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2.
PLoS One ; 16(6): e0252954, 2021.
Article in English | MEDLINE | ID: mdl-34111182

ABSTRACT

The ability of persistent organic pollutants (POPs) with endocrine disrupting properties to interfere with the developing reproductive system is of increasing concern. POPs are transferred from dams to offspring and the high sensitivity of neonates to endocrine disturbances may be caused by underdeveloped systems of metabolism and excretion. The present study aimed to characterize the effect of in utero and lactational exposure to a human relevant mixture of POPs on the female mammary gland, ovarian folliculogenesis and liver function in CD-1 offspring mice. Dams were exposed to the mixture through the diet at Control, Low or High doses (representing 0x, 5000x and 100 000x human estimated daily intake levels, respectively) from weaning and throughout mating, gestation, and lactation. Perinatally exposed female offspring exhibited altered mammary gland development and a suppressed ovarian follicle maturation. Increased hepatic cytochrome P450 enzymatic activities indirectly indicated activation of nuclear receptors and potential generation of reactive products. Hepatocellular hypertrophy was observed from weaning until 30 weeks of age and could potentially lead to hepatotoxicity. Further studies should investigate the effects of human relevant mixtures of POPs on several hormones combined with female reproductive ability and liver function.


Subject(s)
Endocrine Disruptors/toxicity , Liver/physiology , Mammary Glands, Animal/growth & development , Maternal Exposure/adverse effects , Ovarian Follicle/growth & development , Persistent Organic Pollutants/toxicity , Animals , Cytochrome P-450 Enzyme System/metabolism , Female , Humans , Lactation/drug effects , Liver/drug effects , Liver Function Tests , Mammary Glands, Animal/drug effects , Mice , Ovarian Follicle/drug effects , Pregnancy , Up-Regulation
3.
Cell Mol Life Sci ; 78(3): 985-1009, 2021 Feb.
Article in English | MEDLINE | ID: mdl-32447426

ABSTRACT

The membrane lipids diacylglycerol (DAG) and phosphatidic acid (PA) are important second messengers that can regulate membrane transport by recruiting proteins to the membrane and by altering biophysical membrane properties. DAG and PA are involved in the transport from the Golgi apparatus to endosomes, and we have here investigated whether changes in these lipids might be important for regulation of transport to the Golgi using the protein toxin ricin. Modulation of DAG and PA levels using DAG kinase (DGK) and phospholipase D (PLD) inhibitors gave a strong increase in retrograde ricin transport, but had little impact on ricin recycling or degradation. Inhibitor treatment strongly affected the endosome morphology, increasing endosomal tubulation and size. Furthermore, ricin was present in these tubular structures together with proteins known to regulate retrograde transport. Using siRNA to knock down different isoforms of PLD and DGK, we found that several isoforms of PLD and DGK are involved in regulating ricin transport to the Golgi. Finally, by performing lipidomic analysis we found that the DGK inhibitor gave a weak, but expected, increase in DAG levels, while the PLD inhibitor gave a strong and unexpected increase in DAG levels, showing that it is important to perform lipidomic analysis when using inhibitors of lipid metabolism.


Subject(s)
Diacylglycerol Kinase/metabolism , Endosomes/metabolism , Golgi Apparatus/metabolism , Phospholipase D/metabolism , Cell Line, Tumor , Diacylglycerol Kinase/antagonists & inhibitors , Diacylglycerol Kinase/genetics , Diglycerides/metabolism , Endocytosis/drug effects , Enzyme Inhibitors/pharmacology , Humans , Lipidomics/methods , Lipids/analysis , Phospholipase D/antagonists & inhibitors , Phospholipase D/genetics , Protein Isoforms/antagonists & inhibitors , Protein Isoforms/genetics , Protein Isoforms/metabolism , Protein Transport/drug effects , Proteolysis/drug effects , Pyrimidinones/pharmacology , RNA Interference , RNA, Small Interfering/metabolism , Ricin/metabolism , Thiazoles/pharmacology
4.
Environ Int ; 146: 106240, 2021 01.
Article in English | MEDLINE | ID: mdl-33186814

ABSTRACT

Exposure to persistent organic pollutants (POPs), encompassing chlorinated (Cl), brominated (Br) and perfluoroalkyl acid (PFAA) compounds is associated with adverse neurobehaviour in humans and animals, and is observed to cause adverse effects in nerve cell cultures. Most studies focus on single POPs, whereas studies on effects of complex mixtures are limited. We examined the effects of a mixture of 29 persistent compounds (Cl + Br + PFAA, named Total mixture), as well as 6 sub-mixtures on in vitro exposed rat cerebellar granule neurons (CGNs). Protein expression studies of cerebella from in vivo exposed mice offspring were also conducted. The selection of chemicals for the POP mixture was based on compounds being prominent in food, breast milk or blood from the Scandinavian human population. The Total mixture and sub-mixtures containing PFAAs caused greater toxicity in rat CGNs than the single or combined Cl/Br sub-mixtures, with significant impact on viability from 500x human blood levels. The potencies for these mixtures based on LC50 values were Br + PFAA mixture > Total mixture > Cl + PFAA mixture > PFAA mixture. These mixtures also accelerated induced lipid peroxidation. Protection by the competitive N-methyl-D-aspartate (NMDA) receptor antagonist 3-((R)-2-Carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP) indicated involvement of the NMDA receptor in PFAA and Total mixture-, but not Cl mixture-induced toxicity. Gene-expression studies in rat CGNs using a sub-toxic and marginally toxic concentration ((0.4 nM-5.5 µM) 333x and (1 nM-8.2 µM) 500x human blood levels) of the mixtures, revealed differential expression of genes involved in apoptosis, oxidative stress, neurotransmission and cerebellar development, with more genes affected at the marginally toxic concentration. The two important neurodevelopmental markers Pax6 and Grin2b were downregulated at 500x human blood levels, accompanied by decreases in PAX6 and GluN2B protein levels, in cerebellum of offspring mice from mothers exposed to the Total mixture throughout pregnancy and lactation. In rat CGNs, the glutathione peroxidase gene Prdx6 and the regulatory transmembrane glycoprotein gene Sirpa were highly upregulated at both concentrations. In conclusion, our results support that early-life exposure to mixtures of POPs can cause adverse neurodevelopmental effects.


Subject(s)
Environmental Pollutants , Persistent Organic Pollutants , Animals , Cerebellum , Environmental Pollutants/toxicity , Female , Humans , Mice , Neurons , Oxidative Stress , Rats
5.
Front Immunol ; 11: 1417, 2020.
Article in English | MEDLINE | ID: mdl-32754152

ABSTRACT

Severe combined immunodeficiency (SCID) and other T cell lymphopenias can be detected during newborn screening (NBS) by measuring T cell receptor excision circles (TRECs) in dried blood spot (DBS) DNA. Second tier next generation sequencing (NGS) with an amplicon based targeted gene panel using the same DBS DNA was introduced as part of our prospective pilot research project in 2015. With written parental consent, 21 000 newborns were TREC-tested in the pilot. Three newborns were identified with SCID, and disease-causing variants in IL2RG, RAG2, and RMRP were confirmed by NGS on the initial DBS DNA. The molecular findings directed follow-up and therapy: the IL2RG-SCID underwent early hematopoietic stem cell transplantation (HSCT) without any complications; the leaky RAG2-SCID received prophylactic antibiotics, antifungals, and immunoglobulin infusions, and underwent HSCT at 1 year of age. The child with RMRP-SCID had complete Hirschsprung disease and died at 1 month of age. Since January 2018, all newborns in Norway have been offered NBS for SCID using 1st tier TRECs and 2nd tier gene panel NGS on DBS DNA. During the first 20 months of nationwide SCID screening an additional 88 000 newborns were TREC tested, and four new SCID cases were identified. Disease-causing variants in DCLRE1C, JAK3, NBN, and IL2RG were molecularly confirmed on day 8, 15, 8 and 6, respectively after birth, using the initial NBS blood spot. Targeted gene panel NGS integrated into the NBS algorithm rapidly delineated the specific molecular diagnoses and provided information useful for management, targeted therapy and follow-up i.e., X rays and CT scans were avoided in the radiosensitive SCID. Second tier targeted NGS on the same DBS DNA as the TREC test provided instant confirmation or exclusion of SCID, and made it possible to use a less stringent TREC cut-off value. This allowed for the detection of leaky SCIDs, and simultaneously reduced the number of control samples, recalls and false positives. Mothers were instructed to stop breastfeeding until maternal cytomegalovirus (CMV) status was determined. Our limited data suggest that shorter time-interval from birth to intervention, may prevent breast milk transmitted CMV infection in classical SCID.


Subject(s)
Biomarkers/blood , Dried Blood Spot Testing/methods , High-Throughput Nucleotide Sequencing/methods , Neonatal Screening/methods , Severe Combined Immunodeficiency/diagnosis , Cell-Free Nucleic Acids/blood , DNA, Circular/blood , Early Diagnosis , Female , Humans , Infant, Newborn , Male , Prospective Studies
7.
Tidsskr Nor Laegeforen ; 137(20)2017 Oct 31.
Article in Norwegian | MEDLINE | ID: mdl-29094580
8.
Tidsskr Nor Laegeforen ; 137(21)2017 Nov 14.
Article in Norwegian | MEDLINE | ID: mdl-29135185
9.
Tidsskr Nor Laegeforen ; 137(22)2017 11 28.
Article in Norwegian | MEDLINE | ID: mdl-29181925
10.
Biomed Res Int ; 2017: 8472312, 2017.
Article in English | MEDLINE | ID: mdl-28567424

ABSTRACT

The aim of the present study was to evaluate in vitro effects of dietary phytochemicals naringenin, quercetin, and sesamin on the activities of ethoxy- (EROD; CYP1A) and benzyloxy- (BROD; CYP3A) resorufin O-dealkylases after the exposure to the cocktail of persistent organic pollutants (POPs). CD-1 mice were exposed from weaning, through gestation and lactation to a defined mixture of POPs. Hepatic microsomes were prepared from their female offspring at postnatal day 42. Hepatic EROD and BROD activity were evaluated in the presence of quercetin, naringenin, and sesamin at nine concentrations from 5 to 100000 nM. EROD activity was strongly inhibited by quercetin with Ki values from 1.7 to 2.6 µM. BROD activity was inhibited by quercetin with Ki values from 64.9 to 75.3 µM and naringenin with Ki values from 39.3 to 45.8 µM. The IC50 and Ki values did not differ between the groups of mice with different levels of POPs exposure in any of the experimental sets. Sesamin did not inhibit either EROD or BROD. We concluded that the interactions of quercetin and naringenin with CYP1A and CYP3A in mice liver were not affected by the levels of POPs exposure.


Subject(s)
Air Pollutants/toxicity , Cytochrome P-450 Enzyme System/metabolism , Cytochrome P450 Family 1/metabolism , Dioxoles/pharmacology , Flavanones/pharmacology , Lignans/pharmacology , Maternal Exposure/adverse effects , Prenatal Exposure Delayed Effects/enzymology , Quercetin/pharmacology , Animals , Cytochrome P-450 CYP3A , Female , Mice , Microsomes, Liver/enzymology , Microsomes, Liver/pathology , Pregnancy , Prenatal Exposure Delayed Effects/chemically induced , Prenatal Exposure Delayed Effects/pathology
11.
Toxicol Lett ; 259: 95-107, 2016 Sep 30.
Article in English | MEDLINE | ID: mdl-27511595

ABSTRACT

Perfluorinated alkyl acids (PFAAs) are stable chemicals detected in tissue and serum from various species, including humans, and have been linked to adverse health outcomes. Experimental PFAA exposure in rodents has been associated with changes in mammary gland development. The estrogen receptor (ER)-negative human breast epithelial cell line, MCF-10A, can be grown as monolayer, but also has the ability to form three-dimensional acini in vitro, reflecting aspects of mammary glandular morphogenesis. Cells were exposed to five different PFAAs, perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA), perfluorodecanoic acid (PFDA), and perfluoroundecanoic acid (PFUnDA), both in monolayer and acini cultures. In monolayer cultures only the higher concentrations of PFOS, PFNA and PFDA (400-500µM) caused a significant increase in cell death, whereas PFOA and PFUnDA had no effect. Normal acini maturation was negatively impacted by PFOS, PFNA and PFDA already at the lowest concentration tested (0.6µM). Observed effects included loss of organization of the cell clusters and absence of a hollow lumen. Overall, this study demonstrated that PFAAs can interfere with cellular events related to normal development of glandular breast tissue through ER-independent mechanisms.


Subject(s)
Acinar Cells/drug effects , Epithelial Cells/drug effects , Fluorocarbons/toxicity , Mammary Glands, Human/cytology , Acinar Cells/physiology , Cell Culture Techniques , Cell Cycle/drug effects , Cell Death/drug effects , Cell Line , Female , Humans
12.
Biochem Pharmacol ; 110-111: 109-16, 2016 06 15.
Article in English | MEDLINE | ID: mdl-27107807

ABSTRACT

Flavonoids are known to have effects on cytochrome P450 enzymatic activity. However, little effort has been made to examine species differences and the relevance of studies on mammalian and fish microsomes so that extrapolations can be made to humans. Therefore, the effects of several naturally occurring flavonoids on the activity of CYP3A-dependent 7-benzyloxy-4-trifluoromethylcoumarin O-debenzylase (BFCOD) were evaluated in human, pig, mouse, and juvenile rainbow trout sources of hepatic microsomes. Each was exposed to three concentrations (1, 10, and 100µM) of diosmin, naringin, and naringenin. Naringenin competitively inhibited BFCOD activity (Ki values were 24.6µM in human, 15.6µM in pig, and 19.6µM in mouse microsomes). In fish, BFCOD activity was inhibited in a noncompetitive manner (Ki=7µM). Neither diosmin nor naringenin affected BFCOD activity in hepatic microsomes from the studied model organisms. These results suggest that dietary flavonoids potentially inhibit the metabolism of clinical drugs.


Subject(s)
Cytochrome P-450 CYP3A/metabolism , Diosmin/pharmacology , Enzyme Inhibitors/pharmacology , Flavanones/pharmacology , Microsomes, Liver/drug effects , Animals , Citrus/chemistry , Coumarins/metabolism , Cytochrome P-450 CYP3A/genetics , Female , Gene Expression , Humans , Hydrolysis , Kinetics , Liver/drug effects , Liver/enzymology , Male , Mice , Mice, Inbred ICR , Microsomes, Liver/enzymology , Oncorhynchus mykiss , Species Specificity , Swine
13.
DNA Repair (Amst) ; 11(3): 278-85, 2012 Mar 01.
Article in English | MEDLINE | ID: mdl-22209780

ABSTRACT

Reactive oxygen species (ROS) are formed as natural byproducts during aerobic metabolism and readily induce premutagenic base lesions in the DNA. The 8-oxoguanine DNA glycosylase (OGG1) and MutY homolog 1 (MYH) synergistically prevent mutagenesis and cancer formation in mice. Their localization in the mitochondria as well as in the nucleus suggests that mutations in mitochondrial DNA (mtDNA) contribute to the carcinogenesis in the myh⁻/⁻/ogg1⁻/⁻ double knockout mouse. In order to test this hypothesis, we analyzed mtDNA mutagenesis and mitochondrial function in young (1month) and adult (6months) wt and myh⁻/⁻/ogg1⁻/⁻ mice. To our surprise, the absence of OGG1 and MYH had no impact on mtDNA mutation rates in these mice, even at the onset of cancer. This indicates that mtDNA mutagenesis is not responsible for the carcinogenesis of myh⁻/⁻/ogg1⁻/⁻ mice. In line with these results, mitochondrial function was unaffected in the cancerous tissues liver and lung, whereas a significant reduction in respiration capacity was observed in brain mitochondria from the adult myh⁻/⁻/ogg1⁻/⁻ mouse. The reduced respiration capacity correlated with a specific reduction (-25%) in complex I biochemical activity in brain mitochondria. Our results demonstrate that mtDNA mutations are not associated with cancer development in myh⁻/⁻/ogg1⁻/⁻ mice, and that impairment of mitochondrial function in brain could be linked to nuclear DNA mutations in this strain. OGG1 and MYH appear to be dispensable for antimutator function in mitochondria.


Subject(s)
DNA Glycosylases/deficiency , DNA, Mitochondrial/genetics , Mutagenesis/genetics , Mutation/genetics , Neoplasms/enzymology , Neoplasms/genetics , Animals , Cell Respiration/genetics , DNA Damage/genetics , DNA Glycosylases/metabolism , Mice , Mice, Inbred C57BL , Mice, Mutant Strains , Mutation Rate , Organ Specificity/genetics
14.
FEBS J ; 276(10): 2811-21, 2009 May.
Article in English | MEDLINE | ID: mdl-19389114

ABSTRACT

Cockayne syndrome (CS) is a complex, progressive disease that involves neurological and developmental impairment and premature aging. The majority of CS patients have mutations in the CSB gene. The CSB protein is involved in multiple DNA repair pathways and CSB mutated cells are sensitive to a broad spectrum of genotoxic agents. We tested the hypothesis that sensitivity to such genotoxins could be mediated by mitochondrial dysfunction as a consequence of the CSB mutation. mtDNA from csb(m/m) mice accumulates oxidative damage including 8-oxoguanine, and cells from this mouse are hypersensitive to the mitochondrial oxidant menadione. Inhibitors of mitochondrial complexes and the glycolysis inhibitor 2-deoxyglucose kill csb(m/m) cells more efficiently than wild-type cells, via a mechanism that does not correlate with mtDNA damage formation. Menadione depletes cellular ATP, and recovery after depletion is slower in csb(m/m) cells. The bioenergetic alteration in csb(m/m) cells parallels the simpler organization of supercomplexes consisting of complexes I, III and IV in addition to partially disassembled complex V in the inner mitochondrial membrane. Exposing wild-type cells to DNA intercalating agents induces complex alterations, suggesting a link between mtDNA integrity, respiratory complexes and mitochondrial function. Thus, mitochondrial dysfunction may play a role in the pathology of CS.


Subject(s)
Cockayne Syndrome/metabolism , DNA Damage , DNA, Mitochondrial/genetics , Adenosine Triphosphate/metabolism , Cockayne Syndrome/genetics , Cockayne Syndrome/pathology , DNA Helicases , DNA Repair Enzymes , Electrophoresis, Polyacrylamide Gel , Energy Metabolism , Humans , Oxidative Stress , Poly-ADP-Ribose Binding Proteins
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