Hee-Cheol Kim

Autocrine Purinergic Receptor Signaling Is Essential for Macrophage Chemotaxis

Amplification of outside-in chemotactic signaling by inside-out purinergic signaling drives macrophage migration. Self-Help Migration Immune cells such as neutrophils and macrophages migrate to sites of infection or inflammation by following gradients of chemoattractants. These include chemokines, which can be released by other cells at the target site; components of the complement system, such as C5a; and bacterial products, such as the formylated peptide, fMLP. While they navigate along the chemoattractant gradient toward their destination, cells are also exposed to other signals, some of which may compete with the chemoattractant that the cells were already following. Another level of complexity in the regulation of cell migration came from the discovery that migrating neutrophils release adenosine triphosphate (ATP), which then functions in an autocrine fashion through the purinergic receptor P2Y2 to enhance migration; cells deficient in P2Y2 have impaired gradient sensing. Kronlage et al. provide evidence that autocrine ATP signaling is also required for the migration of macrophages in vitro and in an in vivo model. The authors found that more than one type of ATP receptor type as well as metabolites of ATP contributed to the migratory responses of macrophages; furthermore, ATP was not released through pannexin-1 proteins, as has been suggested for neutrophils. Together, these data suggest that autocrine purinergic receptor signaling may play a general role in regulating the chemotactic responses of immune cells. Chemotaxis, the movement of cells along chemical gradients, is critical for the recruitment of immune cells to sites of inflammation; however, how cells navigate in chemotactic gradients is poorly understood. Here, we show that macrophages navigate in a gradient of the chemoattractant C5a through the release of adenosine triphosphate (ATP) and autocrine “purinergic feedback loops” that involve receptors for ATP (P2Y2), adenosine diphosphate (ADP) (P2Y12), and adenosine (A2a, A2b, and A3). Whereas macrophages from mice deficient in pannexin-1 (which is part of a putative ATP release pathway), P2Y2, or P2Y12 exhibited efficient chemotactic navigation, chemotaxis was blocked by apyrase, which degrades ATP and ADP, and by the inhibition of multiple purinergic receptors. Furthermore, apyrase impaired the recruitment of monocytes in a mouse model of C5a-induced peritonitis. In addition, we found that stimulation of P2Y2, P2Y12, or adenosine receptors induced the formation of lamellipodial membrane protrusions, causing cell spreading. We propose a model in which autocrine purinergic receptor signaling amplifies and translates chemotactic cues into directional motility.

Two-photon absorption-controlled multidose drug release: a novel approach for secondary cataract treatment.

Tens of millions of cataract surgeries are done every year and the number is increasing heavily. Posterior capsule opacification is the major postoperative complication with an incidence of 10 to 50% within 5 years, depending on the age of the patient. We present a novel approach for secondary cataract treatment in a noninvasive manner. Photochemically triggered drug release from a polymer enables repeated drug applications for cataract treatment years after implantation of the intraocular lens, just when needed. However, light in the visible spectral range must pass through the lens but must not induce drug release. We demonstrate that two-photon absorption photochemistry is a powerful tool to overcome this problem. With wavelengths in the visible regime, a photochemical reaction that requires energies in the UV is triggered. The high intensities needed for this process never occur in any lighting condition in daily lives, but may be easily obtained with focused laser beams routinely used in ophthalmology. The properties of the therapeutic system are specified and the function is demonstrated by in-vitro cell tests. Noninvasive multidose photochemically triggered drug release from implanted intraocular lenses carrying a drug depot may be a therapeutic as well as an economic choice to established treatments of secondary cataracts.

Synthesis and Photochemistry of Coumarin-Based Self-Assembled Monolayers on Silicon Oxide Surfaces

In this study, we report on a system consisting of self-assembled monolayers (SAMs) formed by 7-(11-trichlorosilylundecyloxy)coumarin on mica and on quartz glass. For the first time, in the absence of an inert atmosphere or a stabilizing matrix, we demonstrate by means of absorption and fluorescence spectroscopy that the photochemical cycloaddition of coumarin head groups is completely reversible in SAMs. Photodimerization and photocleavage were monitored for five cycles of alternating irradiation with light of wavelengths 355 and 254 nm, respectively. SAM formation was analyzed using atomic force microscopy and contact angle measurements. The quantum yield of the single photon absorption induced photocleavage of coumarin dimers in a SAM was determined to be Phi = 0.29. Asymmetrical photocleavage after lactone ring-opening of the coumarin dimer SAM causes a change in contact angle from about 70 degrees to about 55 degrees. This may be observed, for example, as a significant change in surface wettability.

Self-organization of multifunctional surfaces--the fingerprints of light on a complex system.

Nanocomposite patterns and nanotemplates are generated by a single-step bottom-up concept that introduces laser-induced periodic surface structures (LIPSS) as a tool for site-specific reaction control in multicomponent systems. Periodic intensity fluctuations of this photothermal stimulus inflict spatial-selective reorganizations, dewetting scenarios and phase segregations, thus creating regular patterns of anisotropic physicochemical properties that feature attractive optical, electrical, magnetic, and catalytic properties.

Photochemical preparation of sub-wavelength heterogeneous laser-induced periodic surface structures.

Laser-induced periodic surface structures (LIPSS) are a phenomenon caused by interaction of light with solid surfaces. We present a photochemical concept which uses LIPSS-related light intensity patterns for the generation of heterogeneous nanostructures. The process facilitates arbitrary combinations of substrate and LIPSS-pattern materials. An efficient method for the generation of organometallic hybrid-nanowire arrays on porous anodic aluminum oxide is demonstrated.

Molecular origin of the stability of violuric acid radicals at high pH-values

Abstract Violuric acid forms an iminoxy radical by electrochemical oxidation which shows an astonishing lifetime of more than 6 h at pH 2 and still of about 0.5 h at pH 12 in aqueous solution without any observable dimer formation. The stabilization of the radical over such a wide pH-range is related to amino groups in the ring which cause at high pH-values a negatively charged radical shielded from nucleophilic attacks. This finding is based on the comparison of the pH dependence of the radical lifetime of violuric acid and N , N ′ -dimethylvioluric acid which were both isolated in solid form.

Origin of the mediator losses in electrochemical delignification processes: primary and secondary reactions of violuric acid and N,N′-dimethylvioluric acid radicals with lignin model compounds

The use of mediator-based processes for the delignification of wood pulps is of great interest as they promise to be environmentally as well as economically interesting alternatives to the currently dominating chlorine-based processes. Mediator-based processes suffer from the fact that substantial amounts of the mediator molecules are irreversibly lost during the reactions. We have now analyzed in detail the primary and in addition the secondary reactions of the oxime type mediators violuric acid and dimethylvioluric acid with lignin model compounds and softwood pulp in order to find the reasons for the repeatedly reported loss of mediator molecules during enzymatic and electrochemical delignification processes. Using a set of lignin model compounds, representing the various structural subunits in the lignin network, we were able to demonstrate that the loss of mediator is not due to the primary reaction of the mediator radicals with the lignin but occurs in a secondary reaction where the activated lignin subunits react with the mediators in the solution. The primary reaction of the mediator radicals is hydrogen abstraction from phenols and activated aromatic rings which leads to the formation of phenoxyl and phenyl radicals. The secondary reaction is the formation of colored semi-stable covalent adduct molecules from an activated lignin subunit with a mediator molecule in the solution leading to polycyclic N-hydroxy compounds. Only in this secondary reaction mediator molecules are removed from the solution due to covalent attachment to the lignin network. No reaction with the cellulose content in the pulp is observed. The findings reported in this paper point the way to an improved mediator design and appropriately modified processes, where the secondary mediator reactions are suppressed, and removes a major road block towards a technical application of the mediator-based delignification procedures.

High refractive index TiO2-PHEMA hydrogel for ophthalmological applications

A facile route for the synthesis of PHEMA hydrogel incorporating anatase nanoparticles is presented. The hydrogel features a refractive index of 1.527 and high transparency throughout the visual spectral range. Saturn rings have been machined in order to investigate the workability of the material and its potential applicability as intraocular lens implant. The resulting lenses feature high surface quality, foldability, and shape memory which makes them suitable for implantation via extracapsular cataract extraction.

Two-photon absorption-induced drug delivery from polymers for medical applications

Novel polymeric materials carrying a drug depot have been developed which are suitable for fabrication of photochemically modulated drug delivery devices. In order to avoid uncontrolled drug release the drug is covalently attached to the polymer backbone using a photo-active linker. Controlled drug release from the polymer can be accomplished either via single-photon excitation or by two-photon absorption (TPA). In particular the second possibility is of interest for applications where exposure to day light or UV light may not be omitted. One example are polymeric intraocular lenses (IOL), which are implanted instead of the opaque natural lens during cataract surgery. Secondary cataract formation is quite often observed after implantation of polymeric IOLs. In this study the well known cell toxic agent 5-fluorouracil (5FU) attached to a methylmethacrylate-based polymer was investigated as an IOL which can upon photochemical excitation release 5FU in order to treat or to prevent secondary cataract formation. The photochemical cleavage of the linker molecule was analyzed with single- and two-photon excitation. UV/VIS spectroscopy and HPLC analysis confirmed the release of 5FU form the polymer backbone. The diffusion of the drug precursor out from the polymer as well as the hydrolysis of the drug precursor which leads to 5FU formation were investigated in vitro.

Materials for intraocular lenses enabling photo-controlled tuning of focal length in vivo

Typical postoperative complications in cataract surgery are that refractive power and curvature of the implanted intraocular lens (IOL) do not have optimum values, requiring the patient to wear viewing aids. This is mainly because biometric data relevant for calculation of the IOLs shape cannot be determined with the required precision. Hence, there is a need for methods to tune the focal length postoperatively in a non-invasive manner. We have developed polymers where we can induce a change in refractive index by linking or cleaving bonds between a su.ciently large number of side groups of the polymer main chain in a photoinduced cycloaddition or cycloreversion reaction, respectively. These photoreactions lead to a change in refractive index great enough to be interesting for the concept of in vivo tunable IOLs. The photochemical reaction can be triggered by a two-photon process (TPA) using a pulsed laser system, i.e. the energy required for bond breaking is provided by two photons in the visible range. This is important because light in the UV cannot induce undesired changes of the refractive index owing to the strong UV-absorption of the cornea. Undesired changes due to light in the visible range of the spectrum are unlikely to happen because photon density of sun light is much too low for TPA. Due to the excellent spatial resolution that can be achieved with two-photon processes one cannot only modify the refractive index of the entire lens but also selectively in well defined areas enabling to correct for aberrations such as astigmatism. Here, we present new polymers that do not only exhibit a photo induced change of refractive index great enough to induce a change of focal length of more than two diopters in a standard IOL. These new polymers have also significantly improved material properties with respect to the fabrication of the IOL and the TPA-sensitivities and the light energy required to induce the refractive index change.