Identification of Therapeutic Targets of Inflammatory Monocyte Recruitment to Modulate the Allogeneic Injury to Donor Cornea

Purpose: We sought to test the hypothesis that monocytes contribute to the immunopathogenesis of corneal allograft rejection and identify therapeutic targets to inhibit monocyte recruitment. Methods: Monocytes and proinflammatory mediators within anterior chamber samples during corneal graft rejection were quantified by flow cytometry and multiplex protein assays. Lipopolysaccharide or IFN-γ stimulation of monocyte-derived macrophages (MDMs) was used to generate inflammatory conditioned media (CoM). Corneal endothelial viability was tested by nuclear counting, connexin 43, and propidium iodide staining. Chemokine and chemokine receptor expression in monocytes and MDMs was assessed in microarray transcriptomic data. The role of chemokine pathways in monocyte migration across microvascular endothelium was tested in vitro by chemokine depletion or chemokine receptor inhibitors. Results: Inflammatory monocytes were significantly enriched in anterior chamber samples within 1 week of the onset of symptoms of corneal graft rejection. The MDM inflammatory CoM was cytopathic to transformed human corneal endothelia. This effect was also evident in endothelium of excised human cornea and increased in the presence of monocytes. Gene expression microarrays identified monocyte chemokine receptors and cognate chemokines in MDM inflammatory responses, which were also enriched in anterior chamber samples. Depletion of selected chemokines in MDM inflammatory CoM had no effect on monocyte transmigration across an endothelial blood–eye barrier, but selective chemokine receptor inhibition reduced monocyte recruitment significantly. Conclusions: We propose a role for inflammatory monocytes in endothelial cytotoxicity in corneal graft rejection. Therefore, targeting monocyte recruitment offers a putative novel strategy to reduce donor endothelial cell injury in survival of human corneal allografts

Vascular epiphytes of the Atlantic Forest in the Sinos River basin, state of Rio Grande do Sul, Brazil: richness, floristic composition and community structure

The Atlantic Forest, which has a vast epiphytic richness, is a priority area for preservation, listed as one of the five most important world hotspots. Vascular epiphyte richness, composition and community structure were studied in two fragments, one of the ombrophilous (29º43'42"S and 50º22'00"W) and the other of the seasonal (29º40'54"S and 51º06'56"W) forest, both belonging to the Atlantic Forest biome in the Sinos River basin, Rio Grande do Sul, Brazil. In each fragment, 40 trees, divided into four ecological zones, were analyzed. In each zone, the occurrence of the species was recorded, and the importance value of each species was calculated according to the frequency of phorophytes and intervals, and cover scores. The Shannon index was calculated for the two communities. In the fragment of the ombrophilous forest (F1), 30 epiphytic species were recorded, and in the seasonal forest (F2), 25. The highest importance value was found for Microgramma squamulosa (Kaulf.) de la Sota in both fragments. The diversity indexes for F1 (H'=2.72) and F2 (H'=2.55) were similar and reflected the subtropical location of the areas. The decrease in mean richness in both fragments in zone 3 (internal crown) to zone 4 (external crown) may be associated with time and space availability for epiphyte occupation and microclimate variations. Exclusive species were found in the areas, which suggest that a greater number of preserved fragments may result in a greater number of preserved epiphytic species in the Sinos River basin.</p