Photografting and Patterning of Poly(ethylene glycol) Methacrylate Hydrogel on Glass for Biochip Applications.

An efficient procedure for chemical initiator-free, in situ synthesis of a functional polyethylene glycol methacrylate (PEG MA) hydrogel on regular glass substrates is reported. It is demonstrated that self-initiated photografting and photopolymerization driven by UV irradiation can yield tens of nanometer-thick coatings of carboxy-functionalized PEG MA on the aldehyde-terminated borosilicate glass surface. The most efficient formulation for hydrogel synthesis contained methyl methacrylic acid (MAA), 2-hydroxyethyl methacrylate (HEMA), and PEG methacrylate (PEG10MA) monomers (1:1:1). The resulting HEMA/PEG10MA/MAA (HPMAA) coatings had a defined thickness in the range from 11 to 50 nm. The physicochemical properties of the synthesized HPMAA coatings were analyzed by combining water contact angle measurements, stylus profilometry, imaging null ellipsometry, and atomic force microscopy (AFM). The latter technique was employed in the quantitative imaging mode not only for direct probing of the surface topography but also for swelling behavior characterization in the pH range from 4.5 to 8.0. The estimated high swelling ratios of the HPMAA hydrogel (up to 3.2) together with its good stability and resistance to nonspecific protein binding were advantageous in extracellular matrix mimetics via patterning of fibronectin (FN) at a resolution close to 200 nm. It was shown that the fabricated FN micropatterns on HPMAA were equally suitable for single-cell arraying, as well as controlled cell culture lasting at least for 96 h.

Mixed Self-Assembled Monolayers with Terminal Deuterated Anchors: Characterization and Probing of Model Lipid Membrane Formation.

We describe herein a series of self-assembled monolayers (SAMs) on gold designed for adjustable tethering of model lipid membrane phases. The SAMs consist of deuterated aliphatic anchors, HS(CH2)15CONH(CH2CH2O)6CH2CONH-X, where X is either -(CD2)7CD3 or -(CD2)15CD3, dispersed in a stable matrix of protein-repellent molecules, HS(CH2)15CONHCH2CH2OH. The mixed SAMs with variable surface densities of the anchors are thoroughly characterized before and after adsorption of phospholipids by means of ellipsometry, contact angle goniometry, and infrared reflection-absorption spectroscopy (IRRAS). In all cases, the bottom portions of the mixed SAMs (i.e., the h-alkyl thiol segments of the molecules) are arranged in a highly ordered all-trans conformation stabilized by a network of lateral hydrogen bonds. The terminal portions of the anchors (the oligo(ethylene glycol) spacer and deuterated alkyl segments, respectively), however, possess less ordered conformations in the mixed composition regime. For the SAMs containing the longer anchors (-(CD2)15CD3), the contact angle and infrared data point toward partial phase segregation. These findings are in excellent agreement with molecular dynamics simulations by Schulze and Stein. Upon analysis in air, the IRRAS data also indicate strong interaction between the adsorbed phospholipid molecules and the d-alkyl tails of the mixed SAM constituents. In such assemblies are the alkyl tails of the phospholipids aligned perpendicularly with respect to the supporting substrate, regardless of the anchor length. We also probed the in situ formation of a tethered bilayer lipid membrane (tBLM) via fusion of small unilamellar vesicles (SUVs) on the characterized SAMs using a quartz crystal microbalance with dissipation monitoring. Our experiments show that SUVs fuse efficiently of the two mixed SAMs with and without a pre-adsorbed lipid layer. Owing to the defined molecular composition and phase behavior, our SAM platform is attractive for detailed studies of tBLM formation and cell mimetic applications.

Functional fabrication of recombinant human collagen-phosphorylcholine hydrogels for regenerative medicine applications.

The implant-host interface is a critical element in guiding tissue or organ regeneration. We previously developed hydrogels comprising interpenetrating networks of recombinant human collagen type III and 2-methacryloyloxyethyl phosphorylcholine (RHCIII-MPC) as substitutes for the corneal extracellular matrix that promote endogenous regeneration of corneal tissue. To render them functional for clinical application, we have now optimized their composition and thereby enhanced their mechanical properties. We have demonstrated that such optimized RHCIII-MPC hydrogels are suitable for precision femtosecond laser cutting to produce complementing implants and host surgical beds for subsequent tissue welding. This avoids the tissue damage and inflammation associated with manual surgical techniques, thereby leading to more efficient healing. Although we previously demonstrated in clinical testing that RHCIII-based implants stimulated cornea regeneration in patients, the rate of epithelial cell coverage of the implants needs improvement, e.g. modification of the implant surface. We now show that our 500µm thick RHCIII-MPC constructs comprising over 85% water are suitable for microcontact printing with fibronectin. The resulting fibronectin micropatterns promote cell adhesion, unlike the bare RHCIII-MPC hydrogel. Interestingly, a pattern of 30µm wide fibronectin stripes enhanced cell attachment and showed the highest mitotic rates, an effect that potentially can be utilized for faster integration of the implant. We have therefore shown that laboratory-produced mimics of naturally occurring collagen and phospholipids can be fabricated into robust hydrogels that can be laser profiled and patterned to enhance their potential function as artificial substitutes of donor human corneas.

Long-chain alkylthiol assemblies containing buried in-plane stabilizing architectures.

A series of alkylthiol compounds were synthesized to study the formation and structure of complex self-assembled monolayers (SAMs) consisting of interchanging structural modules stabilized by intermolecular hydrogen bonds. The chemical structure of the synthesized compounds, HS(CH(2))(15)CONH(CH(2)CH(2)O)(6)CH(2)CONH-X, where X refers to the extended chains of either -(CH(2))(n)CH(3) or -(CD(2))(n)CD(3), with n = 0, 1, 7, 8, 15, was confirmed by NMR and elemental analysis. The formation of highly ordered, methyl-terminated SAMs on gold from diluted ethanolic solutions of these compounds was revealed using contact angle goniometry, null ellipsometry, cyclic voltammetry, and infrared reflection absorption spectroscopy. The experimental work was complemented with extensive DFT modeling of infrared spectra and molecular orientation. New assignments were introduced for both nondeuterated and deuterated compounds. The latter set of compounds also served as a convenient tool to resolve the packing, conformation, and orientation of the buried and extended modules within the SAM. Thus, it was shown that the lower alkyl portion together with the hexa(ethylene glycol) portion is stabilized by the two layers of lateral hydrogen bonding networks between the amide groups, and they provide a structurally robust support for the extended alkyls. The presented system can be considered to be an extension of the well-known alkyl SAM platform, enabling precise engineering of nanoscopic architectures on the length scale from a few to approximately 60 A for applications such as cell membrane mimetics, molecular nanolithography, and so forth.