Pro-inflammatory cytokines are modified during the multiplication of Trypanosoma cruzi within the placental chorionic villi and are associated with the level of infection via the signaling pathway NF-κB.

PROBLEM: Congenital Trypanosoma cruzi (T. cruzi) infection has been associated with changes in the levels of TNF-α and IFN-γ during the pregnancy. Therefore, we propose to study the participation and dynamics of proinflammatory cytokines in the infection process of placental explants infected by T. cruzi in vitro. METHOD OF STUDY: Chorionic villous explants (CVE) obtained of human term placentas (n = 8) from normal pregnancies were cultured with 105 trypomastigotes/mL of Tulahuen strain DTU VI for 0, 2, 4, 16, 24, 48 and 72 h. Explants were treated with sulfasalazine (SULF) (5 mM) and N-acetyl-cysteine (NAC) (15 mM), as inhibitors molecules of NF-κB pathway, or LPS (1 µg/mL) for 24 and 72 h p.i. Motile trypomastigotes were counted in culture supernatants. Immunohistochemistry and ELISA for TNF-α, IFN-γ, IL-1ß, IL-4, and IL-10 were performed in CVE and culture supernatants respectively. The parasite load was measured by RT-qPCR. RESULTS: T. cruzi invades the chorionic villi from 4 h p.i. increasing significantly its DNA at 48 and 72 h p.i. of culture (parasite multiplication phase). They were detected in stromal cells, which was related to elevation of TNF-α, IL-1ß, IFN-γ, and IL-10. The inhibition of NF-κB activity in the explants decreased the production of the analyzed cytokines, showing elevated levels of T. cruzi DNA during the multiplication phase of the parasite. CONCLUSIONS: Placental tissue modifies the secretion of pro-inflammatory cytokines during the phase of parasite multiplication, but not during the invasion phase, which in turns modifies the level of infection via the signaling pathway NF-κB.

Chagas disease affects the human placental barrier's turnover dynamics during pregnancy.

BACKGROUND: Trypanosoma cruzi crosses the placental barrier and produces the congenital transmission of Chagas disease (CD). Structural alterations of the chorionic villi by this parasite have been described in vitro, but little is known about trophoblast turnover in placentas from women with CD. OBJECTIVE: To analyze the proliferation and fusion processes in placentas from women with CD. METHODS: Archived human term placenta paraffin-embedded blocks were used, from women with CD (CDP), and no pathology (NP). Immunohistochemistry tests were performed for Ki67 to calculate the proliferation index (PI) of cytotrophoblast (CTB) and Syncytin-1, a fusion marker of syncytiotrophoblast (STB). Hematoxylin/Eosin stained sections were employed to analyze STB percentages, STB detachment areas and syncytial knots quantity. Non parametric Student's t-tests were performed (p < 0.05). RESULTS: Syncytial knots and STB detachment significantly increased in placental villi from the CDP group. STB percentage was significantly lower in the CDP group as well as the PI and Syncytin-1 expression significantly decreased in these placentas, compared with control (NP). CONCLUSION: Dynamic of trophoblast turnover is altered in placentas from women with CD. These changes may lead into a gap in the placental barrier possibly allowing the parasite entry into the chorionic villi.

Placental vascularization in middle and late gestation in the pig.

The aims of this study were to determine the changes in the capillary area density in relation to fetal development, to determine immunoexpression of angiogenic factors and to compare their mRNA expression throughout pig gestation. Samples were collected from the maternal-chorioallantoic interface at days 40, 77, 85 and 114 of pregnancy for immunohistochemistry analysis and the measurement of mRNA expression of VEGFA, ANGPT1, ANGPT2, FGF2 and its receptors KDR, TEK, FGFR1, FGFR2respectively. Morphometric measurement of blood vessels was performed. We found a significant increase in capillary area density throughout gestation (P< 0.05). On the maternal side, at day 77, we observed a significant increase in the number of vessels from small vascular areas (P < 0.05) and the vascular area was significantly higher on day 85 (P < 0.05). On the fetal side, the number of vessels and the vascular area increased between days 40 and 77 (P < 0.05) and between days 77 and 114 (P < 0.05), respectively. Immunohistochemical findings revealed intense VEGFA staining and a trend for increased expression towards the end of gestation (P < 0.05). We also demonstrated a high VEGFA, FGF2, FGFR1, ANGPT1 and ANGPT2mRNA expression at day 77 (P < 0.05). In conclusion, our findings suggest that an active angiogenesis would be present even until late-middle gestation at day 77 of pregnancy with the predominance of angiogenic stimulation by VEGFA/KDR, FGF2/FGFR1 and a balance between ANGPT1 and ANGPT2/TEK. Lay summary: Critical moments occur at different stages of placental formation in pigs, where the expression of angiogenic factors, that is, molecules that stimulate the formation of blood vessels must be adequate to promote their development. This exchange is necessary to cover the increasing nutritional demands of fetuses in continuous development. Determining the changes in the area of capillary density in relation to fetal development and the expression of angiogenic factors throughout pregnancy in pigs could contribute to understanding the causes of fetal loss. Placental samples were obtained at gestational days 40, 77, 85 and 114 (n = 7, 10, 7 and 5, respectively). We found that the capillary area density increases accompanying fetal growth with advancing gestation and an increase in capillary area density in late-middle gestation, around day 77, is due to the expansion in the number of small blood vessels on the maternal side. The present findings suggest that an intense angiogenesis would be present even until late-middle gestation at day 77 of pregnancy, with the predominance of angiogenic stimulation by specific molecules that promote this process.

Chagas disease affects the human placental barrier's turnover dynamics during pregnancy

BACKGROUND Trypanosoma cruzi crosses the placental barrier and produces the congenital transmission of Chagas disease (CD). Structural alterations of the chorionic villi by this parasite have been described in vitro, but little is known about trophoblast turnover in placentas from women with CD. OBJECTIVE To analyze the proliferation and fusion processes in placentas from women with CD. METHODS Archived human term placenta paraffin-embedded blocks were used, from women with CD (CDP), and no pathology (NP). Immunohistochemistry tests were performed for Ki67 to calculate the proliferation index (PI) of cytotrophoblast (CTB) and Syncytin-1, a fusion marker of syncytiotrophoblast (STB). Hematoxylin/Eosin stained sections were employed to analyze STB percentages, STB detachment areas and syncytial knots quantity. Non parametric Student's t-tests were performed (p < 0.05). RESULTS Syncytial knots and STB detachment significantly increased in placental villi from the CDP group. STB percentage was significantly lower in the CDP group as well as the PI and Syncytin-1 expression significantly decreased in these placentas, compared with control (NP). CONCLUSION Dynamic of trophoblast turnover is altered in placentas from women with CD. These changes may lead into a gap in the placental barrier possibly allowing the parasite entry into the chorionic villi.

Maternal intake of alpha-lipoic acid prevents development of symptoms associated with a fructose-rich diet in the male offspring in Wistar rats.

The hypothesis was that maternal intake of the antioxidant alpha-lipoid acid (ALA), during the developmental period of the hypothalamic orexigenic neurons, causes a permanent beneficial effect in offspring metabolism. Pregnant Wistar rats were fed with standard diet (food) + ALA (0.4% wt/wt) from day 14 of gestation to day 20 of lactation (n = 4) or food (n = 4). At 3 months of age, male offspring born from ALA-fed rats or controls (CT) were randomly assigned to be fed with food + 10% fructose solution in drinking water (F) or food + tap water (C), resulting in four groups: ALAF, ALAC, CTF, and CTC (n = 5/group). Food intake and body weight (BW) were measured twice a week for 31 days. Metabolites' levels in blood, mRNA expressions of Npy, Agrp (hypothalamus), Fasn, Srebf1, Ppard, and Pparg (liver), and the antioxidant capacity of the liver were determined. Results significance was set at p < 0.05. Average BW gain, daily BW gain, and intraabdominal fat tissue at necropsy were higher in CTF group followed by CTC, ALAF, and ALAC groups. There were no differences between groups in Kcal intake per day. mRNA expressions of hypothalamic and hepatic genes and plasmatic levels of glucose and triglycerides were higher in CTF group followed by ALAF, CTC, and ALAC groups. Fructose intake affected the oxidative capacity of the liver, but this effect was not observed in the ALAF group. In conclusion, maternal ALA intake protected the adult offspring to develop metabolic symptoms associated with high fructose in the drinking water.

Modulatory effects of ghrelin on sperm quality alterations induced by a fructose-enriched diet.

The objectives of this study were: 1) to evaluate the effects of a fructose enriched diet (FED) on rat sperm quality, epididymal function (i.e. oxidative stress and alpha-glucosidase expression) and testosterone concentrations; 2) to determine if the administration of ghrelin (Ghrl), reverses the effects induced by FED. After validating the protocol as an inductor of metabolic syndrome like-symptoms, adult male rats were assigned to one of the following treatments for 8 weeks: FED = 10% fructose enriched in water (v/v); FED + Ghrl = fructose enriched diet plus Ghrl (6 nmol/animal/day, s.c.) from week 6-8; or C = water without fructose (n = 5-10 animals/group). FED significantly decreased sperm concentration and motile sperm count/ml vs C (FED: 19.0 ± 1.6 × 106sperm/ml and 834.6 ± 137.0, respectively vs C: 25.8 ± 2.8 × 106 and 1300.4 ± 202.4, respectively; p < 0.05); ghrelin injection reversed this negative effect (23.5 ± 1.6 × 106sperm/ml and 1381.7 ± 71.3 respectively). FED resulted in hypogonadism, but Ghrl could not normalize testosterone concentrations (C: 1.4 ± 0.1 ng/ml vs FED: 0.8 ± 0.2 ng/ml and FED + Ghrl: 0.6 ± 0.2 ng/ml; p < 0.05). Ghrelin did not reverse metabolic abnormalities secondary to FED. FED did not alter epididymal expression of antioxidants enzymes (superoxido-dismutase, catalase and glutathione peroxidases -Gpx-). Nevertheless, FED + Ghrl significantly increased the expression of Gpx3 (FED + Ghrl: 3.47 ± 0.48 vs FED: 0.69 ± 0.28 and C: 1.00 ± 0.14; p < 0.05). The expression of neutral alpha-glucosidase, which is a marker of epididymal function, did not differ between treatments. In conclusion, the administration of Ghrl modulated the negative effects of FED on sperm quality, possibly by an epididymal increase in Gpx3 expression. However, Ghrl could not neither normalize the metabolism of FED animals, nor reverse hypogonadism.

4-Locus high-resolution HLA allele and haplotype frequencies in admixed population from Nicaragua.

A total of 155 Nicaraguan Mestizos from across the country were genotyped at high-resolution for the human leukocyte antigen loci HLA-A, -B, -C, and -DRB1 using sequence-based typing methods. The respective allele and extended haplotype frequencies, as well as Hardy-Weinberg proportions were calculated. The most frequent extended haplotype identified was A*24:02:01-B*40:02:01-C*03:05-DRB1*04:07:01G, with an estimated frequency of 2.26%. No deviation from Hardy-Weinberg Equilibrium was detected at any of the loci studied. The HLA genotypic data of the population sample reported here are available publicly in the Allele Frequencies Net Database under the population name "Nicaragua Mestizo" and the identifier AFN3610.

Maternal Triclosan consumption alters the appetite regulatory network on Wistar rat offspring and predispose to metabolic syndrome in the adulthood.

The objectives of this study were to evaluate the effects of maternal oral exposure to the antibacterial Triclosan (TCS) during gestation and lactation on the metabolic status of the adult offspring and on the expression of main genes controlling the appetite regulatory network. Pregnant rats were fed ad-libitum with ground food + TCS (1 mg/kg) from day 14 of gestation to day 20 of lactation (n=3) or ground food (n=3). After litter reduction, 12 males and 12 females born from the TCS exposed rats (TCS, n=24) or not (Control, n=24) were used to evaluate monthly body weight, food intake, plasma levels of cholesterol, glucose and triglycerides, and the hypothalamic mRNA expression of agouti-related protein (Agrp), neuropeptide Y (Npy) and propiomelanocortin (Pomc). Body weight for rats in the TCS group was 12.5% heavier for males at 4 months (p<0.001) and 19% heavier for females at 8 months (p=0.01). Food intake was significantly higher for rats in the TCS group at 5 months of age (p<0.01). Cholesterol and glucose levels were significantly higher for rats in the TCS group at 8 months (p<0.05). mRNA expression of Npy and Agrp were significantly increased in hypothalami of rats in the TCS group at 2 months for males or 8 months for females (p<0.05). In conclusion, low doses of oral TCS consumption by the pregnant and lactating dam increase the hypothalamic expression of the orexigenic neuropeptides Npy and Agrp in the offspring and alter their metabolic status during adulthood, resembling development of the metabolic syndrome.