An exacerbated metabolism and mitochondrial reactive oxygen species contribute to mitochondrial alterations and apoptosis in CD4 T cells during the acute phase of Trypanosoma cruzi infection.

Chagas disease caused by Trypanosoma cruzi parasite is an endemic infection in America. It is well known that T. cruzi causes a strong immunosuppression during the acute phase of infection. However, it is not clear whether T. cruzi infection is related to metabolic alterations in CD4 T cells that prevent downstream effector function. Here, we evaluated the CD4 T cell metabolic and mitochondrial profiles from non-infected (NI), acute phase (AP) and chronic phase (CP) T. cruzi infected mice. CD4 T cells from all groups showed increased glucose uptake after stimulation. Moreover, the bioenergetic analysis revealed a rise in glycolysis and a higher oxidative metabolism in CD4 T cells from the AP. These cells showed increased proton leak and uncoupling protein 3 (UCP3) expression that correlated with mitochondrial ROS (mROS) accumulation, mitochondrial membrane potential (MMP) depolarization and expression of PD-1. In addition, CD4 T cells with mitochondrial alteration displayed an activated phenotype, and were less functional and more prone to apoptosis. In contrast, mitochondrial alterations were not observed during in vivo activation of CD4 T cells in a model of OVA-immunization. The Mn-superoxide dismutase (SOD2) expression, which is involved in mROS detoxification, was increased during the AP and CP of infection. Remarkably, the apoptosis observed in CD4 T cells with MMP depolarization was prevented by incubation with N-acetyl cysteine (NAC). Thus, our results showed that infection triggered an exacerbated metabolism together with mROS production in CD4 T cells from the AP of infection. However, antioxidant availability may not be sufficient to avoid mitochondrial alterations rendering these cells more susceptible to apoptosis. Our investigation is the first to demonstrate an association between a disturbed metabolism and an impaired CD4 T cell response during T. cruzi infection.

Potential use of ascorbic acid-based surfactants as skin penetration enhancers.

6-O-Ascorbic acid alkanoates (ASCn) are amphiphilic molecules having physical-chemical properties that depend on the alkyl chain length. The derivatives of low molecular weight (n < 11) have enough aqueous solubility to produce self-assemblies at room temperature ( approximately 25 degrees C), while those with longer alkyl chains possess a critical micellar temperature (CMT) higher than 30 degrees C. At higher temperatures (T degrees > CMT), ASCn aqueous suspensions turn into either micellar solutions or gel phases, depending on the length of the hydrophobic chain. On cooling, coagels are produced, which possess a lamellar structure that exhibit sharp X-ray diffraction patterns and optical birefringence. The semisolid consistency of such coagels is an interesting property to formulate dermatological pharmaceutical dosage forms able to solubilize and stabilize different drugs. The objective of the present study was the evaluation of the enhancing permeation effect of ASCn with different chain lengths and to correlate permeability changes with histological effects. With this purpose, ASCn coagels containing anthralin (antipsoriasic drug) or fluorescein isothiocyanate (FITC, hydrophobic fluorescent marker) were assayed on rat skin (ex vivo) and mice skin (in vivo), respectively. Also, histological studies were performed aimed at detecting some possible side effects of ASCn. No inflammatory cellular response was observed in the skin when ASCn coagels were applied, suggesting non-irritating properties. Light microscopy indicated slight disruption and fragmentation of stratum corneum. The penetration of ASCn through rat skin epidermis was very fast and quantitatively significant. The permeation of anthralin was significantly increased when the drug was vehiculized in ASCn coagels, compared to other pharmaceutical systems. The results indicated that ASC12 seems to have the highest enhancing effect on FITC permeation. ASC12 appears to be the compound that possesses the highest capacity to enhance the penetration of the drugs. Furthermore, it has the highest permeation of the serie.

B cells from aged mice exhibit reduced apoptosis upon B-cell antigen receptor stimulation and differential ability to up-regulate survival signals.

During ageing, autoimmune disorders and the higher susceptibility to infectious have been associated with alterations in the humoral immune response. We report that splenic B lymphocytes from aged mice exhibit lower level of apoptosis induced by B-cell antigen receptor (BCR) ligation in vitro. Respect to B cells from young mice the anti-mu stimulated aged B cells show similar Bcl-2 and Bcl-xL expression but differential kinetic of A1 degradation and a higher level of cFLIP and FAIM. Even though B cells from aged mice show minor Fas expression they exhibit the same susceptibility to anti-Fas induced apoptosis. Aged B cells also present upon BCR stimulation, a higher proliferative response and similar level of activation markers expression than B cells from young mice. These data agree with the observation that aged mice exhibit an increment of T2 and mature B cell subset which rapidly enters cell cycle upon BCR engagement. The diminished apoptosis after activation in aged mice could compromise homeostatic mechanism allowing the persistence of self and non-self antigen specific B cells.

Diminished percentage of antigen bearing cells in the lymph nodes of immune aged rats.

We have demonstrated previously that during experimental autoimmune prostatitis (EAP), aged rats show a diminished humoral autoimmune response. In the present paper we have studied the transport of the autoantigen from the site of injection toward lymphatic organs in rats of different ages with or without EAP. We used as autoantigen prostatic components (rat accessory glands (RAG)) conjugated with fluorescein isothiocyanate (FITC). Studies of flow cytometry, fluorescent microscopy and confocal microscopy show no differences in the percentage of RAG-FITC positive cells or in the localization of the cells in the popliteal lymph nodes of not-immunized young and aged rats. On the other hand, in 18-month-old rats immunized with either RAG or Ovalbumin there were lower levels of specific IgG antibodies and fewer antigen containing cells in the draining lymph nodes than those of 3- or 12-month-old rats. In all groups fluorescent cells were MHC class II positive and some were IgM positive. Our results demonstrate that in immunized 18-month-old rats there is a diminished percentage of cells bearing the antigen in the draining lymph nodes after antigen injection in the skin, related to the levels of specific antibodies able to form antigen-antibody complexes in the periphery.

Age-related alterations in inflammatory response during experimental autoimmune prostatitis.

Experimental autoimmune prostatitis (EAP) is an experimental model of autoimmune disease, developed in Wistar rats against prostatic components. The 12-and 18-month-old rats with EAP show a higher cellular autoimmune response and lower humoral autoimmune response compared to 3-month-old rats. The analysis of NO(.) and O(2)(-) production by peritoneal exudate cells (PECs) resulted in a higher NO(.) and O(2)(-) production in EAP rats at all ages, compared to control animals. PECs from 12- and 18-month-old rats produced more NO(.) and less O(2)(-) than PECs from 3-month-old rats. However, lipopolysacharide (LPS) did not stimulate PECs from aged rats for NO(.) production as much as in 3-month-old rats and thus, turning out in a lower index of LPS-stimulation of PECs from aged rats, compared to 3-month-old rats. Furthermore, the mast cells number in prostates of EAP rats, especially the number of degranulated cells, was higher than in control animals, but no significant differences were found between 3- and 12-month-old control rats. In conclusion, these results show that aging affects differentially the inflammation mediators during EAP.

Changes in the development of experimental autoimmune prostatitis (EAP) by castration in aged rats.

During Experimental Autoimmune Prostatitis (EAP), 12-month-old rats show a higher cellular autoimmune response and lower humoral autoimmune response against prostatic components than 3-month-old rats subjected to the same antigen stimulus. We analyzed if thymus recovery by orchidectomy could affect the development of EAP in 12-month-old rats. Thirty days after gonadectomy, 12-month-old rats showed an increment in the thymic mass and in the thymocytes absolute number, with percentages of the four main cell subpopulations (defined by CD4-CD8 molecules expression) similar to the 3-month-old rats. The DTH response of castrated 12-month-old with EAP were diminished in comparison with sham-castrated 12-month-old rats with EAP, resembling the values observed in 3-month-old rats with EAP. The prostates of castrated 12-month-old rats with EAP did not show inflammatory mononuclear cell infiltration, as did control 3- and 12-month-old rats with EAP. Castration seems to modulate negatively EAP in 12-month-old rats, possibly through the regeneration of thymus after testosterone deprivation.

Effect of aging on experimental autoimmune prostatitis: differential kinetics of development.

We have studied the influence of aging on the kinetics of autoimmune response in Experimental Autoimmune Prostatis (EAP). EAP was induced in 3- and 12-month-old Wistar rats by i.d. immunization with a saline extract of rat male sex accessory glands (RAG), chemically modified, and emulsioned in CFA. After immunization, 12-month-old rats developed a faster and stronger specific DTH response against RAG and mononuclear infiltration in the prostate. The levels of total IgM and IgG against RAG were lower in 12-month-old rats than in 3-month-old rats, with a prevalence of IgG2a, IgG2b, and IgG2c subclasses in both ages. Immunization stimulated slightly the appearance of specific IgG1 to RAG only in 3-month-old rats but in 12-month-old rats there was no specific IgG1 to RAG. On the other hand, normal 12-month-old rats showed higher levels of some natural antibodies and their thymocytes and peripheral lymphocytes had a diminished proliferative capacity compared to 3-month-old rats. These data demonstrated that 12-month-old rats show parameters of an aged immune system and present an exacerbated autoimmune prostatitis compared with 3-month-old rats.

Age-associated changes in lymphoid and antigen-presenting cell functions in mice immunized with Trypanosoma cruzi antigens.

The purpose of these studies was to analyze the role of different immune cell populations in the immune response against Trypanosoma cruzi antigens in aged mice. Mice of different ages (3 and 12 months old) were immunized i.d. with S-105 plus Bordetella pertussis as adjuvant and we compared the activities of the lymph node cells taken from 3- and 12-month-old donor animals to transfer DTH or antibody production to 3-month-old recipients. This study revealed that adherent and non-adherent immune lymph node cells of aged donor animals did not transfer response against the foreign antigen (S-105) whereas 3-month-old non-adherent lymph node cells transferred a DTH response as well as helped the specific antibody production. When total lymph node cells from 3- and 12-month-old mice were mixed, we observed an inhibition of S-105 transferred response indicating a suppressive effect of aged cells on the 3-month-old mice cells. Furthermore, we analyzed the participation of antigen-presenting cells (APC) in the immune response changes related to the previously described aged mice. Peritoneal cavities cells (PC), pulsed in vivo with S-105, obtained from 3- and 12-month-old mice were transferred to normal recipients and a DTH response to S-105 was studied. We observed that the DTH response was lower in the recipients of aged PC with respect to recipients of young PC. The results suggest that APC from aged mice are involved in controlling the cellular immune response to S-105. Age-related changes in immune T cell and APC are discussed in the context of these observations.

Aging: the cellular immune response against autologous and foreign antigens is affected before the humoral response.

In this study we examined the auto- and hetero-immune response in mice of different ages immunized with antigens of Trypanosoma cruzi (S-105). We observed that 20-day- and 12-month-old mice showed decreased response to foreign antigens and increased response to autoantigens, compared with 3-month-old immunized mice. The 6-month-old mice showed hetero- and auto-immune cellular responses similar to those of 12-month-old animals; however, the humoral response was similar to that of 3-month-old animals against either antigen, suggesting that the compartments of the immune response are altered at different moments in the same individual. Immune response against a foreign antigen is correlated with the presence of cellular infiltrate in skeletal and heart muscle whereas no modifications in the tissue are noticed in animals with an autoimmune response. Also, we observed from cell transfer experiments that lymph node cells are involved in the dysregulation that we noticed with aging.