13 research outputs found
Albumin Hydrogels Formed by Electrostatically Triggered Self-Assembly and Their Drug Delivery Capability
Biological
hydrogels are fundamentally biocompatible and have intrinsic
similarities to extracellular matrices in medical applications and
drug delivery systems. Herein we demonstrate the ability to form drug-eluting
protein hydrogels using a novel mechanism that involves the electrostatically
triggered partial denaturation and self-assembly of the protein via
changes in pH. Partial denaturation increases the protein’s
solvent exposed hydrophobic surface area, which then drives self-assembly
of the protein into a hydrogel within 10 min at 37 °C. We describe
the properties of an albumin hydrogel formed by this mechanism. Intrinsic
drug binding properties of albumin to all-trans retinoic acid (atRA)
are conserved through the partial denaturation process, as confirmed
by fluorescence quenching. atRA released from the hydrogel inhibited
smooth muscle cell migration as per an in vitro scratch wound assay.
Atomistic molecular dynamics and potential of mean force calculations
show the preservation and potential creation of new atRA binding sites
with a binding energy of −41 kJ/mol. The resulting hydrogel
is also biocompatible and exhibits rapid postgelation degradation
after its implantation in vivo. This interdisciplinary work provides
a new tool for the development of biocompatible protein hydrogel drug
delivery systems
Albumin Hydrogels Formed by Electrostatically Triggered Self-Assembly and Their Drug Delivery Capability
Biological
hydrogels are fundamentally biocompatible and have intrinsic
similarities to extracellular matrices in medical applications and
drug delivery systems. Herein we demonstrate the ability to form drug-eluting
protein hydrogels using a novel mechanism that involves the electrostatically
triggered partial denaturation and self-assembly of the protein via
changes in pH. Partial denaturation increases the protein’s
solvent exposed hydrophobic surface area, which then drives self-assembly
of the protein into a hydrogel within 10 min at 37 °C. We describe
the properties of an albumin hydrogel formed by this mechanism. Intrinsic
drug binding properties of albumin to all-trans retinoic acid (atRA)
are conserved through the partial denaturation process, as confirmed
by fluorescence quenching. atRA released from the hydrogel inhibited
smooth muscle cell migration as per an in vitro scratch wound assay.
Atomistic molecular dynamics and potential of mean force calculations
show the preservation and potential creation of new atRA binding sites
with a binding energy of −41 kJ/mol. The resulting hydrogel
is also biocompatible and exhibits rapid postgelation degradation
after its implantation in vivo. This interdisciplinary work provides
a new tool for the development of biocompatible protein hydrogel drug
delivery systems
Targeting Heparin to Collagen within Extracellular Matrix Significantly Reduces Thrombogenicity and Improves Endothelialization of Decellularized Tissues
Thrombosis within small-diameter vascular grafts limits the development
of bioartificial, engineered vascular conduits, especially those derived
from extracellular matrix (ECM). Here we describe an easy-to-implement
strategy to chemically modify vascular ECM by covalently linking a
collagen binding peptide (CBP) to heparin to form a heparin derivative
(CBP–heparin) that selectively binds a subset of collagens.
Modification of ECM with CBP–heparin leads to increased deposition
of functional heparin (by ∼7.2-fold measured by glycosaminoglycan
composition) and a corresponding reduction in platelet binding (>70%)
and whole blood clotting (>80%) onto the ECM. Furthermore, addition
of CBP–heparin to the ECM stabilizes long-term endothelial
cell attachment to the lumen of ECM-derived vascular conduits, potentially
through recruitment of heparin-binding growth factors that ultimately
improve the durability of endothelialization in vitro. Overall, our
findings provide a simple yet effective method to increase deposition
of functional heparin on the surface of ECM-based vascular grafts
and thereby minimize thrombogenicity of decellularized tissue, overcoming
a significant challenge in tissue engineering of bioartificial vessels
and vascularized organs
A Thermoresponsive Biodegradable Polymer with Intrinsic Antioxidant Properties
Oxidative stress in tissue can contribute
to chronic inflammation
that impairs wound healing and the efficacy of cell-based therapies
and medical devices. We describe the synthesis and characterization
of a biodegradable, thermoresponsive gel with intrinsic antioxidant
properties suitable for the delivery of therapeutics. Citric acid,
polyÂ(ethylene glycol) (PEG), and poly-<i>N</i>-isopropylacrylamide
(PNIPAAm) were copolymerized by sequential polycondensation and radical
polymerization to produce polyÂ(polyethylene glycol citrate-<i>co</i>-<i>N</i>-isopropylacrylamide) (PPCN). PPCN
was chemically characterized, and the thermoresponsive behavior, antioxidant
properties, morphology, potential for protein and cell delivery, and
tissue compatibility in vivo were evaluated. The PPCN gel has a lower
critical solution temperature (LCST) of 26 °C and exhibits intrinsic
antioxidant properties based on its ability to scavenge free radicals,
chelate metal ions, and inhibit lipid peroxidation. PPCN displays
a hierarchical architecture of micropores and nanofibers, and contrary
to typical thermoresponsive polymers, such as PNIPAAm, PPCN gel maintains
its volume upon formation. PPCN efficiently entrapped and slowly released
the chemokine SDF-1α and supported the viability and proliferation
of vascular cells. Subcutaneous injections in rats showed that PPCN
gels are resorbed over time and new connective tissue formation takes
place without signs of significant inflammation. Ultimately, this
intrinsically antioxidant, biodegradable, thermoresponsive gel could
potentially be used as an injectable biomaterial for applications
where oxidative stress in tissue is a concern
Copper Metal–Organic Framework Nanoparticles Stabilized with Folic Acid Improve Wound Healing in Diabetes
The
successful treatment of chronic nonhealing wounds requires
strategies that promote angiogenesis, collagen deposition, and re-epithelialization
of the wound. Copper ions have been reported to stimulate angiogenesis;
however, several applications of copper salts or oxides to the wound
bed are required, leading to variable outcomes and raising toxicity
concerns. We hypothesized that copper-based metal–organic framework
nanoparticles (Cu-MOF NPs), referred to as HKUST-1, which are rapidly
degraded in protein solutions, can be modified to slowly release Cu<sup>2+</sup>, resulting in reduced toxicity and improved wound healing
rates. Folic acid was added during HKUST-1 synthesis to generate folic-acid-modified
HKUST-1 (F-HKUST-1). The effect of folic acid incorporation on NP
stability, size, hydrophobicity, surface area, and copper ion release
profile was measured. In addition, cytotoxicity and <i>in vitro</i> cell migration processes due to F-HKUST-1 and HKUST-1 were evaluated.
Wound closure rates were assessed using the splinted excisional dermal
wound model in diabetic mice. The incorporation of folic acid into
HKUST-1 enabled the slow release of copper ions, which reduced cytotoxicity
and enhanced cell migration <i>in vitro</i>. <i>In
vivo</i>, F-HKUST-1 induced angiogenesis, promoted collagen deposition
and re-epithelialization, and increased wound closure rates. These
results demonstrate that folic acid incorporation into HKUST-1 NPs
is a simple, safe, and promising approach to control Cu<sup>2+</sup> release, thus enabling the direct application of Cu-MOF NPs to wounds
Repair of critical sized cranial defects with BMP9-transduced calvarial cells delivered in a thermoresponsive scaffold
<div><p>Large skeletal defects caused by trauma, congenital malformations, and post-oncologic resections of the calvarium present major challenges to the reconstructive surgeon. We previously identified BMP-9 as the most osteogenic BMP in vitro and in vivo. Here we sought to investigate the bone regenerative capacity of murine-derived calvarial mesenchymal progenitor cells (iCALs) transduced by BMP-9 in the context of healing critical-sized calvarial defects. To accomplish this, the transduced cells were delivered to the defect site within a thermoresponsive biodegradable scaffold consisting of poly(polyethylene glycol citrate-co-N-isopropylacrylamide mixed with gelatin (PPCN-g). A total of three treatment arms were evaluated: PPCN-g alone, PPCN-g seeded with iCALs expressing GFP, and PPCN-g seeded with iCALs expressing BMP-9. Defects treated only with PPCN-g scaffold did not statistically change in size when evaluated at eight weeks postoperatively (p = 0.72). Conversely, both animal groups treated with iCALs showed significant reductions in defect size after 12 weeks of follow-up (BMP9-treated: p = 0.0025; GFP-treated: p = 0.0042). However, H&E and trichrome staining revealed more complete osseointegration and mature bone formation only in the BMP9-treated group. These results suggest that BMP9-transduced iCALs seeded in a PPCN-g thermoresponsive scaffold is capable of inducing bone formation in vivo and is an effective means of creating tissue engineered bone for critical sized defects.</p></div
Schematic representation of the overall experimental design.
<p>(A) Murine calvarial cells (CALs) were immortalized via retrovirally introduced SV40 large T antigen to produce iCALs. iCALs were then transduced with BMP9 via adenoviral vector and mixed with PPCN scaffolding material. (B) qPCR analysis demonstrating relative expression of BMP9 in iCALs infected with ad-BMP9 (blue bars) compared to control (ad-GFP)-infected cells (red bars). Gene transcript expression was normalized against GAPDH expression. (C) The mixture was subsequently tested in our murine craniofacial defect model. Four millimeter diameter full-thickness calvarial defects were created in the left parietal bone of 8-week-old male athymic (nu/nu) mice. The newly created empty defect reveals the underlying dura mater. PPCN alone or PPCN and adenovirally transduced immortalized calvarial cells were used to fill the defect site.</p
Time-course microCT imaging of the calvarial defects.
<p>At 24–48 hours postoperatively, baseline microCT imaging was performed and analyzed to determine defect volume. Follow-up imaging and analysis was performed at 2, 4, 6, 8, and 12 weeks postoperatively to quantify residual defect volume and new bone ossification. Representative images are shown.</p
Assessment of the Trichrome histology.
<p>Histologic analysis of tissue microsections harvested 12 weeks post-treatment. (A, B) Chondroid matrix and a smaller proportion of mature bone is shown in the defect site of a PPCN + iCAL + AdGFP-treated mouse. (C, D) A visibly higher proportion of mature bone can be appreciated in samples taken from the defect sites treated with PPCN + iCAL + AdBMP9.</p
Histologic evaluation of the BMP9-induced calvarial defect repair.
<p>Histologic analysis of tissue microsections harvested 12 weeks post-treatment. Yellow arrows indicate the original defect borders. (A) The defect site of this AdGFP-treated mouse shows incomplete healing; there is some ingrowth of bone, but fibrous tissue fills part of the defect. (B) Conversely, the defect site of an AdBMP9-treated mouse has been completely bridged with new bone. All sections show no trace of PPCN material, indicating complete resorption.</p