Different Clinical Outcomes of Entamoeba histolytica in Malaysia: Does Genetic Diversity Exist?

The present study was conducted to investigate the clinical outcomes of Entamoeba histolytica infection in symptomatic and asymptomatic Orang Asli (aborigine) communities in Malaysia. Examination was performed on 500 stool samples obtained from Orang Asli communities in 3 different states using formalin-ether concentration, trichrome staining, and single-round PCR techniques. Out of 500 stool samples, single infection of E. histolytica, Entamoeba dispar, and Entamoeba moshkovskii was identified in 3.2%, 13.4%, and 1%, respectively. In addition, 10 samples had mixed infections with E. histolytica and E. dispar. Six samples containing E. dispar were also positive for E. moshkovskii, and only 2 samples had E. histolytica in association with E. dispar and E. moshkovskii. Seventeen E. histolytica-positive samples were from symptomatic subjects, whereas the remaining 11 samples came from asymptomatic subjects. These findings suggest a predominant distribution of pathogenic potential of E. histolytica strains in this community. Therefore, further studies on genotyping of E. histolytica is required, to find out association between E. histolytica genotype and the outcome of the infection.     

Morphological and molecular identification of some closely related Pythium species in Egypt

Twelve isolates of Pythium species (P. aphanidermatum, P. deliense, P. ultimum var. ultimum and P. ultimum var. sporangiiferum) from different hosts were compared from morphological, pathological and molecular viewpoints. Minimum, optimum and maximum temperatures of P. aphanidermatum and P. deliense were similar while those of P. ultimum var. ultimum and P. ultimum var. sporangiiferum were also similar. All tested isolates were highly virulent against cucumber seedlings with 100% damping-off. RAPD data using three different primers revealed that strains of P. ultimum var. ultimum and P. ultimum var. sporangiiferum are distinct from each other. This data can be used to separate those species from P. aphanidermatum and P. deliense. In contrast, RAPD data cannot be used to separate P. aphanidermatum and P. deliense. Sequence analysis of the ribosomal DNA internal transcribed spacers (ITS) was used to establish phylogenetic relationships among the tested isolates.     

The A subunit of type IIb enterotoxin (LT-IIb) suppresses the proinflammatory potential of the B subunit and its ability to recruit and interact with TLR2

The type IIb heat-labile enterotoxin of Escherichia coli (LT-IIb) and its nontoxic pentameric B subunit (LT-IIb-B(5)) display different immunomodulatory activities, the mechanisms of which are poorly understood. We investigated mechanisms whereby the absence of the catalytically active A subunit from LT-IIb-B(5) renders this molecule immunostimulatory through TLR2. LT-IIb-B(5), but not LT-IIb, induced TLR2-mediated NF-kappaB activation and TNF-alpha production. These LT-IIb-B(5) activities were antagonized by LT-IIb; however, inhibitors of adenylate cyclase or protein kinase A reversed this antagonism. The LT-IIb antagonistic effect is thus likely dependent upon the catalytic activity of its A subunit, which causes elevation of intracellular cAMP and activates cAMP-dependent protein kinase A. Consistent with this, a membrane-permeable cAMP analog and a cAMP-elevating agonist, but not catalytically defective point mutants of LT-IIb, mimicked the antagonistic action of wild-type LT-IIb. The mutants moreover displayed increased proinflammatory activity compared with wild-type LT-IIb. Additional mechanisms for the divergent effects on TLR2 activation by LT-IIb and LT-IIb-B(5) were suggested by findings that the latter was significantly stronger in inducing lipid raft recruitment of TLR2 and interacting with this receptor. The selective use of TLR2 by LT-IIb-B(5) was confirmed in an assay for IL-10, which is inducible by both LT-IIb and LT-IIb-B(5) at comparable levels; TLR2-deficient macrophages failed to induce IL-10 in response to LT-IIb-B(5) but not in response to LT-IIb. These differential immunomodulatory effects by LT-IIb and LT-IIb-B(5) have important implications for adjuvant development and, furthermore, suggest that enterotoxic E. coli may suppress TLR-mediated innate immunity through the action of the enterotoxin A subunit.     

Lysophosphatidic acid prevents apoptosis of Caco-2 colon cancer cells via activation of mitogen-activated protein kinase and phosphorylation of Bad

Lysophosphatidic acids (LPA) exert growth factor-like effects through specific G protein-coupled receptors. The presence of different LPA receptors often determines the specific signaling mechanisms and the physiological consequences of LPA in different environments. Among the four members of the LPA receptor family, LPA(2) has been shown to be overexpressed in colon cancer suggesting that the signaling by LPA(2) may potentiate growth and survival of tumor cells. In this study, we examined the effect of LPA on survival of colon cancer cells using Caco-2 cells as a cell model system. LPA rescued Caco-2 cells from apoptosis elicited by the chemotherapeutic drug, etoposide. This protection was accompanied by abrogation of etoposide-induced stimulation of caspase activity via a mechanism dependent on Erk and PI3K. In contrast, perturbation of cellular signaling mediated by the LPA(2) receptor by knockdown of a scaffold protein NHERF2 abrogated the protective effect of LPA. Etoposide decreased the expression of Bcl-2, which was reversed by LPA. Etoposide decreased the phosphorylation level of the proapoptotic protein Bad in an Erk-dependent manner, without changing Bad expression. We further show that LPA treatment resulted in delayed activation of Erk. These results indicate that LPA protects Caco-2 cells from apoptotic insult by a mechanism involving Erk, Bad, and Bcl-2.     

MicroRNA-21 as a predictor and prognostic factor for trastuzumab therapy in human epidermal growth factor receptor 2-positive metastatic breast cancer

Breast cancer is the second most common cancer diagnosed worldwide. Human epidermal growth factor receptor 2 (HER2)-positive breast cancer represents about 20% to 30% of all breast cancers. Trastuzumab is used in the treatment of HER2-positive breast cancer. MicroRNA-21 (miR-21) is an oncomiR that acts by inhibiting many tumor-suppressor genes. We analyzed the relative expression levels of serum miR-21 in 20 HER2-positive metastatic breast cancer patients before and after 3 months of treatment with trastuzumab. miR-21 levels decreased with a high significant difference after trastuzumab therapy (P = 0.001). Although miR-21 expression levels were lower in responders than in nonresponders, the difference was not statistically significant ( P = 0.6). Our results demonstrated a significant negative correlation between its basal expression, expression levels after treatment, and time to progression ( P = 0.03 and 0.01, respectively). These results make miR-21 a potential prognostic factor for HER2-positive metastatic breast cancer patients. Additionally, it can be an interesting potential target in therapy using antisense oligonucleotides for miR-21.     

Effect of Cystoseira barbata and Dictyota dichotoma-algae on reproduction and protein pattern of Biomphalaria alexandrina snails

Controlling the snail intermediate host of schistosomiasis is one of the strategies used to reduce disease incidence. In this study screening tests were carried out on the action of dried powder of eight algal species against Biomphalaria alexandrina snails. The most effective powders were from Dictyota dichotoma and Cystoseira barbata with LC₅₀ values of 560 and 2200?ppm, respectively. Prolonged exposure to sublethal concentrations of these two powders caused adverse effects on snails' reproduction especially at LC₂₅. Hatchability of exposed eggs was lowest at LC₂₅ of C. barbata, and LC₁₅ of D. dichotoma. The most pronounced effects on the hermaphrodite gland of snails were caused by high concentrations of both algal suspensions, where ova were either degenerated or deformed and there were detrimental effects on all spermatogenesis stages. The electrophoretic separation of haemolymph proteins showed that the concentration with most effect was the LC₁₅ for both tested algae as they resulted in more bands than the control and also an elevation in total protein content.     

Excess Linoleic Acid Increases Collagen I/III Ratio and “Stiffens” the Heart Muscle Following High Fat Diets

Controversy exists on the benefits versus harms of n-6 polyunsaturated fatty acids (n-6 PUFA). Although n-6 PUFA demonstrates anti-atherosclerotic properties, survival following cardiac remodeling may be compromised. We hypothesized that n-6 PUFA like linoleic acid (LA) or other downstream PUFAs like γ-linolenic acid or arachidonic acid alter the transforming growth factor-β (TGFβ)-collagen axis in the heart. Excess dietary LA increased the collagen I/III ratio in the mouse myocardium, leading to cardiac “stiffening” characterized by impaired transmitral flow indicative of early diastolic dysfunction within 5 weeks. In vitro, LA under TGFβ1 stimulation increased collagen I and lysyl oxidase (LOX), the enzyme that cross-links soluble collagen resulting in deposited collagen. Overexpression of fatty acid desaturase 2 (fads2), which metabolizes LA to downstream PUFAs, reduced collagen deposits, LOX maturation, and activity with LA, whereas overexpressing fads1, unrelated to LA desaturation, did not. Furthermore, fads2 knockdown by RNAi elevated LOX activity and collagen deposits in fibroblasts with LA but not oleic acid, implying a buildup of LA for aggravating such pro-fibrotic effects. As direct incubation with γ-linolenic acid or arachidonic acid also attenuated collagen deposits and LOX activity, we concluded that LA itself, independent of other downstream PUFAs, promotes the pro-fibrotic effects of n-6 PUFA. Overall, these results attempt to reconcile opposing views of n-6 PUFA on the cardiovascular system and present evidence supporting a cardiac muscle-specific effect of n-6 PUFAs. Therefore, aggravation of the collagen I/III ratio and cardiac stiffening by excess n-6 PUFA represent a novel pathway of cardiac lipotoxicity caused by high n-6 PUFA diets.     

The novel steroidal alkaloids dendrogenin A and B promote proliferation of adult neural stem cells

Dendrogenin A (DDA) and dendrogenin B (DDB) are new aminoalkyl oxysterols which display re-differentiation of tumor cells of neuronal origin at nanomolar concentrations. We analyzed the influence of dendrogenins on adult mice neural stem cell proliferation, sphere formation and differentiation. DDA and DDB were found to have potent proliferative effects in neural stem cells. Additionally, they induce neuronal outgrowth from neurospheres during in vitro cultivation. Taken together, our results demonstrate a novel role for dendrogenins A and B in neural stem cell proliferation and differentiation which further increases their likely importance to compensate for neuronal cell loss in the brain.