Activated fibroblasts modify keratinocyte stem niche through TET1 and IL-6 to promote their rapid transformation in a mouse model of prenatal arsenic exposure.

Early life exposure to environmental pollutants such as arsenic (As) can increase the risk of cancers in the offspring. In an earlier study, we showed that only prenatal As exposure significantly increases epidermal stem cell proliferation and accelerates skin tumorigenesis in BALB/c mouse offspring. In the present work, we have examined the role of As-conditioned dermal fibroblasts (DFs) in creating pro-tumorigenic niches for Keratinocyte stem cells (KSCs) in the offspring. DFs isolated from prenatally exposed animals showed increased levels of activation markers (α-SMA, Fibronectin, Collagen IV), induction of ten-eleven translocation methylcytosine dioxygenase 1(TET1), and secreted high levels of niche modifying IL-6. This led to enhanced proliferation, migration, and survival of KSCs. Increased IL-6 production in As-conditioned fibroblast was driven through TET1 mediated 5-mC to 5-hmC conversion at -698/-526 and -856/-679 region on its promoter. IL-6 further acted through downstream activation of JAK2-STAT3 signaling, promoting epithelial-to-mesenchymal transition (EMT) in KSCs. Inhibition of pSTAT3 induced by IL-6 reduced the EMT process in KSCs resulting in a significant decrease in their proliferation, migration, and colony formation. Our results indicate that IL-6 produced by prenatally conditioned fibroblasts plays a major role in regulating the KSC niche and promoting skin tumor development in As-exposed offspring.

"How to Release or Not Release, That Is the Question." A Review of Interleukin-1 Cellular Release Mechanisms in Vascular Inflammation.

Cardiovascular disease remains the leading cause of death worldwide, characterized by atherosclerotic activity within large and medium-sized arteries. Inflammation has been shown to be a primary driver of atherosclerotic plaque formation, with interleukin-1 (IL-1) having a principal role. This review focuses on the current state of knowledge of molecular mechanisms of IL-1 release from cells in atherosclerotic plaques. A more in-depth understanding of the process of IL-1's release into the vascular environment is necessary for the treatment of inflammatory disease processes, as the current selection of medicines being used primarily target IL-1 after it has been released. IL-1 is secreted by several heterogenous mechanisms, some of which are cell type-specific and could provide further specialized targets for therapeutic intervention. A major unmet challenge is to understand the mechanism before and leading to IL-1 release, especially by cells in atherosclerotic plaques, including endothelial cells, vascular smooth muscle cells, and macrophages. Data so far indicate a heterogeneity of IL-1 release mechanisms that vary according to cell type and are stimulus-dependent. Unraveling this complexity may reveal new targets to block excess vascular inflammation.

Prenatal arsenic exposure alters keratinocyte stem cell fate through persistent activation of IGF2R-MAPK cascade leading to aggravated skin carcinogenesis in mice offspring.

Chronic exposure to arsenic (As) promotes skin carcinogenesis in humans and potentially disturbs resident stem cell dynamics, particularly during maternal and early life exposure. In the present study, we demonstrate how only prenatal arsenic exposure disturbs keratinocyte stem cell (KSC) conditioning using a BALB/c mice model. Prenatal As exposure alters the normal stemness (CD34, KRT5), differentiation (Involucrin), and proliferation (PCNA) program in skin of offspring with progression of age as observed at 2, 10, and 18 weeks. Primary KSCs isolated from exposed animal at Day-2 showed increased survival (Bax:Bcl-xL, TUNEL assay), proliferation (BrdU), and differentiation (KRT5, Involucrin) potential through the activation of pro-carcinogenic IGF2R-MAPK cascade (IGF2R-G(α)q-MEK1-ERK1/2). This was associated with reduced enrichment of histone H3K27me3 and its methylase, EZH2 along with increased binding of demethylase, KDM6A at Igf2r promoter. Altered KSCs conditioning through disturbed Igf2r imprint contributed to impaired proliferation and differentiation and an aggravated tumor response in offspring.

Analysis of gut bacteriome of in utero arsenic-exposed mice using 16S rRNA-based metagenomic approach.

Introduction: Approximately 200 million people worldwide are affected by arsenic toxicity emanating from the consumption of drinking water containing inorganic arsenic above the prescribed maximum contaminant level. The current investigation deals with the role of prenatal arsenic exposure in modulating the gut microbial community and functional pathways of the host. Method: 16S rRNA-based next-generation sequencing was carried out to understand the effects of in utero 0.04 mg/kg (LD) and 0.4 mg/kg (HD) of arsenic exposure. This was carried out from gestational day 15 (GD-15) until the birth of pups to understand the alterations in bacterial diversity. Results: The study focused on gestational exposure to arsenic and the altered gut microbial community at phyla and genus levels, along with diversity indices. A significant decrease in firmicutes was observed in the gut microbiome of mice treated with arsenic. Functional analysis revealed that a shift in genes involved in crucial pathways such as insulin signaling and non-alcoholic fatty liver disease pathways may lead to metabolic diseases in the host. Discussion: The present investigation may hypothesize that in utero arsenic exposure can perturb the gut bacterial composition significantly as well as the functional pathways of the gestationally treated pups. This research paves the way to further investigate the probable mechanistic insights in the field of maternal exposure environments, which may play a key role in epigenetic modulations in developing various disease endpoints in the progeny.

Prenatal exposure to arsenic promotes sterile inflammation through the Polycomb repressive element EZH2 and accelerates skin tumorigenesis in mouse.

Prenatal and postnatal life stress could be a potent programmer of phenotype or disease state of an individual in the later life. Prenatal arsenic exposure has been shown to promote developmental defects, low birth weight, immunotoxicity and is associated with various cancers including skin cancer in adulthood. To investigate the association between prenatal arsenic exposure and adult life skin carcinogenesis, we used a two-stage cutaneous carcinogenesis model in which BALB/c mice were prenatally exposed to 0.04 mg/kg and 0.4 mg/kg arsenic (As). Exposure to arsenic was sufficient to shorten the tumor latency period and promote epidermal hyperplasia in the offspring upon challenge with dimethylbenz[a]/12-O-tetradecanoylphorbol-13-acetate (DMBA/TPA). The levels of inflammatory and tissue microenvironment remodeling factors such as IL-1α and TNF-α were persistently elevated in the skin, and their inhibition through diacerein led to a significant decrease in the tumor response, suggesting their role in tumorigenesis. While there was overexpression of multiple epigenetic regulators at tissue level, we found decreased enrichment of Polycomb repressive complex 2 (PRC2) member EZH2 and H3K27me3 mark at the upstream of the affected inflammatory genes. The higher expression of the inflammatory genes suggests the gene specific selective nature of EZH2 repression which was also associated with increased binding of the activator KDM6a (demethylase). Further, arsenic conditioned basal keratinocytes cells (BKCs) showed increased migration and proliferation along with higher expression of tumor associated cytokines. Inhibition of EZH2 in the BKCs lead to their further upregulation suggesting that BKCs might be the potential cell type for the interaction of EZH2 and inflammatory cytokines. The present study provides new evidence for the role of PRC2 group regulators in inflammatory conditioning and development of skin cancer in offspring prenatally exposed to arsenic.

Prenatal arsenic exposure induces immunometabolic alteration and renal injury in rats.

Arsenic (As) exposure is progressively associated with chronic kidney disease (CKD), a leading public health concern present worldwide. The adverse effect of As exposure on the kidneys of people living in As endemic areas have not been extensively studied. Furthermore, the impact of only prenatal exposure to As on the progression of CKD also has not been fully characterized. In the present study, we examined the effect of prenatal exposure to low doses of As 0.04 and 0.4 mg/kg body weight (0.04 and 0.4 ppm, respectively) on the progression of CKD in male offspring using a Wistar rat model. Interestingly, only prenatal As exposure was sufficient to elevate the expression of profibrotic (TGF-ß1) and proinflammatory (IL-1α, MIP-2α, RANTES, and TNF-α) cytokines at 2-day, 12- and 38-week time points in the exposed progeny. Further, alteration in adipogenic factors (ghrelin, leptin, and glucagon) was also observed in 12- and 38-week old male offspring prenatally exposed to As. An altered level of these factors coincides with impaired glucose metabolism and homeostasis accompanied by progressive kidney damage. We observed a significant increase in the deposition of extracellular matrix components and glomerular and tubular damage in the kidneys of 38-week-old male offspring prenatally exposed to As. Furthermore, the overexpression of TGF-ß1 in kidneys corresponds with hypermethylation of the TGF-ß1 gene-body, indicating a possible involvement of prenatal As exposure-driven epigenetic modulations of TGF-ß1 expression. Our study provides evidence that prenatal As exposure to males can adversely affect the immunometabolism of offspring which can promote kidney damage later in life.

MicroRNA-129-5p-regulated microglial expression of the surface receptor CD200R1 controls neuroinflammation.

CD200R1 is an inhibitory surface receptor expressed in microglia and blood macrophages. Microglial CD200R1 is known to control neuroinflammation by keeping the microglia in resting state, and therefore, tight regulation of its expression is important. CCAAT/enhancer-binding protein ß (CEBPß) is the known regulator of CD200R1 transcription. In the present study, our specific intention was to find a possible posttranscriptional regulatory mechanism of CD200R1 expression. Here we investigated a novel regulatory mechanism of CD200R1 expression following exposure to an environmental stressor, arsenic, combining in silico analysis, in vitro, and in vivo experiments, as well as validation in human samples. The in silico analysis and in vitro studies with primary neonatal microglia and BV2 microglia revealed that arsenic demethylates the promoter of a microRNA, miR-129-5p, thereby increasing its expression, which subsequently represses CD200R1 by binding to its 3'-untranslated region and shuttling the CD200R1 mRNA to the cytoplasmic-processing body in mouse microglia. The role of miR-129-5p was further validated in BALB/c mouse by stereotaxically injecting anti-miR-129. We found that anti-miR-129 reversed the expression of CD200R1, as well as levels of inflammatory molecules IL-6 and TNF-α. Experiments with a CD200R1 siRNA-induced loss-of-function mouse model confirmed an miR-129-5p→CD200R1→IL-6/TNF-α signaling axis. These main findings were replicated in a human cell line and validated in human samples. Taken together, our study revealed miR-129-5p as a novel posttranscriptional regulator of CD200R1 expression with potential implications in neuroinflammation and related complications.

Perinatal exposure to silver nanoparticles reprograms immunometabolism and promotes pancreatic beta-cell death and kidney damage in mice.

Silver nanoparticles (AgNPs) are extensively utilized in food, cosmetics, and healthcare products. Though the effects of AgNPs exposure on adults are well documented, the long-term effects of gestational/perinatal exposure upon the health of offspring have not been addressed. Herein, we show that only perinatal exposure to AgNPs through the mother could lead to chronic inflammation in offspring which persists till adulthood. Further, AgNPs exposure altered offspring's immune responses against environmental stresses. AgNPs exposed offspring showed an altered response in splenocyte proliferation assay when challenged to lipopolysaccharide, concanavalin-A, AgNPs, or silver ions. Perinatal AgNPs exposure affected metabolic parameters (resistin, glucagon-like peptide-1, leptin, insulin) and upregulated JNK/P38/ERK signaling in the pancreas. We observed pancreatic damage, reduced insulin level, and increased blood glucose levels. Further, we observed renal damage, particularly to tubular and glomerular regions as indicated by histopathology and electron microscopy. Our study thus shows that only perinatal exposure to AgNPs could induce persistent inflammation, alter immune responses against foreign antigens and metabolism which may contribute to pancreatic and renal damage later in life.

Oral subchronic exposure to silver nanoparticles causes renal damage through apoptotic impairment and necrotic cell death.

Silver nanoparticles (AgNPs) are one of the most widely used nanomaterials. Following oral exposure, AgNPs can accumulate in various organs including kidneys where they show gender specific accumulation. There is limited information on their effect on renal system following long-term animal exposure especially at the ultramicroscopic and molecular level. In this study, we have assessed the effect of 60 days oral AgNPs treatment on kidneys of female Wistar rats at doses of 50 ppm and 200 ppm that are below previously reported lowest observed adverse effect level (LOAEL). AgNPs treatment led to decrease in kidney weight and some loss of renal function as seen by increased levels of serum creatinine and early toxicity markers such as KIM-1, clusterin and osteopontin. We also observed significant mitochondrial damage, loss of brush border membranes, pronounced swelling of podocytes and degeneration of their foot processes using transmission electron microscopy (TEM). These symptoms are similar to those seen in nephrotic syndrome and 'Minimal change disease' of kidney where few changes are visible under light microscopy but significant ultrastructural damage is observed. Prolonged treatment of AgNPs also led to the activation of cell proliferative, survival and proinflammatory factors (Akt/mTOR, JNK/Stat and Erk/NF-κB pathways and IL1ß, MIP2, IFN-γ, TNF-α and RANTES) and dysfunction of normal apoptotic pathway. Our study shows how long term AgNPs exposure may promote ultrastructural damage to kidney causing inflammation and expression of cell survival factors. These changes, in the long term, could lead to inhibition of the beneficial apoptotic pathway and promotion of necrotic cell death in kidneys.

Mercury exposure induces cytoskeleton disruption and loss of renal function through epigenetic modulation of MMP9 expression.

Mercury is one of the major heavy metal pollutants occurring in elemental, inorganic and organic forms. Due to ban on most inorganic mercury containing products, human exposure to mercury generally occurs as methylmercury (MeHg) by consumption of contaminated fish and other sea food. Animal and epidemiological studies indicate that MeHg affects neural and renal function. Our study is focused on nephrotoxic potential of MeHg. In this study, we have shown for the first time how MeHg could epigenetically modulate matrix metalloproteinase 9(MMP9) to promote nephrotoxicity using an animal model of sub chronic MeHg exposure. MeHg caused renal toxicity as was seen by increased levels of serum creatinine and expression of early nephrotoxicity markers (KIM-1, Clusterin, IP-10, and TIMP). MeHg exposure also correlated strongly with induction of MMP9 mRNA and protein in a dose dependent manner. Further, while induction of MMP9 promoted cytoskeleton disruption and loss of cell-cell adhesion (loss of F-actin, Vimentin and Fibronectin), inhibition of MMP9 was found to reduce these disruptions. Mechanistic studies by ChIP analysis showed that MeHg modulated MMP9 by promoting demethylation of its regulatory region to increase its expression. Bisulfite sequencing identified critical CpGs in the first exon of MMP9 which were demethylated following MeHg exposure. ChIP studies also showed loss of methyl binding protein, MeCP2 and transcription factor PEA3 at the demethylated site confirming decreased CpG methylation. Our studies thus show how MeHg could epigenetically modulate MMP9 to promote cytoskeleton disruption leading to loss of renal function.