Unprecedented genetic variability of PFam54 paralogs among Eurasian Lyme borreliosis-causing spirochetes.

Lyme borreliosis (LB) is the most common vector-borne disease in the Northern Hemisphere caused by spirochetes belonging to the Borrelia burgdorferi sensu lato (Bbsl) complex. Borrelia spirochetes circulate in obligatory transmission cycles between tick vectors and different vertebrate hosts. To successfully complete this complex transmission cycle, Bbsl encodes for an arsenal of proteins including the PFam54 protein family with known, or proposed, influences to reservoir host and/or vector adaptation. Even so, only fragmentary information is available regarding the naturally occurring level of variation in the PFam54 gene array especially in relation to Eurasian-distributed species. Utilizing whole genome data from isolates (n = 141) originated from three major LB-causing Borrelia species across Eurasia (B. afzelii, B. bavariensis, and B. garinii), we aimed to characterize the diversity of the PFam54 gene array in these isolates to facilitate understanding the evolution of PFam54 paralogs on an intra- and interspecies level. We found an extraordinarily high level of variation in the PFam54 gene array with 39 PFam54 paralogs belonging to 23 orthologous groups including five novel paralogs. Even so, the gene array appears to have remained fairly stable over the evolutionary history of the studied Borrelia species. Interestingly, genes outside Clade IV, which contains genes encoding for proteins associated with Borrelia pathogenesis, more frequently displayed signatures of diversifying selection between clades that differ in hypothesized vector or host species. This could suggest that non-Clade IV paralogs play a more important role in host and/or vector adaptation than previously expected, which would require future lab-based studies to validate.

Gene-expression profiling of laser-dissected islets and studies in deficient mice reveal chemokines as differential driving force of type 1 diabetes.

Although type 1 diabetes (T1D) results from the autoimmune destruction of the insulin-producing ß-cells, its treatment is largely restricted to exogenous insulin administration. Only few therapies targeting the autoaggressive immune system have been introduced into clinical practice or are considered in clinical trials. Here, we provide a gene expression profile of the islet microenvironment obtained by laser-dissection microscopy in an inducible mouse model. Thereby, we have identified novel targets for immune intervention. Increased gene expression of most inflammatory proteins was apparent at day 10 after T1D induction and largely paralleled the observed degree of insulitis. We further focused on genes involved in leukocyte migration, including chemokines and their receptors. Besides the critical chemokine CXCL10, we found several other chemokines upregulated locally in temporary or chronic manner. Localization of the chemokine ligand/receptor pairs to the islet microenvironment has been confirmed by RNAscope. Interference with the CXCL16-CXCR6 and CX3CL1-CX3CR1 axes, but not the CCL5-CCR1/3/5 axis, resulted in reduced insulitis and lower T1D incidence. Further, we found that the receptors for the differentially expressed chemokines CXCL10, CXCL16 and CX3CL1 are distributed unevenly among islet autoantigen-specific T cells, which explains why the interference with just one chemokine axis cannot completely abrogate insulitis and T1D.

Contaminant biomonitoring augmented with a qPCR array indicates hepatic mRNA gene expression effects in wild-collected seabird embryos.

Birds can bioaccumulate persistent contaminants, and maternal transfer to eggs may expose embryos to concentrations sufficient to cause adverse effects during sensitive early-life stages. However, using tissue residue concentrations alone to infer whether contaminant effects are occurring suffers from uncertainty, and efficient, sensitive biomarkers remain limited in wildlife. We studied relationships between whole embryo contaminant concentrations (total mercury, organochlorine pesticides, perfluoroalkyl substances, polychlorinated biphenyls, and halogenated flame retardants) together with mRNA expression in embryonic liver tissue from a Pacific Ocean seabird, the rhinoceros auklet (Cerorhinca monocerata). Fresh eggs were collected, incubated under controlled conditions, and from the pre-hatch embryo, hepatic RNA was extracted for qPCR array analysis to measure gene expression (2-∆Cq), while the remaining embryo was analyzed for contaminant residues. Contaminant and gene expression data were assessed with a combination of multivariate approaches and linear models. Results indicated correlations between embryonic total mercury and several genes such as sepp1, which encodes selenoprotein P. Correlation between the biotransformation gene cyp1a4 and the C7 perfluoroalkyl carboxylic acid PFHpA was also evident. This study demonstrates that egg collection from free-living populations for contaminant biomonitoring programs can relate chemical residues to in ovo mRNA gene expression effects in embryo hepatic tissue.

Evaluation of human skin response to solar-simulated radiation in an ex vivo model: Effects and photoprotection of L-Carnosine.

Sunlight, and more specifically the UV component, induces several skin damages, including sunburns, erythema and photoaging. The purpose of this work is to set up an ex vivo human skin model to assess the capacity of active principles in protecting skin from the deleterious effects of solar radiation. Ex vivo human skin biopsies were cultured in an air-liquid interface and exposed to solar-simulated radiation (SSR, 300-750 nm). L-Carnosine (0.2% and 2%) was applied topically to be tested as photoprotective compound. The effect on oxidative stress induction, photoaging and skin transcriptional profile was assessed by evaluating reactive oxygen species, advanced glycosylation end products formation and gene expression changes. In our model, SSR increases ROS production and AGE accumulation and affects the expression of genes related to oxidative stress, pigmentation, immunity, inflammation and photoaging. Among these pathways, 11 genes were selected as biomarkers to evaluate the skin solar radiation response. Results showed that L-Carnosine provides effective prevention against solar radiation damages reducing ROS, AGEs and mitigating the modulation of the selected biomarker genes. In conclusion, we report that our ex vivo skin model is a valuable system to assess the consequences of solar light exposure and the capacity of topically applied L-Carnosine to counteract them.

Functional redundancy in response to runoff input upholds microbial community in hydrocarbon-contaminated land-sea continuum.

An experimental approach mimicking the land-sea continuum in microcosms was developed in order to determine the effect of the terrigenous inputs by soil runoff on the microbial functional potential in hydrocarbon (HC) contaminated marine coastal sediment. We hypothesized that the coalescent event increases the functional potential of microbial communities in marine coastal sediments, influencing the fate of HC in marine coastal ecosystems. The microbial functional potential including the HC degradation ability was assessed by DNA-array to compare the sediment receiving or not terrigenous inputs. The removal of HC and the functional gene richness in sediment was unchanged with the terrigenous inputs. However, the gene variants (GVs) composition was modified indicating functional redundancy. In addition, functional indicators including GVs related to sulfite reduction, denitrification and polyaromatic degradation were identified in higher proportion in sediment receiving terrigenous inputs. The terrigenous inputs modified the functional co-occurrence networks, showing a reorganization of the GVs associations with an increase of the network complexity. Different keystone GVs ensuring similar functions were identified in networks with or without terrigenous inputs, further confirming functional redundancy. We argue that functional redundancy maintains the structure of microbial community in hydrocarbon-contaminated land-sea continuum mixing zone. Our results provide helpful functional information for the monitoring and management of coastal environment affected by human land-based activities.

[Interferon-related gene array in predicting the efficacy of interferon therapy in chronic hepatitis B].

This study aims to clarify host factors of IFN treatment in the treatment of chronic hepatitis B (CHB) patients by screening the differentially expressed genes of IFN pathway CHB patients with different response to interferon (IFN) therapy. Three cases were randomly selected in IFN-responding CHB patients (Rs), non-responding CHB patients (NRs) and healthy participants, respectively. The human type I IFN response RT 2 profiler PCR array was used to detect the expression levels of IFN-related genes in peripheral blood monocytes (PBMCs) from healthy participants and CHB patients before and after Peg-IFN-α 2a treatment. The results showed that more differentially expressed genes appeared in Rs group than NRs group after IFN treatment. Comparing with healthy participants, IFNG, IL7R, IRF1, and IRF8 were downregulated in both Rs and NRs group before IFN treatment; CXCL10, IFIT1, and IFITM1 were upregulated in the Rs; IL13RA1 and IFI35 were upregulated in the NRs, while IFRD2, IL11RA, IL4R, IRF3, IRF4, PYHIN1, and ADAR were downregulated. The expression of IL15, IFI35 and IFI44 was downregulated by 4.09 ( t = 10.58, P < 0.001), 5.59 ( t = 3.37, P = 0.028) and 10.83 ( t = 2.8, P = 0.049) fold in the Rs group compared with the NRs group, respectively. In conclusion, IFN-response-related gene array is able to evaluate IFN treatment response by detecting IFN-related genes levels in PBMC. High expression of CXCL10, IFIT1 and IFITM1 before treatment may suggest satisfied IFN efficacy, while high expression of IL13RA1, IL15, IFI35 and IFI44 molecules and low expression of IFRD2, IL11RA, IL4R, IRF3, IRF4, PYHIN1 and ADAR molecules may be associated with poor IFN efficacy.

Interferon-related gene array in predicting the efficacy of interferon therapy in chronic hepatitis B / 生物医学工程学杂志

This study aims to clarify host factors of IFN treatment in the treatment of chronic hepatitis B (CHB) patients by screening the differentially expressed genes of IFN pathway CHB patients with different response to interferon (IFN) therapy. Three cases were randomly selected in IFN-responding CHB patients (Rs), non-responding CHB patients (NRs) and healthy participants, respectively. The human type I IFN response RT 2 profiler PCR array was used to detect the expression levels of IFN-related genes in peripheral blood monocytes (PBMCs) from healthy participants and CHB patients before and after Peg-IFN-α 2a treatment. The results showed that more differentially expressed genes appeared in Rs group than NRs group after IFN treatment. Comparing with healthy participants, IFNG, IL7R, IRF1, and IRF8 were downregulated in both Rs and NRs group before IFN treatment; CXCL10, IFIT1, and IFITM1 were upregulated in the Rs; IL13RA1 and IFI35 were upregulated in the NRs, while IFRD2, IL11RA, IL4R, IRF3, IRF4, PYHIN1, and ADAR were downregulated. The expression of IL15, IFI35 and IFI44 was downregulated by 4.09 ( t = 10.58, P < 0.001), 5.59 ( t = 3.37, P = 0.028) and 10.83 ( t = 2.8, P = 0.049) fold in the Rs group compared with the NRs group, respectively. In conclusion, IFN-response-related gene array is able to evaluate IFN treatment response by detecting IFN-related genes levels in PBMC. High expression of CXCL10, IFIT1 and IFITM1 before treatment may suggest satisfied IFN efficacy, while high expression of IL13RA1, IL15, IFI35 and IFI44 molecules and low expression of IFRD2, IL11RA, IL4R, IRF3, IRF4, PYHIN1 and ADAR molecules may be associated with poor IFN efficacy.

Genomic Diversity, Virulence Gene, and Prophage Arrays of Bovine and Human Shiga Toxigenic and Enteropathogenic Escherichia coli Strains Isolated in Hungary.

Escherichia coli belonging to the enterohemorrhagic (EHEC), Shiga toxin-producing (STEC) and atypical enteropathogenic (aEPEC) pathotypes are significant foodborne zoonotic pathogens posing serious health risks, with healthy cattle as their main reservoir. A representative sampling of Hungarian cattle farms during 2017-2018 yielded a prevalence of 6.5 and 5.8% for STEC and aEPEC out of 309 samples. The draft genomes of twelve STEC (of them 9 EHEC) and four aEPEC of bovine origin were determined. For comparative purposes, we also included 3 EHEC and 2 aEPEC strains of human origin, as well four commensal isolates and one extraintestinal pathogenic E. coli (ExPEC) obtained from animals in a final set of 26 strains for a WGS-based analysis. Apart from key virulence genes, these isolates harbored several additional virulence genes with arrays characteristic for the site of isolation. The most frequent insertion site of Shiga toxin (stx) encoding prophages was yehV for the Stx1 prophage and wrbA and sbcB for Stx2. For O157:H7 strains, the locus of enterocyte effacement pathogenicity island was present at the selC site, with integration at pheV for other serotypes, and pheU in the case of O26:H11 strains. Several LEE-negative STEC and aEPEC as well as commensal isolates carried additional prophages, with an average of ten prophage regions per isolate. Comparative phylogenomic analysis showed no clear separation between bovine and human lineages among the isolates characterized in the current study. Similarities in virulence gene arrays and close phylogenetic relations of bovine and human isolates underline the zoonotic potential of bovine aEPEC and STEC and emphasize the need for frequent monitoring of these pathogens in livestock.

Cold acclimation diversity in Arabidopsis thaliana: CRISPR/Cas9 as a tool to fine analysis of Tandem Gene Arrays, application to CBF genes.

In this period of climate change, it is of major importance to increase knowledge about the mechanisms by whose plants adapt to their environment. Tandem gene arrays (TAG) are overrepresented in the pool of tandem duplicates involved in stress response and are consequently of special interest. Nevertheless, until recently, addressing questions about individual genes or fine regulations in such structures was very difficult due to the close arrangement of the genome, almost preventing the production of targeted simple or multiple mutants. The CRISPR/Cas9 new tool offers new opportunities as the setting of gene deletion strategies in various genetic backgrounds. Here, we used this technology on the cold acclimation CBF pathway in different accessions of Arabidopsis thaliana with the same set of guide RNAs. Deleted lines free of T-DNA have been produced for simple or multiple copies of CBF genes and evaluated for cold tolerance after acclimation. Expression levels of CBF genes and five COR genes have also been analyzed. Our data suggest first that two or three missing CBF genes are necessary to induce a strong reduction in cold tolerance and secondly that most deletions have a low impact on the expression of remaining CBF copies which contradicts the previous hypothesis in the literature. Our results thus show that the CRISPR/Cas9 deletion strategy is a useful performance tool to investigate how tandem gene arrays work.

Molecular diagnosis of ABMR with or without donor-specific antibody in kidney transplant biopsies: Differences in timing and intensity but similar mechanisms and outcomes.

We studied the clinical, histologic, and molecular features distinguishing DSA-negative from DSA-positive molecularly defined antibody-mediated rejection (mABMR). We analyzed mABMR biopsies with available DSA assessments from the INTERCOMEX study: 148 DSA-negative versus 248 DSA-positive, compared with 864 no rejection (excluding TCMR and Mixed). DSA-positivity varied with mABMR stage: early-stage (EABMR) 56%; fully developed (FABMR) 70%; and late-stage (LABMR) 58%. DSA-negative patients with mABMR were usually sensitized, 60% being HLA antibody-positive. Compared with DSA-positive mABMR, DSA-negative mABMR was more often C4d-negative; earlier by 1.5 years (average 2.4 vs. 3.9 years); and had lower ABMR activity and earlier stage in molecular and histology features. However, the top ABMR-associated transcripts were identical in DSA-negative versus DSA-positive mABMR, for example, NK-associated (e.g., KLRD1 and GZMB) and IFNG-inducible (e.g., PLA1A). Genome-wide class comparison between DSA-negative and DSA-positive mABMR showed no significant differences in transcript expression except those related to lower intensity and earlier time of DSA-negative ABMR. Three-year graft loss in DSA-negative mABMR was the same as DSA-positive mABMR, even after adjusting for ABMR stage. Thus, compared with DSA-positive mABMR, DSA-negative mABMR is on average earlier, less active, and more often C4d-negative but has similar graft loss, and genome-wide analysis suggests that it involves the same mechanisms. SUMMARY SENTENCE: In 398 kidney transplant biopsies with molecular antibody-mediated rejection, the 150 DSA-negative cases are earlier, less intense, and mostly C4d-negative, but use identical molecular mechanisms and have the same risk of graft loss as the 248 DSA-positive cases.

Evolving the surveillance and workup of heart transplant rejection: A real-world analysis of the Molecular Microscope Diagnostic System.

The Molecular Microscope Diagnostic System (MMDx) analyzes RNA transcripts of transplanted heart tissue to differentiate among T cell-mediated rejection (TCMR), antibody-mediated rejection (AMR), injury, and healthy tissue. However, little is known about its performance in relation to other modalities in a real-world heart transplant population. We evaluated whether MMDx performs in agreement with other validated modalities. Two hundred and twenty-eight corresponding endomyocardial biopsies (EMBx) and MMDx specimens from 135 adult heart transplant patients were retrospectively reviewed with correlating donor-derived cell-free DNA (dd-cfDNA). Rejection was classified on EMBx in 29 specimens (TCMR ≥ 2R and/or AMR ≥ 1), on MMDx in 56 specimens, and in 74 values with dd-cfDNA ≥0.20%. Despite moderate agreement between EMBx and MMDx (84% agreement, Cohen's kappa, 0.48, p < .001), systematic differences were observed (McNemar's test, p < .001) where MMDx classified 32 of 37 discordant cases as rejection. MMDx and dd-cfDNA demonstrated slight agreement (72% agreement, Cohen's kappa, 0.39, p < .001); however, systematic differences were also apparent where MMDx classified 12 of 50 discordant specimens as rejection when dd-cfDNA was <0.20% (McNemar's test, p < .001). Our findings provide insight on the performance of MMDx relative to other modalities in a heart transplant cohort and have implications on the surveillance and workup of allograft rejection in heart transplantation.

Hyperbaric Oxygen Therapy after Mid-Cervical Spinal Contusion Injury.

Hyperbaric oxygen (HBO) therapy is frequently used to treat peripheral wounds or decompression sickness. Evidence suggests that HBO therapy can provide neuroprotection and has an anti-inflammatory impact after neurological injury, including spinal cord injury (SCI). Our primary purpose was to conduct a genome-wide screening of mRNA expression changes in the injured spinal cord after HBO therapy. An mRNA gene array was used to evaluate samples taken from the contused region of the spinal cord following a lateralized mid-cervical contusion injury in adult female rats. HBO therapy consisted of daily, 1-h sessions (3.0 ATA, 100% O2) initiated on the day of SCI and continued for 10 days. Gene set enrichment analyses indicated that HBO upregulated genes in pathways associated with electron transport, mitochondrial function, and oxidative phosphorylation, and downregulated genes in pathways associated with inflammation (including cytokines and nuclear factor kappa B [NF-κB]) and apoptotic signaling. In a separate cohort, spinal cord histology was performed to verify whether the HBO treatment impacted neuronal cell counts or inflammatory markers. Compared with untreated rats, there were increased NeuN positive cells in the spinal cord of HBO-treated rats (p = 0.004). We conclude that HBO therapy, initiated shortly after SCI and continued for 10 days, can alter the molecular signature of the lesioned spinal cord in a manner consistent with a neuroprotective impact.

Utilizing Two Borrelia bavariensis Isolates Naturally Lacking the PFam54 Gene Array To Elucidate the Roles of PFam54-Encoded Proteins.

Lyme borreliosis is the most common vector-borne disease in the Northern Hemisphere, caused by spirochetes belonging to the Borrelia burgdorferi sensu lato species complex, which are transmitted by ixodid ticks. B. burgdorferi sensu lato species produce a family of proteins on the linear plasmid 54 (PFam54), some of which confer the functions of cell adhesion and inactivation of complement, the first line of host defense. However, the impact of PFam54 in promoting B. burgdorferi sensu lato pathogenesis remains unclear because of the hurdles to simultaneously knock out all PFam54 proteins in a spirochete. Here, we describe two Borrelia bavariensis strains, PBN and PNi, isolated from patients naturally lacking PFam54 but maintaining the rest of the genome with greater than 95% identity to the reference B. bavariensis strain, PBi. We found that PBN and PNi less efficiently survive in human serum than PBi. Such defects were restored by introducing two B. bavariensis PFam54 recombinant proteins, BGA66 and BGA71, confirming the role of these proteins in providing complement evasion of B. bavariensis. Further, we found that all three strains remain detectable in various murine tissues 21 days post-subcutaneous infection, supporting the nonessential role of B. bavariensis PFam54 in promoting spirochete persistence. This study identified and utilized isolates deficient in PFam54 to associate the defects with the absence of these proteins, building the foundation to further study the role of each PFam54 protein in contributing to B. burgdorferi sensu lato pathogenesis. IMPORTANCE To establish infections, Lyme borreliae utilize various means to overcome the host's immune system. Proteins encoded by the PFam54 gene array play a role in spirochete survival in vitro and in vivo. Moreover, this gene array has been described in all currently available Lyme borreliae genomes. By investigating the first two Borrelia bavariensis isolates naturally lacking the entire PFam54 gene array, we showed that both patient isolates display an increased susceptibility to human serum, which can be rescued in the presence of two PFam54 recombinant proteins. However, both isolates remain infectious to mice after intradermal inoculation, suggesting the nonessential role of PFam54 during the long-term, but may differ slightly in the colonization of specific tissues. Furthermore, these isolates show high genomic similarity to type strain PBi (>95%) and could be used in future studies investigating the role of each PFam54 protein in Lyme borreliosis pathogenesis.

The molecular phenotypes of injury, steatohepatitis, and fibrosis in liver transplant biopsies in the INTERLIVER study.

To extend previous molecular analyses of rejection in liver transplant biopsies in the INTERLIVER study (ClinicalTrials.gov #NCT03193151), the present study aimed to define the gene expression selective for parenchymal injury, fibrosis, and steatohepatitis. We analyzed genome-wide microarray measurements from 337 liver transplant biopsies from 13 centers. We examined expression of genes previously annotated as increased in injury and fibrosis using principal component analysis (PCA). PC1 reflected parenchymal injury and related inflammation in the early posttransplant period, slowly regressing over many months. PC2 separated early injury from late fibrosis. Positive PC3 identified a distinct mildly inflamed state correlating with histologic steatohepatitis. Injury PCs correlated with liver function and histologic abnormalities. A classifier trained on histologic steatohepatitis predicted histologic steatohepatitis with cross-validated AUC = 0.83, and was associated with pathways reflecting metabolic abnormalities distinct from fibrosis. PC2 predicted histologic fibrosis (AUC = 0.80), as did a molecular fibrosis classifier (AUC = 0.74). The fibrosis classifier correlated with matrix remodeling pathways with minimal overlap with those selective for steatohepatitis, although some biopsies had both. Genome-wide assessment of liver transplant biopsies can not only detect molecular changes induced by rejection but also those correlating with parenchymal injury, steatohepatitis, and fibrosis, offering potential insights into disease mechanisms for primary diseases.

Mechanistic Target of Rapamycin Complex 2 Regulation of the Primary Human Trophoblast Cell Transcriptome.

Mechanistic Target of Rapamycin Complex 2 (mTORC2) regulates placental amino acid and folate transport. However, the role of mTORC2 in modulating other placental functions is largely unexplored. We used a gene array following the silencing of rictor to identify genes regulated by mTORC2 in primary human trophoblast (PHT) cells. Four hundred and nine genes were differentially expressed; 102 genes were down-regulated and 307 up-regulated. Pathway analyses demonstrated that inhibition of mTORC2 resulted in increased expression of genes encoding for pro-inflammatory IL-6, VEGF-A, leptin, and inflammatory signaling (SAPK/JNK). Furthermore, down-regulated genes were functionally enriched in genes involved in angiogenesis (Osteopontin) and multivitamin transport (SLC5A6). In addition, the protein expression of leptin, VEGFA, IL-6 was increased and negatively correlated to mTORC2 signaling in human placentas collected from pregnancies complicated by intrauterine growth restriction (IUGR). In contrast, the protein expression of Osteopontin and SLC5A6 was decreased and positively correlated to mTORC2 signaling in human IUGR placentas. In conclusion, mTORC2 signaling regulates trophoblast expression of genes involved in inflammation, micronutrient transport, and angiogenesis, representing novel links between mTOR signaling and multiple placental functions necessary for fetal growth and development.

Excitotoxic stimulation activates distinct pathogenic and protective expression signatures in the hippocampus.

Excitotoxic events underlying ischaemic and traumatic brain injuries activate degenerative and protective pathways, particularly in the hippocampus. To understand opposing pathways that determine the brain's response to excitotoxicity, we used hippocampal explants, thereby eliminating systemic variables during a precise protocol of excitatory stimulation. N-methyl-d-aspartate (NMDA) was applied for 20 min and total RNA isolated one and 24 h later for neurobiology-specific microarrays. Distinct groups of genes exhibited early vs. delayed induction, with 63 genes exclusively reduced 24-h post-insult. Egr-1 and NOR-1 displayed biphasic transcriptional modulation: early induction followed by delayed suppression. Opposing events of NMDA-induced genes linked to pathogenesis and cell survival constituted the early expression signature. Delayed degenerative indicators (up-regulated pathogenic genes, down-regulated pro-survival genes) and opposing compensatory responses (down-regulated pathogenic genes, up-regulated pro-survival genes) generated networks with temporal gene profiles mirroring coexpression network clustering. We then used the expression profiles to test whether NF-κB, a potent transcription factor implicated in both degenerative and protective pathways, is involved in the opposing responses. The NF-κB inhibitor MG-132 indeed altered NMDA-mediated transcriptional changes, revealing components of opposing expression signatures that converge on the single response element. Overall, this study identified counteracting avenues among the distinct responses to excitotoxicity, thereby suggesting multi-target treatment strategies and implications for predictive medicine.

TNMplot.com: A Web Tool for the Comparison of Gene Expression in Normal, Tumor and Metastatic Tissues.

Genes showing higher expression in either tumor or metastatic tissues can help in better understanding tumor formation and can serve as biomarkers of progression or as potential therapy targets. Our goal was to establish an integrated database using available transcriptome-level datasets and to create a web platform which enables the mining of this database by comparing normal, tumor and metastatic data across all genes in real time. We utilized data generated by either gene arrays from the Gene Expression Omnibus of the National Center for Biotechnology Information (NCBI-GEO) or RNA-seq from The Cancer Genome Atlas (TCGA), Therapeutically Applicable Research to Generate Effective Treatments (TARGET), and The Genotype-Tissue Expression (GTEx) repositories. The altered expression within different platforms was analyzed separately. Statistical significance was computed using Mann-Whitney or Kruskal-Wallis tests. False Discovery Rate (FDR) was computed using the Benjamini-Hochberg method. The entire database contains 56,938 samples, including 33,520 samples from 3180 gene chip-based studies (453 metastatic, 29,376 tumorous and 3691 normal samples), 11,010 samples from TCGA (394 metastatic, 9886 tumorous and 730 normal), 1193 samples from TARGET (1 metastatic, 1180 tumorous and 12 normal) and 11,215 normal samples from GTEx. The most consistently upregulated genes across multiple tumor types were TOP2A (FC = 7.8), SPP1 (FC = 7.0) and CENPA (FC = 6.03), and the most consistently downregulated gene was ADH1B (FC = 0.15). Validation of differential expression using equally sized training and test sets confirmed the reliability of the database in breast, colon, and lung cancer at an FDR below 10%. The online analysis platform enables unrestricted mining of the database and is accessible at TNMplot.com.

Transcriptomic Changes Associated with Loss of Cell Viability Induced by Oxysterol Treatment of a Retinal Photoreceptor-Derived Cell Line: An In Vitro Model of Smith-Lemli-Opitz Syndrome.

Smith-Lemli-Opitz Syndrome (SLOS) results from mutations in the gene encoding the enzyme DHCR7, which catalyzes conversion of 7-dehydrocholesterol (7DHC) to cholesterol (CHOL). Rats treated with a DHCR7 inhibitor serve as a SLOS animal model, and exhibit progressive photoreceptor-specific cell death, with accumulation of 7DHC and oxidized sterols. To understand the basis of this cell type specificity, we performed transcriptomic analyses on a photoreceptor-derived cell line (661W), treating cells with two 7DHC-derived oxysterols, which accumulate in tissues and bodily fluids of SLOS patients and in the rat SLOS model, as well as with CHOL (negative control), and evaluated differentially expressed genes (DEGs) for each treatment. Gene enrichment analysis and compilation of DEG sets indicated that endoplasmic reticulum stress, oxidative stress, DNA damage and repair, and autophagy were all highly up-regulated pathways in oxysterol-treated cells. Detailed analysis indicated that the two oxysterols exert their effects via different molecular mechanisms. Changes in expression of key genes in highlighted pathways (Hmox1, Ddit3, Trib3, and Herpud1) were validated by immunofluorescence confocal microscopy. The results extend our understanding of the pathobiology of retinal degeneration and SLOS, identifying potential new druggable targets for therapeutic intervention into these and other related orphan diseases.

Donor genetic variants as risk factors for thrombosis after liver transplantation: A genome-wide association study.

Thrombosis after liver transplantation substantially impairs graft- and patient survival. Inevitably, heritable disorders of coagulation originating in the donor liver are transmitted by transplantation. We hypothesized that genetic variants in donor thrombophilia genes are associated with increased risk of posttransplant thrombosis. We genotyped 775 donors for adult recipients and 310 donors for pediatric recipients transplanted between 1993 and 2018. We determined the association between known donor thrombophilia gene variants and recipient posttransplant thrombosis. In addition, we performed a genome-wide association study (GWAS) and meta-analyzed 1085 liver transplantations. In our donor cohort, known thrombosis risk loci were not associated with posttransplant thrombosis, suggesting that it is unnecessary to exclude liver donors based on thrombosis-susceptible polymorphisms. By performing a meta-GWAS from children and adults, we identified 280 variants in 55 loci at suggestive genetic significance threshold. Downstream prioritization strategies identified biologically plausible candidate genes, among which were AK4 (rs11208611-T, p = 4.22 × 10-05 ) which encodes a protein that regulates cellular ATP levels and concurrent activation of AMPK and mTOR, and RGS5 (rs10917696-C, p = 2.62 × 10-05 ) which is involved in vascular development. We provide evidence that common genetic variants in the donor, but not previously known thrombophilia-related variants, are associated with increased risk of thrombosis after liver transplantation.