Viral hemorrhagic fevers are characterized by enhanced permeability. One of the most affected target organs of hantavirus-induced hemorrhagic fever with renal syndrome is the kidney, and an infection often results in acute renal failure. To study the underlying cellular effects leading to kidney dysfunction, we infected human renal cell types in vitro that are critical for the barrier functions of the kidney, and we examined kidney biopsy specimens obtained from hantavirus-infected patients. We analyzed the infection and pathogenic effects in tubular epithelial and glomerular endothelial renal cells and in podocytes. Both epithelial and endothelial cells and podocytes were susceptible to hantavirus infection in vitro. The infection disturbed the structure and integrity of cell-to-cell contacts, as demonstrated by redistribution and reduction of the tight junction protein ZO-1 and the decrease in the transepithelial resistance in infected epithelial monolayers. An analysis of renal biopsy specimens from hantavirus-infected patients revealed that the expression and the localization of the tight junction protein ZO-1 were altered compared to renal biopsy specimens from noninfected individuals. Both tubular and glomerular cells were affected by the infection. Furthermore, the decrease in glomerular ZO-1 correlates with disease severity induced by glomerular dysfunction. The finding that different renal cell types are susceptible to hantaviral infection and the fact that infection results in the breakdown of cell-to-cell contacts provide useful insights in hantaviral pathogenesis.

Several non-animal methods are now available to address the key events leading to skin sensitization as defined by the adverse outcome pathway. The KeratinoSens assay addresses the cellular event of keratinocyte activation and is a method accepted under OECD TG 442D. In this study, the results of an inter-laboratory evaluation of the "me-too" LuSens assay, a bioassay that uses a human keratinocyte cell line harboring a reporter gene construct composed of the rat antioxidant response element (ARE) of the NADPH:quinone oxidoreductase 1 gene and the luciferase gene, are described. Earlier in-house validation with 74 substances showed an accuracy of 82% in comparison to human data. When used in a battery of non-animal methods, even higher predictivity is achieved. To meet European validation criteria, a multicenter study was conducted in 5 laboratories. The study was divided into two phases, to assess 1) transferability of the method, and 2) reproducibility and accuracy. Phase I was performed by testing 8 non-coded test substances; the results showed a good transferability to naïve laboratories even without on-site training. Phase II was performed with 20 coded test substances (performance standards recommended by OECD, 2015). In this phase, the intra- and inter-laboratory reproducibility as well as accuracy of the method was evaluated. The data demonstrate a remarkable reproducibility of 100% and an accuracy of over 80% in identifying skin sensitizers, indicating a good concordance with in vivo data. These results demonstrate good transferability, reliability and accuracy of the method thereby achieving the standards necessary for use in a regulatory setting to detect skin sensitizers.