Although the existence of limbal stem cell (LSC) niche is accepted, precise knowledge its three-dimensional (3D) architecture remains incomplete. The LSC was explored on freshly excised and organ-cultured corneoscleral rims from human donors (n = 47), pigs 15) mice 27) with full-field optical coherence microscopy (FFOCM). Limbal crypt features were detected in 90% rims, extending between palisades Vogt as radially oriented rectangular (74% eyes) and/or rounded (23% forms, often branching off...

Abstract We aimed to evaluate efficiency and safety of transplantation limbal stem cells (LSC) cultured on human amniotic membrane with no feeders compare LSC tissue transplantation. Thirty eyes stage III deficiency were treated autologous (autoLSC) or allogeneic (alloLSC) (prospective phase II clinical trial; average follow-up time, 72 months) (autoLT) (alloLT) (retrospective control group; 132 between 1993 2014. The 5-year graft survival defined by absence recurrence the signs was 71% for...

Second-harmonic generation (SHG) microscopy is currently the preferred technique for visualizing collagen in intact tissues, but usual implementations struggle to reveal fibrils oriented out of imaging plane. Recently, an advanced SHG modality, circular dichroism (CD-SHG), has been proposed specifically highlight out-of-plane fibrils. In this study, we present a theoretical analysis CD-SHG signals that goes beyond electric dipolar approximation account chirality. We demonstrate magnetic...

We uncover the significance of a previously unappreciated structural feature in corneal stroma, important to its biomechanics. Vogt striae are known clinical indicator keratoconus, and consist dark, vertical lines crossing depth. However we detected stromal most corneas, not only keratoconus. observed with multiple imaging modalities 82% 118 human pathology-specific differences. Striae generally depart from anchor points at Descemet's membrane posterior stroma obliquely V-shape, whereas...

Diagnosis of bacterial endophthalmitis (BE) often fails due to: (1) insufficient volumes vitreous fluid (VF) and aqueous humour (AH); (2) lack sensitivity culture; (3) antibiotic treatments; (4) polymerase chain reaction (PCR) cross-contamination; (5) limitations on the interpretation real-time PCR melting curve. We developed a fast (f-real-t) to improve performance laboratory diagnosis BE.The following samples were processed after adding an internal control: phosphate buffered saline (PBS);...

Corneal scarring associated with various corneal conditions is a leading cause of blindness worldwide. The present study aimed to test the hypothesis that stromal stem cells have therapeutic effect and are able restore extracellular matrix organization transparency in vivo. We first developed mouse model scar induced by liquid nitrogen (N2 ) application. then reversed injecting or human injured cornea. To characterize this cells, we used combination vivo (slit lamp, optical coherence...

Purpose The prognosis of people infected with Fungi especially immunocompromised depends on rapid and accurate diagnosis to capitalize time administration specific treatments. However, cultures produce false negative results nucleic-acid amplification techniques require complex post-amplification procedures differentiate relevant fungal types. objective this work was develop a new diagnostic strategy based real-time polymerase-chain reaction high-resolution melting analysis (PCR-HRM) that a)...

Controlling both organizations and transparency of dense collagen scaffolds.

To search for gold-standard histology indicators using alternative imaging modalities in keratoconic corneas.Prospective observational case-control study. Fourteen corneas and 20 normal (10 vivo healthy subjects 10 ex donor corneas) were examined. Images of taken by spectral domain optical coherence tomography (SD-OCT) confocal microscopy (IVCM) before keratoplasty. The same removed corneal buttons imaged after keratoplasty with full-field (FFOCM) then fixed sent histology. Controls...

Second harmonic generation (SHG) enables in situ imaging of fibrillar collagen architecture connective tissues. Recently, Circular Dichroism SHG (CD-SHG) microscopy has been implemented to take advantage chirality improve 3D visualization. It measures the normalized difference signal obtained upon excitation by left versus right circular polarizations. However, CD-SHG is not well characterized yet, and quite different values are reported literature. Here, we identify two major artifacts that...

<h3>Background:</h3> Acanthamoeba keratitis (AK) is a sight-threatening infection, and none of the current diagnosis tests are able to detect in one reaction low levels vast majority strains associated with pathology. The goal this work was validate new tool for detection American Type Cell Collection (ATCC) referenced <i>Acanthamoeba</i> monitoring simultaneously DNA extraction yields PCR inhibitors. Performances were assessed on corneal scrapings. <h3>Methods:</h3> Primers selected region...

Background: Propionibacteriaceae ( Propioni ) are anaerobic bacteria associated with human and animal infections. Present-day methods of diagnosis for unsatisfactory due to a lack sensitivity culture, time required culture results (3 14 days) difficulties in interpreting SYBR Green real-time PCR results. The goal this work was validate new rapid sensitive test the infections (endophthalmitis, corneal ulcers others). Material methods: DNA extracted using MagNA Pure isolation kit (Roche),...

Purpose: Cholera toxin and isoproterenol (β-adrenergic receptor agonist) are largely used to enhance cell proliferation. The aim of the study was assess effects cholera on growth differentiation cells cultured from human superficial limbal explants.Methods: Limbal epithelial were explantsin basal medium either supplemented with or for 3 weeks. Growth kinetics morphometry studied by light confocal microscopy. Progenitor differentiated markers immunocytochemistry, flow cytometry, Colony...

To provide quantitative parameters for assessment of human donor corneal stroma by imaging stromal features diseased and normal corneas with full-field optical coherence microscopy (FFOCM), using confocal (CM) histology as reference techniques.Bowman's layer (BL) thickness keratocyte density were assessed ex vivo in 23 27 pathological (keratoconus other disorders) FFOCM, CM histology. Stromal backscattering was FFOCM. Additionally, 10 CM.In the logarithm normalized reflectivity a linear...

Purpose: To evaluate the incidence, clinical and microbiological characteristics risk factors of infectious keratitis in patients with limbal stem cell deficiency (LSCD).Methods: Retrospective comparative case series 35 severe LSCD.Results: The mean follow-up time was 46 months. Infectious were mainly caused by Gram positive bacteria (94%). Only 7 infections (37%) healed under fortified adapted antibiotics. In 8 cases (42%), amniotic membrane transplantation required 4 (21%) «à chaud»...

Epithelial and stromal stem cells are required to maintain corneal transparency. The aim of the study was develop a new method isolate grow both (SSC) epithelial limbal (LSC) from small human biopsies under culture conditions in accordance with safety requirements mandatory for clinical use humans. Superficial explants were retrieved donor corneo-scleral rims. Human dissociated by digestion collagenase A, either after scraping or no scraping. Isolated cultured Essential 8 medium (E8), E8...

The aims of this study were to determine whether human limbal explant cultures without feeder cells result in expansion epithelial progenitors and estimate the optimal time for progenitor cells. Limbal explants from ten corneas cultured 7, 9, 11, 14, 18, 21 days. two enzymatically dissociated or directly 14 Progenitor characterized by their ability form colonies, immunocytochemistry, quantitative real-time polymerase chain reaction. Colonies identified after 18 days culture, but not number...

The quality of donor corneal stroma, which makes up about 90% total thickness, is likely to be one the main, if not major, limiting factor(s) for success deep anterior lamellar and penetrating keratoplasty. These are surgical procedures that involve replacing part or all diseased layers, respectively, by donated tissue, graft, taken from a recently deceased individual. However, means evaluate stromal grafts in eye banks limited lack capability high-resolution quantitative assessment disease...