The adoption of a new technology into basic research, and industrial and clinical settings requires rigorous testing to build confidence in the reproducibility, reliability, robustness, and relevance of these models. Tissue chips are promising new technology, they have the potential to serve as a valuable tool in biomedical research, as well as pharmaceutical development with regards to testing for efficacy and safety. The principal goals of this study were to validate a previously established proximal tubule tissue chip model in an independent laboratory and to extend its utility to testing of nephrotoxic compounds. Here, we evaluated critical endpoints from the tissue chip developer laboratory, focusing on biological relevance (long-term viability, baseline protein and gene expression, ammoniagenesis, and vitamin D metabolism), and toxicity biomarkers. Tissue chip experiments were conducted in parallel with traditional 2D culture conditions using two different renal proximal tubule epithelial cell sources. The results of these studies were then compared to the findings reported by the tissue chip developers. While the overall transferability of this advanced tissue chip platform was a success, the reproducibility with the original report was greatly dependent on the cell source. This study demonstrates critical importance of developing microphysiological platforms using renewable cell sources.

RIPK3, a key mediator of necroptosis, has been implicated in the host defense against viral infection primary in immune cells. However, gene expression analysis revealed that RIPK3 is abundantly expressed not only in immune organs but also in the gastrointestinal tract, particularly in the small intestine. We found that orally inoculated Listeria monocytogenes, a bacterial foodborne pathogen, efficiently spread and caused systemic infection in Ripk3-deficient mice while almost no dissemination was observed in wild-type mice. Listeria infection activated the RIPK3-MLKL pathway in cultured cells, which resulted in suppression of intracellular replication of Listeria. Surprisingly, Listeria infection–induced phosphorylation of MLKL did not result in host cell killing. We found that MLKL directly binds to Listeria and inhibits their replication in the cytosol. Our findings have revealed a novel functional role of the RIPK3-MLKL pathway in nonimmune cell-derived host defense against Listeria invasion, which is mediated through cell death–independent mechanisms.

Cell-based assays are an attractive option to measure gene expression response to exposure, but the cost of whole-transcriptome RNA sequencing has been a barrier to the use of gene expression profiling for in vitro toxicity screening. In addition, standard RNA sequencing adds variability due to variable transcript length and amplification. Targeted probe-sequencing technologies such as TempO-Seq, with transcriptomic representation that can vary from hundreds of genes to the entire transcriptome, may reduce some components of variation. Analyses of high-throughput toxicogenomics data requires renewed attention to read-calling algorithms and simplified dose-response modeling for datasets with relatively few samples. Using data from induced pluripotent stem cell-derived cardiomyocytes treated with chemicals at varying concentrations, we describe here and make available a pipeline for handling expression data generated by TempO-Seq to align reads, clean and normalize raw count data, identify differentially-expressed genes and calculate transcriptomic concentration-response points of departure. The methods are extensible to other forms of concentration-response gene-expression data, and we discuss the utility of the methods for assessing variation in susceptibility and the diseased cellular state.

Background: Maternal diet during pregnancy has been shown to influence the child neuro-developmental outcomes. Studies examining effects of dietary patterns on offspring behavior are sparse. Objective: Determine if maternal adherence to a Mediterranean diet is associated with child behavioral outcomes assessed early in life, and to evaluate the role of differentially methylated regions (DMRs) regulating genomically imprinted genes in these associations. Methods: Among 325 mother/infant pairs, we used regression models to evaluate the association between tertiles of maternal periconceptional Mediterranean diet adherence (MDA) scores derived from a Food Frequency Questionnaire, and social and emotional scores derived from the Infant Toddler Social and Emotional Assessment (ITSEA) questionnaire in the second year of life. Methylation of nine genomically imprinted genes was measured to determine if MDA was associated with CpG methylation. Results: Child depression was significantly associated with maternal MDA (Bonferroni-corrected p = 0.048). While controlling for false-discovery, compared to offspring of women with the lowest MDA tertile, those with MDA scores in middle and high MDA tertiles had decreased odds for atypical behaviors (OR (95%CI) = 0.40 (0.20, 0.78) for middle and 0.40 (0.17, 0.92) for highest tertile)), for maladaptive behaviors (0.37 (0.18, 0.72) for middle tertile and 0.42 (0.18, 0.95) for highest tertile) and for an index of autism spectrum disorder behaviors (0.46 (0.23, 0.90) for middle and 0.35 (0.15, 0.80) for highest tertile). Offspring of women with the highest MDA tertile were less likely to exhibit depressive (OR=0.28 (0.12, 0.64)) and anxiety (0.42 (0.18, 0.97)) behaviors and increased odds of social relatedness (2.31 (1.04, 5.19)) behaviors when compared to low MDA mothers. Some associations varied by sex. Perinatal MDA score was associated with methylation differences for imprinted control regions of PEG10/SGCE (females: Beta (95%CI) = 1.66 (0.52, 2.80) - Bonferroni-corrected p = 0.048; males: -0.56 (-1.13, -0.00)), as well as both MEG3 and IGF2 in males (0.97 (0.00, 1.94)) and -0.92 (-1.65, -0.19) respectively. Conclusions: In this ethnically diverse cohort, maternal adherence to a Mediterranean diet in early pregnancy was associated with favorable neurobehavioral outcomes in early childhood and with sex-dependent methylation differences of MEG3, IGF2, and SGCE/PEG10 DMRs.