Trichomonas vaginalis Exosomes Deliver Cargo to Host Cells and Mediate Host∶Parasite Interactions

Trichomonas vaginalis is a common sexually transmitted parasite that colonizes the human urogential tract where it remains extracellular and adheres to epithelial cells. Infections range from asymptomatic to highly inflammatory, depending on the host and the parasite strain. Here, we use a combination of methodologies including cell fractionation, immunofluorescence and electron microscopy, RNA, proteomic and cytokine analyses and cell adherence assays to examine pathogenic properties of T. vaginalis. We have found that T.vaginalis produces and secretes microvesicles with physical and biochemical properties similar to mammalian exosomes. The parasite-derived exosomes are characterized by the presence of RNA and core, conserved exosomal proteins as well as parasite-specific proteins. We demonstrate that T. vaginalis exosomes fuse with and deliver their contents to host cells and modulate host cell immune responses. Moreover, exosomes from highly adherent parasite strains increase the adherence of poorly adherent parasites to vaginal and prostate epithelial cells. In contrast, exosomes from poorly adherent strains had no measurable effect on parasite adherence. Exosomes from parasite strains that preferentially bind prostate cells increased binding of parasites to these cells relative to vaginal cells. In addition to establishing that parasite exosomes act to modulate host∶parasite interactions, these studies are the first to reveal a potential role for exosomes in promoting parasite∶parasite communication and host cell colonization.     

Ventilatory support: a dynamical systems approach

Misunderstanding of the dynamical behavior of the ventilatory system, especially under assisted ventilation, may explain the problems encountered in ventilatory support monitoring. Proportional assist ventilation (PAV) that theoretically gives a breath by breath assistance presents instability with high levels of assistance. We have constructed a mathematical model of interactions between three objects: the central respiratory pattern generator modelled by a modified Van der Pol oscillator, the mechanical respiratory system which is the passive part of the system and a controlled ventilator that follows its own law. The dynamical study of our model shows the existence of two crucial behaviors, i.e. oscillations and damping, depending on only two parameters, namely the time constant of the mechanical respiratory system and a cumulative interaction index. The same result is observed in simulations of spontaneous breathing as well as of PAV. In this last case, increasing assistance leads first to an increase of the tidal volume (V_T), a further increase in assistance inducing a decrease in V_T, ending in damping of the whole system to an attractive fixed point. We conclude that instabilities observed in PAV may be explained by the different possible dynamical behaviors of the system rather than changes in mechanical characteristics of the respiratory system.     

Characterization of Lung Fibroblasts More than Two Decades after Mustard Gas Exposure

Purpose

In patients with short-term exposure to the sulfur mustard gas, the delayed cellular effects on lungs have not been well understood yet. The lung pathology shows a dominant feature consistent with obliterative bronchiolitis, in which fibroblasts play a central role. This study aims to characterize alterations to lung fibroblasts, at the cellular level, in patients with delayed respiratory complications after short-term exposure to the sulfur mustard gas.

Methods

Fibroblasts were isolated from the transbronchial biopsies of patients with documented history of exposure to single high-dose sulfur mustard during 1985–7 and compared with the fibroblasts of control subjects.

Results

Compared with controls, patients’ fibroblasts were thinner and shorter, and showed a higher population doubling level, migration capacity and number of filopodia. Sulfur mustard decreased the in vitro viability of fibroblasts and increased their sensitivity to induction of apoptosis, but did not change the rate of spontaneous apoptosis. In addition, higher expression of alpha smooth muscle actin showed that the lung''s microenvironment in these patients is permissive for myofibroblastic differentiation.

Conclusions

These findings suggest that in patients under the study, the delayed pulmonary complications of sulfur mustard should be considered as a unique pathology, which might need a specific management by manipulation of cellular components.     

A Simple Dynamic Model of Respiratory Pump

To study the interaction of forces that produce chest wall motion, we propose a model based on the lever system of Hillman and Finucane (J Appl Physiol 63(3):951–961, 1987) and introduce some dynamic properties of the respiratory system. The passive elements (rib cage and abdomen) are considered as elastic compartments linked to the open air via a resistive tube, an image of airways. The respiratory muscles (active) force is applied to both compartments. Parameters of the model are identified in using experimental data of airflow signal measured by pneumotachography and rib cage and abdomen signals measured by respiratory inductive plethysmography on eleven healthy volunteers in five conditions: at rest and with four level of added loads. A breath by breath analysis showed, whatever the individual and the condition are, that there are several breaths on which the airflow simulated by our model is well fitted to the airflow measured by pneumotachography as estimated by a determination coefficient R ² ≥ 0.70. This very simple model may well represent the behaviour of the chest wall and thus may be useful to interpret the relative motion of rib cage and abdomen during quiet breathing.     

RTVP-1 expression is regulated by SRF downstream of protein kinase C and contributes to the effect of SRF on glioma cell migration

Gliomas are characterized by increased infiltration into the surrounding normal brain tissue. We recently reported that RTVP-1 is highly expressed in gliomas and plays a role in the migration of these cells, however the regulation of RTVP-1 expression in these cells is not yet described. In this study we examined the role of PKC in the regulation of RTVP-1 expression and found that PMA and overexpression of PKCα and PKCε increased the expression of RTVP-1, whereas PKCδ exerted an opposite effect. Using the MatInspector software, we identified a SRF binding site on the RTVP-1 promoter. Chromatin immunoprecipitation (ChIP) assay revealed that SRF binds to the RTVP-1 promoter in U87 cells, and that this binding was significantly increased in response to serum addition. Moreover, silencing of SRF blocked the induction of RTVP-1 expression in response to serum. We found that overexpression of PKCα and PKCε increased the activity of the RTVP-1 promoter and the binding of SRF to the promoter. In contrast, overexpression of PKCδ blocked the increase in RTVP-1 expression in response to serum and the inhibitory effect of PKCδ was abrogated in cells expressing a SRFT160A mutant. SRF regulated the migration of glioma cells and its effect was partially mediated by RTVP-1. We conclude that RTVP-1 is a PKC-regulated gene and that this regulation is at least partly mediated by SRF. Moreover, RTVP-1 plays a role in the effect of SRF on glioma cell migration.     

Roles of tyrosine phosphorylation and cleavage of protein kinase Cdelta in its protective effect against tumor necrosis factor-related apoptosis inducing ligand-induced apoptosis

Protein kinase Cdelta (PKCdelta) regulates cell apoptosis in a cell- and stimulus-specific manner. Here, we studied the role of PKCdelta in the apoptotic effect of TRAIL in glioma cells. We found that transfection of the cells with a PKCdelta kinase-dead mutant (K376R) or with a small interfering RNA targeting the PKCdelta mRNA increased the apoptotic effect of tumor necrosis factor-related apoptosis inducing ligand (TRAIL), whereas overexpression of PKCdelta decreased it. PKCdelta acted downstream of caspase 8 and upstream of cytochrome c release from the mitochondria. TRAIL induced cleavage of PKCdelta within 2-3 h of treatment, which was abolished by caspase 3, 8, and 9 inhibitors. The cleavage of PKCdelta was essential for its protective effect because overexpression of a caspase-resistant mutant (PKCdeltaD327A) did not protect glioma cells from TRAIL-induced apoptosis but rather increased it. TRAIL induced translocation of PKCdelta to the perinuclear region and the endoplasmic reticulum and phosphorylation of PKCdelta on tyrosine 155. Using a PKCdeltaY155F mutant, we found that the phosphorylation of PKCdelta on tyrosine 155 was essential for the cleavage of PKCdelta in response to TRAIL and for its translocation to the endoplasmic reticulum. In addition, phosphorylation of PKCdelta on tyrosine 155 was necessary for the activation of AKT in response to TRAIL. Our results indicate that PKCdelta protects glioma cells from the apoptosis induced by TRAIL and implicate the phosphorylation of PKCdelta on tyrosine 155 and its cleavage as essential factors in the anti-apoptotic effect of PKCdelta.