Three-dimensional mapping of the orientation of collagen corneal lamellae in healthy and keratoconic human corneas using SHG microscopy

SHG image acquired with sagittal optical sectioning (A) of a healthy cornea and (B) of a keratoconic cornea. Scale bars: 30 μm. Keratoconus is an eye disorder that causes the cornea to take an abnormal conical shape, thus impairing its refractive functions and causing blindness. The late diagnosis of keratoconus is among the principal reasons for corneal surgical transplantation. This pathology is characterized by a reduced corneal stiffness in the region immediately below Bowman's membrane, probably due to a different lamellar organization, as suggested by previous studies. Here, the lamellar organization in this corneal region is characterized in three dimensions by means of second-harmonic generation (SHG) microscopy. In particular, a method based on a three-dimensional correlation analysis allows to probe the orientation of sutural lamellae close to the Bowman's membrane, finding statistical differences between healthy and keratoconic samples. This method is demonstrated also in combination with an epi-detection scheme, paving the way for a potential clinical ophthalmic application of SHG microscopy for the early diagnosis of keratoconus

Preparation and photoacoustic analysis of cellular vehicles containing gold nanorods

Gold nanorods are attractive for a range of biomedical applications, such as the photothermal ablation and the photoacoustic imaging of cancer, thanks to their intense optical absorbance in the near-infrared window, low cytotoxicity and potential to home into tumors. However, their delivery to tumors still remains an issue. An innovative approach consists of the exploitation of the tropism of tumor-associated macrophages that may be loaded with gold nanorods in vitro. Here, we describe the preparation and the photoacoustic inspection of cellular vehicles containing gold nanorods. PEGylated gold nanorods are modified with quaternary ammonium compounds, in order to achieve a cationic profile. On contact with murine macrophages in ordinary Petri dishes, these particles are found to undergo massive uptake into endocytic vesicles. Then these cells are embedded in biopolymeric hydrogels, which are used to verify that the stability of photoacoustic conversion of the particles is retained in their inclusion into cellular vehicles. We are confident that these results may provide new inspiration for the development of novel strategies to deliver plasmonic particles to tumors

Observation of an improved healing process in superficial skin wounds after irradiation with a blue-LED haemostatic device

The healing process of superficial skin wounds treated with a blue-LED haemostatic device is studied. Four mechanical abrasions are produced on the back of 10 Sprague Dawley rats: two are treated with the blue-LED device, while the other two are left to naturally recover. Visual observations, non-linear microscopic imaging, as well as histology and immunofluorescence analyses are performed 8 days after the treatment, demonstrating no adverse reactions neither thermal damages in both abraded areas and surrounding tissue. A faster healing process and a better-recovered skin morphology are observed: the treated wounds show a reduced inflammatory response and a higher collagen content. Blue LED induced photothermal effect on superficial abrasions

Stable, Monodisperse, and Highly Cell-Permeating Nanocochleates from Natural Soy Lecithin Liposomes

(1) Background: Andrographolide (AN), the main diterpenoid constituent of Andrographis paniculata, has a wide spectrum of biological activities. The aim of this study was the development of nanocochleates (NCs) loaded with AN and based on phosphatidylserine (PS) or phosphatidylcholine (PC), cholesterol and calcium ions in order to overcome AN low water solubility, its instability under alkaline conditions and its rapid metabolism in the intestine. (2) Methods: The AN-loaded NCs (AN–NCs) were physically and chemically characterised. The in vitro gastrointestinal stability and biocompatibility of AN–NCs in J77A.1 macrophage and 3T3 fibroblasts cell lines were also investigated. Finally, the uptake of nanocarriers in macrophage cells was studied. (3) Results: AN–NCs obtained from PC nanoliposomes were suitable nanocarriers in terms of size and homogeneity. They had an extraordinary stability after lyophilisation without the use of lyoprotectants and after storage at room temperature. The encapsulation efficiency was 71%, while approximately 95% of AN was released in PBS after 24 h, with kinetics according to the Hixson–Crowell model. The in vitro gastrointestinal stability and safety of NCs, both in macrophages and 3T3 fibroblasts, were also assessed. Additionally, NCs had extraordinary uptake properties in macrophages. (4) Conclusions: NCs developed in this study could be suitable for both AN oral and parental administration, amplifying its therapeutic value

Extracellular chaperones prevent Aβ42-induced toxicity in rat brains

Alzheimer\u27s disease (AD) is a progressive neurodegenerative disorder characterised by cognitive decline, formation of the extracellular amyloid β (Aβ42) plaques, neuronal and synapse loss, and activated microglia and astrocytes. Extracellular chaperones, which are known to inhibit amyloid fibril formation and promote clearance of misfolded aggregates, have recently been shown to reduce efficiently the toxicity of HypF-N misfolded oligomers to immortalised cell lines, by binding and clustering them into large species. However, the role of extracellular chaperones on Aβ oligomer toxicity remains unclear, with reports often appearing contradictory. In this study we microinjected into the hippocampus of rat brains Aβ42 oligomers pre-incubated for 1 h with two extracellular chaperones, namely clusterin and α2-macroglobulin. The chaperones were found to prevent Aβ42-induced learning and memory impairments, as assessed by the Morris Water Maze test, and reduce Aβ42-induced glia inflammation and neuronal degeneration in rat brains, as probed by fluorescent immunohistochemical analyses. Moreover, the chaperones were able to prevent Aβ42 colocalisation with PSD-95 at post-synaptic terminals of rat primary neurons, suppressing oligomer cytotoxicity. All such effects were not effective by adding pre-formed oligomers and chaperones without preincubation. Molecular chaperones have therefore the potential to prevent the early symptoms of AD, not just by inhibiting Aβ42 aggregation, as previously demonstrated, but also by suppressing the toxicity of Aβ42 oligomers after they are formed. These findings elect them as novel neuroprotectors against amyloid-induced injury and excellent candidates for the design of therapeutic strategies against AD