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Thioredoxin-interacting protein (TXNIP), also known as Vitamin-D upregulated protein-1 (VDUP-1), interacts with thioredoxin to regulate redox responses and participates in diverse disorders including metabolic, cardiovascular, inflammatory and malignant diseases. Psoriasis is characterized by chronic skin inflammation and an aberrant pattern of keratinocyte differentiation. Clinically, psoriasis is associated with various cardiometabolic comorbidities but studies on TXNIP's biological role in skin disorders are limited. In this study, we investigated TXNIP expression in psoriasis and its regulation in normal human epidermal keratinocytes (NHEKs), and then explored how TXNIP regulated skin keratinocyte differentiation to determine its role in psoriasis pathogenesis. Our immunohistochemical study demonstrated extensive TXNIP expression in the upper and lower epidermis of psoriasis compared to predominant TXNIP expression in the basal layer of normal skin. 1, 25-dihydroxyvitamin D

Poly(ADP-ribose) polymerase-1 (PARP-1) plays an essential role in DNA repair by catalyzing the polymerization of ADP-ribose unit to target proteins. Several studies have shown that PARP-1 can regulate inflammatory responses in various disease models. The intracellular Nod-like receptor NLRP3 has emerged as the most crucial innate immune receptor because of its broad specificity in mediating immune response to pathogen invasion and danger signals associated with cellular damage. In our study, we found NLRP3 stimuli-induced caspase-1 maturation and IL-1β production were impaired by PARP-1 knockout or PARP-1 inhibition in bone marrow-derived macrophages (BMDM). The step 1 signal of NLRP3 inflammasome activation was not affected by PARP-1 deficiency. Moreover, ATP-induced cytosolic ROS production was lower in Parp-1−/− BMDM, resulting in the decreased inflammasome complex assembly. PARP-1 can translocate to cytosol upon ATP stimulation and trigger the PARylation modification on NLRP3, leading to NLRP3 inflammasome assembly. PARP-1 was also a bridge between NLRP3 and thioredoxin-interacting protein (TXNIP) and participated in NLRP3/TXNIP complex formation for inflammasome activation. Overall, PARP-1 positively regulates NLRP3 inflammasome activation via increasing ROS production and interaction with TXNIP and NLRP3, leading to PARylation of NLRP3. Our data demonstrate a novel regulatory mechanism for NLRP3 inflammasome activation by PARP-1. Therefore, PARP-1 can serve as a potential target in the treatment of IL-1β associated inflammatory diseases. Supplementary Information The online version contains supplementary material available at 10.1007/s00018-022-04138-z.

CoMut plot is widely used in cancer research publications as a visual summary of mutational landscapes in cancer cohorts. This summary plot can inspect gene mutation rate and sample mutation burden with their relevant clinical details, which is a common first step for analyzing the recurrence and co-occurrence of gene mutations across samples. The cBioPortal and iCoMut are two web-based tools that allow users to create intricate visualizations from pre-loaded TCGA and ICGC data. For custom data analysis, only limited command-line packages are available now, making the production of CoMut plots difficult to achieve, especially for researchers without advanced bioinformatics skills. To address the needs for custom data and TCGA/ICGC data comparison, we have created CoMutPlotter, a web-based tool for the production of publication-quality graphs in an easy-of-use and automatic manner. We introduce a web-based tool named CoMutPlotter to lower the barriers between complex cancer genomic data and researchers, providing intuitive access to mutational profiles from TCGA/ICGC projects as well as custom cohort studies. A wide variety of file formats are supported by CoMutPlotter to translate cancer mutation profiles into biological insights and clinical applications, which include Mutation Annotation Format (MAF), Tab-separated values (TSV) and Variant Call Format (VCF) files. In summary, CoMutPlotter is the first tool of its kind that supports VCF file, the most widely used file format, as its input material. CoMutPlotter also provides the most-wanted function for comparing mutation patterns between custom cohort and TCGA/ICGC project. Contributions of COSMIC mutational signatures in individual samples are also included in the summary plot, which is a unique feature of our tool. CoMutPlotter is freely available at http://tardis.cgu.edu.tw/comutplotter .

UVB dosage is generally regarded as the most critical factor that determines the severity of UVB-induced skin erythema. However, recent studies have demonstrated that different UV irradiances induce varying biological responses in mouse skin even at constant UV doses. UVB-induced inflammasome activation is particularly observed in human skin keratinocytes, which are classified as immunocompetent cells, but not in mouse skin keratinocytes, which do not express sufficient inflammasome complex components. In human skin UVB-induced sunburn reactions, NLRP1 inflammasome activation critically mediates the inflammatory responses. Here, we employed primary human skin keratinocytes to explore the impact of different irradiances of a constant UVB dosage on inflammasome activation and related inflammatory responses. Our findings indicated that low-irradiance UVB induced relatively stronger NLRP1 inflammasome activation, which manifested as more active IL-1β, IL-18 release, and enhanced procaspase-1 cleavage compared to high-irradiance UVB at the same dose. Irradiance did not influence cell lysis or the expression of inflammasome complex proteins including NLRP1, proIL-1β, proIL-18, procaspase-1, and ASC. The UVB-induced TNF-α and cyclooxygenase-2 expression was also relatively higher in keratinocytes exposed to low-irradiance UVB. Low-irradiance UVB also increased reactive oxygen species production. UVB-triggered signaling analysis revealed that low-irradiance UVB resulted in more prominent p38 and JNK activation. Therefore, our findings indicated that, in addition to the role of total dosage, irradiance crucially modulates UVB-elicited inflammation in human skin keratinocytes, thus providing novel insights into human skin photobiology.

UVB can induce inflammatory responses contributing to diverse skin damage. UVB-triggered inflammasome activation of human keratinocytes underlies UVB-induced skin sunburn reaction. Pleiotropic functions of spleen tyrosine kinase (Syk) have rendered it as a potential therapeutic target. In immunocytes, Syk modulates immunoreceptor signaling and NLRP3 inflammasome activation. In skin, Syk mediates EGFR signaling, regulates keratinocyte differentiation and is involved in inflammatory disorders. However, roles of Syk in UVB-induced inflammasome activation in keratinocytes remain elusive. We investigated roles of keratinocyte Syk in UVB-triggered photo-responses. Primary normal human epidermal keratinocytes (NHEKs) isolated from skin were used. Syk knockdown or Syk inhibitor R406 was applied to investigate functions of keratinocyte Syk in UVB photobiology. The possible in vivo role of Syk was evaluated by checking UVB-induced skin damage in R406-treated mice. UVB was able to induce Syk phosphorylation in NHEKs that could be regulated by reactive oxygen species (ROS) generation and EGFR. Syk knockdown or Syk inhibitor (R406) treatment reduced UVB-triggered apoptosis-associated speck-like protein containing a caspase-recruitment domain (ASC) crosslinking, procaspase-1 cleavage, active IL-1β formation, and gasdermin D activation, indicating roles of Syk in UVB-triggered inflammasome activation in keratinocytes. UVB-induced production of IL-8, TNF-α, ROS, and phosphorylation of JNK and p38 were attenuated after Syk knockdown or inhibition. R406 ameliorated UVB-induced mouse skin damage, including erythema and transepidermal water loss (TEWL). Thus, Syk participated in UVB-induced inflammasome activation and inflammatory response in vitro and in vivo, suggesting potential photo-protective effects of Syk inhibition in UVB-induced skin inflammation.

UV irradiation can injure the epidermis, resulting in sunburn, inflammation, and cutaneous tissue disorders. Previous studies demonstrate that EGFR in keratinocytes can be activated by UVB and contributes to inflammation. Poly (ADP-ribose) polymerase-1 (PARP-1) is a nuclear enzyme and plays an essential role in DNA repair under moderate stress. In this study, we set out to understand how PARP-1 regulates UVB irradiation-induced skin injury and interplays with EGFR to mediate the inflammation response. We found that PARP-1 deficiency exacerbated the UVB-induced inflammation, water loss, and back skin damage in mice. In human primary keratinocytes, UVB can activate PARP-1 and enhance DNA damage upon PARP-1 gene silencing. Moreover, PARP-1 silencing and PARP inhibitor olaparib can suppress UVB-induced COX-2 and MMP-1 expression, but enhance TNF-α and IL-8 expression. In addition, EGFR silencing or EGFR inhibition by gefitinib can decrease UVB-induced COX-2, TNF-α, and IL-8 expression, suggesting EGFR activation via paracrine action can mediate UVB-induced inflammation responses. Immunoblotting data revealed that PARP-1 inhibition decreases UVB-induced EGFR and p38 activation. Pharmacological inhibition of p38 also dramatically led to the attenuation of UVB-induced inflammatory gene expression. Of note, genetic ablation of PARP-1 or EGFR can attenuate UVB-induced ROS production, and antioxidant NAC can attenuate UVB-induced EGFR-p38 signaling axis and PARP-1 activation. These data suggest the regulatory loops among EGFR, PARP-1, and ROS upon UVB stress. PARP-1 not only serves DNA repair function but also orchestrates interactions to EGFR transactivation and ROS production, leading to p38 signaling for inflammatory gene expression in keratinocytes.

Differential diagnosis of intrahepatic cholangiocarcinoma (iCCA) from its histologic mimickers, especially metastatic adenocarcinomas of gastric and pancreatic origin, is a great challenge for pathologists. In this study, through bioinformatics analysis of data from The Cancer Genome Atlas and Gene Expression Omnibus, we identified C-reactive protein (CRP) as a candidate marker to differentiate iCCA from other adenocarcinomas and validated its diagnostic performance by immunohistochemistry in a large cohort of clinical samples including 103 iCCAs, 384 other adenocarcinomas, and 34 liver metastases of various origins. The sensitivity and specificity of CRP expression in the diagnosis of iCCA were 75.7% and 91.1% when using tissue microarrays and 93.3% and 88.2% when using whole tissue sections, respectively. We also compared the diagnostic performance of CRP with N-cadherin, a previously reported marker for iCCA. The sensitivity and specificity of N-cadherin were 54.4% and 92.2% when using tissue microarrays and 80.0% and 88.2% when using whole tissue sections, respectively. The sensitivity of CRP was higher than that of N-cadherin, whereas their specificity was similar. CRP expression was associated with mass-forming gross type (P

Cancer is a genetic disease caused by somatic mutations; however, the understanding of the causative biological processes generating these mutations is limited. A cancer genome bears the cumulative effects of mutational processes during tumor development. Deciphering mutational signatures in cancer is a new topic in cancer research. The Wellcome Trust Sanger Institute (WTSI) has categorized 30 reference signatures in the COSMIC database based on the analyses of ∼10 000 sequencing datasets from TCGA and ICGC. Large cohorts and bioinformatics skills are required to perform the same analysis as WTSI. The quantification of known signatures in custom cohorts is not possible under the current framework of the COSMIC database, which motivates us to construct a database for mutational signatures in cancers and make such analyses more accessible to general researchers. mSignatureDB (http://tardis.cgu.edu.tw/msignaturedb) integrates R packages and in-house scripts to determine the contributions of the published signatures in 15 780 individual tumors from 73 TCGA/ICGC cancer projects, making comparison of signature patterns within and between projects become possible. mSignatureDB also allows users to perform signature analysis on their own datasets, quantifying contributions of signatures at sample resolution, which is a unique feature of mSignatureDB not available in other related databases.

Background Galectin-3 is widely expressed in many immunocytes and epithelial cells including skin keratinocytes. Galectin-3 can regulate immunological or inflammatory processes and plays a proinflammatory role in some disease models. Galectin-3 has a role in disorders related to ultraviolet (UV) photodamage such as apoptosis, skin squamous cell carcinoma and basal cell carcinoma. However, the evidence of galectin-3 in UVB-induced skin inflammation is still limited and the underlying molecular mechanism remains elusive. Objective We aimed to investigate the effects of galectin-3 in human epidermal keratinocytes and in mice after UVB irradiation. Methods Primary human epidermal keratinocytes with galectin-3 knockdown were used as the in vitro model. ELISA, QPCR, and western blotting were applied to evaluate the released cytokine, mRNA and protein expression. Histologic analysis, measurement of erythema and transepidermal water loss (TEWL) were applied to evaluate UVB-induced skin damage in galectin-3 knockout mice. Results In UVB-irradiated human keratinocytes, galectin-3 knockdown downregulated the UVB-induced ASC crosslinking, cleavage of caspase-1, and formation of active IL-1β. Galectin-3 knockdown also decreased UVB-induced production of reactive oxygen species, p38 phosphorylation, and COX2 expression in human keratinocytes. After four days of UVB irradiation, galectin-3 knockout mice showed reduced gross erythema, histologic features of tissue inflammation, quantified levels of erythema and TEWL compared to wild type mice. The skin tissue lysate also showed less expression of active IL-1β and COX2 in galectin-3 knockout mice. Conclusion Galectin-3 may play a positive regulatory role in UVB-induced skin inflammation.

High throughput sequencing technologies have been an increasingly critical aspect of precision medicine owing to a better identification of disease targets, which contributes to improved health care cost and clinical outcomes. In particular, disease-oriented targeted enrichment sequencing is becoming a widely-accepted application for diagnostic purposes, which can interrogate known diagnostic variants as well as identify novel biomarkers from panels of entire human coding exome or disease-associated genes. We introduce a workflow named VAReporter to facilitate the management of variant assessment in disease-targeted sequencing, the identification of pathogenic variants, the interpretation of biological effects and the prioritization of clinically actionable targets. State-of-art algorithms that account for mutation phenotypes are used to rank the importance of mutated genes through visual analytic strategies. We established an extensive annotation source by integrating a wide variety of biomedical databases and followed the American College of Medical Genetics and Genomics (ACMG) guidelines for interpretation and reporting of sequence variations. In summary, VAReporter is the first web server designed to provide a “one-stop” resource for individual’s diagnosis and large-scale cohort studies, and is freely available at http://rnd.cgu.edu.tw/vareporter .

The treatment of L929 fibrosarcoma cells with zVAD has been shown to induce necroptosis. However, whether autophagy is involved or not in this event remains controversial. In this study, we re-examined the role of autophagy in zVAD-induced cell death in L929 cells and further elucidated the signaling pathways triggered by caspase inhibition and contributing to autophagic death. First, we found that zVAD can stimulate LC3-II formation, autophagosome and autolysosome formation, and ROS accumulation. Antioxidants, beclin 1 or Atg5 silencing, and class III PtdIns3K inhibitors all effectively blocked ROS production and cell death, suggesting ROS accumulation downstream of autophagy contributes to cell necrosis. zVAD also stimulated PARP activation, and the PARP inhibitor DPQ can reduce zVAD-induced cell death, but did not affect ROS production, suggesting the increased ROS leads to PARP activation and cell death. Notably, our data also indicated the involvement of Src-dependent JNK and ERK in zVAD-induced ROS production and autophagic death. We found caspase 8 is associated with c-Src at the resting state, and upon zVAD treatment this association was decreased and accompanied by c-Src activation. In conclusion, we confirm the autophagic death in zVAD-treated L929 cells, and define a new molecular pathway in which Src-dependent ERK and JNK activation can link a signal from caspase inhibition to autophagy, which in turn induce ROS production and PARP activation, eventually leading to necroptosis. Thus, in addition to initiating proteolytic activity for cell apoptosis, inactivated caspase 8 also functions as a signaling molecule for autophagic death.

The alkylating agent N-methyl-N'-nitro-N'-nitrosoguanidine (MNNG) can cause excess DNA strand breaks that lead to poly(ADP-ribose)polymerase-1 (PARP-1) overactivation and cell death (parthanatos). However, the detail mechanism of MNNG-induced parthanatos was not well-investigated. In this study, we used MNNG-treated mouse embryonic fibroblasts (MEFs) to elucidate the signaling pathways of MNNG-induced parthanatos. We found that MNNG-induced cell death accompanied by rapid PARP-1 activation, c-Jun N-terminal kinase (JNK) activation, biphasic reactive oxygen species (ROS) production and intracellular calcium increase. The early ROS production occurring at 1 min and peaking at 5-15 min after MNNG treatment partially resulted from NADPH oxidase. In contrast, the late phase of ROS production occurring at 30 min and time-dependently increasing up to 6h after MNNG treatment was generated by mitochondria. The antioxidant, NAC can abrogate all phenomena caused by MNNG. Results indicate that the calcium rise was downstream of early ROS production, and was involved in PARP-1 and JNK activation. Moreover, the PARP inhibitor was able to reduce MNNG-induced late-phase ROS production, calcium elevation, and cell death. Results further indicated the involvement of RIP1 in sustained ROS production and calcium increase. We characterized the interactive roles of ROS, calcium, JNK, and RIP1 in MNNG-induced cell death. We found that in addition to the alkylating property previously demonstrated, ROS production triggered by MNNG results in enhanced DNA damage and PARP-1 activation. Moreover, intracellular calcium elevation and ROS production have mutual amplification effects and thus contribute to PARP-1-mediated parthanatos.

Ahead of Print article withdrawn by publisher. A previous report showed that the pan-caspase inhibitor zVAD can induce necrosis accompanied by autophagosome formation in L929 fibrosarcoma cells. Such autophagic cell death relies on caspase 8 inhibition and ROS accumulation. Since the connection of these molecules is still poorly understood, we explored the underlying signaling cascades in this event. First, we confirmed zVAD can stimulate LC3 cleavage, beclin 1 gene expression, autophagosome formation, and ROS accumulation in L929 cells. Antioxidants, Beclin 1 or Atg5 silencing, and class III PtdIns3K inhibitors all effectively blocked ROS production and cell death, suggesting ROS accumulation downstream of autophagy contributes to cell necrosis. zVAD also stimulated PARP activation, and the PARP inhibitor DPQ can reduce zVAD-induced cell death, but not affect ROS production, suggesting the increased ROS leads to PARP activation and cell death. Notably, our data also indicated the involvement of Src-dependent JNK and ERK in zVAD-induced autophagic cell death. We found caspase 8 is associated with c-Src at resting state, and upon zVAD treatment this association is decreased and accompanied by c-Src activation. These results provide new insight into the nonenzymatic function of caspase 8. In addition to initiating proteolytic activity for cell apoptosis, inactivated caspase 8 also functions as a signaling molecule for autophagic cell death.