CD4+CD69+ T cells and CD4+CD25+FoxP3+ Treg cells imbalance in peripheral blood, spleen and peritoneal lavage from pristane-induced systemic lupus erythematosus (SLE) mice.

BACKGROUND: Adaptive immune cells, including CD4+CD69+ and CD4+CD25+FoxP3+ regulatory T (Treg) cells, are important for maintaining immunological tolerance. In human systemic lupus erythematosus (SLE), CD4+CD25+FoxP3+ Treg cells are reduced, whereas CD69 expression is increased, resulting in a homeostatic immune imbalance that may intensify autoreactive T cell activity. To analyze the mechanisms implicated in autotolerance failure, we evaluated CD4+CD69+ and CD4+CD25+FoxP3+ T cells and interleukin profiles in a pristane-induced SLE experimental model. METHODS: For lupus induction, 26 female Balb/c mice received a single intraperitoneal 0.5 ml dose of pristane, and 16 mice received the same dose of saline. Blood and spleen samples were collected from euthanized mice 90 and 120 days after pristane or saline inoculation. Mononuclear cells from peripheral blood (PBMC), peritoneal lavage (PL) and splenocytes were obtained by erythrocyte lysis and cryopreserved for further evaluation by flow cytometry using the GuavaEasyCyte TM HT. After thawing, cells were washed and stained with monoclonal antibodies against CD3, CD4, CD8, CD25, CD28, CD69, FoxP3, CD14 and Ly6C (BD Pharmingen TM). Interleukins were quantified using Multiplex® MAP. The Mann-Whitney test and the Pearson coefficient were used for statistical analysis, and p < 0.05 considered significant. RESULTS: Compared with the controls, SLE-induced animals presented increased numbers of CD4+CD69+ T cells in the blood on T90 and T120 (p = 0.022 and p = 0.008) and in the spleen on T120 (p = 0.049), but there were decreased numbers in the PL (p = 0.049) on T120. The percentage of Treg was lower in blood (p < 0.005 and p < 0.012) on T90 and T120, in spleen (p = 0.043) on T120 and in PL (p = 0.001) on T90. Increased numbers of CD4 + CD69+ T cells in the PL were positively associated with high IL-2 (p = 0.486) and IFN-γ (p = 0.017) levels, whereas reduced Treg cells in the blood were negatively correlated with TNFα levels (p = 0.043) and positively correlated with TGFß1 (p = 0.038). CONCLUSION: Increased numbers of CD4+CD69+ T cells and reduced numbers of CD4+CD25+FoxP3+ Treg cells with an altered interleukin profile suggests loss of autotolerance in pristane-induced lupus mice, which is similar to human lupus. Therefore, this model is useful in evaluating mechanisms of cellular activation, peripheral tolerance and homeostatic immune imbalance involved in human SLE.

CD4+CD69+ T cells and CD4+CD25+FoxP3+ Treg cells imbalance in peripheral blood, spleen and peritoneal lavage from pristane-induced systemic lupus erythematosus (SLE) mice

Abstract Background: Adaptive immune cells, including CD4+CD69+ and CD4+CD25+FoxP3+ regulatory T (Treg) cells, are important for maintaining immunological tolerance. In human systemic lupus erythematosus (SLE), CD4+CD25+FoxP3+ Treg cells are reduced, whereas CD69 expression is increased, resulting in a homeostatic immune imbalance that may intensify autoreactive T cell activity. To analyze the mechanisms implicated in autotolerance failure, we evaluated CD4+CD69+ and CD4+CD25+FoxP3+ T cells and interleukin profiles in a pristane-induced SLE experimental model. Methods: For lupus induction, 26 female Balb/c mice received a single intraperitoneal 0.5 ml dose of pristane, and 16 mice received the same dose of saline. Blood and spleen samples were collected from euthanized mice 90 and 120 days after pristane or saline inoculation. Mononuclear cells from peripheral blood (PBMC), peritoneal lavage (PL) and splenocytes were obtained by erythrocyte lysis and cryopreserved for further evaluation by flow cytometry using the GuavaEasyCyte TM HT. After thawing, cells were washed and stained with monoclonal antibodies against CD3, CD4, CD8, CD25, CD28, CD69, FoxP3, CD14 and Ly6C (BD Pharmingen TM). Interleukins were quantified using Multiplex® MAP. The Mann-Whitney test and the Pearson coefficient were used for statistical analysis, and p < 0.05 considered significant. Results: Compared with the controls, SLE-induced animals presented increased numbers of CD4+CD69+ T cells in the blood on T90 and T120 (p = 0.022 and p = 0.008) and in the spleen on T120 (p = 0.049), but there were decreased numbers in the PL (p = 0.049) on T120. The percentage of Treg was lower in blood (p < 0.005 and p < 0.012) on T90 and T120, in spleen (p = 0.043) on T120 and in PL (p = 0.001) on T90. Increased numbers of CD4+ CD69+ T cells in the PL were positively associated with high IL-2 (p = 0.486) and IFN-γ (p = 0.017) levels, whereas reduced Treg cells in the blood were negatively correlated with TNFα levels (p = 0.043) and positively correlated with TGFβ1 (p = 0.038). Conclusion: Increased numbers of CD4+CD69+ T cells and reduced numbers of CD4+CD25+FoxP3+ Treg cells with an altered interleukin profile suggests loss of autotolerance in pristane-induced lupus mice, which is similar to human lupus. Therefore, this model is useful in evaluating mechanisms of cellular activation, peripheral tolerance and homeostatic immune imbalance involved in human SLE.

Langerin (CD207)-positive cells in leprosy: Possible implications for pathogenesis of the disease with special emphasis on dermal immunoreactivity.

Leprosy is a disease caused by Mycobacterium leprae, which is characterized by two distinct poles, the tuberculoid pole and the lepromatous pole, depending on the immune response to the bacillus. Langerin-positive cells are dendritic cells that appear to play an essential role in the development of the disease. These cells are specialized in the processing and presentation of antigens, exerting an important function in the activation of the immune system. To evaluate the expression of langerin-positive cells (CD207+) in skin lesion fragments of patients with a diagnosis of M. leprae infection and to associate the expression of these cells with the polar forms of the disease. Langerin-positive cells were detected in larger numbers in lesions of patients with the tuberculoid form compared to those with the lepromatous form. The presence of a larger number of these cells in patients with the tuberculoid form suggests an important participation of langerin-positive cells, capturing antigens and favoring an effective immune response to infection with M. leprae.

Inflammatory and Anti-inflammatory Responses Co-exist Inside Lung Granuloma of Fatal Cases of Coccidioidomycosis: A Pilot Report.

Coccidioidomycosis is a fungal disease caused by Coccidioides immitis or Coccidioides posadasii. These fungi are endemic in the southern USA and northern Mexico. Immunocompromised patients are susceptible to develop severe forms of this fungal infection. Cytokines play an important role in controlling the fungal infection, but little is known about the predominant immunological environment in human lung tissue from fatal cases. Our aim was to analyze the pro-inflammatory and anti-inflammatory cytokines and monocyte/macrophages markers (CD14 and CD206) in the granulomas of six fatal cases of coccidioidomycosis. Cytokines and surface markers were higher in coccidioidomycosis cases when compared to control (P < 0.05). CD14 positive cells were increased inside the coccidioidal granuloma when compared to the outside (P < 0.05). No differences were found in the number of CD206+ cells inside the granuloma when compared to the outer population (P > 0.05). Interestingly, an analysis of stain intensity signals showed an increased signaling of CD14, CD206, IL-10 and TNFα inside the granuloma when compared to the outside (P < 0.05). iNOS and IL-12 gene expression were not detected in coccidioidomycosis cases, while IL-10, IL-6 and TGFß gene expression were detected, but the differences when compared to healthy lungs were not significant (P > 0.05). TNFα gene expression was lower in coccidioidomycosis cases when compared to healthy lung (P = 0.05). In conclusion, pro- and anti-inflammatory responses co-exist inside of the granulomas of fatal cases of coccidioidomycosis and the absent of iNOS and IL-12 gene expression may be related with patient's outcome.

Functional and phenotypic evaluation of eosinophils from patients with the acute form of paracoccidioidomycosis.

BACKGROUND: Eosinophilia is a typical finding of the acute/juvenile form of paracoccidioidomycosis (PCM), a systemic mycosis endemic in Latin America. This clinical form is characterized by depressed cellular immune response and production of Th2 cytokines. Moreover, it has been shown that the increased number of eosinophils in peripheral blood of patients returns to normal values after antifungal treatment. However, the role of eosinophils in PCM has never been evaluated. This study aimed to assess the phenotypic and functional characteristics of eosinophils in PCM. METHODS/PRINCIPAL FINDINGS: In 15 patients with the acute form of the disease, we detected expression of MBP, CCL5 (RANTES) and CCL11 (eotaxin) in biopsies of lymph nodes and liver. In addition, there were higher levels of chemokines and granule proteins in the peripheral blood of patients compared to controls. Isolation of eosinophils from blood revealed a higher frequency of CD69+ and TLR2+ eosinophils in patients compared to controls, and a lower population of CD80+ cells. We also evaluated the fungicidal capacity of eosinophils in vitro. Our results revealed that eosinophils from PCM patients and controls exhibit similar ability to kill P. brasiliensis yeast cells, although eosinophils of patients were less responsive to IL-5 stimulation than controls. CONCLUSION/PRINCIPAL FINDINGS: In conclusion, we suggest that eosinophils might play a role in the host response to fungi and in the pathophysiology of PCM by inducing an intense and systemic inflammatory response in the initial phase of the infection.

Recombinant BCG expressing a PspA-PdT fusion protein protects mice against pneumococcal lethal challenge in a prime-boost strategy.

Pneumococcal proteins have been evaluated as genetically-conserved potential vaccine candidates. We have previously demonstrated that a fragment of PspA in fusion with PdT (rPspA-PdT) induced protective immune responses in mice. However, purified proteins have shown poor immunogenicity and often require the combination with strong adjuvants and booster doses. Here, we investigated the use of a Bacillus Calmette-Guérin (BCG) strain, a well-established prophylactic vaccine for tuberculosis with known adjuvant properties, for delivery of the PspA-PdT fusion protein. Immunization of mice in a prime-boost strategy, using rPspA-PdT as a boost, demonstrated that rBCG PspA-PdT/rPspA-PdT was able to induce an antibody response against both proteins, promoting an IgG1 to IgG2 antibody isotype shift. Sera from rBCG PspA-PdT/rPspA-PdT immunized mice showed antibodies able to bind to the pneumococcal surface and promoted higher complement deposition when compared with WT-BCG/rPspA-PdT or a single dose of rPspA-PdT. In addition, these antisera inhibited the cytolytic activity of Ply. Production of interleukin-6 (IL-6), gamma interferon (IFN-γ), and tumor necrosis factor alpha (TNF-α) was increased in splenocytes culture. Furthermore, a higher expression of CD69 early activation molecule was observed on splenic CD4+ T cells from mice immunized with rBCG PspA-PdT before and after the protein booster dose. Finally, immunization with rBCG PspA-PdT/rPspA-PdT protected mice against pneumococcal lethal challenge. These results support the further investigation of recombinant BCG strains to express pneumococcal proteins, which could be administered in early stages of life and lead to protective pneumococcal immunity in infants and children.

Clinical and immunologic impact of CCR5 blockade in graft-versus-host disease prophylaxis.

Graft-versus-host disease (GVHD) is a major cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Lymphocyte trafficking via chemokine receptors such as CCR5 plays a critical role in alloreactive responses, and previous data suggest that CCR5 blockade with maraviroc results in a low incidence of visceral GVHD. However, the full scope of clinical and immunologic effects of CCR5 blockade in HSCT has not been described. We compared a cohort of patients enrolled on a trial of reduced-intensity allo-HSCT with standard GVHD prophylaxis plus maraviroc to a contemporary control cohort receiving standard GVHD prophylaxis alone. Maraviroc treatment was associated with a lower incidence of acute GVHD without increased risk of disease relapse, as well as reduced levels of gut-specific markers. At day 30, maraviroc treatment increased CCR5 expression on T cells and dampened T-cell activation in peripheral blood without impairing early immune reconstitution or increasing risk for infections. Patients who developed acute GVHD despite maraviroc prophylaxis showed increased T-cell activation, naive T-cell skewing, and elevated serum CXCL9 and CXCL10 levels. Collectively, these data suggest that maraviroc effectively protects against GVHD by modulating alloreactive donor T-cell responses, and that CXCR3 signaling may be an important resistance mechanism to CCR5 blockade in GVHD.

Dendritic cell chimerism in oral mucosa of transplanted patients affected by graft-versus-host disease.

OBJECTIVE: Graft-versus-host disease (GVHD) is one of the main complications after haematopoietic stem cell transplantation. Clinical features of GVHD include either an acute (aGVHD) or a chronic (cGVHD) condition that affects locations such as the oral mucosa. While the involvement of the host's dendritic cells (DCs) has been demonstrated in aGVHD, the origin (donor/host) and mechanisms underlying oral cGVHD have not been completely elucidated. In this study, we intend to determine the origin of DCs present in mucosal tissue biopsies from the oral cavity of transplanted patients affected by cGVHD. METHODS: We purified DCs, from oral biopsies of three patients with cGVHD, through immunobeads and subsequently performed DNA extraction. The origin of the obtained DCs was determined by PCR amplification of 13 informative short tandem repeat (STR) alleles. We also characterised the DCs phenotype and the inflammatory infiltrate from biopsies of two patients by immunohistochemistry. RESULTS: Clinical and histological features of the biopsies were concordant with oral cGVHD. We identified CD11c-, CD207- and CD1a-positive cells in the epithelium and beneath the basal layer. Purification of DCs from the mucosa of patients affected by post-transplantation cGVHD was >95%. PCR-STR data analysis of DCs DNA showed that 100% of analysed cells were of donor origin in all of the evaluated patients. CONCLUSION: Our results demonstrate that resident DCs isolated from the oral tissue of allotransplanted patients affected by cGVHD are originated from the donor. Further research will clarify the role of DCs in the development and/or severity of oral cGVHD.

Quantitative and functional analysis of CD69(+) NKG2D(+) T regulatory cells in healthy subjects.

T regulatory (Treg) cells have a key role in immune homeostasis and the pathogenesis of chronic inflammatory and autoimmune diseases. CD69 is an early leukocyte activation molecule that under steady state conditions is detected in a small proportion of lymphocytes in peripheral blood and lymphoid tissues. Although it has been reported that a subset of CD69(+) T cells behaves as Treg lymphocytes, the possible relationship between CD69(+) Treg cells and CD4(+)NKG2D(+) T lymphocytes, which also exert immunosuppressive activity, has not been explored. In this study, we analyzed the expression of CD69 and NKG2D by T lymphocytes from the peripheral blood of twenty-five healthy subjects by multi-parametric flow cytometry analysis, and their suppressive activity by an assay of inhibition of lymphocyte activation (CD40L expression) and proliferation (carboxyfluorescein partition assay). We found a very small percentage of CD4(+)CD69(+)NKG2D(+) T cells (median 0.002%, Q1-Q3, 0.001-0.004%), which also expressed TGF-ß (Latency Associated Peptide or LAP) and IL-10, in all samples analyzed. These cells exerted an important in vitro suppressive effect on both activation and proliferation of T effector cells. Our data suggest that at very small numbers, CD4(+)CD69(+)NKG2D(+) lymphocytes seem to exert a relevant functional immune-regulatory role in healthy subjects.

Interleukin-18 increases TLR4 and mannose receptor expression and modulates cytokine production in human monocytes.

Interleukin-18 is a proinflammatory cytokine belonging to the interleukin-1 family of cytokines. This cytokine exerts many unique biological and immunological effects. To explore the role of IL-18 in inflammatory innate immune responses, we investigated its impact on expression of two toll-like receptors (TLR2 and TLR4) and mannose receptor (MR) by human peripheral blood monocytes and its effect on TNF-α, IL-12, IL-15, and IL-10 production. Monocytes from healthy donors were stimulated or not with IL-18 for 18 h, and then the TLR2, TLR4, and MR expression and intracellular TNF-α, IL-12, and IL-10 production were assessed by flow cytometry and the levels of TNF-α, IL-12, IL-15, and IL-10 in culture supernatants were measured by ELISA. IL-18 treatment was able to increase TLR4 and MR expression by monocytes. The production of TNF-α and IL-10 was also increased by cytokine treatment. However, IL-18 was unable to induce neither IL-12 nor IL-15 production by these cells. Taken together, these results show an important role of IL-18 on the early phase of inflammatory response by promoting the expression of some pattern recognition receptors (PRRs) that are important during the microbe recognition phase and by inducing some important cytokines such as TNF-α and IL-10.

Immunophenotypic characterization and distribution of dendritic cells in odontogenic cystic lesions.

OBJECTIVE: To analyze the expression and distribution patterns of mature dendritic cells (mDCs) and immature DCs (imDCs) in radicular cysts (RCs), dentigerous cysts (DtCs), and keratocystic odontogenic tumors (KCOTs). MATERIALS AND METHODS: Forty-nine odontogenic cystic lesions (OCLs) (RCs, n = 20; DtCs, n = 15; KCOTs, n = 14) were assessed using the following markers: S100, CD1a and CD207 for imDCs; and CD83 for mDCs. RESULTS: Almost all cases were S100, CD1a, and CD207 positive, whereas 63% were CD83 positive. RCs presented greater number of immunostained cells, followed by DtCs, and KCOTs. The number of S100+ cells was greater than both CD1a+ and CD207+ cells (P < 0.001), which showed approximately similar amounts, followed by lower number of CD83+ cells (P < 0.001) in each OCL type. Different from S100+ cells, both CD1a+ and CD207+ cells on the epithelium (P < 0.05) and CD83+ cells on the capsule (P < 0.05) were preferentially observed. In RCs, significant correlation was found between the thickness epithelium with S100+ and CD1a+ cells, and between the degree of inflammation with CD83+ cells. CONCLUSIONS: Dendritic cell populations in OCLs can be phenotypically heterogeneous, and it could represent distinct lineages and/or functional stages. It is suggested that besides DC-mediated immune cell interactions, DC-mediated tissue differentiation and maintenance in OCLs should also be considered.

Human antibody and memory B and T-cell responses after primary and booster immunisation against Neisseria meningitidis B.

Vaccination against disease aims at the induction of long-lasting cellular and humoral immune responses. Few studies have addressed the mechanisms by which meningococcal vaccines generate and sustain immunological memory. The goal of this study was to investigate the development of long-term humoral and cellular memory to Neisseria meningitidis serogroup B (MenB) in health subjects after immunisation with the Cuban outer membrane protein (OMP) vaccine (VA-MENGOC-BC). The results showed that three doses of vaccine were necessary to induce a detectable memory B-cell response (mean of 0.46%) which became undetectable 6 months later. After boosting, only 2 of 5 individuals responded with an increase in memory B-cell frequencies (values of 0.15% and 0.34%). Bactericidal and opsonic antibody levels were higher after primary immunisation (log(2) mean and median of 4.7 and 1212, respectively) when compared with post-booster response (log(2) mean of 2.6 and median of 285, respectively). Together, these data suggest a failure of vaccine to induce long-term memory B-cell and serological memory in adults. However, we observed a significant and functional memory T-cell response specially after boosting, with a predominance of activated (CD69(+)) central memory T-cell (CD4(+)CD45(-)CCR7(+)) response. Therefore, this study suggests that vaccination with the MenB vaccine induced the generation and activation of memory T-cells but failed to maintain the memory B-cell population at a stable size and/or function.

Purification and biological effects of a C-type lectin isolated from Bothrops moojeni

Snake venom proteins from the C-type lectin family have very distinct biological activities despite their highly conserved primary structure, which is homologous to the carbohydrate recognition region of true C-type lectins. We purified a lectin-like protein (BmLec) from Bothrops moojeni venom and investigated its effect on platelet aggregation, insulin secretion, antibacterial activity, and isolated kidney cells. The BmLec was purified using two chromatographic steps: affinity chromatography and reverse phase high performance liquid chromatography (HPLC). BmLec showed a dose-dependent platelet aggregation and significantly decreased the bacterial growth rate in approximately 15%. During scanning electron microscopy, the profile of Xanthomonas axonopodis pv. passiflorae treated with lectin disclosed a high vesiculation and membrane rupture. BmLec induced a strong and significant increase in insulin secretion at 2.8 and 16.7 mM glucose concentrations, and this effect was seen in the presence of EGTA in both experiments. BmLec (10 ìg/mL) increased the perfusion pressure, renal vascular resistance and urinary flow. The glomerular filtration rate and percentages of sodium, potassium and chloride tubular transport were reduced at 60 minutes of perfusion. Renal alterations caused by BmLec were completely inhibited by indomethacin in all evaluated parameters. In conclusion, the C-type lectin isolated from Bothrops moojeni affected platelet aggregation, insulin secretion, antibacterial activity and isolated kidney function.(AU)

Presence of HIV-1 DNA in spermatozoa from HIV-positive patients: changes in the semen parameters.

Although very inefficient, sexual transmission of HIV-1 is responsible for more than 80% of infections worldwide. Yet, the presence of HIV in spermatozoa has been a matter of debate. The aim of this study was to evaluate the presence of HIV nucleic acids and the distribution of mannose receptors in sperm cells, and to determine the semen parameters and cytokine levels in ejaculates from HIV-positive patients. The presence of non-seminal cells in purified sperm was revealed by light microscopy, flow cytometry and RT-PCR. HIV nucleic acids were evaluated by nested PCR; the distributions of mannose receptors on the surface of the sperm and cytokine levels in ejaculates were determined by fluorescence microscopy and flow cytometry respectively. Sperm characteristics were determined by conventional methods. HIV DNA was detected in 69.2% of purified sperm from HIV-positive men; in contrast all purified sperm were negative for HIV RNA. The distribution of mannose receptors and cytokine levels in HIV-1-positive men were similar to uninfected individuals. Using the Principal Component Analysis (PCA) method, it was possible to determine that semen parameters of HIV-positive men exhibit different distributions compared to HIV-negative individuals. Finally, these results indicate that viral DNA is present in purified sperm from HIV-positive men and that HIV infection of spermatozoa could be associated with lower seminal parameters as demonstrated by the PCA method. The similar distribution of mannose receptors between infected and uninfected individuals suggests that sperm cells from infected individuals interact normally with oocytes.

On the quaternary structure of a C-type lectin from Bothrops jararacussu venom--BJ-32 (BjcuL).

BJ-32 (also known as BjcuL) is a C-type lectin from the venom of Bothrops jararacussu with specificity for beta-galactosides and a remarkable ability to agglutinate several species of trypanosomatids. Our objective was to study the oligomerization state of native BJ-32 by using different biophysical and computational methods. Small-angle X-ray light scattering (SAXS) experiments disclosed a compact, globular protein with a radius of gyration of 36.72+/-0.04A and molecular weight calculated as 147.5+/-2.0kDa. From analytical ultracentrifugation analysis, it was determined that the BJ-32 sedimentation profile fits nicely to a decamer model. The analysis of the intrinsic emitted fluorescence spectra for BJ-32 solutions indicated that association of subunits in the decamer is accompanied by changes in the environment of Tryptophan residues. Both ab initio and comparative models of BJ-32 supported the resemblance of the decamer in the crystallographic structure from a close homologue, the rattlesnake venom lectin (RSL) from Crotalus atrox.

Snake venomics and antivenomics of the arboreal neotropical pitvipers Bothriechis lateralis and Bothriechis schlegelii.

We report the comparative proteomic characterization of the venoms of two related neotropical arboreal pitvipers from Costa Rica of the genus Bothriechis, B. lateralis (side-striped palm pit viper) and B. schlegelii (eyelash pit viper). The crude venoms were fractionated by reverse-phase HPLC, followed by analysis of each chromatographic fraction by SDS-PAGE, N-terminal sequencing, MALDI-TOF mass fingerprinting, and collision-induced dissociation tandem mass spectrometry of tryptic peptides. The venom proteomes of B. lateralis and B. schlegelii comprise similar number of distinct proteins belonging, respectively, to 8 and 7 protein families. The two Bothriechis venoms contain bradykinin-potentiating peptides (BPPs), and proteins from the phospholipase A 2 (PLA 2), serine proteinase, l-amino acid oxidase (LAO), cysteine-rich secretory protein (CRISP), and Zn (2+)-dependent metalloproteinase (SVMP) families, albeit each species exhibit different relative abundances. Each venom also contains unique components, for example, snake venom vascular endothelial growth factor (svVEGF) and C-type lectin-like molecules in B. lateralis, and Kazal-type serine proteinase inhibitor-like proteins in B. schlegelii. Using a similarity coefficient, we estimate that the similarity of the venom proteins between the two Bothriechis taxa may be <10%, indicating a high divergence in their venom compositions, in spite of the fact that both species have evolved to adapt to arboreal habits. The major toxin families of B. lateralis and B. schlegelii are SVMP (55% of the total venom proteins) and PLA 2 (44%), respectively. Their different venom toxin compositions provide clues for rationalizing the distinct signs of envenomation caused by B. schlegelii and B. lateralis. An antivenomic study of the immunoreactivity of the Instituto Clodomiro Picado (ICP) polyvalent antivenom toward Bothriechis venoms revealed that l-amino acid oxidase and SVMPs represent the major antigenic protein species in both venoms. Our results provide a ground for rationalizing the reported protection of the ICP polyvalent antivenom against the hemorrhagic, coagulant, defibrinating, caseinolytic and fibrin(ogen)olytic activities of Bothriechis ( schlegelii, lateralis) venoms. However, these analyses also evidenced the limited recognition capability of the polyvalent antivenom toward a number of Bothriechis venom components, predominantly BPPs, svVEGF, Kazal-type inhibitors, some PLA 2 proteins, some serine proteinases, and CRISP molecules.

[Hematodermic CD4+ CD56+ neoplasm in childhood]. / Neoplasia hematodérmica CD4+ CD56+ en la infancia.

Hematodermic CD4+ CD56+ neoplasm with plasmacytoid dendritic cell phenotype is a rare and aggressive neoplasm recently recognized by the WHO-EORTC classification. It generally appears in elderly adults, exceptionally in childhood. We present a 12-year-old girl with severe mental retardation, genetic clinical features and multiple nodular cutaneous lesions on legs and arms. Histologically the nodules showed diffuse dermal infiltrate of medium and small cells and expression of CD4, CD56, CD43, S100 and plasmacytoid dendritic markers: CD123, BDCA-2 under flow cytometry study. Peripheral blood and bone marrow were not involved. Clinical remission of cutaneous lesions was observed after two weeks of acute lymphoblastic leukemia therapy.

Neoplasia hematodérmica CD4+ CD56+ en la infancia / Hematodermic CD4+ CD56+ neoplasm in childhood

La neoplasia hematodérmica CD4+ CD56+ con fenotipo de célula dendrítica plasmocitoide es una rara y agresiva neoplasia recientemente reconocida por la WHO-EORTC classification. Afecta adultos de edad media y ancianos, siendo muy pocos los casos descriptos en niños. Presentamos el caso de una niña de 12 años con grave retraso mental, estigmas genéticos y múltiples lesiones cutáneas localizadas en miembros inferiores y superiores. Histológicamente se observó un infiltrado dérmico difuso de células pequeñas y medianas con expresión de CD4, CD56, CD43 y S100 así como de marcadores dendríticos plasmocitoides: CD 123 y BDCA-2 confirmados por citometría de flujo, sin compromiso de sangre periférica ni médula ósea. Cumpliendo dos semanas de tratamiento para leucemia linfoblástica aguda evolucionó con remisión clínica de las lesiones cutaneas.

Hematodermic CD4+ CD56+ neoplasm with plasmacytoid dendritic cell phenotype is a rare and aggressive neoplasm recently recognized by the WHO-EORTC classification. It generally appears in elderly adults, exceptionally in childhood. We present a 12-year-old girl with severe mental retardation, genetic clinical features and multiple nodular cutaneous lesions on legs and arms. Histologically the nodules showed diffuse dermal infiltrate of medium and small cells and expression of CD4, CD56, CD43, S100 and plasmacytoid dendritic markers: CD123, BDCA-2 under flow cytometry study. Peripheral blood and bone marrow were not involved. Clinical remission of cutaneous lesions was observed after two weeks of acute lymphoblastic leukemia therapy.

Gut immune stimulation by non pathogenic Gram(+) and Gram(-) bacteria. Comparison with a probiotic strain.

We analyzed the gut immune stimulation induced by Gram-positive bacteria: non probiotic Lactobacillus acidophilus CRL 1462 and Lactobacillus acidophilus A9; two potentially probiotic strains: L. acidophilus CRL 924 and Lactobacillusdelbrueckii subsp. bulgaricus CRL 423; comparatively with a probiotic strain: Lactobacillus casei CRL 431. We also studied Gram-negative bacteria: Escherichia coli 129 and E. coli 13-7 in BALB/c mice. All the strains increased the number of IgA+ cells. We analyzed the cytokines IFNgamma, TNFalpha, IL-17, IL-12, IL-6 and MIP-1alpha. The Gram(+) strains increased the number of IL-10+ cells. Gram(-) strains did not increase IL-10+ cells, but they increased the number of IL-12+ cells. The probiotic strain increased mainly IFNgamma and TNFalpha. In the study of the receptors TLR-2, TLR-4 and CD-206, we demonstrated that only the probiotic strain increased the number of CD-206+ cells. All the Gram(+) strains increased the number of TLR-2+ cells and the Gram(-) strains of the TLR-4+ cells. The probiotic strain induced the release of IL-6 by a preparation enriched in intestinal epithelial cells (IEC). Gram(+) and Gram(-) bacteria activated different immune receptors and induced a different cytokine profile. The probiotic strain showed a great activity on the immune cells and the enriched population in IEC, activating mainly cells of the innate immune system.

Some aspects of the venom proteome of the Colubridae snake Philodryas olfersii revealed from a Duvernoy's (venom) gland transcriptome.

We investigated the putative toxins of Philodryas olfersii (Colubridae), a representative of a family of snakes neglected in venom studies despite their growing medical importance. Transcriptomic data of the venom gland complemented by proteomic analysis of the gland secretion revealed the presence of major toxin classes from the Viperidae family, including serine proteases, metalloproteases, C-type lectins, Crisps, and a C-type natriuretic peptide (CNP). Interestingly, the phylogenetic analysis of the CNP precursor showed it as a linker between two related precursors found in Viperidae and Elapidae snakes. We suggest that these precursors constitute a monophyletic group derived from the vertebrate CNPs.