Effect of the standardized Cimicifuga foetida extract on Hsp 27 expression in the MCF-7 cell line

Cimicifuga foetida, an Asian Cimicifuga species, has been employed as a cooling and detoxification agent in traditional Chinese medicine since ancient times. For this herb, two cycloartane triterpene glycosides isolated from the rhizomes have demonstrated cytotoxicity on rat tumor and human cancer cell lines. Since human Hsp27 is increased in various human cancers and exhibits cytoprotective activity that affects tumorigenesis and the susceptibility of tumours to cancer treatment, the purpose of this research was to study the expression of this protein in MCF-7 breast cancer cells. To accomplish this aim, MCF-7 cells were exposed to different concentrations of Cimicifuga foetida extract showing a reduction in cell number measured by the sulforhodamine assay. In addition, the expression of Hsp-27 mRNA detected by RT-PCR and Hsp-27 protein detected by immnofluorescence was present in all conditions, except when using the highest concentration of Cimicifuga foetida extract (2,000 jig /L). We conclude that Hsp 27 expression at 2,000 jig /L Cimicifuga foetida extract is diminished. This is the first report showing the Hsp-27 expression after exposure to Cimicifuga foetida extract in MCF-7 cells.     

A Scoping Review and Prevalence Analysis of Soil-Transmitted Helminth Infections in Honduras

Background

Honduras is endemic for soil-transmitted helminth (STH) infections, but critical information gaps still remain on the prevalence and intensity of these infections as well as on their spatial distribution at subnational levels.

Objectives

Firstly, to review the research activity on STH infections in Honduras and secondly, to carry out a national prevalence analysis and map the geographical distribution of these infections in children.

Methods

A systematic search was conducted of the published and grey literature to identify scientific work on the impact and prevalence of STH infections done between May 1930 and June 30, 2012. International databases and Honduran journals were searched. Grey literature was gleaned from local libraries and key informants. Select studies conducted between 2001 and 2012 were used to produce prevalence maps and to investigate association between STH prevalence and socio-economic and environmental factors.

Results

Of 257 identified studies, 211 (21.4% peer-reviewed) were retained for analysis and categorized as clinical research (10.9%), treatment efficacy studies (8.1%) or epidemiological studies (81%). Prevalence analysis and geographical mapping included 36 epidemiological studies from Honduras''s 18 departments and 23% of its municipalities. Overall STH prevalence was >50% in 40.6% of municipalities. Prevalences above 20% for each trichuriasis, ascariasis, and hookworm infection were found in 68%, 47.8%, and 7.2% of studied municipalities, respectively. Municipalities with lower human development index, less access to of potable water, and with higher annual precipitation showed higher STH prevalences.

Conclusions

This is the first study to provide a comprehensive historic review of STH research activity and prevalence in Honduras, revealing important knowledge gaps related to infection risk factors, disease burden, and anti-parasitic drug efficacy, among others. Our decade-long prevalence analysis reveals geographical differences in STH prevalence and these findings suggest that differential intervention strategies might be necessary in Honduras for the control of these infections.     

Incidence of Human Taenia solium Larval Infections in an Ecuadorian Endemic Area: Implications for Disease Burden Assessment and Control

Background

Human cysticercosis is a zoonotic disease causing severe health disorders and even death. While prevalence data become available worldwide, incidence rate and cumulative incidence figures are lacking, which limits the understanding of the Taenia solium epidemiology.

Methodology/Principal findings

A seroepidemiological cohort study was conducted in a south-Ecuadorian community to estimate the incidence rate of infection with and the incidence rate of exposure to T. solium based on antigen and antibody detections, respectively. The incidence rate of infection was 333.6 per 100,000 person-years (95% CI: [8.4–1,858] per 100,000 person-years) contrasting with a higher incidence rate of exposure 13,370 per 100,000 person-years (95% CI: [8,730–19,591] per 100,000 person-years). The proportion of infected individuals remained low and stable during the whole study year while more than 25% of the population showed at least one antibody seroconversion/seroreversion during the same time period.

Conclusions/Significance

Understanding the transmission of T. solium is essential to develop ad hoc cost-effective prevention and control programs. The estimates generated here may now be incorporated in epidemiological models to simulate the temporal transmission of the parasite and the effects of control interventions on its life cycle. These estimates are also of high importance to assess the disease burden since incidence data are needed to make regional and global projections of morbidity and mortality related to cysticercosis.     

Biochemical and enzymatic characterization of a partially purified casein kinase-1 like activity from Trypanosoma cruzi

Two protein kinase activities that use casein as a substrate, Q-I and Q-II, were identified in the epimastigote stage of Trypanosoma cruzi upon chromatography on Q-Sepharose. Q-I was purified further through concanavalin A-sepharose (Q-I*) to remove any trace of the contaminating protease cruzipain. The optimal activity for Q-I* was obtained at pH 8.0, 25 degreesC, 5 mM MgCl(2) and 75 mM NaCl. The size and pI of Q-I* were determined to be 33-36 kDa and 9.6, respectively. When two selective peptide substrates for casein kinases (CKs) (P1: RRKDLHDDEEDEAMSITA for CK1 and P2: RRRADDSDDDDD for CK2) were used, Q-I* was shown to specifically phosphorylate P1. Kinetic studies showed that Q-I* has a K(m) of 5.3 +/- 0.34 mg/ml for casein, 157.6 +/- 5.3 microM for P1 and 35.9 +/- 3.9 microM for ATP. The enzyme was inhibited by N-(2-amino-ethyl)-5-chloroisoquinoline-8-sulfonamide (CKI-7) or 1-(5-chloroisoquinoline-8-sulfonyl) (CKI-8), two inactivators of mammalian CKs. CKI-7 behaved as a competitive inhibitor with respect to ATP, with a K(I) of 75-100 microM. Treatment with high concentrations of polylysine or heparin also resulted in a significant inhibition of Q-I*. Two well-known activators of mammalian CKs, spermine and spermidine, were also tested. Spermine and spermidine activated Q-I* in a dose-dependent manner. Based on the following characteristics: (1) the ionic strength required for elution from anion-exchange resins; (2) its molecular size and monomeric structure; (3) pI; (4) high level of specificity for P1; (5) inactivation by CKI-7 and CKI-8; and (6) insensitivity to GTP and low concentrations of heparin, we conclude that Q-I* belongs to the CK1 family of protein kinases.     

Naegleria fowleri: enolase is expressed during cyst differentiation

Cysts of Naegleria fowleri present an external single-layered cyst wall. To date, little information exists on the biochemical components of this cyst wall. Knowledge of the cyst wall composition is important to understand its resistance capacity under adverse environmental conditions. We have used of a monoclonal antibody (B4F2 mAb) that specifically recognizes enolase in the cyst wall of Entamoeba invadens. By Western blot assays this antibody recognized in soluble extracts of N. fowleri cysts a 48-kDa protein with similar molecular weight to the enolase reported in E. invadens cysts. Immunofluorescence with the B4F2 mAb revealed positive cytoplasmic vesicles in encysting amebas, as well as a positive reaction at the cell wall of mature cysts. Immunoelectron microscopy using the same monoclonal antibody confirmed the presence of enolase in the cell wall of N. fowleri cysts and in cytoplasmic vesicular structures. In addition, the B4F2 mAb had a clear inhibitory effect on encystation of N. fowleri.     

Depletion of highly abundant proteins in blood plasma by ammonium sulfate precipitation for 2D-PAGE analysis

Ammonium sulfate precipitation (ASP) was explored as a method for depleting some highly abundant proteins from blood plasma, in order to reduce the dynamic range of protein concentration and to improve the detection of low abundance proteins by 2D-PAGE. 40% ammonium sulfate saturation was chosen since it allowed depleting 39% albumin and 82% α-1-antitrypsin. ASP-depletion showed high reproducibility in 2D-PAGE analysis (4.2% variation in relative abundance of albumin), similar to that offered by commercial affinity-depletion columns. Besides, it allowed detecting 59 spots per gel, very close to the number of spots detected in immuno-affinity-depleted plasma. Thus, ASP at 40% saturation is a reliable depletion method that may help in proteomic analysis of blood plasma. Finally, ASP-depletion seems to be complementary to hydrophobic interaction chromatography (HIC)-depletion, and therefore an ASP-step followed by a HIC-step could probably deplete the most highly abundant plasma proteins, thus improving the detection of low abundance proteins by 2D-PAGE.     

Leishmania repression of host translation through mTOR cleavage is required for parasite survival and infection

The protozoan parasite Leishmania alters the activity of its host cell, the macrophage. However, little is known about the effect of Leishmania infection on host protein synthesis. Here, we show that the Leishmania protease GP63 cleaves the mammalian/mechanistic target of rapamycin (mTOR), a serine/threonine kinase that regulates the translational repressor 4E-BP1. mTOR cleavage results in the inhibition of mTOR complex 1 (mTORC1) and concomitant activation of 4E-BP1 to promote Leishmania proliferation. Consistent with these results, pharmacological activation of 4E-BPs with rapamycin, results in a dramatic increase in parasite replication. In contrast, genetic deletion of 4E-BP1/2 reduces parasite load in macrophages ex vivo and decreases susceptibility to cutaneous leishmaniasis in vivo. The parasite resistant phenotype of 4E-BP1/2 double-knockout mice involves an enhanced type I IFN response. This study demonstrates that Leishmania evolved a survival mechanism by activating 4E-BPs, which serve as major targets for host translational control.     

Retinal degeneration progression changes lentiviral vector cell targeting in the retina

In normal mice, the lentiviral vector (LV) is very efficient to target the RPE cells, but transduces retinal neurons well only during development. In the present study, the tropism of LV has been investigated in the degenerating retina of mice, knowing that the retina structure changes during degeneration. We postulated that the viral transduction would be increased by the alteration of the outer limiting membrane (OLM). Two different LV pseudotypes were tested using the VSVG and the Mokola envelopes, as well as two animal models of retinal degeneration: light-damaged Balb-C and Rhodopsin knockout (Rho-/-) mice. After light damage, the OLM is altered and no significant increase of the number of transduced photoreceptors can be obtained with a LV-VSVG-Rhop-GFP vector. In the Rho-/- mice, an alteration of the OLM was also observed, but the possibility of transducing photoreceptors was decreased, probably by ongoing gliosis. The use of a ubiquitous promoter allows better photoreceptor transduction, suggesting that photoreceptor-specific promoter activity changes during late stages of photoreceptor degeneration. However, the number of targeted photoreceptors remains low. In contrast, LV pseudotyped with the Mokola envelope allows a wide dispersion of the vector into the retina (corresponding to the injection bleb) with preferential targeting of Müller cells, a situation which does not occur in the wild-type retina. Mokola-pseudotyped lentiviral vectors may serve to engineer these glial cells to deliver secreted therapeutic factors to a diseased area of the retina.