Fulvio Ratto

Photothermally-induced disordered patterns of corneal collagen revealed by SHG imaging.

The loss of organization of the corneal collagen lattice induced by photothermal effects was analyzed by using second-harmonic generation (SHG) imaging. Porcine cornea samples were treated with low-power laser irradiation in order to get localized areas of tissue disorganization. The disorder induced within the irradiated area of corneal stroma was quantified by means of Discrete Fourier Transform, auto-correlation and entropy analyses of the SHG images. Polarization modulated SHG measurements allowed to probe the changes in the structural anisotropy of sub-micron hierarchical levels of the stromal collagen. Our results emphasize the great potential of the SHG imaging to detect subtle modifications in the collagen assembly. The proposed analytical methods may be used to track several genetic, pathologic, accidental or surgical-induced disorder states of biological tissues.

Chitosan Films Doped with Gold Nanorods as Laser-Activatable Hybrid Bioadhesives

Chitosan is a linear chain polysaccharide comprising glucosamine and N -acetyl glucosamine residues joined together by β -1,4-glycosidic bonds. It is derived as an acids-soluble material by deacetylation of chitin, which is commonly found in the outer skeleton of crustaceans and in the cell walls of fungi. A variety of fundamental properties such as excellent biocompatibility and biodegradability make chitosan a very attractive material for biomedical applications including wound dressing, tissue engineering and drug delivery. [ 1 , 2 ] The peculiar tendency of chitosan to form films with high mechanical strength, good elasticity and rather slow biodegradation is among the principal reasons why this material is gaining a primary role in medical technology. [ 2–4 ] Examples of applications of chitosan films are the localized chemotherapy based on the local delivery and sustained release of antineoplastic agents after cancer removal [ 5 ] and the tissue repair by closing chronic, accidental, and surgical wounds. [ 4 , 6 ] In order to enable these applications, the chitosan films need to be secured to the target, which may be achieved by suturing. [ 7 ] However, in many cases, suturing may be hardly feasible such as for poorly accessible or delicate body regions (e.g., intracranial, myocardial, etc.). In addition metals and synthetic materials in use for implantation can interfere with the functional rehabilitation of the site due to foreign-body reactions, causing infl ammation and abnormal wound healing. [ 8 ]

Gold nanorods as new nanochromophores for photothermal therapies

Results and perspectives on the biomedical exploitation of gold nanorods with plasmon resonances in the near infrared window are reported. The authors describe experimental studies of laser-activated nanoparticles in the direct welding of connective tissues, which may become a valuable technology in biomedicine. In particular, colloidal gold nanorods excited by diode laser radiation at 810 nm were used to mediate functional photothermal effects and weld eyes lens capsules and arteries. The preparation of biopolymeric matrices including gold nanorods is also described, as well as preliminary tests for their application in the closure of wounds in vessels and tendons. Finally, the use of these nanoparticles for future applications in the diagnosis, imaging and therapy of cancer is discussed.

Photothermal effects in connective tissues mediated by laser-activated gold nanorods

We report a study on the application of laser-activated nanoparticles in the direct welding of connective tissues, which may become a valuable technology in biomedicine. We use colloidal gold nanorods as new near-infrared chromophores to mediate functional photothermal effects in the eye lens capsules. Samples obtained ex vivo from porcine eyes are treated to simulate heterotransplants with 810-nm diode laser radiation in association with a stain of gold nanorods of aspect ratio approximately 4. This stain is applied at the interface between a patch of capsule from a donor eye and the capsule of a recipient eye. Then, by administration of laser pulses of 40 msec and approximately 100-140 J/cm(2), we achieved the local denaturation of the endogenous collagen filaments, which reveals that the treated area reached temperatures above 50 degrees C. The thermal damage is confined within 50-70 mum in a radial distance from the irradiated area.

Laser tissue welding in ophthalmic surgery

Laser welding of ocular tissues is an alternative technique or adjunct to conventional suturing in ophthalmic surgery. It is based on the photothermal interaction of laser light with the main components of the extracellular matrix of connective tissues. The advantages of the welding procedure with respect to standard suturing and stapling are reduced operation times, lesser inflammation, faster healing and increased ability to induce tissue regeneration. The procedure we set up is based on the use of an infrared diode laser in association with the topical application of the chromophore Indocyanine Green. Laser light may be delivered either continuously or in pulses, thus identifying two different techniques that have been applied clinically in various types of transplants of the cornea.

In vivo carotid artery closure by laser activation of hyaluronan-embedded gold nanorods

We prove the first application of near-infrared-absorbing gold nanorods (GNRs) for in vivo laser closure of a rabbit carotid artery. GNRs are first functionalized with a biopolymeric shell and then embedded in hyaluronan, which gives a stabilized and handy laser-activable formulation. Four rabbits undergo closure of a 3-mm longitudinal incision performed on the carotid artery by means of a 810-nm diode laser in conjunction with the topical application of the GNRs composite. An effective surgery is obtained by using a 40-W/cm(2) laser power density. The histological and electron microscopy evaluation after a 30-day follow-up demonstrates complete healing of the treated arteries with full re-endothelization at the site of GNRs application. The absence of microgranuloma formation and/or dystrophic calcification is evidence that no host reaction to nanoparticles interspersed through the vascular tissue occurred. The observation of a reshaping and associated blue shift of the NIR absorption band of GNRs after laser treatment supports the occurrence of a self-terminating process, and thus of additional safety of the minimally invasive laser procedure. This study underlines the feasibility of using GNRs for in vivo laser soldering applications, which represents a step forward toward the introduction of nanotechnology-based therapies in minimally invasive clinical practices.

Thermal Transitions of Fibrillar Collagen Unveiled by Second-Harmonic Generation Microscopy of Corneal Stroma

The thermal transitions of fibrillar collagen are investigated with second-harmonic generation polarization anisotropy microscopy. Second-harmonic generation images and polarization anisotropy profiles of corneal stroma heated in the 35-80°C range are analyzed by means of a theoretical model that is suitable to probe principal intramolecular and interfibrillar parameters of immediate physiological interest. Our results depict the tissue modification with temperature as the interplay of three destructuration stages at different hierarchical levels of collagen assembly including its tertiary structure and interfibrillar alignment, thus supporting and extending previous findings. This method holds the promise of a quantitative inspection of fundamental biophysical and biochemical processes and may find future applications in real-time and postsurgical functional imaging of collagen-rich tissues subjected to thermal treatments.

Emerging concepts of laser-activated nanoparticles for tissue bonding.

We report recent achievements and future perspectives of minimally invasive bonding of biological tissues triggered by laser light. In particular, we review new advancements in the biomedical exploitation of near-infrared absorbing gold nanoparticles as an original solution for the photothermal closure of surgical incisions. Advanced concepts of laser tissue bonding involving the application of hybrid nanocomposites obtained by inclusion of nanochromophores into biopolymer scaffolds are also introduced. The perspectives of tissue bonding are discussed in the following aspects: (1) tissue bonding with highly-stabilized nanochromophores, (2) enhanced tissue bonding with patterned nanocomposites, (3) real-time monitoring of temperature distributions, (4) tracking of tissue regeneration based on the optical resonances of gold nanoparticles.

Size dependent biological profiles of PEGylated gold nanorods

The perspective of introducing plasmonic particles for applications in biomedical optics is receiving much interest. However, their translation into clinical practices is delayed by various factors, which include a poor definition of their biological interactions. Here, we describe the preparation and the biological profiles of gold nanorods belonging to five different size classes with average effective radii between ∼5 and 20 nm and coated with polyethylene glycol (PEG). All these particles exhibit decent stability in the presence of representative proteins, low cytotoxicity and satisfactory compatibility with intravenous administration, in terms of their interference with blood tissue. However, the suspension begins to become unstable after a few days of exposure to blood proteins. Moreover, the cytotoxicity is a little worse for smaller particles, probably because their purification is more critical, while undesirable interactions with the mononuclear phagocyte system are minimal in the intermediate size range. Overall, these findings hold implications of practical relevance and suggest that PEGylated gold nanorods may be a versatile platform for a variety of biomedical applications.

Composition of Ge — Si – islands in the growth of Ge on Si — 111 – by x-ray spectromicroscopy

The stoichiometry of Ge∕Si islands grown on Si(111) substrates at temperatures ranging from 460to560°C was investigated by x-ray photoemission electron microscopy (XPEEM). By developing a specific analytical framework, quantitative information on the surface Ge∕Si stoichiometry was extracted from laterally resolved XPEEM Si 2p and Ge 3d spectra, exploiting the chemical sensitivity of the technique. Our data show the existence of a correlation between the base area of the self-assembled islands and their average surface Si content: the larger the lateral dimensions of the 3D structures, the higher their relative Si concentration. The deposition temperature determines the characteristics of this relation, pointing to the thermal activation of kinetic diffusion processes.