HiPSC-derived 3D neural models reveal neurodevelopmental pathomechanisms of the Cockayne Syndrome B.

Cockayne Syndrome B (CSB) is a hereditary multiorgan syndrome which-through largely unknown mechanisms-can affect the brain where it clinically presents with microcephaly, intellectual disability and demyelination. Using human induced pluripotent stem cell (hiPSC)-derived neural 3D models generated from CSB patient-derived and isogenic control lines, we here provide explanations for these three major neuropathological phenotypes. In our models, CSB deficiency is associated with (i) impaired cellular migration due to defective autophagy as an explanation for clinical microcephaly; (ii) altered neuronal network functionality and neurotransmitter GABA levels, which is suggestive of a disturbed GABA switch that likely impairs brain circuit formation and ultimately causes intellectual disability; and (iii) impaired oligodendrocyte maturation as a possible cause of the demyelination observed in children with CSB. Of note, the impaired migration and oligodendrocyte maturation could both be partially rescued by pharmacological HDAC inhibition.

Vemurafenib acts as an aryl hydrocarbon receptor antagonist: Implications for inflammatory cutaneous adverse events.

BACKGROUND: In recent years, the BRAF inhibitor vemurafenib has been successfully established in the therapy of advanced melanoma. Despite its superior efficacy, the use of vemurafenib is limited by frequent inflammatory cutaneous adverse events that affect patients' quality of life and may lead to dose reduction or even cessation of anti-tumor therapy. To date, the molecular and cellular mechanisms of vemurafenib-induced rashes have remained largely elusive. METHODS: In this study, we deployed immunohistochemistry, RT-qPCR, flow cytometry, lymphocyte activation tests, and different cell-free protein-interaction assays. RESULTS: We here demonstrate that vemurafenib inhibits the downstream signaling of the canonical pathway of aryl hydrocarbon receptor (AhR) in vitro, thereby inducing the expression of proinflammatory cytokines (eg, TNF) and chemokines (eg, CCL5). In line with these results, we observed an impaired expression of AhR-regulated genes (eg, CYP1A1) and an upregulation of the corresponding proinflammatory genes in vivo. Moreover, results of lymphocyte activation tests showed the absence of drug-specific T cells in respective patients. CONCLUSION: Taken together, we obtained no hint of an underlying sensitization against vemurafenib but found evidence suggesting that vemurafenib enhances proinflammatory responses by inhibition of canonical AhR signaling. Our findings contribute to our understanding of the central role of the AhR in skin inflammation and may point toward a potential role for topical AhR agonists in supportive cancer care.

Keratinocytes Counteract UVB-Induced Immunosuppression in Mice through HIF-1a Signaling.

The transcription factor HIF-1a regulates cellular metabolism under hypoxia but also immune responses and UVB-induced skin reactions. In keratinocytes (KCs), HIF-1a is an environmental sensor orchestrating the adaptation to environmental changes. In this study, we investigated the role of HIF-1a in KCs for skin reactions to acute and chronic UVB exposure in mice. The function of HIF-1a in KCs under UVB exposure was analyzed in KC-specific HIF-1a conditional knockout (cKO) mice. cKO mice were hypersensitive to acute high-dose UVB irradiation compared with wild-type mice, displaying increased cell death and delayed barrier repair. After chronic low-dose UVB treatment, cKO mice also had stronger epidermal damage but reduced infiltration of dermal macrophages and T helper cells compared with wild-type mice. Irradiated cKO mice revealed accumulation of regulatory lymphocytes in dorsal skin-draining lymph nodes compared with wild-type and unirradiated mice. This was reflected by an augmented IL-10 release of lymph node cells and a weaker contact hypersensitivity reaction to DNFB in UVB-exposed cKO mice than in wild-type and unirradiated controls. In summary, we found that KC-specific HIF-1a expression is crucial for adaptation to UVB exposure and inhibits the development of UVB-induced immunosuppression in mice. Therefore, HIF-1a signaling in KCs could ameliorate photoaging-related skin disorders.

Unraveling the differential impact of PAHs and dioxin-like compounds on AKR1C3 reveals the EGFR extracellular domain as a critical determinant of the AHR response.

Polycyclic aromatic hydrocarbons (PAHs), dioxin-like compounds (DLCs) and structurally-related environmental pollutants may contribute to the pathogenesis of various diseases and disorders, primarily by activating the aryl hydrocarbon receptor (AHR) and modulating downstream cellular responses. Accordingly, AHR is considered an attractive molecular target for preventive and therapeutic measures. However, toxicological risk assessment of AHR-modulating compounds as well as drug development is complicated by the fact that different ligands elicit remarkably different AHR responses. By elucidating the differential effects of PAHs and DLCs on aldo-keto reductase 1C3 expression and associated prostaglandin D2 metabolism, we here provide evidence that the epidermal growth factor receptor (EGFR) substantially shapes AHR ligand-induced responses in human epithelial cells, i.e. primary and immortalized keratinocytes and breast cancer cells. Exposure to benzo[a]pyrene (B[a]P) and dioxin-like polychlorinated biphenyl (PCB) 126 resulted in a rapid c-Src-mediated phosphorylation of EGFR. Moreover, both AHR agonists stimulated protein kinase C activity and enhanced the ectodomain shedding of cell surface-bound EGFR ligands. However, only upon B[a]P treatment, this process resulted in an auto-/paracrine activation of EGFR and a subsequent induction of aldo-keto reductase 1C3 and 11-ketoreduction of prostaglandin D2. Receptor binding and internalization assays, docking analyses and mutational amino acid exchange confirmed that DLCs, but not B[a]P, bind to the EGFR extracellular domain, thereby blocking EGFR activation by growth factors. Finally, nanopore long-read RNA-seq revealed hundreds of genes, whose expression is regulated by B[a]P, but not by PCB126, and sensitive towards pharmacological EGFR inhibition. Our data provide novel mechanistic insights into the ligand response of AHR signaling and identify EGFR as an effector of environmental chemicals.

Correction to: The AHR represses nucleotide excision repair and apoptosis and contributes to UV-induced skin carcinogenesis.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

A Protective Role of Aryl Hydrocarbon Receptor Repressor in Inflammation and Tumor Growth.

The aryl hydrocarbon receptor (AhR) is known for mediating the toxicity of environmental pollutants such as dioxins and numerous dioxin-like compounds, and is associated with the promotion of various malignancies, including lymphoma. The aryl hydrocarbon receptor repressor (AhRR), a ligand-independent, transcriptionally inactive AhR-like protein is known to repress AhR signaling through its ability to compete with the AhR for dimerization with the AhR nuclear translocator (ARNT). While AhRR effectively blocks AhR signaling, several aspects of the mechanism of AhRR's functions are poorly understood, including suppression of inflammatory responses and its putative role as a tumor suppressor. In a transgenic mouse that overexpresses AhRR (AhRR Tg) we discovered that these mice suppress 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)- and inflammation-induced tumor growth after subcutaneous challenge of EL4 lymphoma cells. Using mouse embryonic fibroblasts (MEF) we found that AhRR overexpression suppresses the AhR-mediated anti-apoptotic response. The AhRR-mediated inhibition of apoptotic resistance was associated with a suppressed expression of interleukin (IL)-1ß and cyclooxygenase (COX)-2, which was dependent on activation of protein kinase A (PKA) and the CAAT-enhancer-binding protein beta (C/EBPß). These results provide mechanistic insights into the role of the AhRR to suppress inflammation and highlight the AhRR as a potential therapeutic target to suppress tumor growth.

The AHR represses nucleotide excision repair and apoptosis and contributes to UV-induced skin carcinogenesis.

Ultraviolet B (UVB) radiation induces mutagenic DNA photoproducts, in particular cyclobutane pyrimidine dimers (CPDs), in epidermal keratinocytes (KC). To prevent skin carcinogenesis, these DNA photoproducts must be removed by nucleotide excision repair (NER) or apoptosis. Here we report that the UVB-sensitive transcription factor aryl hydrocarbon receptor (AHR) attenuates the clearance of UVB-induced CPDs in human HaCaT KC and skin from SKH-1 hairless mice. Subsequent RNA interference and inhibitor studies in KC revealed that AHR specifically suppresses global genome but not transcription-coupled NER. In further experiments, we found that the accelerated repair of CPDs in AHR-compromised KC depended on a modulation of the p27 tumor suppressor protein. Accordingly, p27 protein levels were increased in AHR-silenced KC and skin biopsies from AHR-/- mice, and critical for the improvement of NER. Besides increasing NER activity, AHR inhibition was accompanied by an enhanced occurrence of DNA double-strand breaks triggering KC apoptosis at later time points after irradiation. The UVB-activated AHR thus acts as a negative regulator of both early defense systems against carcinogenesis, NER and apoptosis, implying that it exhibits tumorigenic functions in UVB-exposed skin. In fact, AHR-/- mice developed 50% less UVB-induced cutaneous squamous cell carcinomas in a chronic photocarcinogenesis study than their AHR+/+ littermates. Taken together, our data reveal that AHR influences DNA damage-dependent responses in UVB-irradiated KC and critically contributes to skin photocarcinogenesis in mice.