Post-ischemic inactivation of HIF prolyl hydroxylases in endothelium promotes maladaptive kidney repair by inducing glycolysis.

Ischemic acute kidney injury (AKI) is common in hospitalized patients and increases the risk for chronic kidney disease (CKD). Impaired endothelial cell (EC) functions are thought to contribute in AKI to CKD transition, but the underlying mechanisms remain unclear. Here, we identify a critical role for endothelial oxygen sensing prolyl hydroxylase domain (PHD) enzymes 1-3 in regulating post-ischemic kidney repair. In renal endothelium, we observed compartment-specific differences in the expression of the three PHD isoforms in both mice and humans. We found that post-ischemic concurrent inactivation of endothelial PHD1, PHD2, and PHD3 but not PHD2 alone promoted maladaptive kidney repair characterized by exacerbated tissue injury, fibrosis, and inflammation. Single-cell RNA-seq analysis of the post-ischemic endothelial PHD1, PHD2 and PHD3 deficient (PHDTiEC) kidney revealed an endothelial glycolytic transcriptional signature, also observed in human kidneys with severe AKI. This metabolic program was coupled to upregulation of the SLC16A3 gene encoding the lactate exporter monocarboxylate transporter 4 (MCT4). Strikingly, treatment with the MCT4 inhibitor syrosingopine restored adaptive kidney repair in PHDTiEC mice. Mechanistically, MCT4 inhibition suppressed pro-inflammatory EC activation reducing monocyte-endothelial cell interaction. Our findings suggest avenues for halting AKI to CKD transition based on selectively targeting the endothelial hypoxia-driven glycolysis/MCT4 axis.

Chemical inhibition of oxygen-sensing prolyl hydroxylases impairs angiogenic competence of human vascular endothelium through metabolic reprogramming.

Endothelial cell (EC) metabolism has emerged as a driver of angiogenesis. While hypoxia inactivates the oxygen sensors prolyl-4 hydroxylase domain-containing proteins 1-3 (PHD1-3) and stimulates angiogenesis, the effects of PHDs on EC functions remain poorly defined. Here, we investigated the impact of chemical PHD inhibition by dimethyloxalylglycine (DMOG) on angiogenic competence and metabolism of human vascular ECs. DMOG reduced EC proliferation, migration, and tube formation capacities, responses that were associated with an unfavorable metabolic reprogramming. While glycolytic genes were induced, multiple genes encoding sub-units of mitochondrial complex I were suppressed with concurrent decline in nicotinamide adenine dinucleotide (NAD+) levels. Importantly, the DMOG-induced defects in EC migration could be partially rescued by augmenting NAD+ levels through nicotinamide riboside or citrate supplementation. In summary, by integrating functional assays, transcriptomics, and metabolomics, we provide insights into the effects of PHD inhibition on angiogenic competence and metabolism of human vascular ECs.

Gestational exposure to silver nanoparticles enhances immune adaptation and protection against streptozotocin-induced diabetic nephropathy in mice offspring.

Silver nanoparticles (AgNPs) possess unique antimicrobial properties. As a result, they are being increasingly used in a wide range of applications. Several studies have shown detrimental effects of AgNPs exposure, including inflammation, accumulation, and cellular damage to different organs. However, the effect of AgNPs exposure during gestation, a critical and susceptible period of human development, on pregnant females and its long-term effects on offspring's health has not been studied. Therefore, we conducted a long-term study where we assessed the effect of gestational AgNPs exposure on pregnant mice and followed their offspring until the age of 12 months. Gestational exposure to AgNPs induced systemic inflammation in the pregnant mice at gestational day (GD) 18. Interestingly, developing fetuses exposed to AgNPs, showed anti-inflammatory conditions as indicated by reduced expression of inflammatory genes in fetal organs at GD 18 and reduced serum levels of TNF-α, IFN-γ, IL-17A, IL-6, and MCP-1 in AgNPs exposed pups at postnatal day (PD) 2. Surprisingly, post-weaning, AgNPs exposed offspring showed a heightened immune activation as shown by upregulation of inflammatory cytokines at PD 28, which persisted till late in life. Moreover, we observed metabolic alterations which persisted until adulthood in mice. To understand the impact of long-term immunometabolic changes on the progression of diabetes and kidney diseases under stressed conditions, we exposed offspring to streptozotocin which revealed a protective role of low-dose gestational AgNPs exposure against streptozotocin-induced diabetes and associated nephropathy.

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.

Role of Endothelial Prolyl-4-Hydroxylase Domain Protein/Hypoxia-Inducible Factor Axis in Acute Kidney Injury.

Ischemia reperfusion injury (IRI) results from a cessation or restriction of blood supply to an organ followed by reestablishment of perfusion and reoxygenation. In the kidney, IRI due to transplantation, cardiac surgery with cardiopulmonary bypass, and other major vascular surgeries contributes to acute kidney injury (AKI), a clinical condition associated with significant morbidity and mortality in hospitalized patients. In the postischemic kidney, endothelial damage promotes inflammatory responses and leads to persistent hypoxia of the renal tubular epithelium. Like other cell types, endothelial cells respond to low oxygen tension by multiple hypoxic signaling mechanisms. Key mediators of adaptation to hypoxia are hypoxia-inducible factors (HIF)-1 and -2, transcription factors whose activity is negatively regulated by prolyl-hydroxylase domain proteins 1 to 3 (PHD1 to PHD3). The PHD/HIF axis controls several processes determining injury outcome, including ATP generation, cell survival, proliferation, and angiogenesis. Here, we discuss recent advances in our understanding of the endothelial-derived PHD/HIF signaling and its effects on postischemic AKI.

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.

Hypoxic preconditioning protects against ischemic kidney injury through the IDO1/kynurenine pathway.

Prolonged cellular hypoxia leads to energetic failure and death. However, sublethal hypoxia can trigger an adaptive response called hypoxic preconditioning. While prolyl-hydroxylase (PHD) enzymes and hypoxia-inducible factors (HIFs) have been identified as key elements of oxygen-sensing machinery, the mechanisms by which hypoxic preconditioning protects against insults remain unclear. Here, we perform serum metabolomic profiling to assess alterations induced by two potent cytoprotective approaches, hypoxic preconditioning and pharmacologic PHD inhibition. We discover that both approaches increase serum kynurenine levels and enhance kynurenine biotransformation, leading to preservation of NAD+ in the post-ischemic kidney. Furthermore, we show that indoleamine 2,3-dioxygenase 1 (Ido1) deficiency abolishes the systemic increase of kynurenine and the subsequent renoprotection generated by hypoxic preconditioning and PHD inhibition. Importantly, exogenous administration of kynurenine restores the hypoxic preconditioning in the context of Ido1 deficiency. Collectively, our findings demonstrate a critical role of the IDO1-kynurenine axis in mediating hypoxic preconditioning.

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.

Developmental Exposure to Endocrine Disrupting Chemicals and Its Impact on Cardio-Metabolic-Renal Health.

Developmental origin of health and disease postulates that the footprints of early life exposure are followed as an endowment of risk for adult diseases. Epidemiological and experimental evidence suggest that an adverse fetal environment can affect the health of offspring throughout their lifetime. Exposure to endocrine disrupting chemicals (EDCs) during fetal development can affect the hormone system homeostasis, resulting in a broad spectrum of adverse health outcomes. In the present review, we have described the effect of prenatal EDCs exposure on cardio-metabolic-renal health, using the available epidemiological and experimental evidence. We also discuss the potential mechanisms of their action, which include epigenetic changes, hormonal imprinting, loss of energy homeostasis, and metabolic perturbations. The effect of prenatal EDCs exposure on cardio-metabolic-renal health, which is a complex condition of an altered biological landscape, can be further examined in the case of other environmental stressors with a similar mode of action.

Inhibition of Endothelial PHD2 Suppresses Post-Ischemic Kidney Inflammation through Hypoxia-Inducible Factor-1.

BACKGROUND: Prolyl-4-hydroxylase domain-containing proteins 1-3 (PHD1 to PHD3) regulate the activity of the hypoxia-inducible factors (HIFs) HIF-1 and HIF-2, transcription factors that are key regulators of hypoxic vascular responses. We previously reported that deficiency of endothelial HIF-2 exacerbated renal ischemia-reperfusion injury, whereas inactivation of endothelial PHD2, the main oxygen sensor, provided renoprotection. Nevertheless, the molecular mechanisms by which endothelial PHD2 dictates AKI outcomes remain undefined. METHODS: To investigate the function of the endothelial PHD2/HIF axis in ischemic AKI, we examined the effects of endothelial-specific ablation of PHD2 in a mouse model of renal ischemia-reperfusion injury. We also interrogated the contribution of each HIF isoform by concurrent endothelial deletion of both PHD2 and HIF-1 or both PHD2 and HIF-2. RESULTS: Endothelial deletion of Phd2 preserved kidney function and limited transition to CKD. Mechanistically, we found that endothelial Phd2 ablation protected against renal ischemia-reperfusion injury by suppressing the expression of proinflammatory genes and recruitment of inflammatory cells in a manner that was dependent on HIF-1 but not HIF-2. Persistence of renoprotective responses after acute inducible endothelial-specific loss of Phd2 in adult mice ruled out a requirement for PHD2 signaling in hematopoietic cells. Although Phd2 inhibition was not sufficient to induce detectable HIF activity in the kidney endothelium, in vitro experiments implicated a humoral factor in the anti-inflammatory effects generated by endothelial PHD2/HIF-1 signaling. CONCLUSIONS: Our findings suggest that activation of endothelial HIF-1 signaling through PHD2 inhibition may offer a novel therapeutic approach against ischemic AKI.

Accelerated and scarless wound repair by a multicomponent hydrogel through simultaneous activation of multiple pathways.

Scarless healing of injury remains a clinical challenge because of its complicated and overlapping phases of inflammation, clearing, and regeneration. Curcumin has been already established as a potential wound healing agent for normal and diabetic-impaired wounds. Herein, the question has been addressed whether a well-known antioxidant cerium oxide nanoparticle (CNP) can potentiate the activity of curcumin to promote a cellular program for scarless healing. In this study, we have developed a biocompatible poly (acrylamide) hydrogel (PAGE)-based dressing material comprising of CNP and curcumin (ACC) and tested its wound healing activity in an animal model of acute wound. Characterization of the CNP- and curcumin-entrapped hydrogel dressing (ACC) demonstrated high loading efficiency and sustained release of curcumin. In a full-thickness acute wound healing model of rat, a single application of ACC dressing demonstrated higher wound healing efficacy (78%) and negligible scarring compared to dressings containing only curcumin or CNP in 7 days. Enhanced cell proliferation, higher collagen content, advanced wound maturity, re-epithelialization, and granulation tissue formation were observed using the combination of curcumin and CNP (ACC). Study of cellular mechanisms identified MCP-1 and TGF-ß as the key drivers of differential and accelerated healing observed in the ACC group. These, coupled with the upregulation of growth-related signaling pathways (HER2/ErbB2, TGF-ß-Smad2/3, MAPK/ERK, AKT, and VEGF), promoted almost scarless healing in animals treated with ACC. The optimized combination of curcumin and CNP used in our study shows distinct advantage and can be a better agent for complete wound healing.

Sub-acute oral exposure of zinc oxide nanoparticles causes alteration in iron homeostasis through acute phase response: A protective effect by surface modification.

Zinc oxide nanoparticles (ZnO NPs) are one of the most widely used nanomaterials. Following oral exposure, these NPs can accumulate in various organs and induce the toxicity due to their physiochemical characteristics. In present study to reduce the toxicity, surface engineered ZnO NPs (c-ZnO NPs) were in-situ synthesized by using polyacrylamide grafted guar gum (PAm-g-GG) polymer in alkaline media. Further, the comparative effect of bared ZnO NPs (b-ZnO NPs) and c-ZnO NPs were assessed on secondary target organ liver and kidneys of Swiss mice at doses of 10, 50 and 300 mg/kg following 28 days repeated oral treatment. The b-ZnO NPs were incited severe damages in liver and kidney tissue than c-ZnO NPs as seen by transmission electron microscopy and histopathology. The increased levels of serum biomarkers (AST, ALT, ALP, creatinine, uric acid, and urea) were also observed, that remarking a disturbance in the function of liver and kidney. After sub-acute oral treatment of b-ZnO NPs, the hepatic pro-inflammatory cytokines (IL-6, TNF-α, and MMP-9) were up-regulated that causes the activation of acute phase response (APR). We also observed significantly increased in expression of hepatic acute phase proteins (hepcidin and haptoglobin) and altered interlinked iron (Fe) signaling biomarkers (hephaestin, TF, TFR-1, LDH, and ferroportin). This study emphasizes that exposure to ZnO NPs may cause inflammation mediated APR through ultra-structural damage of tissue that could escort the progression of anemia. Nevertheless, the capping with PAm-g-GG in c- ZnO NPs has reduced the toxicity by altering the surface reactive property of ZnO NPs.

Association between PAHs biomarkers and kidney injury biomarkers among kitchen workers with microalbuminuria: A cross-sectional pilot study.

BACKGROUND: To study the association between kidney injury biomarkers and urinary OH-PAH metabolites in kitchen workers, with microalbuminuria. METHODS: A cross-sectional pilot study was conducted among 120 male kitchen workers in a mega kitchen located at Coimbatore, India. Personal and sub-clinical details of study subjects were collected using a questionnaire. Albumin, creatinine, and albumin-creatinine ratio (ACR) were measured using urine dipstick test for the determination of microalbuminuria. Urinary hydroxylated PAHs metabolites (1-NAP, 9-HF, 3-HF, 2-HF, 9-PHN, and 1-OHP) were measured using GC-MS/MS and urinary kidney biomarkers (uNGAL, uCyst-C, uKIM-1, uOPN, and uTIMP-1) were measured using Multiplex Reader. RESULTS: Concentrations of urinary PAHs metabolites (1-NAP, 3-HF, 2-HF, 9-PHN, and 1-OHP) and kidney biomarkers (uKIM-1, uTIMP-1, uCyst-C and uNGAL) were significantly higher among kitchen workers with MAU compared to non-kitchen workers with MAU. Urinary kidney biomarkers viz., uKIM-1, uTIMP-1, uCyst-C, uNGAL, and uOPN showed higher median concentration among the kitchen workers with MAU compared to kitchen workers without MAU. Significant positive correlation was observed for 9-HF with uKIM-1 and uTIMP-1 and 1-OHP with uKIM-1. ACR was also well correlated with urinary kidney biomarkers. ROC analysis showed higher sensitivity and specificity for uKIM-1, uCyst-C, and uNGAL as biomarkers for early prediction of acute kidney injury among kitchen workers. CONCLUSIONS: The PAHs exposure among kitchen workers can lead to kidney injury. This was evident from the association of OH-PAHs and kidney injury biomarkers in kitchen workers with microalbuminuria.

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.

Impaired lysosomal activity mediated autophagic flux disruption by graphite carbon nanofibers induce apoptosis in human lung epithelial cells through oxidative stress and energetic impairment.

BACKGROUND: Graphite carbon nanofibers (GCNF) have emerged as a potential alternative of carbon nanotubes (CNT) for various biomedical applications due to their superior physico-chemical properties. Therefore in-depth understanding of the GCNF induced toxic effects and underlying mechanisms in biological systems is of great interest. Currently, autophagy activation by nanomaterials is recognized as an emerging toxicity mechanism. However, the association of GCNF induced toxicity with this form of cell death is largely unknown. In this study, we have assessed the possible mechanism; especially the role of autophagy, underlying the GCNF induced toxicity. METHODS: Human lung adenocarcinoma (A549) cells were exposed to a range of GCNF concentrations and various cellular parameters were analyzed (up to 48 h). Transmission electron microscopy, immunofluorescent staining, western blot and quantitative real time PCR were performed to detect apoptosis, autophagy induction, lysosomal destabilization and cytoskeleton disruption in GCNF exposed cells. DCFDA assay was used to evaluate the reactive oxygen species (ROS) production. Experiments with N-acetyl-L-cysteine (NAC), 3-methyladenine (3-MA) and LC3 siRNA was carried out to confirm the involvement of oxidative stress and autophagy in GCNF induced cell death. Comet assay and micronucleus (MN) assay was performed to assess the genotoxicity potential. RESULTS: In the present study, GCNF was found to induce nanotoxicity in human lung cells through autophagosomes accumulation followed by apoptosis via intracellular ROS generation. Mechanistically, impaired lysosomal function and cytoskeleton disruption mediated autophagic flux blockade was found to be the major cause of accumulation rather than autophagy induction which further activates apoptosis. The whole process was in line with the increased ROS level and their pharmacological inhibition leads to mitigation of GCNF induced cell death. Moreover the inhibition of autophagy attenuates apoptosis indicating the role of autophagy as cell death process. GCNF was also found to induce genomic instability. CONCLUSION: Our present study demonstrates that GCNF perturbs various interrelated signaling pathway and unveils the potential nanotoxicity mechanism of GCNF through targeting ROS-autophagy-apoptosis axis. The current study is significant to evaluate the safety and risk assessment of fibrous carbon nanomaterials prior to their potential use and suggests caution on their utilization for biomedical research.

Photo-induced biosynthesis of silver nanoparticles from aqueous extract of Dunaliella salina and their anticancer potential.

The synthesis of silver nanoparticles (AgNPs) via green route, using biological entities is an area of interest, because one of the potential applications in the nanomedicine. In the present study, we have developed photo-induced, ecofriendly, low cost method for biosynthesis of the stable silver nanoparticles using aqueous extract of Dunaliella salina (AED) which act as both reducing as well as stabilizing agent. Biosynthesis of the AgNPs was optimized as: sunlight exposure (30min), AED (5% (v/v)) and AgNO3 (4mM). Biosynthesis of AgNPs was monitored by using UV-Vis spectroscopy which exhibited sharp SPR band at 430nm after 30min of bright sunlight exposure. SEM and TEM analyses confirmed the presence of spherical AgNPs with average size of 15.26nm. Crystalline nature of AgNPs was confirmed by SAED and XRD analyses where Braggs reflection pattern at (111), (200), (220) and (311) corresponded to face centered cubic crystal lattice of metallic silver. FTIR analysis revealed the involvement of various functional groups present in AED. AFM analysis confirmed the average surface roughness of synthesized AgNPs as 8.48nm. AgNPs were also screened for anticancer potential using assay of calcein AM/PI, Annexin/PI and cancer biomarkers against cancer cell line (MCF-7), while normal cell line (MCF-10A) were kept as control. Interestingly, anticancer potential was comparable to the known anticancer drug (Cisplatin), and was not detrimental to the normal cell line. Therefore, such green synthesized AgNPs may be explored as anticancer agent.

Fresh Water Cyanobacteria Geitlerinema sp. CCC728 and Arthrospira sp. CCC729 as an Anticancer Drug Resource.

An increasing number of cancer patients worldwide, especially in third world countries, have raised concern to explore natural drug resources, such as the less explored fresh water filamentous cyanobacteria. Six strains of cyanobacteria (Phormidium sp. CCC727, Geitlerinema sp. CCC728, Arthrospira sp. CCC729, Phormidium sp. CCC731, Phormidium sp. CCC730, and Leptolyngbya sp. CCC732) were isolated (paddy fields and ponds in the Banaras Hindu University, campus) and five strains screened for anticancer potential using human colon adenocarcinoma (HT29) and human kidney adenocarcinoma (A498) cancer cell lines. Geitlerinema sp. CCC728 and Arthrospira sp. CCC729 were the most potent as determined by examination of morphological features and by inhibition of growth by graded concentrations of crude extracts and thin-layer chromatography (TLC) eluates. Cell cycle analysis and multiplex assays using cancer biomarkers also confirmed Geitlerinema sp. CCC728 and Arthrospira sp. CCC729 as cancer drug resources. Apoptotic studies in the cells of A498 (cancer) and MCF-10A (normal human epithelial) exposed to crude extracts and TLC fractions revealed no significant impact on MCF-10A cells emphasizing its importance in the development of anticancer drug. Identification of biomolecules from these extracts are in progress.

Arsenic exposure causes epigenetic dysregulation of IL-8 expression leading to proneoplastic changes in kidney cells.

Prolonged arsenic exposure has been shown to cause several detrimental effects in adults. However its effects following prenatal exposure are not well defined at the epigenetic level, particularly in terms of changes which may predispose an individual to adult malignancies. In this work, we have studied the effect of arsenic exposure on renal system using human embryonic kidney cells and prenatally exposed animals and identified Interleukin-8(IL-8) and its homologue (CINC-1) as mediators of arsenic induced renal toxicity. We further show that embryonic kidney cells are more responsive to arsenic leading to higher induction of IL-8 as compared to adult cells due to DNA methylation and histone acetylation (H3 acetylation) changes in the IL-8 promoter. Through bisulfite analysis of the IL-8 promoter, we have also identified an arsenic modulated CpG site at -168 bases upstream of transcription start site. This CpG is associated with C/EBP and CREB binding sites in the IL-8 promoter and its demethylation by arsenic coupled with increased H3 histone acetylation and CBP/P300 recruitment could lead to induction of IL-8. Our study shows how epigenetic modulation of IL-8 by arsenic could contribute to increased cell migratory and proliferative capabilities, cell cycle dysregulation and renal toxicity.