Trypanosoma cruzi uses several strategies to survive in different hosts. A key step in the life-cycle of this parasite is metacyclogenesis, which involves various morphological, biochemical, and ...genetic changes that induce the differentiation of non-pathogenic epimastigotes into pathogenic metacyclic trypomastigotes. During metacyclogenesis, T. cruzi displays distinct morphologies and ultrastructural features, which have not been fully characterized.
We performed a temporal description of metacyclogenesis using different microscopy techniques that resulted in the identification of three intermediate forms of T. cruzi: intermediates I, II and III. Such classification was based on morphological and ultrastructural aspects as the location of the kinetoplast in relation to the nucleus, kinetoplast shape and kDNA topology. Furthermore, we suggested that metacyclic trypomastigotes derived from intermediate forms that had already detached from the substrate. We also found that changes in the kinetoplast morphology and kDNA arrangement occurred only after the repositioning of this structure toward the posterior region of the cell body. These changes occurred during the later stages of differentiation. In contrast, changes in the nucleus shape began as soon as metacyclogenesis was initiated, while changes in nuclear ultrastructure, such as the loss of the nucleolus, were only observed during later stages of differentiation. Finally, we found that kDNA networks of distinct T. cruzi forms present different patterns of DNA topology.
Our study of T. cruzi metacyclogenesis revealed important aspects of the morphology and ultrastructure of this intriguing cell differentiation process. This research expands our understanding of this parasite's fascinating life-cycle. It also highlights the study of T. cruzi as an important and exciting model system for investigating diverse aspects of cellular, molecular, and evolutionary biology.
Heparan sulfate proteoglycans (HSPGs) act as signaling co‐receptors by interaction of their sulfated glycosaminoglycan chains with numerous signaling molecules. In breast cancer, the function of ...heparan sulfate 2‐O‐sulfotransferase (HS2ST1), the enzyme mediating 2‐O‐sulfation of HS, is largely unknown. Hence, a comparative study on the functional consequences of HS2ST1 overexpression and siRNA knockdown was performed in the breast cancer cell lines MCF‐7 and MDA‐MB‐231. HS2ST1 overexpression inhibited Matrigel invasion, while its knockdown reversed the phenotype. Likewise, cell motility and adhesion to fibronectin and laminin were affected by altered HS2ST1 expression. Phosphokinase array screening revealed a general decrease in signaling via multiple pathways. Fluorescent ligand binding studies revealed altered binding of fibroblast growth factor 2 (FGF‐2) to HS2ST1‐expressing cells compared with control cells. HS2ST1‐overexpressing cells showed reduced MAPK signaling responses to FGF‐2, and altered expression of epidermal growth factor receptor (EGFR), E‐cadherin, Wnt‐7a, and Tcf4. The increased viability of HS2ST1‐depleted cells was reduced to control levels by pharmacological MAPK pathway inhibition. Moreover, MAPK inhibitors generated a phenocopy of the HS2ST1‐dependent delay in scratch wound repair. In conclusion, HS2ST1 modulation of breast cancer cell invasiveness is a compound effect of altered E‐cadherin and EGFR expression, leading to altered signaling via MAPK and additional pathways.
Our results suggest that, in breast cancer, high levels of HS2ST1 result in structural changes in heparan sulfate and altered growth factor binding, which leads to attenuated signaling through the MAPK and additional pathways. Reduced signaling and expression of E‐cadherin and epidermal growth factor receptor (EGFR) is associated with reduced viability, adhesion, migration, and invasion of breast cancer cells.
Metastasis is the leading cause of cancer-related deaths. Despite this relevance, there is no specific therapy targeting metastasis. The interaction of the tumor cell with platelets, forming ...microemboli is crucial for successful hematogenous dissemination. Heparin disrupts it by a P-selectin-mediated event. However, its clinical use for this purpose is hindered by the requirement of high doses, leading to anticoagulant-related side effects. In this study, we obtained a low-anticoagulant heparin through the fractionation of a pharmaceutical bovine heparin. This derivative was referred to as LA-hep and we investigated its efficacy in inhibiting metastases and explored its capacity of suppressing the interaction between tumor cells and platelets. Our data revealed that LA-hep is as efficient as porcine unfractionated heparin in attenuating lung metastases from melanoma and colon adenocarcinoma cells in an assay with a single intravenous administration. It also prevents platelet arrest shortly after cell injection in wild-type mice and suppresses melanoma-platelets interaction in vitro. Moreover, LA-hep blocks P-selectin's direct binding to tumor cells and platelet aggregation, providing further evidence for the role of P-selectin as a molecular target. Even in P-selectin-depleted mice which developed a reduced number of metastatic foci, both porcine heparin and LA-hep further inhibited metastasis burden. This suggests evidence of an additional mechanism of antimetastatic action. Therefore, our results indicate a dissociation between the heparin anticoagulant and antimetastatic effects. Considering the simple and highly reproducible methodology used to purify LA-hep along with the data presented here, LA-hep emerges as a promising drug for future use in preventing metastasis in cancer patients.
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●Introduction of a low anticoagulant heparin (LA-hep) obtained from bovine heparin and devoid of bleeding effect.●A detailed characterization of LA-hep, including structure, average molecular weight, and its impact on in vivo hemostasis, is shown.●LA-hep demonstrated a remarkable capacity to inhibit metastatic dissemination in vivo.●LA-hep's antimetastatic activity is primarily based on the inhibition of tumor cell-platelet contact mediated by P-selectin.●LA-hep, safer than standard heparin, emerges as a promising candidate for the development of antitumoral therapies.
The aim of this study was to evaluate whether dysregulation of molecules involved in FOXO1-dependent insulin signaling in the liver is associated with de novo lipogenesis (DNL) and altered lipid ...metabolism in severely obese subjects.
Observational retrospective study.
We considered 71 obese subjects (age 20-68 years; body mass index (BMI)>40 kg m(-2) or BMI>35 kg m(-2) in the presence of metabolic complications) classified into three groups according to liver histology: normal liver (n=12), simple steatosis (n=27) and nonalcoholic steatohepatitis (NASH; n=32). Key nodes in insulin signaling and gene expression of molecules implicated in insulin-dependent glucoregulatory pathway and DNL were evaluated by quantitative real-time PCR and western blotting.
Patients with steatosis had decreased phosphorylation of the insulin kinase AKT1, mediating insulin receptor signaling, and the transcription factor FOXO1, which was therefore more active mediating insulin resistance at transcriptional level. Despite no changes in insulin receptor substrate (IRS)1 mRNA levels, the mRNA and protein levels of the FOXO1 target IRS2 increased progressively with the severity of steatosis from normal liver to NASH. IRS2 expression was correlated with the severity of steatosis, dyslipidemia and liver damage. In patients with NASH, upregulation of IRS2 was associated with preserved activation of AKT2, mediating the stimulating effect of insulin on DNL, and overexpression of its target sterol regulatory element-binding protein 1c (SREBP1c), inducing DNL at transcriptional level. Both FOXO1 and SREBP1c overexpression converged on upregulation of glucokinase, providing substrates for DNL, in NASH patients.
Differential regulation of IRS1 and IRS2 and of their downstream effectors AKT1 and AKT2 is consistent with upregulation of FOXO1 and may justify the paradoxical state of insulin resistance relative to the glucoregulatory pathway and augmented insulin sensitivity of the liporegulatory pathway typical of steatosis and the metabolic syndrome in obese patients.
Androgens have important cardiometabolic actions in males, but their metabolic role in females is unclear. To determine the physiologic androgen receptor (AR)‐dependent actions of androgens on ...atherogenesis in female mice, we generated female AR‐knockout (ARKO) mice on an atherosclerosis‐prone apolipoprotein E (apoE)‐deficient background. After 8 weeks on a high‐fat diet, but not on a normal chow diet, atherosclerosis in aorta was increased in ARKO females (+59% vs. control apoE‐deficient mice with intact AR gene). They also displayed increased body weight (+18%), body fat percentage (+62%), and hepatic triglyceride levels, reduced insulin sensitivity, and a marked atherogenic dyslipidemia (serum cholesterol, +52%). Differences in atherosclerosis, body weight, and lipid levels between ARKO and control mice were abolished in mice that were ovariectomized before puberty, consistent with a protective action of ovarian androgens mediated via the AR. Furthermore, the AR agonist dihydrotestosterone reduced atherosclerosis (‐41%; thoracic aorta), subcutaneous fat mass (‐44%), and cholesterol levels (‐35%) in ovariectomized mice, reduced hepatocyte lipid accumulation in hepatoma cells in vitro, and regulated mRNA expression of hepatic genes pivotal for lipid homeostasis. In conclusion, we demonstrate that the AR protects against diet‐induced atherosclerosis in female mice and propose that this is mediated by modulation of body composition and lipid metabolism.—Fagman, J. B., Wilhelmson, A. S., Motta, B. M., Pirazzi, C., Alexanderson, C., De Gendt, K., Verhoeven, G., Holmäng, A., Anesten, F., Jansson, J.‐O., Levin, M., Borén, J., Ohlsson, C., Krettek, A., Romeo, S., Tivesten, A. The androgen receptor confers protection against diet‐induced atherosclerosis, obesity, and dyslipidemia in female mice. FASEB J. 29, 1540‐1550 (2015). www.fasebj.org
•Definition of a numerical transient model of a historical building (case study).•Evaluation of the thermal energy needs and adaptive comfort analysis of the case study.•Overall retrofit of the ...historical building considered.•Energy, economic and environmental analysis of absorption and vapor compression heat pump systems, proposed for the retrofit of the case study.
In this work a numerical investigation of the energy needs of the Italian historical building Castle of Zena (XXIII century) and a feasibility study for the retrofit of its HVAC plant is presented.
About the building envelope behaviour two types of numerical analyses have been performed. First of all, free floating conditions (no temperature control) have been considered, in order to evaluate the building envelope performance through the adaptive comfort approach. During almost the 50% of the summer season the internal temperature is above the upper limit of the comfort range, therefore a cooling plant is needed. Secondly, an ideal temperature control has been considered, in order to calculate the annual energy needs for space heating (around 164kWh/m2/year) and cooling (around 5kWh/m2/year).
Regarding the HVAC retrofit, due to the historical constraints, it has been decided to use a fan coil emission system linked to a heat pump appliance. Four combination have been analysed, combining two type of heat pump (compression and absorption) with two type of heat source (air and water). Neglecting the control, emission, storage and distribution sub-system energy needs, there are two systems with the lowest primary energy consumption: the EHP-WS and the GAHP-WS (around 130kWh/m2/year), followed by the GAHP-AS (around 147kWh/m2/year) and the EHP-AS (around 180kWh/m2/year). From the economic point of view the EHP-WS has the shortest pay-back time, 7 years, also thanks to the use of the existing well. However, excluding the water source, only the GAHP is economically feasible. Lastly, a significant reduction of greenhouse gas emission (CO2) could be obtained replacing the EHP-AS with the GAHP-AS (−26%), the EHP-WS (−28%) or the GAHP-WS (−34%).
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