In silico investigation of the role of miRNAs in a possible developmental origin of prostate cancer in F1 and F2 offspring of mothers exposed to a phthalate mixture.

A previous study using miRNA sequencing revealed that exposure to a mixture of phthalates during pregnancy and lactation dysregulated rno-miR-184 and rno-miR-141-3p in the ventral prostate (VP) of offspring. Here, rno-miR-184 and rno-miR-141-3 expressions were obtained by RT-qPCR in the VP of F1 males as well as in F2 offspring, aiming to establish a relationship with possible oncogenic targets through in silico analyses with multigenerational approach. Additionally, some targets were measured by western blots to highlight a possible relationship between the deregulated miRNAs and some of their targets. VP samples from rats exposed to a mixture of phthalates maternally during pregnancy and lactation (GD10 to PND21-F1) and VP from offspring (F2) were examined. The phthalate mixture at both concentrations (20 µg and 200 mg/kg/day) increased the expression of both miRNAs in the F1 (PND22 and 120) and F2 (descendants of F1-treated males) prostate. Target prediction analysis revealed that both microRNAs are responsible for modulating the expression and synthesis of 40 common targets. A phthalate target association analysis and the HPA database showed an interesting relationship among these possible miRNAs modulated targets with prostate adenocarcinoma and other oncogenic processes. Western blots showed alteration in P63, P53, WNT5, and STAT3 expression, which are targeted by the miRNAs, in the VP of F1/F2 males. The data draw attention to the epigenetic modulation in the prostate of descendants exposed to phthalates and adds to one of the few currently found in the literature to point to microRNAs signature as biomarkers of exposure to plasticizers.

Synergistic interactions between PBDEs and PCBs in human neuroblastoma cells.

Polybrominated diphenyl ethers (PBDEs) and polychlorinated biphenyls (PCBs) are ubiquitous environmental pollutants. Exposure to these chemicals has been associated with developmental neurotoxicity, endocrine dysfunction, and reproductive disorders. Humans and wildlife are generally exposed to a mixture of these environmental pollutants, highlighting the need to evaluate the potential effects of combined exposures. In this study, we investigated the cytotoxic effects of the combined exposure to two PBDEs and two PCBs in a human neuronal cell line. 2,2',4,4'-Tetrabromodiphenyl ether, 2,2',4,4',5-pentabromodiphenyl ether, PCB-126 (3,3',4,4',5-pentachlorobiphenyl; a dioxin-like PCB), and PCB-153 (2,2',4,4',5,5'-hexachlorobiphenyl; a non-dioxin-like PCB) were chosen, because their concentrations are among the highest in human tissues and the environment. The results suggest that the nature of interactions is related to the PCB structure. Mixtures of PCB-153 and both PBDEs had a prevalently synergistic effect. In contrast, mixtures of each PBDE congener with PCB-126 showed additive effects at threshold concentrations, and synergistic effects at higher concentrations. These results emphasize the concept that the toxicity of xenobiotics may be affected by possible interactions, which may be of significance given the common coexposures to multiple contaminants.

Integrated transcriptome and proteome analysis indicates potential biomarkers of prostate cancer in offspring of pregnant rats exposed to a phthalate mixture during gestation and lactation.

As the second leading cause of death for cancer among men worldwide, prostate cancer (PCa) prevention and detection remain a critical challenge. One aspect of PCa research is the identification of common environmental agents that may increase the risk of initiation and progression of PCa. Endocrine disrupting chemicals (EDCs) are strong candidates for risk factors, partially because they alter essential pathways for prostate gland development and oncogenesis. Phthalates correspond to a set of commercially used plasticizers that humans are exposed to ubiquitously. Here, we show that maternal exposure to a phthalate mixture interferes with the expression profile of mRNA and proteins in the ventral prostate of offspring and increases the susceptibility to prostate adenocarcinomas in aged animals. The data highlight Ubxn11, Aldoc, Kif5c, Tubb4a, Tubb3, Tubb2, Rab6b and Rab3b as differentially expressed targets in young and adult offspring descendants (PND22 and PND120). These phthalate-induced targets were enriched for pathways such as: dysregulation in post-translational protein modification (PTPM), cell homeostasis, HSP90 chaperone activity, gap junctions, and kinases. In addition, the Kif5c, Tubb3, Tubb2b and Tubb4a targets were enriched for impairment in cell cycle and GTPase activity. Furthermore, these targets showed strong relationships with 12 transcriptional factors (TF), which regulate the phosphorylation of eight protein kinases. The correlation of TF-kinases is associated with alterations in immune system, RAS/ErbB/VEGF/estrogen/HIF-1 signaling pathways, cellular senescence, cell cycle, autophagy, and apoptosis. Downregulation of KIF5C, TUBB3 and RAB6B targets is associated with poor prognosis in patients diagnosed with adenocarcinoma. Collectively, this integrative investigation establishes the post-transcriptional mechanisms in the prostate that are modulated by maternal exposure to phthalate mixture during gestation and lactation.

Can ureteral stents with a different format reduce patients' symptomatology? A single-blinded, randomized, controlled study comparing smooth-walled and grooved ureteral stents.

INTRODUCTION: The true benefits of perioperative JJ stent placement are being widely studied due to its known side effects. However, no consensus has been reached in the literature regarding the best type of stent. This prospective, randomized, single-blinded study therefore aimed to compare the symptomatology associated with two JJ stent designs: smooth-walled and grooved. MATERIALS AND METHODS: The study prospectively recruited 42 patients who underwent JJ stent placement between July 2019 and August 2020. The patients were randomly divided into two groups according to the JJ stent design used: the smooth-walled stent (control) and grooved stent (intervention) groups. After surgery, all patients completed the Portuguese-validated Ureteral Stent Symptom Questionnaire at three timepoints (days 7 and 30 post-surgical procedure, and day 30 post-stent removal). RESULTS: No significant differences in gender, age, median body mass index, laterality, type of surgical procedure (flexible, semi-rigid or mixed ureteroscopy) were found. Smooth-walled JJ stents were associated with a higher incidence of flank pain (52.38% vs. 10%, P = .006) and suprapubic pain (57.14% vs. 30%, P = .04) on the 7th post-procedure day. Linear mixed regression showed significantly lower flank (P < .001) and suprapubic pain (P < .01), and significantly better sexual performance in the intervention group (P = .03). CONCLUSIONS: Ureteral stent with a grooved format are associated with a lower incidence of flank and suprapubic pain and had less impact on the sexual performance of patients.

Polyphenols and health: what compounds are involved?

On the basis of prospective, cross-sectional and intervention studies linking polyphenols to human health, several experimental papers in the literature have tried to evaluate the molecular mechanisms involved in their bioactivity. Polyphenols are reported to in vitro inhibit cancer cell proliferation, reduce vascularisation, protect neurons, stimulate vasodilation and improve insulin secretion, but are often studied as aglycones or as sugar conjugates and at non-physiological concentration. However, it is now well established that polyphenols undergo substantial metabolism after being ingested by humans in dietary relevant amount and that concentrations of plasma metabolites after a normal dietary intake rarely exceed nmol/L. This viewpoint intends to highlight that uncritical judgements made on the basis of the published literature, particularly about toxicity and bioactivity, may sometimes have been misled and misleading and to conclude that i) bioavailability values reported in the literature for phenolic compounds should be strongly reconsidered in the light of the large number of newly identified circulating and excreted metabolites, with particular attention to colonic ring-fission products which are obviously contributing much more than expected to the percentage of their absorption; ii) it is phenolic metabolites, formed in the small intestine and hepatic cells, and low molecular weight catabolic products of the colonic microflora to travel around the human body in the circulatory system or reach body tissues to elicit bioactive effects. Understanding these compounds certainly carries interest for drug-discovery but also for dietary prevention of disease.

Muscarinic receptors prevent oxidative stress-mediated apoptosis induced by domoic acid in mouse cerebellar granule cells.

In mouse cerebellar granule neurons (CGNs) low concentrations of domoic acid (DomA) induce apoptotic cell death, which is mediated by oxidative stress; apoptosis is more pronounced in CGNs from Gclm (-/-) mice, which lack the modifier subunit of glutamate cysteine ligase (GCL) and have very low GSH levels. By activating M(3) muscarinic receptors, the cholinergic agonist carbachol inhibits DomA-induced apoptosis, and the anti-apoptotic action of carbachol is more pronounced in CGNs from Gclm (+/+) mice. Carbachol does not prevent DomA-induced increase in reactive oxygen species, suggesting that its anti-apoptotic effect is downstream of reactive oxygen species production. Carbachol inhibits DomA-induced activation of Jun N-terminal (JNK) and p38 kinases, increased translocation to mitochondria of the pro-apoptotic protein Bax, and activation of caspase-3. Carbachol activates extracellular signal-regulated kinases 1/2 (ERK1/2) MAPK and phospahtidylinositol-3 kinase (PI3K) in CGNs from both genotypes. However, while the protective effect of carbachol is mediated by ERK1/2 MAPK in CGNs from both mouse genotypes, inhibitors of PI3K are only effective at antagonizing the action of carbachol in CGNs from Gclm (+/+) mice. In CGNs from both Gclm (+/+) and (-/-) mice, carbachol induces a MAPK-dependent increase in the level of the anti-apoptotic protein Bcl-2. In contrast, carbachol causes a PI3K-dependent increase in GCL activity and of GSH levels only in CGNs from Gclm (+/+) mice. Such increase in GCL is not because of a transcriptionally-mediated increase in glutamate cysteine ligase catalytic subunit or glutamate cysteine ligase modifier subunit, but rather to an increase in the formation of the GCL holoenzyme. The results indicate that multiple pathways may contribute to the protective action of carbachol toward DomA-induced apoptosis. Compromised GCLM expression, which is also found in a common genetic polymorphism in humans, leads to lower GSH levels, which can exacerbate the neurotoxicity of DomA, and decreases the anti-apoptotic effectiveness of muscarinic agonists.

Xestospongins: potent membrane permeable blockers of the inositol 1,4,5-trisphosphate receptor.

Xestospongins (Xe's) A, C, D, araguspongine B, and demethylxestospongin B, a group of macrocyclic bis-1-oxaquinolizidines isolated from the Australian sponge, Xestospongia species, are shown to be potent blockers of IP3-mediated Ca2+ release from endoplasmic reticulum vesicles of rabbit cerebellum. XeC blocks IP3-induced Ca2+ release (IC50 = 358 nM) without interacting with the IP3-binding site, suggesting a mechanism that is independent of the IP3 effector site. Analysis of Pheochromocytoma cells and primary astrocytes loaded with Ca2+-sensitive dye reveals that XeC selectively blocks bradykinin- and carbamylcholine-induced Ca2+ efflux from endoplasmic reticulum stores. Xe's represent a new class of potent, membrane permeable IP3 receptor blockers exhibiting a high selectivity over ryanodine receptors. Xe's are a valuable tool for investigating the structure and function of IP3 receptors and Ca2+ signaling in neuronal and nonneuronal cells.

Apoptosis induced by domoic acid in mouse cerebellar granule neurons involves activation of p38 and JNK MAP kinases.

In mouse cerebellar granule neurons (CGNs) the marine neurotoxin domoic acid (DomA) induces neuronal cell death, either by apoptosis or by necrosis, depending on its concentration, with apoptotic damage predominating in response to low concentrations (100 nM). DomA-induced apoptosis is due to selective activation of AMPA/kainate receptors, and is mediated by DomA-induced oxidative stress, leading to mitochondrial dysfunction and activation of caspase-3. The p38 MAP kinase and the c-Jun NH2-terminal protein kinase (JNK) have been shown to be preferentially activated by oxidative stress. Here we report that DomA increases p38 MAP kinase and JNK phosphorylation, and that this effect is more pronounced in CGNs from Gclm (-/-) mice, which lack the modifier subunit of glutamate-cysteine ligase, have very low glutathione (GSH) levels, and are more sensitive to DomA-induced apoptosis than CGNs from wild-type mice. The increased phosphorylation of JNK and p38 kinase was paralleled by a decreased phosphorylation of Erk 1/2. The AMPA/kainate receptor antagonist NBQX, but not the NMDA receptor antagonist MK-801, prevents DomA-induced activation of p38 and JNK kinases. Several antioxidants (GSH ethyl ester, catalase and phenylbutylnitrone) also prevent DomA-induced phosphorylation of JNK and p38 MAP kinases. Inhibitors of p38 (SB203580) and of JNK (SP600125) antagonize DomA-induced apoptosis. These results indicate the importance of oxidative stress-activated JNK and p38 MAP kinase pathways in DomA-induced apoptosis in CGNs.

Role of autophagy in environmental neurotoxicity.

Human exposure to neurotoxic pollutants (e.g. metals, pesticides and other chemicals) is recognized as a key risk factor in the pathogenesis of neurodegenerative disorders. Emerging evidence indicates that an alteration in autophagic pathways may be correlated with the onset of the neurotoxicity resulting from chronic exposure to these pollutants. In fact, autophagy is a natural process that permits to preserving cell homeostasis, through the seizure and degradation of the cytosolic damaged elements. However, when an excessive level of intracellular damage is reached, the autophagic process may also induce cell death. A correct modulation of specific stages of autophagy is important to maintain the correct balance in the organism. In this review, we highlight the critical role that autophagy plays in neurotoxicity induced by the most common classes of environmental contaminants. The understanding of this mechanism may be helpful to discover a potential therapeutic strategy to reduce side effects induced by these compounds.

Cholesterol homeostasis in the developing brain: a possible new target for ethanol.

Cholesterol is an essential component of cell membranes and plays an important role in signal transduction. This brief overview presents evidence from the literature that ethanol may affect cholesterol homeostasis and that, in the developing brain, this may be involved in its developmental neurotoxicity. The effects caused by inborn errors of cholesterol synthesis and by in utero ethanol exposure present several similarities in humans (eg, Smith-Lemli-Opitz syndrome and fetal alcohol syndrome), as well as in animal models. Ethanol has a cholesterol-reducing effect on the cardiovascular system, and a protective effect against Alzheimer's disease, whose pathogenesis has been linked to altered cholesterol homeostasis in the brain. In vitro, ethanol affects several functions that are mediated by cholesterol and important for brain development, such as glial cell proliferation, synaptogenesis, neuronal survival and neurite outgrowth. The brain contains high levels of cholesterol, mostly synthesized in situ. Astrocytes produce large amounts of cholesterol that can be released by these cells and utilized by neurons to form synapses. Ethanol up-regulates the cholesterol transporter ATP binding cassette A1 and cholesterol efflux from primary astrocyte cultures without affecting cholesterol synthesis.

Diet-brain connections: role of neurotoxicants.

In certain cases, the consumption of food or beverages can lead to intoxication and disease. Such food-induced intoxications may be due to microbial toxins, to toxic substances naturally occurring in some foods, or to contaminants or residues of various kinds. Some of these agents have neurotoxic properties and may contribute to the etiology of certain psychiatric disorders or neurodegenerative diseases. This paper reviews a selected number of dietary neurotoxicants that naturally, or as a result of human interventions, find their way into food or beverages, and have been associated with neurotoxic outcomes in humans. Chosen examples include domoic acid, a phycotoxin associated with amnesic shellfish poisoning; ß-N-oxalylamine-l-alanine (l-BOAA), present in chickling peas and believed to be responsible for neurolathyrism; and two alcohols, methanol and ethanol, which can cause severe neurotoxic effects in adults and the developing fetus.

Paraoxonase (PON1) gene in mice: sequencing, chromosomal localization and developmental expression.

Serum paraoxonase hydrolyses the toxic metabolites of several organophosphorus insecticides, as well as the nerve agents soman and sarin. We have previously shown that elevated serum paraoxonase levels protect mice against organophosphate toxicity. In the present study, we determined the cDNA sequence and chromosomal location of the mouse paraoxonase gene, as well as its developmental expression in mice and rats. The mouse cDNA encodes a protein of 355 amino acids and shows 81% identity with the human sequence. In situ hybridization demonstrated that the mouse paraoxonase gene maps to chromosome 6, a region conserved with the paraoxonase region of chromosome 7q21-22 in humans. Serum paraoxonase activities toward three substrates, paraoxon, chlorpyrifos-oxon and diazoxon, were very low at birth and increased with age reaching adult levels at 20 days in mice and 25 days in rats. The increase of serum paraoxonase activity in developing animals correlates well with the increased resistance to organophosphate poisoning that has been reported in previous studies.

Catalytic efficiency determines the in-vivo efficacy of PON1 for detoxifying organophosphorus compounds.

Human paraoxonase (PON1) is a polymorphic, high-density lipoprotein (HDL)-associated esterase that hydrolyzes the toxic metabolites of several organophosphorus (OP) insecticides and nerve agents. The activity polymorphism is determined by a Gln/Arg (Q/R) substitution at position 192. Injection of purified PON1 protects animals from OP poisoning. In the present study, we investigated the in-vivo function of PON1 for detoxifying organophosphorus insecticides in PON1-knockout mice that were challenged via dermal exposure with diazoxon, diazinon and paraoxon. PON1-knockout mice were extremely sensitive to diazoxon. Doses (2 and 4 mg/kg) that caused no cholinesterase (ChE) inhibition in wild-type mice were lethal to the knockout mice, which also showed slightly increased sensitivity to the parent compound diazinon. Surprisingly, these knockout mice did not show increased sensitivity to paraoxon. In-vitro assays indicated that the PON1R192 isoform hydrolyzed diazoxon less rapidly than did the PON1Q192 isoform. In-vivo analysis, where PON1-knockout mice received the same amount of either PON1(192) isoform via intraperitoneal (i.p.) injection 4 h prior to exposure, showed that both isoforms provided a similar degree of protection against diazoxon, while PON1R192 conferred better protection against chlorpyrifos-oxon than PON1Q192. Injection of purified rabbit PON1 or either human PON1(192) isoform did not protect PONI-knockout mice from paraoxon toxicity, nor did over-expression of the human PON1R192 transgene in wild-type mice. Kinetic analysis of the two human PON1(192) isoforms revealed that the catalytic efficiency (Vmax/Km) determines the in-vivo efficacy of PON1 for organophosphorus detoxication. The results indicate that PON1 plays a major role in the detoxication of diazoxon and chlorpyrifos oxon but not paraoxon.

Effects of ethanol on calcium homeostasis in the nervous system: implications for astrocytes.

Ethanol is a major health concern, with neurotoxicity occurring after both in utero exposure and adult alcohol abuse. Despite a large amount of research, the mechanism(s) underlying the neurotoxicity of ethanol remain unknown. One of the cellular aspects that has been investigated in relationship to the neuroteratogenicity and neurotoxicity of ethanol is the maintenance of calcium homeostasis. Studies in neuronal cells and other cells have shown that ethanol can alter intracellular calcium levels and affect voltage and receptor-operated calcium channels, as well as G protein-mediated calcium responses. Despite increasing evidence of the important roles of glial cells in the nervous systems, few studies exist on the potential effects of ethanol on calcium homeostasis in these cells. This brief review discusses a number of reported effects of alcohol on calcium responses that may be relevant to astrocytes' functions.

Linear versus nonlinear models for hemoglobin adduct formation by acrylamide and its metabolite glycidamide: implications for risk estimation.

Hemoglobin cysteine adduct levels formed by acrylamide (AA) and its epoxide metabolite glycidamide (GA) as previously determined (Bergmark et al., Toxicol. Appl. Pharmacol., 111: 352-363, 1991) in rats given single injections of AA were used to estimate tissue doses, D = integral of Cdt (area under the concentration curve in the blood compartment), of the two compounds. The data were adapted to linear or nonlinear kinetic models, where the latter model accounted for the Michaelis-Menten kinetics of the metabolic conversion of AA to GA. In the linear model, the first-order rates, k*, of elimination from all processes were estimated to be 0.50 and 0.48 h-1 for AA and GA, respectively. In the nonlinear model, the parametrical values Vmax = 19.1 h-1 and Km = 66 microM for the in vivo metabolic conversion of AA to GA, and k1 = 0.21 h-1 and k2 = 0.48 h-1 for the first-order rates of elimination from all other processes of AA and GA, respectively, were found to give the best fit to the exact dosimetric expressions [formula: see text] Using Equation B, it was estimated that the percentage of AA converted to GA approaches 58% when [AA]o, the initial concentration of AA, approaches zero. The implications for high-to-low-dose extrapolation of toxic effects of the derived mathematical relationships between administered dose and tissue dose are discussed.

A genetic polymorphism of MAO-B modifies the association of cigarette smoking and Parkinson's disease.

In a population-based case-control study, we found a reversal of the association of cigarette smoking with Parkinson's disease (PD) in relation to the monoamine oxidase B intron 13 genetic polymorphism. A reduced PD risk related to pack-years of smoking was detected for persons with the G allele, whereas an opposite effect was found among persons with the A allele. These results indicate an unexplained interaction between cigarette smoking and this genetic polymorphism.

The effects of ethanol on glial cell proliferation: relevance to the fetal alcohol syndrome.

Exposure to ethanol during pregnancy is detrimental to brain development. Individuals affected by the Fetal Alcohol Syndrome present a number of central nervous system dysfunctions including microencephaly and mental retardation. Studies on the mechanisms of ethanol's developmental neurotoxicity have focused on its interaction with neurons; however, emerging evidence is suggesting that ethanol can significantly affect glial cells as well. A number of in vitro studies have shown that ethanol can inhibit the proliferation of various glial cells (mostly primary astrocytes or astrocytoma cells) at relatively high concentrations (100-200 mM). On the other hand, proliferation induced by some, but not all mitogens, is inhibited by low concentrations (10-50 mM) of ethanol. These inhibitory effects of ethanol may contribute to its developmental neurotoxicity observed following in vivo exposure. Animal models have indeed shown that ethanol causes microencephaly when given during the brain growth spurt, a period of brain development characterized by astroglial proliferation and maturation.

A comparison of the antinociceptive responses to the GABA-receptor agonists THIP and baclofen.

The antinociceptive action of the gamma-aminobutyric acid (GABA) agonists THIP and baclofen was evaluated in mice using hot-plate (48 and 55 degrees C) and tail-immersion (50 degrees C) procedures. It was found that atropine reversed antinociception induced by THIP but not that induced by baclofen in the 48 degrees C test, whereas the anticholinergic drug blocked the response to both GABA agonists when the stimulus was provided by a 55 degrees C hot-plate. Atropine methylnitrate, mecamylamine, picrotoxin and bicuculline had no effect on antinociception induced by THIP or baclofen. Prior treatment with haloperidol enhanced only the response to baclofen on the 55 degrees C hot-plate. A reciprocal cross-tolerance was found between THIP and baclofen in the tail-immersion assay, although only THIP exhibited cross-tolerance to morphine. These results suggest that while the analgesic response to THIP and baclofen is partially mediated by a common system, the two agents act by independent mechanisms as well.

Identification of m3, m4 and m5 subtypes of muscarinic receptor mRNA in human blood mononuclear cells.

In this study we made use of the Reverse transcription-polymerase chain reaction to analyze the expression of mRNA for the five subtypes of muscarinic acetylcholine receptors in human blood mononuclear cells. mRNA for m3, m4 and m5 subtypes was detected, while mRNA for m1 and m2 muscarinic receptors was not found. Similar results were obtained for three different healthy human subjects studied. Interestingly, the m5 subtype was expressed at higher levels in blood mononuclear cells than in cerebral cortex. To our knowledge this is the first time that m5 muscarinic receptor mRNA has been found outside of the central nervous system.

Lack of m2 muscarinic acetylcholine receptor mRNA in rat lymphocytes.

The presence of muscarinic acetylcholine receptors on lymphocytes has been demonstrated by radioligand binding experiments. Although the specific subtype(s) of muscarinic acetylcholine receptors expressed in lymphocytes is still unknown, some reports suggest the presence of the m2 subtype. In this study we analyzed the expression of m2 subtype mRNA in rat mononuclear cells, B lymphocytes and T lymphocytes by Northern blot hybridization and reverse transcription-polymerase chain reaction. Positive signals for the presence of m2 mRNA were found in rat heart, brainstem, cerebral cortex, corpus striatum and hippocampus, which were used as positive controls. On the other hand, no expression of m2 was detected in lymphocytes. These results indicate that mRNA for the m2 subtype is absent in rat lymphocytes and that one or more other subtypes may be responsible for the reported results in binding experiments.