottom of hydrogels) images of fibroblasts (blue: nuclei, green: F-actin, scale bars: 50 μm). (b, c) The ratio of cell numbers distributed in the hydrogels determined by the quantification analysis for 150 × 150 μm2 on confocal images on days 4 and 10 (n = 7).
(a) IVIS in vivo images of transplanted fibroblast cells in supramolecular hydrogels for 21 days. (b) Relative fluorescence intensity reflecting the cell proliferation of transplanted cells for 21 days (n = 4).
(a) IVIS in vivo images of transplanted fibroblast cells in supramolecular hydrogels for 21 days. (b) Relative fluorescence intensity reflecting the cell proliferation of transplanted cells for 21 days (n = 4).
Biomimetic Supramolecular Drug Delivery Hydrogels for Accelerated Skin Tissue Regeneration Sang Hoon Jeong, Mungu Kim, Tae Yeon Kim, Hyunsik Choi, and Sei Kwang Hahn ACS Biomaterials Science & Engineering Article ASAP DOI: 10.1021/acsbiomaterials.1c00705
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