No abstract

The control of corneal transparency depends on the integrity of its endothelial monolayer, which is considered nonregenerative in adult humans. In pathological situations, endothelial cell (EC) loss, not offset by mitosis, can lead to irreversible corneal edema and blindness. However, the hypothesis of a slow, clinically insufficient regeneration starting from the corneal periphery remains debatable. The authors have re-evaluated the microanatomy of the endothelium in order to identify structures likely to support this homeostasis model. Whole endothelia of 88 human corneas (not stored, and stored in organ culture) with mean donor age of 80 6 12 years were analyzed using an original flatmounting technique. In 61% of corneas, cells located at the extreme periphery (last 200 lm of the endothelium) were organized in small clusters with two to three cell layers around Hassall-Henle bodies. In 68% of corneas, peripheral ECs formed centripetal rows 830 6 295 lm long, with Descemet membrane furrows visible by scanning electron microscopy. EC density was significantly higher in zones with cell rows. When immunostained, ECs in the extreme periphery exhibited lesser differentiation (ZO-1, Actin, Na/ K ATPase, CoxIV) than ECs in the center of the cornea but preferentially expressed stem cell markers (Nestin, Telomerase, and occasionally breast cancer resistance protein) and, in rare cases, the proliferation marker Ki67. Stored corneas had fewer cell clusters but more Ki67-positive ECs. We identified a novel anatomic organization in the periphery of the human corneal endothelium, suggesting a continuous slow centripetal migration, throughout life, of ECs from specific niches.

Skin dose is one of the key issues for clinical dosimetry in radiation therapy. Currently planning computer systems are unable to accurately predict dose in the buildup region, leaving ambiguity as to the dose levels actually received by the patient's skin during radiotherapy. This is one of the prime reasons why in vivo measurements are necessary to estimate the dose in the buildup region. A newly developed metal-oxide-semiconductor-field-effect-transistor (MOSFET) detector designed specifically for dose measurements in rapidly changing dose gradients was introduced for accurate in vivo skin dosimetry. The feasibility of this detector for skin dose measurements was verified in comparison with plane parallel ionization chamber and radiochromic films. The accuracy of a commercial treatment planning system (TPS) in skin dose calculations for intensity-modulated radiation therapy treatment of nasopharyngeal carcinoma was evaluated using MOSFET detectors in an anthropomorphic phantom as well as on the patients. Results show that this newly developed MOSFET detector can provide a minimal but highly reproducible intrinsic buildup of 7 mg cm(-2) corresponding to the requirements of personal surface dose equivalent Hp (0.07). The reproducibility of the MOSFET response, in high sensitivity mode, is found to be better than 2% at the phantom surface for the doses normally delivered to the patients. The MOSFET detector agrees well with the Attix chamber and the EBT Gafchromic film in terms of surface and buildup region dose measurements, even for oblique incident beams. While the dose difference between MOSFET measurements and TPS calculations is within measurement uncertainty for the depths equal to or greater than 0.5 cm, an overestimation of up to 8.5% was found for the surface dose calculations in the anthropomorphic phantom study. In vivo skin dose measurements reveal that the dose difference between the MOSFET results and the TPS calculations was on average -7.2%, ranging from -4.3% to -9.2%. The newly designed MOSFET detector encapsulated into a thin water protective film has a minimal reproducible intrinsic buildup recommended for skin dosimetry. This feature makes it very suitable for routine IMRT QA and accurate in vivo skin dosimetry.

Corneal endothelial cells (CECs) are terminally differentiated cells, specialized in regulating corneal hydration and transparency. They are highly polarized flat cells that separate the cornea from the aqueous humor. Their apical surface, in contact with aqueous humor is hexagonal, whereas their basal surface is irregular. We characterized the structure of human CECs in 3D using confocal microscopy of immunostained whole corneas in which cells and their interrelationships remain intact. Hexagonality of the apical surface was maintained by the interaction between tight junctions and a submembraneous network of actomyosin, braced like a drum. Lateral membranes, which support enzymatic pumps, presented complex expansions resembling interdigitated foot processes at the basal surface. Using computer-aided design and drafting software, we obtained a first simplified 3D model of CECs. By comparing their expression with those in epithelial, stromal and trabecular corneal cells, we selected 9 structural or functional proteins for which 3D patterns were specific to CECs. This first 3D map aids our understanding of the morphologic and functional specificity of CECs and could be used as a reference for characterizing future cell therapy products destined to treat endothelial dysfunctions.

This study demonstrates that thin human corneal lamellae could have high potential as carriers in next-generation therapy for endothelial dysfunctions.