No abstract

Our preliminary results show that topical treatment with ADSC seems to improve corneal wound healing.

Infectious diseases remain an important global health problem. The interaction of a wide range of pathogen bacteria with host cells from many different tissues is frequently mediated by proteoglycans. These compounds are ubiquitous complex molecules which are not only involved in adherence and colonization, but can also participate in other steps of pathogenesis. To overcome the problem of microbial resistance to antibiotics new therapeutic agents could be developed based on the characteristics of the interaction of pathogens with proteoglycans.

Morphological and functional alterations of peripheral somatosensory neurons during the aging process lead to a decline of somatosensory perception. Here, we analyze the changes occurring with aging in trigeminal ganglion (TG), TRPM8-expressing cold thermoreceptor neurons innervating the mouse cornea, which participate in the regulation of basal tearing and blinking and have been implicated in the pathogenesis of dry eye disease (DED). TG cell bodies and axonal branches were examined in a mouse line (TRPM8 -EYFP) expressing a fluorescent reporter. In 3 months old animals, about 50% of TG cold thermoreceptor neurons were intensely fluorescent, likely providing strongly fluorescent axons and complex corneal nerve terminals with ongoing activity at 34°C and low-threshold, robust responses to cooling. The remaining TRPM8 corneal axons were weakly fluorescent with nonbeaded axons, sparsely ramified nerve terminals, and exhibited a low-firing rate at 34°C, responding moderately to cooling pulses as do weakly fluorescent TG neurons. In aged (24 months) mice, the number of weakly fluorescent TG neurons was strikingly high while the morphology of TRPM8 corneal axons changed drastically; 89% were weakly fluorescent, unbranched, and often ending in the basal epithelium. Functionally, 72.5% of aged cold terminals responded as those of young animals, but 27.5% exhibited very low-background activity and abnormal responsiveness to cooling pulses. These morpho-functional changes develop in parallel with an enhancement of tear's basal flow and osmolarity, suggesting that the aberrant sensory inflow to the brain from impaired peripheral cold thermoreceptors contributes to age-induced abnormal tearing and to the high incidence of DED in elderly people.

The aim of our study was to assess the clinical effectiveness of topical adipose derived stem cell (ADSC) treatment in laser induced corneal wounds in mice by comparing epithelial repair, inflammation, and histological analysis between treatment arms. Corneal lesions were performed on both eyes of 40 mice by laser induced photorefractive keratectomy. All eyes were treated with topical azythromycin bid for three days. Mice were divided in three treatment groups (n = 20), which included: control, stem cells and basic serum; which received topical treatment three times daily for five consecutive days. Biomicroscope assessments and digital imaging were performed by two masked graders at 30, 54, 78, 100, and 172 h to analyze extent of fluorescein positive epithelial defect, corneal inflammation, etc. Immunohistochemical techniques were used in fixed eyes to assess corneal repair markers Ki67, α Smooth Muscle Actin (α-SMA) and E-Cadherin. The fluorescein positive corneal lesion areas were significantly smaller in the stem cells group on days 1 (p < 0.05), 2 (p < 0.02) and 3. The stem cell treated group had slightly better and faster re-epithelization than the serum treated group in the initial phases. Comparative histological data showed signs of earlier and better corneal repair in epithelium and stromal layers in stem cell treated eyes, which showed more epithelial layers and enhanced wound healing performance of Ki67, E-Cadherin, and α-SMA. Our study shows the potential clinical and histological advantages in the topical ADSC treatment for corneal lesions in mice.

PURPOSE. To define the characteristics and time course of the morphologic and functional changes experienced by corneal sensory nerves after photorefractive keratectomy (PRK).METHODS. Unilateral corneal excimer laser photoablation was performed in 54 anesthetized 3-to 6-month-old mice; 11 naïve animals served as control. Mice were killed 0, 3, 7, 15, and 30 days after PRK. Excised eyes were placed in a recording chamber superfused at 348C. Electrical nerve impulse activity of single sensory terminals was recorded with a micropipette applied onto the corneal surface. Spontaneous and stimulus-evoked (cold, heat, mechanical, and chemical stimuli) nerve terminal impulse (NTI) activity was analyzed. Corneas were fixed and stained with anti-b-Tubulin III antibody to measure nerve density and number of epithelial nerve penetration points of regenerating subbasal leashes.RESULTS. Nerve fibers and NTI activity were absent in the injured area between 0 and 7 days after PRK, when sparse regenerating nerve sprouts appear. On day 15, subbasal nerve density reached half the control value and abnormally responding cold-sensitive terminals were recorded inside the lesion. Thirty days after PRK, nerve density was almost restored, active cold thermoreceptors were abundant, and polymodal nociceptor activity first reappeared.CONCLUSIONS. Morphologic regeneration of subbasal corneal nerves started shortly after PRK ablation and was substantially completed 30 days later. Functional recovery appears faster in cold terminals than polymodal terminals, possibly reflecting an incomplete damage of the more extensively branched cold-sensitive axon terminals. Evolution of postsurgical discomfort sensations quality may be associated with the variable regeneration pattern of each fiber type.

The epithelium of the cornea is continuously exposed to pathogens, and adhesion to epithelial cells is regarded as an essential first step in bacterial pathogenesis. In this article, the involvement of glycosaminoglycans in the adhesion of various pathogenic bacteria to corneal epithelial cells is analyzed. All microorganisms use glycosaminoglycans as receptors, but arranged in different patterns depending on the Gram-type of the bacterium. The heparan sulfate chains of syndecans are the main receptors, though other molecular species also seem to be involved, particularly in Gram-negative bacteria. Adherence is inhibited differentially by peptides, including heparin binding sequences, indicating the participation of various groups of Gram-positive, and -negative adhesins. The length of the saccharides produces a major effect, and low molecular weight chains inhibit the binding of Gram-negative microorganisms but increase the adherence of Gram-positives. Pathogen adhesion appears to occur preferentially through sulfated domains, and is very dependent on N- and 6-O-sulfation of the glucosamine residue and, to a lesser extent, 2-O sulfation of uronic acid. These data show the differential use of corneal receptors, which could facilitate the development of new anti-infective strategies.

Most of the PG core proteins underwent significant changes in cultured keratoconic cells and corneas. With regard to GAG chains, the polymerization of the chains and their chemical modification was modified in way that depended on the specific type of GAG involved.