Protective role of purified cysteine proteinases against Fasciola gigantica infection in experimental animals

Fascioliasis is one of the public health problems in the world. Cysteine proteinases (CP) released by Fasciola gigantica play a key role in parasite feeding, migration through host tissues, and in immune evasion. There has been some evidence from several parasite systems that proteinases might have potential as protective antigens against parasitic infections. Cysteine proteinases were purified and tested in vaccine trials of sheep infected with the liver fluke. Multiple doses (2 mg of CP in Freund's adjuvant followed by 3 booster doses 1 mg each at 4 week intervals) were injected intramuscularly into sheep 1 week prior to infect orally with 300 F. gigantica metacercariae. All the sheep were humanely slaughtered 12 weeks after the first immunization. Changes in the worm burden, ova count, and humoral and cellular responses were evaluated. Significant reduction was observed in the worm burden (56.9%), bile egg count (70.7%), and fecel egg count (75.2%). Immunization with CP was also found to be associated with increases of total IgG, IgG(1), and IgG(2) (P<0.05). Data showed that the serum cytokine levels of pro-inflammatory cytokines, IL-12, IFN-γ, and TNF-α, revealed significant decreases (P<0.05). However, the anti-inflammatory cytokine levels, IL-10, TGF-β, and IL-6, showed significant increases (P<0.05). In conclusion, it has been found that CP released by F. gigantica are highly important candidates for a vaccine antigen because of their role in the fluke biology and host-parasite relationships.     

Relation between augmentation index and adiponectin during one-year metformin treatment for nonalcoholic steatohepatosis: effects beyond glucose lowering?

ABSTRACT: BACKGROUND: Insulin resistance (IR) is the major driving force behind development and progression of atherosclerosis in patients with nonalcoholic fatty liver disease (NAFLD). Therefore, correction of IR is a relevant therapeutic target. We performed the current trial to evaluate whether 12- month metformin therapy improves vascular stiffness in patients with NAFLD and to assess if this improvement is associated with change in glucose control, insulin resistance or circulating adiponectin. METHODS: In randomized, placebo controlled study, 63 patients with NAFLD were assigned to one of two groups: Group 1 received daily metformin; Group 2 received placebo. Central aortic augmentation index (AI) was performed using SphygmoCor (version 7.1, AtCor Medical, Sydney, Australia) at baseline, at 4-and 12-month treatment period. Metabolic parameters, insulin resistance markers and serum adiponectin levels were determined. RESULTS: In placebo group: AI did not improve during the treatment period. Liver function and adiponectin levels did not change during the study. In multiple linear regression analysis, the independent predictors of arterial stiffness improvement were metformin treatment and increase in circulating adiponectin levels. Among metformin treated patients: AI decreased significantly during the study. ALP and ALT decreased during initial 4-month treatment period, however raised to the pretreatment levels after 12 months. Serum adiponectin level tended to increase during treatment period with metformin. CONCLUSIONS: Metformin treatment was associated with significant decrease in AI during one year treatment in NAFLD patients. These beneficial vascular effects was associated with exposure to metformin per se as well as change in adiponectin levels suggesting that metformin may mediate its vascular effects via glicemic control-independent mechanisms.     

Functional characterization of circulating tumor cells with a prostate-cancer-specific microfluidic device

Cancer metastasis accounts for the majority of cancer-related deaths owing to poor response to anticancer therapies. Molecular understanding of metastasis-associated drug resistance remains elusive due to the scarcity of available tumor tissue. Isolation of circulating tumor cells (CTCs) from the peripheral blood of patients has emerged as a valid alternative source of tumor tissue that can be subjected to molecular characterization. However, issues with low purity and sensitivity have impeded adoption to clinical practice. Here we report a novel method to capture and molecularly characterize CTCs isolated from castrate-resistant prostate cancer patients (CRPC) receiving taxane chemotherapy. We have developed a geometrically enhanced differential immunocapture (GEDI) microfluidic device that combines an anti-prostate specific membrane antigen (PSMA) antibody with a 3D geometry that captures CTCs while minimizing nonspecific leukocyte adhesion. Enumeration of GEDI-captured CTCs (defined as intact, nucleated PSMA+/CD45− cells) revealed a median of 54 cells per ml identified in CRPC patients versus 3 in healthy donors. Direct comparison with the commercially available CellSearch® revealed a 2–400 fold higher sensitivity achieved with the GEDI device. Confocal microscopy of patient-derived GEDI-captured CTCs identified the TMPRSS2:ERG fusion protein, while sequencing identified specific androgen receptor point mutation (T868A) in blood samples spiked with only 50 PC C4-2 cells. On-chip treatment of patient-derived CTCs with docetaxel and paclitaxel allowed monitoring of drug-target engagement by means of microtubule bundling. CTCs isolated from docetaxel-resistant CRPC patients did not show any evidence of drug activity. These measurements constitute the first functional assays of drug-target engagement in living circulating tumor cells and therefore have the potential to enable longitudinal monitoring of target response and inform the development of new anticancer agents.     

The expression of interleukin-8 and interleukin-8 receptors in endometrial carcinoma

Interleukin-8 (IL-8) is a pro-inflammatory cytokine which exerts its effects via binding to 2 receptors, CXCR1 and CXCR2 and is known to promote angiogenesis, mitogenesis and motogenesis in cancer. IL-8 is over expressed in endometrial carcinoma, but the expression of CXCR1 and CXCR2 in endometrial carcinoma has not been previously investigated. The aim of this study was to determine the expression of IL-8 receptors in endometrial carcinoma. The expression of CXCR1 and CXCR2 was studied in endometrial carcinomas and normal endometrium by immunohistochemistry in 101 tumours. IL-8 and IL-8 receptor expression was also studied by Real-time quantitative PCR (RT-qPCR) in 17 tumours in comparison to normal endometrium. The expression profile was correlated to the clinico-pathological features of the tumours. Immunohistochemistry showed CXCR1 and CXCR2 were expressed in all cases of endometrial carcinoma, with CXCR1 showing stronger expression. There was a statistically significant correlation between CXCR2 staining intensity and tumour grade (P=0.012) and disease free survival (P=0.015) independently. On RT-qPCR, 14/17, 15/17 and 16/17 tumours showed significant increase in IL-8, CXCR1 and CXCR2 expression levels in comparison to normal endometrium, with median fold increase of 42-fold, 51-fold and 27-fold, respectively. This is the first report of the expression of IL-8 receptors in endometrial carcinoma and the results show an association between IL-8 and IL-8 receptors and the pathogenesis of endometrial carcinoma, and represent potential prognostic biomarkers and therapeutic targets.     

The Revised Classification of Eukaryotes

This revision of the classification of eukaryotes, which updates that of Adl et al. [J. Eukaryot. Microbiol. 52 (2005) 399], retains an emphasis on the protists and incorporates changes since 2005 that have resolved nodes and branches in phylogenetic trees. Whereas the previous revision was successful in re‐introducing name stability to the classification, this revision provides a classification for lineages that were then still unresolved. The supergroups have withstood phylogenetic hypothesis testing with some modifications, but despite some progress, problematic nodes at the base of the eukaryotic tree still remain to be statistically resolved. Looking forward, subsequent transformations to our understanding of the diversity of life will be from the discovery of novel lineages in previously under‐sampled areas and from environmental genomic information.     

On impulse response functions computed from dynamic contrast-enhanced image data by algebraic deconvolution and compartmental modeling

Concentration-time courses measured by dynamic contrast-enhanced (DCE) imaging can be described by a convolution of the arterial input with an impulse response function, Q_T(t), characterizing tissue microcirculation. Data analysis is based on two different approaches: computation of Q_T(t) by algebraic deconvolution (AD) and subsequent evaluation according to the indicator dilution theory (IDT) or parameterization of Q_T(t) by analytical expressions derived by compartmental modeling. Pitfalls of both strategies will be addressed in this study.Tissue data acquired by DCE-CT in patients with head-and-neck cancer and simulated by a reference model (MMID4) were analyzed by a two-compartment model (TCM), a permeability-limited two-compartment model (PL-TCM) and AD. Additionally, MMID4 was used to compute the ‘true’ response function that corresponds to the simulated tumor data.TCM and AD yielded accurate fits, whereas PL-TCM performed worse. Nevertheless, the corresponding response functions diverge markedly. The response curves obtained by TCM decrease exponentially in the early perfusion phase and overestimate the tissue perfusion, Q_T(0). AD also resulted in response curves starting with a negative slope and not – as the ‘true’ response function in accordance with the IDT – with a horizontal plateau. They are thus not valid responses in the sense of the IDT that can be used unconditionally for parameter estimation.Response functions differing considerably in shape can result in virtually identical tissue curves. This non-uniqueness makes a strong argument not to use algebraic but rather analytical deconvolution to reduce the class of solutions to representatives that are in accordance with a-priori knowledge. To avoid misinterpretations and systematic errors, users must be aware of the pitfalls inherent to the different concepts.     

The Polymorphic Pseudokinase ROP5 Controls Virulence in Toxoplasma gondii by Regulating the Active Kinase ROP18

Secretory polymorphic serine/threonine kinases control pathogenesis of Toxoplasma gondii in the mouse. Genetic studies show that the pseudokinase ROP5 is essential for acute virulence, but do not reveal its mechanism of action. Here we demonstrate that ROP5 controls virulence by blocking IFN-γ mediated clearance in activated macrophages. ROP5 was required for the catalytic activity of the active S/T kinase ROP18, which phosphorylates host immunity related GTPases (IRGs) and protects the parasite from clearance. ROP5 directly regulated activity of ROP18 in vitro, and both proteins were necessary to avoid IRG recruitment and clearance in macrophages. Clearance of both the Δrop5 and Δrop18 mutants was reversed in macrophages lacking Irgm3, which is required for IRG function, and the virulence defect was fully restored in Irgm3^−/− mice. Our findings establish that the pseudokinase ROP5 controls the activity of ROP18, thereby blocking IRG mediated clearance in macrophages. Additionally, ROP5 has other functions that are also Irgm3 and IFN-γ dependent, indicting it plays a general role in governing virulence factors that block immunity.