Inhibition of multidrug resistance proteins by MK571 restored the erectile function in obese mice through cGMP accumulation.

BACKGROUND: Intracellular levels of cyclic nucleotides can also be controlled by the action of multidrug resistance protein types 4 (MRP4) and 5 (MRP5). To date, no studies evaluated the role of their inhibition in an animal model of erectile dysfunction (ED). OBJECTIVES: To evaluate the effect of a 2-week treatment with MK571, an inhibitor of the efflux of cyclic nucleotides in the ED of obese mice. MATERIALS AND METHODS: Mice were divided in three groups: (i) lean, (ii) obese, and (iii) obese + MK571. The corpus cavernosum (CC) were isolated, and concentration-response curves to acetylcholine (ACh), sodium nitroprusside (SNP), and tadalafil in addition to electrical field stimulation (EFS) were carried out in phenylephrine pre-contracted tissues. Expression of ABCC4 and ABCC5, intracellular levels of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), the protein levels for pVASPSer157 and pVASPSer239 , and the intracavernous pressure (ICP) were also determined. The intracellular and extracellular (supernatant) ratios in CC from obese and lean stimulated with a cGMP-increasing substance (BAY 58-2667) in the absence and presence of MK571 (20 µM, 30 min) were also assessed. RESULTS: The treatment with MK571 completely reversed the lower relaxing responses induced by EFS, ACh, SNP, and tadalafil observed in obese mice CC in comparison with untreated obese mice. Cyclic GMP and p-VASPSer239 expression were significantly reduced in CC from obese groups. MK571 promoted a sixfold increase in cGMP without interfering in the protein expression of p-VASPSer239 . Neither the cAMP levels nor p-VASPSer157 were altered in MK571-treated animals. The ICP was ∼50% lower in obese than in the lean mice; however, the treatment with MK571 fully reversed this response. Expressions of ABCC4 and ABCC5 were not different between groups. The intra/extracellular ratio of cGMP was similar in CC from obese and lean mice stimulated with BAY 58-2667. CONCLUSIONS: The MRPs inhibition by MK571 favored the accumulation of cGMP in the smooth muscle cells, thus improving the smooth muscle relaxation and the erectile function in obese mice.

Anoxygenic photosynthesis linked to Neoarchean iron formations in Carajás (Brazil).

Microbial activity is often invoked as a direct or indirect contributor to the precipitation of ancient chemical sedimentary rocks such as Precambrian iron formations (IFs). Determining a specific metabolic pathway from the geological record remains a challenge, however, due to a lack of constraints on the initial conditions and microbially induced redox reactions involved in the formation of iron oxides. Thus, there is ongoing debate concerning the role of photoferrotrophy, that is the process by which inorganic carbon is fixed into organic matter using light as an energy source and Fe(II) as an electron donor, in the deposition of IFs. Here, we examine ~2.74-Ga-old Neoarchean IFs and associated carbonates from the Carajás Mineral Province, Brazil, to reconstruct redox conditions and to infer the oxidizing mechanism that allowed one of the world's largest iron deposits to form. The absence of cerium (Ce) anomalies reveals that conditions were pervasively anoxic during IF deposition, while unprecedented europium (Eu) anomalies imply that Fe was supplied by intense hydrothermal activity. A positive and homogeneous Fe isotopic signal in space and time in these IFs indicates a low degree of partial oxidation of Fe(II), which, combined with the presence of 13 C-depleted organic matter, points to a photoautotrophic metabolic driver. Collectively, our results argue in favor of reducing conditions during IF deposition and suggest anoxygenic photosynthesis as the most plausible mechanism responsible for Fe oxidation in the Carajás Basin.

Impact of gestational low-protein intake on embryonic kidney microRNA expression and in nephron progenitor cells of the male fetus.

BACKGROUND: Here, we have demonstrated that gestational low-protein (LP) intake offspring present lower birth weight, reduced nephron numbers, renal salt excretion, arterial hypertension, and renal failure development compared to regular protein (NP) intake rats in adulthood. We evaluated the expression of various miRNAs and predicted target genes in the kidney in gestational 17-days LP (DG-17) fetal metanephros to identify molecular pathways involved in the proliferation and differentiation of renal embryonic or fetal cells. METHODS: Pregnant Wistar rats were classified into two groups based on protein supply during pregnancy: NP (regular protein diet, 17%) or LP diet (6%). Renal miRNA sequencing (miRNA-Seq) performed on the MiSeq platform, RT-qPCR of predicted target genes, immunohistochemistry, and morphological analysis of 17-DG NP and LP offspring were performed using previously described methods. RESULTS: A total of 44 miRNAs, of which 19 were up and 25 downregulated, were identified in 17-DG LP fetuses compared to age-matched NP offspring. We selected 7 miRNAs involved in proliferation, differentiation, and cellular apoptosis. Our findings revealed reduced cell number and Six-2 and c-Myc immunoreactivity in metanephros cap (CM) and ureter bud (UB) in 17-DG LP fetuses. Ki-67 immunoreactivity in CM was 48% lesser in LP compared to age-matched NP fetuses. Conversely, in LP CM and UB, ß-catenin was 154%, and 85% increased, respectively. Furthermore, mTOR immunoreactivity was higher in LP CM (139%) and UB (104%) compared to that in NP offspring. TGFß-1 positive cells in the UB increased by approximately 30% in the LP offspring. Moreover, ZEB1 metanephros-stained cells increased by 30% in the LP offspring. ZEB2 immunofluorescence, although present in the entire metanephros, was similar in both experimental groups. CONCLUSIONS: Maternal protein restriction changes the expression of miRNAs, mRNAs, and proteins involved in proliferation, differentiation, and apoptosis during renal development. Renal ontogenic dysfunction, caused by maternal protein restriction, promotes reduced reciprocal interaction between CM and UB; consequently, a programmed and expressive decrease in nephron number occurs in the fetus.

Menthol ameliorates voiding dysfunction in types I and II diabetic mouse model.

AIMS: Overactive bladder (OAB) is one of the most common complications of both type 1 (T1DM) and type 2 diabetes mellitus (T2DM). In healthy conditions, menthol infused intravesically reduces the threshold for initiating micturition reflex, but no study evaluated its effects in diabetic conditions. Therefore, we have used mouse models of T1DM and T2DM to evaluate the effects of menthol on cystometric alterations and increased bladder contractility in vitro. METHODS: For T1DM induction, male C57BL6 mice were injected with streptozotocin (STZ) and evaluated after 4 weeks. For T2DM induction, mice were fed with high-fat diet (HFD) for 12 weeks to induce obesity. Urodynamic profiles were assessed by filling cystometry through the infusion of menthol (100 µM for 30 min) or vehicle (DMSO 0.1%). Contractile responses to carbachol, potassium chloride (KCl), and electrical-field stimulation (EFS) were measured in isolated bladders after 20 min incubation with menthol (100 µM) or vehicle. RESULTS: Filling cystometry showed that STZ-injected mice exhibited higher bladder capacity, threshold pressure, and non-voiding contractions (NVCs), which were significantly reduced by menthol infusion. The increased voiding frequency in STZ group were unaffected by menthol. In HFD-fed obese mice menthol significantly attenuated the increased threshold pressure and NVC frequency, but unaffected the changes of voiding frequency. In both STZ-injected and HFD-fed mice, incubation of isolated bladders with menthol normalized the enhanced contractile responses to carbachol, KCl, and EFS stimulation. CONCLUSIONS: Menthol may be a potential pharmacological option for the treatment of OAB as a consequence of T1DM and T2DM.

Impaired dipsogenic and renal response to repetitive intracerebroventricular angiotensin II (AngII) injections in rats.

The role of the central nervous system (CNS) in the control of blood pressure and hydrosaline homeostasis has been demonstrated by several studies. While circulating angiotensin II (AngII) tends to retain sodium by a direct renal action as well as through aldosterone release, stimulation of brain AngII receptors has been reported to induce natriuresis. Repetitive intracerebroventricular AngII injection was recently demonstrated to be capable of leading to desensitisation of the dipsogenic effect of AngII stimuli. The aim of the current study was to investigate a possible central desensitisation to AngII stimuli by observing the effects of a low-concentration solution of AngII on the dipsogenic and natriuretic mechanisms in conscious rats, compared with appropriate age-matched 0.15 M NaCl-injected subjects, as evaluated by lithium clearance. The present report confirmed earlier reports on the potent natriuretic and dipsogenic effects of central AngII receptor stimulation. Natriuresis is mediated by a decrease in sodium reabsorption in the proximal and post-proximal tubule segments of the nephron. The current findings lend further support to the idea that AngII, in the CNS, is instrumental in the regulation of body fluid homeostasis. The magnitude of the dipsogenic and renal response to AngII was significantly decreased by repetitive stimulus.

Hypothalamic SOCS-3 expression and the effect of intracerebroventricular angiotensin II injection on water intake and renal sodium handling in SHR.

In rats, the acute central dipsogenic and natriuretic action of angiotensin II (AngII) seems to be independent of the hemodynamic effects of the peptide; however, in genetically hypertensive models, this relationship has not yet been investigated. It has been demonstrated that AngII induces the suppressor of cytokine signaling (SOCS-3) expression in the brain that, in turn, modulates further activation of the pathway, leading to desensitization to AngII stimuli with regard to its dipsogenic effect. This study investigates age-related Janus kinase (JAK-2) and SOCS-3 hypothalamic expression, by immunoblotting, and the involvement of SOCS-3 expression in urinary sodium handling and dipsogenic response in spontaneously hypertensive rats (SHR), compared with age-matched Wistar-Kyoto (WKy) rats. The intracerebroventricular (i.c.v.) application of AngII significantly enhanced the dipsogenic response, reduced C(Cr), and reciprocally promoted increased absolute and fractional rates of excretion of sodium in WKy rats. The central AngII-induced dipsogenic effect in WKy and SHR was significantly attenuated by prior i.c.v. administration of DUP753. In addition, the magnitude of the dipsogenic and renal response to AngII was significantly attenuated in age-matched SHR. Blocking of hypothalamic SOCS-3 expression by an antisense oligonucleotide resulted in partial reversal of the refractory nature of AngII in thirst responses in SHR. The altered centrally applied AngII response in SHR associated with increased hypothalamic JAK-2/SOCS-3 expression may suggest that abnormal regulation of the central angiotensin pathways may contribute to dysfunction of water-electrolyte homeostasis in SHR.