Protein variation in blood-dwelling schistosome worms generated by differential splicing of micro-exon gene transcripts

Schistosoma mansoni is a well-adapted blood-dwelling parasitic helminth, persisting for decades in its human host despite being continually exposed to potential immune attack. Here, we describe in detail micro-exon genes (MEG) in S. mansoni, some present in multiple copies, which represent a novel molecular system for creating protein variation through the alternate splicing of short (<= 36 bp) symmetric exons organized in tandem. Analysis of three closely related copies of one MEG family allowed us to trace several evolutionary events and propose a mechanism for micro-exon generation and diversification. Microarray experiments show that the majority of MEGs are up-regulated in life cycle stages associated with establishment in the mammalian host after skin penetration. Sequencing of RT-PCR products allowed the description of several alternate splice forms of micro-exon genes, highlighting the potential use of these transcripts to generate a complex pool of protein variants. We obtained direct evidence for the existence of such pools by proteomic analysis of secretions from migrating schistosomula and mature eggs. Whole-mount in situ hybridization and immunolocalization showed that MEG transcripts and proteins were restricted to glands or epithelia exposed to the external environment. The ability of schistosomes to produce a complex pool of variant proteins aligns them with the other major groups of blood parasites, but using a completely different mechanism. We believe that our data open a new chapter in the study of immune evasion by schistosomes, and their ability to generate variant proteins could represent a significant obstacle to vaccine development.

In vivo uptake of a haem analogue Zn protoporphyrin IX by the human malaria parasite P. falciparum-infected red blood cells

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Cytochemical characterization of eosinophilic leukocytes circulating in the blood of the turtle (Chrysemys dorbignih)

Eosinophils and neutrophils are granulocytic leukocytes that are present in the blood of most vertebrates. Studies have been performed on lower vertebrates to understand the biological roles of the cells in defense mechanisms and to establish phylogenetic studies and new experimental models. Whether these 2 cell types exist in reptiles is a matter of controversy. In the blood of turtles there are 2 types of granulocytes that exhibit eosinophilia, one of them with round cytoplasmic granules and the other with elongated cytoplasmic granules. It has been suggested that these cells may be eosinophils in different stages of maturation but they also may be distinct cell types, i.e. eosinophils and neutrophils. In the present study, we characterized the 2 types of granulocytes that are present in the blood of Chrysemys dorbignih, using cytochemical techniques. Type I eosinophils showed activity of nonspecific esterase, peroxidase activity that is resistant to KCN, and basic proteins. Type II eosinophils exhibited activity of trimetaphosphatase, alkaline phosphatase, nonspecific esterase, peroxidase that is sensitive to KCN, and basic proteins. These observations indicate the existence of 2 distinct cell types in the blood of Chrysemys dorbignih, type I and type II eosinophils, that correspond to eosinophils and heterophils (neutrophils) of mammals and other vertebrates.

Description of gamontogonic and sporogonic stages of Hepatozoon spp. (Apicomplexa, Hepatozoidae) from Caudisoma durissa terrifica (Serpentes, Viperidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Ultrastructure features of the midgut of the female adult Amblyomma cajennense ticks Fabricius, 1787 (Acari: Ixodidae) in several feeding stages and subjected to three infestations

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Grading and staging chronic hepatitis C and its relation to genotypes and epidemiological factors in brazilian blood donors

Progression of chronic hepatitis C is known to be associated with some factors, but influence of HCV genotypes is still controversial. Association between HCV genotypes and other risk factors was examined to determine which factors are associated with progression of infection. One hundred consecutive anti-HCV positive volunteer blood donors were evaluated for several risk factors, examined for HCV genotypes, and submitted to hepatic biopsy and biochemical exams.HCV genotyping were carried out in 89 patients and hepatic biopsy in 78. Transmission routes were found to be illicit intravenous drug use (26%), Gluconergan® use in a non-safe manner (48%) and blood transfusion (15%). HCV genotype was 1 in 45%, 3 in 40%, and it was not associated with the stage of fibrosis or with inflammatory activity. There was no significant association of factors related to infection, chronic alcohol use, or duration of illness, with progression of the lesion. There was a significant association of aminotransferase levels and the fibrosis stage. Univariate analysis showed that the age at contamination, patient's age, GT-gamma, and aminotransferase levels over three times the upper normal limits, were associated with fibrosis stages 2 to 4. Multivariate analysis detected age (odds ratio=1.19), and GT-gamma (odds ratio=2.02) as independent factors.

DEVELOPMENT OF BLOOD-PRESSURE AND CARDIAC REFLEXES IN THE FROG PSEUDIS-PARADOXSUS

Systemic arterial blood pressure and heart rate (f(H)) were measured in unanesthetized, unrestrained larvae and adults of the paradoxical frog, Pseudis paradoxus from São Paulo State in Brazil. Four developmental groups were used, representing the complete transition from aquatic larvae to primarily air-breathing adults. f(H) (49-66 beats/min) was not significantly affected by development, whereas mean arterial blood pressure was strongly affected, being lowest in the stage 37-39 larvae (10 mmHg), intermediate in the stage 44-45 larvae (18 mmHg), and highest in the juveniles and adults (31 and 30 mmHg, respectively). Blood pressure was not significantly correlated with body mass, which was greatest in the youngest larvae and smallest in the juveniles. In the youngest larvae studied (stages 37-39), lung ventilation was infrequent, causing a slight decrease in arterial blood pressure but no change in heart rate. Lung ventilation was more frequent in stages 44-45 larvae and nearly continuous in juveniles and adults floating at the surface. Bradycardia during both forced and voluntary diving was observed in almost every advanced larva, juvenile, and adult but in only one of four young larvae. Developmentally related changes in blood pressure were not complete until metamorphosis, whereas diving bradycardia was present at an earlier stage.

Tissue stages of Hepatozoon canis in naturally infected dogs from São Paulo State, Brazil

A total of 222 dogs were examined by blood smear examination and Hepatozoon canis infection was detected in 13 dogs (5.9%). Five H. canis-infected dogs were necropsied to observe tissue stages in the organs. Fragments of spleen, liver, lungs, heart, kidneys, lymph nodes, bone marrow and skeletal muscles were used to made touch-impression smears. No macroscopic lesions were found in the organs. Two dogs had gamonts within polymorphonuclear cells and schizonts in various stages of development within the spleen and the bone marrow. Nevertheless, no mature meronts were found.

Morphology and histoenzymology of eosinophilic granulocytes in the circulating blood of the turtle (Chrysemys dorbignih)

The studies on the characterization of eosinophils and neutrophils/ heterophils of turtles are contradictory. Some authors have pointed out the existence of two distinct cell types: eosinophils and heterophils. Other authors have proposed that eosinophils and heterophils may be the same cells in different stages of maturation. These interpretations are based only on a morphological analysis. In the blood of the turtle (Chrysemys dorbignih), a South American freshwater species, there are two types of granulocytes with eosinophilic staining pattern: the first with round cytoplasmic granules and the second with ellipsoidal cytoplasmic granules. In the present study by using histoenzymological methods for the analyses of enzymological cellular content, we found that the cells with round cytoplasmic granules were positive for nonspecific esterase and the cells with ellipsoidal granules were positives for acid phosphatase, alkaline phosphatase, nonspecific esterase and peroxidase. The results show that these cells are distinct cells and that the cells with ellipsoidal cytoplasmic granules have the same histoenzymological characteristics as the neutrophils/heterophils of mammalians and other vertebrates.

Characterization of blood mononuclear phagocytes in Phrynops hilarii (Chelonia Chelidae)

The localization of peroxidase activity in different cell regions is used as a criterion for the classification of the stage of maturation of mammalian mononuclear phagocytes with a positive peroxidase reaction indicating the presence of monoblasts, promonocytes, monocytes and macrophages. In this study it was evaluated the peroxidase activity of blood mononuclear phagocytes of this turtle detected at different stages of differentiation. The present observations suggest that, in turtles, the differentiation of mononuclear phagocytes occur in the blood circulation, in contrast to animals, where only are monocytes in circulating blood and macrophage differentiation occurs in other body compartments. © 2007 Sociedad Chilena de Anatomía.

Morphology of the midgut of Rhipicephalus sanguineus (Latreille, 1806) (Acari: Ixodidae) adult ticks in different feeding stages

The intestinal epithelial cells of ticks are fundamental for their full feeding and reproductive success, besides being considered important sites for the development of pathogens. Rhipicephalus sanguineus ticks are known for their great medical and veterinary importance, and for this reason, the knowledge of their intestinal morphology may provide relevant subsidies for the control of these animals, either by direct acaricidal action over these cells or by the production of vaccines. Therefore, this study aimed to describe the midgut morphology of male and female R. sanguineus ticks in different feeding stages, by means of histological analysis. Significant differences were observed between the genders, and such alterations may refer mainly to the distinct demands for nutrients, much higher in females, which need to develop and carry out the egg-laying process. In general, the midgut is coated by a thin muscle layer and presents a pseudostratified epithelium, in which two basic types of cells can be observed, connected to a basal membrane - generative or stem and digestive cells. The latter was classified as follows: residual, deriving from the phase anterior to ecdysis; pinocytic, with vesicles containing liquid or pre-digested components of blood; phagocytic, with entire cells or remnants of nuclear material inside cytoplasmic vesicles; and mature, free in the lumen. Digestion is presumably intracellular and asynchronous and corresponds to a process which starts with the differentiation of generative cells into pinocytic digestive cells, which subsequently start to phagocytize intact blood cells and finally detach from the epithelium, being eliminated with feces. © 2012 Springer-Verlag Berlin Heidelberg.

Cardiovascular alterations at different stages of hypertension development during ethanol consumption: Time-course of vascular and autonomic changes

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

High sodium intake during postnatal phases induces an increase in arterial blood pressure in adult rats

Coordenadoria de Aperfeiçoamento em Pesquisa (CAPES)

Patients With Endometriosis of the Rectosigmoid Have a Higher Percentage of Natural Killer Cells in Peripheral Blood

Study Objective: To estimate the concentration of natural killer (NK) cells in the peripheral blood in patients with and without endometriosis. Design: Case-control study (Canadian Task Force classification II-2). Setting: Tertiary referral hospital. Patients: One hundred fifty-five patients who had undergone videolaparoscopy were divided into 2 groups: those with endometriosis (n = 100) and those without endometriosis (n = 55). Interventions: The percentage of NK cells relative to peripheral lymphocytes was quantified at flow cytometry in 155 patients who had undergone laparoscopy. In addition to verifying the presence of endometriosis, stage of disease and the sites affected were also evaluated. Measurements and Main Results: The mean (SD) percentage of NK cells was higher (15.3% [9.8%]) in patients with endometriosis than in the group without the disease (10.6% [5.8%]) (p < .001). The percentage of NK cells was highest (19.8 [10.3%]) in patients with advanced stages of endometriosis and in those in whom the rectosigmoid colon was affected. In a statistical model of probability, the association of this marker (NK cells >= 11%) with the presence of symptoms such as pain and intestinal bleeding during menstruation and the absence of previous pregnancy yielded a 78% likelihood of the rectosigmoid colon being affected. Conclusion: Compared with patients without endometriosis, those with endometriosis demonstrate a higher concentration of peripheral NK cells. The percentage of NK cells is greater, primarily in patients with advanced stages of endometriosis involving the rectosigmoid colon. Therefore, it may serve as a diagnostic marker for this type of severe endometriosis, in particular if considered in conjunction with the symptoms. Journal of Minimally Invasive Gynecology (2012) 19, 317-324 (C) 2012 AAGL. All rights reserved.

Multiple Intravenous Administrations of Human Umbilical Cord Blood Cells Benefit in a Mouse Model of ALS

Background: A promising therapeutic strategy for amyotrophic lateral sclerosis (ALS) is the use of cell-based therapies that can protect motor neurons and thereby retard disease progression. We recently showed that a single large dose (25x10(6) cells) of mononuclear cells from human umbilical cord blood (MNC hUCB) administered intravenously to pre-symptomatic G93A SOD1 mice is optimal in delaying disease progression and increasing lifespan. However, this single high cell dose is impractical for clinical use. The aim of the present pre-clinical translation study was therefore to evaluate the effects of multiple low dose systemic injections of MNC hUCB cell into G93A SOD1 mice at different disease stages. Methodology/Principal Findings: Mice received weekly intravenous injections of MNC hUCB or media. Symptomatic mice received 10(6) or 2.5x10(6) cells from 13 weeks of age. A third, pre-symptomatic, group received 10(6) cells from 9 weeks of age. Control groups were media-injected G93A and mice carrying the normal hSOD1 gene. Motor function tests and various assays determined cell effects. Administered cell distribution, motor neuron counts, and glial cell densities were analyzed in mouse spinal cords. Results showed that mice receiving 10(6) cells pre-symptomatically or 2.5x10(6) cells symptomatically significantly delayed functional deterioration, increased lifespan and had higher motor neuron counts than media mice. Astrocytes and microglia were significantly reduced in all cell-treated groups. Conclusions/Significance: These results demonstrate that multiple injections of MNC hUCB cells, even beginning at the symptomatic disease stage, could benefit disease outcomes by protecting motor neurons from inflammatory effectors. This multiple cell infusion approach may promote future clinical studies.