Transition metals (TM) are essential microelements with various biological functions demanded in tissue regeneration applications. Little is known about therapeutic potential of TM within soft hydrogel biomaterials. The soluble TM, such as Zn, Cu, Mn and Co, were stably incorporated into gelatin network during cryogelation. TM content in the resultant cryogels varied from 0.1×10 3 to 11.8×10 3 ppm, depending on the initial TM type and concentration in reaction solution. Zn was uniformly complexed with the gelatin scaffold according to elemental imaging, increasing the swelling of polymer walls and the G'/G'' values and also decreasing the size of cryogel macro-pores Zn-doped cryogels supported migration of human skin fibroblasts (HSF); only upper Zn content of 11.8×10 3 ppm in the scaffold caused c.a. 50% inhibition of cell growth. Zn ions solubilized in culture medium were more active towards HSF (IC50 ≈ 0.3 mM). Treatment of splinted full-skin excisional wounds in rats with the Zn-doped and non-doped cryogels showed that Zn considerably promotes d passing inflammatory/proliferation phases of healing process, inducing more intense dermis formation and structuration. The results show the feasibility of development of cryogel based formulations with different TM and support high phase-specific ability of the Zn-gelatin cryogels to repair acute wounds.
Oligopeptides
are versatile cell modulators resembling pleiotropic activities of
ECM proteins and growth factors. Studying the role of cell-instructive
peptide signals within 3D scaffolds, yet poorly known, requires effective
approaches to introducing bioactive sequences into appropriate materials.
We synthesized RGD and GHK motif based peptides 1 and 2 linked to the terminal adamantyl group (Ad) and their fluorescent
derivatives 3 and 4. Poly(hydroxyethyl methacrylate)
(pHEMA) cryogels with additional PEG/β-cyclodextrin (CD) units
were prepared as an inert macroporous scaffold capable to bind the
adamantylated peptides via affinity CD-Ad complexation. According
to toluidine blue staining, the CD moieties were effectively and stably
incorporated in the pHEMA cryogels at nanomolar amounts per milligram
of material. The CD component gradually increased the thickness and
swelling ability of the polymer walls of cryogels, resulting in a
noticeable decrease in macropore size and modulation of viscoelastic
properties. The labeled peptides exhibited fast kinetics of specific
binding to the CD-modified cryogels and were simultaneously immobilized
by coincubation. The peptide loading approached ca. 0.31 mg per cm2 of cryogel sheet. A well-defined mitogenic effect of the
immobilized peptides (2 < 1≪ 1 + 2) was revealed toward 3T3 and PC-12 cells. The synergistic
action of RGD and GHK peptides induced a profound change in cell behavior/morphology
attributed to a growth-factor-like activity of the composition. Altogether,
our results provide an effective procedure for the preparation of
CD-modified pHEMA cryogels and their uniform in situ functionalization
with bioactive peptide(s) of interest and an informative study of
cellular responses in the functionalized scaffolds.
A new self-assembled formulation of methylprednisolone succinate (MPS) based on a carboxylated trifunctional block copolymer of ethylene oxide and propylene oxide (TBC-COOH) was developed. TBC-COOH and MPS associated spontaneously at increased concentrations in aqueous solutions to form almost monodisperse mixed micelles (TBC-COOH/MPS) with a hydrodynamic diameter of 19.6 nm, zeta potential of -27.8 mV and optimal weight ratio ∼1:6.3. Conditions for the effective formation of TBC-COOH/MPS were elucidated by comparing copolymers and glucocorticoids with different structure. The micellar structure of TBC-COOH/MPS persisted upon dilution, temperature fluctuations and interaction with blood serum components. TBC-COOH increased antiradical activity of MPS and promoted its intrinsic cytotoxicity in vitro attributed to enhanced cellular availability of the mixed micelles. Intracellular transportation and hydrolysis of MPS were analyzed using optimized liquid chromatography tandem mass spectrometry with multiple reaction monitoring which showed increased level of both MPS and methylprednisolone in neuronal cells treated with the formulated glucocorticoid. Our results identify TBC-COOH/MPS as an advanced in situ prepared nanoformulation and encourage its further investigation for a potential local glucocorticoid therapy.
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