Antiparasitic Activity of Two Natural Terpenes from Salvia cuspidata against Leishmania amazonensis.

Leishmaniasis is a neglected disease caused by flagellated parasites of the Leishmania genus affecting more than 10 million people worldwide. Current treatments for leishmaniasis involve the administration of poorly tolerated drugs with toxic side effects in patients. There is an imperative necessity for novel compounds to treat this disease. One of the most used strategies in the search for different antiparasitic compounds is the screening of purified plant molecules. The diterpenes 12-hydroxy-11,14-diketo-6,8,12-abietatrien-19,20-olide (HABTO) and 5-epi-icetexone (ICTX) isolated from Salvia cuspidata were shown to be effective against Leishmania amazonensis in vitro and in vivo. They displayed an antiproliferative effect against L. amazonensis promastigotes. They also induce an increase in ROS levels and affect the mitochondrial activity of parasites. HABTO and ICTX in an in vivo model of cutaneous leishmaniasis decrease footpad swelling, parasite load, and splenic index. Moreover, they induce significant reduction in the O.D. of total anti-Leishmania IgG and IgG1 subtype antibody responses against L. amazonensis compared to the PBS group but maintain high levels of IgG2a. This suggests that in HABTO- or ICTX-treated mice, there is a slowdown in the progression of the disease. These terpenes could be considered as possible novel antileishmanial agents against L. amazonensis and thus treat cutaneous leishmaniasis.

Placental leukocyte infiltration accompanies gestational changes induced by hyperthyroidism.

In brief: The endocrine and immunological disruption induced by hyperthyroidism could alter gestation, placenta, and fetal development. This study suggests an immunological role of thyroid hormones in gestation. Abstract: Thyroid dysfunctions lead to metabolic, angiogenic, and developmental alterations at the maternal-fetal interface that cause reproductive complications. Thyroid hormones (THs) act through their nuclear receptors that interact with other steroid hormone receptors. Currently, immunological regulation by thyroid status has been characterized to a far less extent. It is well known that THs exert regulatory function on immune cells and modulate cytokine expression, but how hyperthyroidism (hyper) modulates placental immunological aspects leading to placental alterations is unknown. This work aims to throw light on how hyper modulates immunological and morphological placental aspects. Control and hyper (induced by a daily s.c. injection of T4 0.25 mg/kg) Wistar rats were mated 8 days after starting T4 treatment and euthanized on days 19 (G19) and 20 (G20) of pregnancy. We removed the placenta to perform qPCR, flow cytometry, immunohistochemistry, Western blot and histological analysis, and amniotic fluid and serum to evaluate hormone levels. We observed that hyper increases the fetal number, fetal weight, and placental weight on G19. Moreover, hyper induced an endocrine imbalance with higher serum corticosterone and changed placental morphology, specifically the basal zone and decidua. These changes were accompanied by an increased mRNA expression of glucocorticoid receptor and monocyte chemoattractant protein-1, an increased mRNA and protein expression of prolactin receptor, and an increase in CD45+ infiltration. Finally, by in vitro assays, we evidenced that TH induced immune cell activation. In summary, we demonstrated that hyper modulates immunological and morphological placental aspects and induces fetal phenotypic changes, which could be related to preterm labor observed in hyper.

Therapeutic effect of Prosopis strombulifera (LAM) BENTH aqueous extract on a murine model of cutaneous leishmaniasis.

Background and aim: Prosopis strombulifera (Lam.) Benth is a rhizomatous shrub native from different zones of Argentine Republic. P. strombulifera aqueous extract (PsAE) has different effects and several biological activities have been reported. The goal of this study was to analyze the activity of PsAE on a murine model of cutaneous leishmaniasis caused by Leishmania amazonensis. Experimental procedure: PsAE was orally administered at 150 mg/animal/day on BALB/c mice infected in the right footpad (RFP) with 1 × 105 promastigotes of L. amazonensis. As a chemotherapeutic control of treatment, animals receive a commercial form of meglumine antimoniate (MA) (Glucantime®, Aventis, Paris, France). Results and conclusion: We observe that the size of RFP lesions of infected mice without treatment showed a grade of inflammation, ulceration and necrosis at the site of infection much greater than that observed with PsAE or MA treatment. Moreover, PsAE was capable of decreasing parasite burden and splenic index. Furthermore, PsAE treated mice showed a significant decrease in O.D. of total anti-Leishmania IgG antibody responses against L. amazonensis. This decrease was similar to those observed when the reference drug, MA, was used. This would indicate that PsAE treatment inhibits or delays disease progression in mice. In conclusion, our findings suggest that PsAE could be a potential candidate to be used, as a new therapeutic strategy, to treat cutaneous leishmaniasis caused by L. amazonensis.

UHPLC-Q/Orbitrap/MS/MS fingerprinting and antitumoral effects of Prosopis strombulifera (LAM.) BENTH. queous extract on allograft colorectal and melanoma cancer models.

The aqueous extract of the Argentinean native plant, Prosopis strombulifera (PsAE), presents cytotoxicity against human cancer cell lines by inducing cytostasis, necrosis and apoptosis; with diminution of clonogenic survival; without genotoxic effects nor oral animal toxicity. Until now, the chemical extract composition and its in vivo antitumoral properties remain unknown; these studies are the aim of the current work. The PsAE was characterized by chemical fingerprinting and the metabolome was identified by tandem UHPLC-PDA-HESI-Q-orbitrap® mass spectrometry. Colorectal tumors were induced by DMH administration and melanomas resulted from B16-F0 S.C. cells injection; then, animals were treated orally with PsEA. To correlate in vivo results with in vitro cytotoxicity, B16-F0 cell were cultured to determine: cell proliferation and viability by dye exclusion assays, MTT and CFSE dilution; cell cycle distribution by flow cytometry; and immunoblotting of p21cip1, PCNA, cleaved caspase 3, cleaved PARP and TUBA1A. Based on UHPLC-OT-MS and PDA analysis, twenty-six compounds were identified, including: 5 simple organic acids, 4 phenolic acids, 4 procyanidins, 11 flavonoids, and 2 oxylipins. On C57BL6 mice, PsAE significantly increases the median survival on colorectal cancer and reduces the final volume and weight of melanomas. Over cultured cells, the treatment induce over-expression of p21, cytostasis by G2/M cell cycle arrest and apoptosis; while, on in vivo melanomas, treatment up-regulates p21 and slightly decreases PCNA. In conclusion, PsAE is composed by phenolic compounds which demonstrate cytotoxic and antitumoral properties when is orally administrated. Presented results support future research of PsAE as a potential phytomedicine for cancer treatment.

Mutant huntingtin affects endocytosis in striatal cells by altering the binding of AP-2 to membranes.

Clathrin-mediated endocytosis plays an important role in the maintenance of neuronal integrity in the synaptic terminals. Here we studied the effect of anomalous polyglutamine expansion in huntingtin on the interaction of coat proteins with membranes, in areas of mouse brain or in cultured striatal cells. We observed that this anomaly induces a redistribution of AP-2, but not other coat proteins, from the membrane to the cytosol in the striatum, and in the cultured striatal cells. It was also noted that huntingtin associates with AP-2, and that this association decreases due to the mutation in huntingtin. This decreased receptor-mediated endocytosis, measured by the internalization of transferrin in the mutated cells. It was also confirmed that huntingtin mutation made the cells more vulnerable to the action of quinolinic acid, with an increasing degradation of the AP-2 alpha subunits. On the basis of these results, we conclude that abnormal polyglutamine expansion in huntingtin affects clathrin-mediated endocytosis, and may be one of the pathogenic mechanisms of neurodegeneration.

Expression of NHERF1 in colonic tumors induced by 1,2-dimethylhydrazine in rats is independent of plasma ovarian steroids.

In normal embryonic fibroblasts, the Na(+)/H(+) exchanger regulator factor 1 (NHERF1) stabilizes E-cadherin/ß-catenin binding and the lack of NHERF1 expression promotes cell transformation thus acting as a tumor suppressor gene. We here tested the hypothesis that NHERF1 could act as a tumor suppressor gene in colon cancer as a mediator of estrogens' protective actions in colon carcinogenesis. We studied the expression and localization of NHERF1 and ß-catenin by immunohistochemistry in colonic tumors induced by 1,2 dimethylhidrazine (DMH) in Sprague-Dawley rats. One group of the rats treated with the carcinogen was ovariectomized (OVX) in the middle of the tumor induction, simulating a human menopausal condition. We observed a protective role of estrogens in colon cancer, as non-ovariectomized rats (DMH) had a reduced tumor area compared with the ovariectomized group (DMH + OVX; mean ± SE) 28.98 ± 4.65 vs. 67.58 ± 8.69 (p < 0.00380). Despite the lack of plasma estrogen stimulation, we found abundant expression of NHERF1 in colon tumors from ovariectomized rats. NHERF1 was mainly localized in the cytoplasm of the adenocarcinoma cells and lost the apical localization previously reported in normal colon tissue. We also detected expression of NHERF1 by western blot in the SW48, CACO-2, and HT29 colon cancer cell lines. Non-estrogenic factors in plasma or the tumor microenvironment may regulate NHERF1 expression in transformed colon epithelial cells. Further studies are required to understand the regulation of NHERF1 expression in colon cancer tissue.

Estrogens regulate the expression of NHERF1 in normal colon during the reproductive cycle of Wistar rats.

In breast cancer cell lines, the Na(+)/H(+) exchanger regulator factor 1 (NHERF1) gene is regulated at the transcriptional level by estrogens, the protein expression levels correlate with the presence of estrogen receptors and the effect is blocked by anti-estrogens. However, there is limited information regarding the regulation of NHERF1 by estrogens in normal colon tissue. The NHERF1 protein has an important role in the maintenance of the intestine ultrastructure. NHERF1-deficient mice showed defects in the intestinal microvilli as well as molecular alterations in brush border membrane proteins. Here, we have studied the expression of NHERF1 in normal rat colon and uterus during the reproductive cycle of Wistar rats. We found that NHERF1 expression in rat colon during the estral cycle is modified by estrogen levels: higher expression of NHERF1 was observed during the proestrous and estrous stages and lower expression in diestrous 1 when estrogen levels decreased. In uterus, NHERF1 was expressed in the apical region of the luminal epithelium and glands in all stages of the estral cycle, and in both colon and uterus, the expression was independent of the proliferation status. Our results show that NHERF1 expression is regulated by estrogens in colon during the rat estral cycle.