Angiogenic and prothrombotic markers in extensive slow-flow vascular malformations: implications for antiangiogenic/antithrombotic strategies.

BACKGROUND: Venous and combined malformations are slow-flow haemodynamically inactive lesions that are present at birth and worsen slowly with advancing age, showing no tendency towards involution. The pathogenesis of vascular anomalies has not been fully elucidated, but their formation and progression are closely related to angiogenesis. Localized intravascular coagulation associated with venous or combined malformations is characterized by low fibrinogen, high D-dimers, and normal platelet count. OBJECTIVES: To assess the relationship of angiogenic factors with prothrombotic and endothelial damage/dysfunction markers in patients with extensive slow-flow vascular malformations. METHODS: A 2-year study (2005-2007) included 31 consecutive patients with extensive slow-flow vascular malformations from one centre. RESULTS: Serum levels of the endothelial receptor tyrosine kinase TIE-2, matrix metalloproteinase (MMP)-9 and angiopoietin (Ang)-2 and plasma levels of D-dimer, plasminogen activator inhibitor type 1 (PAI-1), tissue-type plasminogen activator and von Willebrand factor (vWf) were significantly increased in patients compared with healthy controls, whereas serum levels of vascular endothelial growth factor (VEGF)-C, VEGF-D, MMP-2, Ang-1, platelet-derived growth factor (PDGF)-AB and PDGF-BB were significantly decreased in patients compared with controls. A strong positive correlation was present between Ang-1 and PDGF-AB levels (r = 0.63, P < 0.001), between PDGF-AB and PDGF-BB levels (r = 0.67, P < 0.001), and between fibrinogen and PAI-1 levels (r = 0.41, P = 0.031). A strong negative correlation was present between Ang-1 and vWf levels (r = -0.48, P = 0.006), between D-dimer and fibrinogen levels (r = -0.71, P < 0.001), and between PDGF-AB and vWf levels (r = -0.42, P = 0.017). CONCLUSIONS: These findings suggest that angiogenic, coagulation and endothelial damage/dysfunction markers are possibly linked in pathogenesis of extensive slow-flow vascular malformations, and might have therapeutic implications.

[Prolonged complete clinical remission in patients with severe pemphigus vulgaris after cycles of intravenous cyclophosphamide]. / Remisión clínica completa prolongada en pacientes con pénfigo vulgar grave después del tratamiento con ciclos intravenosos de ciclofosfamida.

BACKGROUND: Corticosteroids are the systemic treatment of choice in patients with pemphigus vulgaris, but chronic administration is associated with side effects. Intravenous treatment with cyclophosphamide can improve the clinical signs of pemphigus vulgaris. MATERIAL AND METHODS: We prospectively studied 8 patients diagnosed with pemphigus vulgaris. Six of these had mucocutaneous pemphigus vulgaris and 2 had mucosal pemphigus vulgaris. Treatment consisted of 10 cycles of cyclophosphamide at a dose of 10-15 mg/kg separated by 15 days, while maintaining the initial corticosteroid and immunosuppressant dose. Clinical efficacy was assessed and the anti-epidermal intercellular substance (EIS) and anti-desmoglein (DSG) 3 and 1 antibody titers were monitored (by indirect immunofluorescence and enzyme-linked immunosorbent assay, respectively). RESULTS: All patients with pemphigus vulgaris responded excellently to treatment. Five of the 8 patients achieved complete remission of pemphigus lesions after 10 cycles of cyclophosphamide. In the other 3 patients, the skin lesions disappeared a few weeks after the last cycle of cyclophosphamide. A substantial reduction in immuno suppressant dose was possible in all patients. Furthermore, an improved immunologic response was observed in all cases after cyclophosphamide treatment, with decreased anti-DSG1 and anti-DSG3 antibody titers and well as decreased circulating anti-EIS antibody titers. During the mean 15.1 month follow-up (range, 1-25 months), no new lesions appeared and no side effects of cyclophosphamide therapy were reported. CONCLUSIONS: Fortnightly cycles of intravenous cyclophosphamide may be a useful therapeutic option in patients with severe pemphigus vulgaris. A reduction of corticosteroid dose was possible with this therapeutic approach and the cumulative cyclophosphamide dose was lower than with daily oral administration. Our findings also show that the therapeutic approach induces clinical and immunologic remission in most patients.

Remisión clínica completa prolongada en pacientes con pénfigo vulgar grave después del tratamiento con ciclos intravenosos de ciclofosfamida / Prolonged Complete Clinical Remission in Patients With Severe Pemphigus Vulgaris After Cycles of Intravenous Cyclophosphamide

Introducción: Los corticosteroides son el tratamiento sistémico de elección en los pacientes con pénfigo vulgar (PV). Su administración crónica produce efectos secundarios. La administración de ciclofosfamida (CF) intravenosa puede mejorar las manifestaciones clínicas del PV. Material y método: Estudiamos prospectivamente 8 pacientes diagnosticados de PV. Seis de los pacientes presentaban PV cutáneo-mucoso (PVCM) y 2 casos PV mucoso (PVM). El tratamiento consistió en 10 ciclos quincenales de CF en dosis de 10-15 mg/kg, manteniendo la dosis inicial de corticosteroides y de inmunosupresor. Se evaluó la eficacia clínica y la evolución del título de anticuerpos anti-sustancia intercelular epidérmica (SIE) (inmunofluorescencia indirecta) y específicamente anti-desmogleína (Dsg) 1 y 3 (ELISA). Resultados: Todos los pacientes con PV presentaron una excelente respuesta al tratamiento. Cinco de los 8 pacientes presentaron una remisión completa de las lesiones de PV después de los 10 ciclos de CF. En los otros 3 pacientes las lesiones cutáneas desparecieron unas semanas después del último ciclo de CF. En todos los pacientes se redujo de forma importante la dosis de inmunosupresores. Además, en todos los casos se observó una mejoría en la respuesta inmunológica después del tratamiento con CF, con disminución en el título de anticuerpos frente a las Dsg 1 y 3, así como del título de anticuerpos circulantes frente a la SIE. Tras un seguimiento medio de 15,1 meses (1-25 meses) ningún paciente ha presentado nuevas lesiones de PV. A su vez, no se han observado efectos secundarios por la CF. Conclusiones: La administración de pulsos quincenales de CF intravenosa puede ser una opción terapéutica útil en pacientes con PV grave. Este esquema terapéutico permite disminuir la dosis de corticosteroides con una menor dosis acumulada de CF que en la administración oral diaria. Además, nuestros resultados muestran que este esquema terapéutico se acompaña de una remisión clínica e inmunológica en la mayoría de los pacientes (AU)

Background: Corticosteroids are the systemic treatment of choice in patients with pemphigus vulgaris, but chronic administration is associated with side effects. Intravenous treatment with cyclophosphamide can improve the clinical signs of pemphigus vulgaris. Material and methods: We prospectively studied 8 patients diagnosed with pemphigus vulgaris. Six of these had mucocutaneous pemphigus vulgaris and 2 had mucosal pemphigus vulgaris. Treatment consisted of 10 cycles of cyclophosphamide at a dose of 10-15 mg/kg separated by 15 days, while maintaining the initial corticosteroid and immunosuppressant dose. Clinical efficacy was assessed and the anti-epidermal intercellular substance (EIS) and anti-desmoglein (DSG) 3 and 1 antibody titers were monitored (by indirect immunofluorescence and enzyme-linked immunosorbent assay, respectively). Results: All patients with pemphigus vulgaris responded excellently to treatment. Five of the 8 patients achieved complete remission of pemphigus lesions after 10 cycles of cyclophosphamide. In the other 3 patients, the skin lesions disappeared a few weeks after the last cycle of cyclophosphamide. A substantial reduction in immunosuppressant dose was possible in all patients. Furthermore, an improved immunologic response was observed in all cases after cyclophosphamide treatment, with decreased anti-DSG1 and anti-DSG3 antibody titers and well as decreased circulating anti-EIS antibody titers. During the mean 15.1 month follow-up (range, 1-25 months), no new lesions appeared and no side effects of cyclophosphamide therapy were reported. Conclusions: Fortnightly cycles of intravenous cyclophosphamide may be a useful therapeutic option in patients with severe pemphigus vulgaris. A reduction of corticosteroid dose was possible with this therapeutic approach and the cumulative cyclophosphamide dose was lower than with daily oral administration. Our findings also show that the therapeutic approach induces clinical and immunologic remission in most patients (AU)

Ear, nose and throat manifestations in pemphigus vulgaris.

BACKGROUND: Pemphigus vulgaris (PV) is an autoimmune disease characterized by mucocutaneous intraepithelial blisters and pathogenic autoantibodies against desmoglein 3. There are two clinical forms: mucosal (MPV) and mucocutaneous (MCPV). The frequency of ear, nose and throat (ENT) involvement in PV is not clearly defined. Only a few isolated individual cases have been reported. OBJECTIVES: The objective of our study was to determine the incidence of ENT involvement in patients with PV. PATIENTS: We studied prospectively all 16 patients diagnosed with PV and treated in the Department of Dermatology of the University Clinic of Navarra between 2001 and 2005. They were 10 cases of MPV and six cases of MCPV. All patients were evaluated for ENT manifestations by endoscopic examination. RESULTS: Of the 16 patients, 13 presented with throat symptoms (81%), 12 pharyngeal (75%) and seven laryngeal symptoms (44%). Fourteen patients (88%) had active PV lesions on endoscopic evaluation (eight patients had active lesions on both pharyngeal and laryngeal mucosa, four had PV lesions only on laryngeal mucosa and two had PV lesions on pharyngeal mucosa). Laryngeal lesions were most commonly present in MPV patients. The frequency of nasal symptoms (38%) was lower than active PV lesions (62%) found on ENT examination. Oral symptoms and oral active PV lesions were the most frequent findings (94%). Only three patients with MCPV showed erosions on the external auditory canal. CONCLUSIONS: As ENT endoscopy allows more extensive areas of mucosa to be examined than simple visual inspection, we recommend that it be included in the examination of all patients with PV. By obtaining more complete information concerning the extent of the disease, a more accurate diagnosis can be made, better choice of drug and dose may be decided and, ultimately, response to treatment may be improved.

BY55/CD160 cannot be considered a cytotoxic marker in cytomegalovirus-specific human CD8(+) T cells.

CD160/BY55 is a glucosyl-phosphatidylinositol (GPI)-anchored cell membrane receptor that is expressed primarily in natural killer (NK) cells. Its presence in CD8(+) T lymphocytes is considered to be a marker of cytotoxic activity, although there are few data in this regard. In the present work, we analysed the expression of CD160 in subpopulations of cytomegalovirus (CMV)-specific CD8(+) T cells. Subpopulations were defined by CD28 and CD57 expression and exhibited varying degrees of differentiation and cytotoxic potential, as evaluated by the expression of perforin, interferon (IFN)-gamma and interleukin (IL)-7Ralpha/CD127. We included subjects with different intensities of anti-viral immune response. Results showed that the terminally differentiated CD28(-) CD57(+) subset displaying the highest level of perforin expressed CD160 at a level similar to that of memory CD28(+) CD57(-)perforin(-) cells. A comparison of the expression of perforin in CD160(+) cells versus CD160(-) cells showed that expression was significantly higher in the absence of CD160. Interestingly, the CMV-specific CD8(+) T cell subset from a patient with ongoing CMV reactivation did not begin to express CD160 until day +92 of the follow-up period. Taken together, our data show that CD160 cannot be considered a cytotoxic marker in CMV-specific CD8(+) T cells.

[Past, present and future of anti-idiotype vaccination]. / Pasado, presente y futuro de la vacunación anti-idiotipo.

Cancer vaccines are conceived as therapeutic tools, in contrast to the prophylactic vaccines used to fight against infectious diseases. Among the most potent therapeutic vaccines, anti-idiotype vaccination is directed against the tumor idiotype, the only well-characterized tumor antigen displayed in neoplastic B-cells. Anti-idiotype vaccines have demonstrated clinical benefit against follicular lymphoma and are currently being evaluated in two different phase III clinical trials. Additional emerging strategies, which include the use of dendritic cells and the production of vaccines via molecular means will surely allow us to draw important conclusions concerning the treatment of cancer patients.

Anti-inflammatory cytokines induce lipopolysaccharide tolerance in human monocytes without modifying toll-like receptor 4 membrane expression.

Toll-like receptor 4 (TLR4) participates in innate immunity by detecting lipopolysaccharides (LPS) of Gram-negative bacterial cell walls. TLR4 macrophage expression in mice is modulated by LPS. This fact constitutes, at least partially, the molecular basis for LPS tolerance. Very recently, the effect of interferon-gamma (IFN-gamma), a pro-inflammatory cytokine, has been described on TLR4 membrane expression of human monocytes. IFN-gamma up-regulates TLR4 expression and antagonizes the LPS-induced TLR4 down-regulation. These data prompted us to study the expression of membrane TLR4 in human mono- cytes in which LPS tolerance was induced by LPS and by anti-inflammatory cytokines [interleukin-10 (IL-10) and transforming growth factor beta1 (TGFbeta1)]. Data concerning this latter model, and more specifically, the effect of anti-inflammatory cytokines over TLR4 expression, are not available at present. We show here that membrane TLR4 expression in human monocytes falls after LPS exposure. The effect was prolonged for 12 h, but then expression returned to normal levels. The incubation of human monocytes with IL-10, TGFbeta1 or a mixture of both induces no alterations in membrane TLR4 expression. However, these cytokines are able to substitute the tolerizing LPS exposure in order to induce LPS tolerance. Our data help to achieve a better understanding of the way cytokines control the cellular expression of TLR.

[Active immunotherapy in the treatment of haematological neoplasias]. / Inmunoterapia activa en el tratamiento de neoplasias hematológicas.

The continuous search for therapeutic approaches that improve the conventional treatments of neoplasms, together with an improved understanding of the immune system, has led in recent years to the development of Immunotherapy. Basically, a distinction can be made between two forms of immunotherapy: passive immunotherapy, which consists in the transfer of antibodies or cells previously generated in vitro that are directed against the tumour, and active immunotherapy, which attempts to activate in vivo the immune system and induce it to elaborate a specific response against the tumor antibodies. Hematological neoplasms, specifically some B lymphomas, express in their membrane an immunoglobulin that is considered a specific antigen of the tumour, which is why these diseases have become the ideal target for immunotherapy treatments. There are many alternatives, ranging from protein vaccines, which have already shown clinical benefits, to those of the second generation, which make use of the new techniques of molecular biology to increase the efficacy of the vaccines and obtain their production in a quicker and less costly way, but with which there are not yet definitive clinical results.

Inmunoterapia activa en el tratamiento de neoplasias hematológicas / Active immunotherapy in the treatment of haematological neoplasias

La continua búsqueda de abordajes terapéuticos que mejoren los tratamientos convencionales de las enfermedades neoplásicas junto con el mejor conocimiento del sistema inmunitario ha llevado en los últimos años al desarrollo de la inmunoterapia. Básicamente se pueden distinguir dos formas de inmunoterapia: la inmunoterapia pasiva, que consiste en la transferencia de anticuerpos o células previamente generados in vitro que se dirigen contra el tumor, y la inmunoterapia activa, que pretende activar in vivo el sistema inmunitario e inducirlo a elaborar una respuesta específica contra los antígenos tumorales. Las neoplasias hematológicas, concretamente algunos linfomas B, expresan en su membrana una inmunoglobulina que se considera un verdadero antígeno específico de tumor; por eso estas enfermedades se han convertido en la diana ideal de los tratamientos de inmunoterapia. Las alternativas son muchas, desde las vacunas proteicas que ya han demostrado beneficios clínicos, hasta las de segunda generación, que aprovechan las nuevas técnicas de biología molecular para aumentar la eficacia de las vacunas y conseguir su producción de forma más rápida y menos costosa, pero con las que todavía no hay resultados clínicos definitivos (AU)

[Toll type receptors: molecular bases of the relationship between innate and adaptation responses of the immune system]. / Receptores tipo Toll: bases moleculares de la relación entre respuestas innatas y adaptativas del sistema inmunitario.

Microbes have on their surface molecular patterns that are common among a broad range of pathogens. These patterns are recognized by a wide variety of cellular receptors, the most important of which are a family of transmembrane proteins termed "Toll-like receptors" (TLR). TLRs are pattern-recognition receptors that have key roles in detecting pathogens and initiating inflammatory responses. The receptor of Gram negative bacterial LPS, TLR4, is the best characterized member of the TLR family. So far, ten mammalian toll-like receptors (TLR1-TLR10) have been identified. Recent studies revealed that the TLR signaling pathway is a critical mediator of sepsis. An understanding of TLRs and their signaling pathway will reveal a therapeutic target in sepsis and other immune mediated diseases.

Valor de la detección de autoanticuerpos en la patología glomerular / Value of detecting autoantibodies in glomerular pathology

No disponible

The increase of IFN-gamma production through aging correlates with the expanded CD8(+high)CD28(-)CD57(+) subpopulation.

The use of flow cytometry to detect intracellular cytokines at the single cell level has the potential to quantify cytokine production together with the possibility of phenotypic identification of the cell population concerned. The unbalanced presence of intracellular cytokines produced by T cells has been recognized in some pathological conditions. To better address this issue, we studied the production of IFN-gamma and IL-4 in CD4(+) and CD8(+high) T cells in healthy donors of a broad range of age (17-62 years). Given that an increase of IFN-gamma and IL-4 with aging had been reported by some authors in healthy controls, we have performed a multivariate analysis to assess the intrinsic role of aging or of other external factors, such as chronic antigenic exposures (i.e., viruses), over the cytokine production of phenotypically characterized T cells. In this respect we show that, mainly in CD8(+high) T cells, the production of IFN-gamma is directly correlated with age. Besides, the cytokine production correlates with the CD8(+high)CD28(-)CD57(+) T-cell population, which we have recently reported elevated in aged individuals. Perhaps this T-cell subpopulation plays a regulatory role as a Tc1 response in aging individuals.

Etiopatogenia de la artritis reumatoide / Etiopathogenesis of the rheumatoid arthritis

La etiopatogenia de la artritis reumatoide permanece sin resolver. Se supone que sobre una base genética predisponente actuarían factores ambientales, quizá infecciosos, que desencadenarían el proceso inflamatorio. Sin embargo, se desconoce si existe un desencadenante infeccioso y si la enfermedad es primariamente autoinmune o inflamatoria. Aunque el papel preciso de los linfocitos T como iniciadores o perpetuadores del proceso inflamatorio es aún controvertido, se piensa que los linfocitos T CD4+ reconocen antígenos y estimulan a otras poblaciones celulares como linfocitos B, macrófagos y fiibroblastos sinoviales para producir mediadores inflamatorios. La destrucción articular resulta de la proliferación de la capa intima sinovial formándose el ‘pannus’ que al desarrollarse invade el cartílago y hueso adyacente. Los sinoviocitos tipo fibroblástico y los macrófagos constituyen los elementos celulares predominantes del ‘pannus’ invasor (AU)

The etiopathogenesis of rheumatoid arthritis remains unresolved. Based on a genetic predisposition an exogenous agent, perhaps infectious, is supposed to trigger the inflammatory process. Nonetheless it is not known whether there is an infectious trigger and if the disease is primarily autoimmune or inflammatory. Although the precise role of T cells as initiators or perpetuators of inflammatory process is still controversial, the CD4+ T cells are thought to recognize antigens and stimulate B cells, macrophages, and synovial fibroblasts to produce inflammatory mediators. Joint destruction results from the proliferation of the synovial intimal layer to form a pannus that overgrows and invades adjacent cartilage and bone. Fibroblast-like synoviocytes and macrophages are the predominant cellular components of the invading pannus (AU)

CD5 and CD23 expression on B cells in peripheral blood and synovial fluid of rheumatoid arthritis patients: relationship with interleukin-4, soluble CD23 and tumour necrosis factor alpha levels.

METHODS: We have studied in peripheral blood (PB) and synovial fluid (SF) of 31 patients diagnosed with rheumatoid arthritis (RA), the expression of CD5 and CD23 antigens on B cells, and the levels of soluble CD23 (sCD23), interleukin-4 (IL-4) and tumour necrosis factor alpha (TNF-alpha). We have also correlated the results with the disease activity index. RESULTS: CD5+ B cells are expanded in SF and, moreover, show higher expression of CD23 than CD5 - B cells. Twelve patients had detectable levels of IL-4 in plasma and 10 in SF (nine patients in both samples); the absence of IL-4 was related to a higher expression of CD23 on CD5 + B cells and with higher levels of sCD23. A negative correlation was found in SF between TNF-alpha and sCD23 levels. CONCLUSION: There is no correlation between disease activity index and the different parameters studied (expression of CD5 and CD23 on B cells, sCD23, IL-4 and TNF-alpha levels) either in plasma/PB or in SF.

Cytokine flow cytometry differentiates the clinical status of multiple sclerosis (MS) patients.

In this study we have examined intracellular cytokines in peripheral blood mononuclear cells (PBMC) of MS patients by flow cytometry (cytokine flow cytometry). MS progressive patients showed an increased number of cells producing interferon-gamma (IFN-gamma) after activation with phorbol 12-myristate 13-acetate and ionomycin, compared with patients with clinically inactive forms (P < 0001) and with healthy controls (P = 0001). These cells belonged to the CD4+ and CD8+ subsets in similar proportions. Clinically inactive patients showed a lower level of cells producing IL-2 than controls (P = 0.03) and active MS patients (P = 0.03). Most IL-2-producing cells were CD4+ lymphocytes, although a small part of the IL-2 was also produced by CD8+ cells. The percentage of cells producing simultaneously IL-2 and IFN-gamma was increased in active MS and they were mainly CD4+ lymphocytes. No differences in the production of IL-4 were observed between groups. However, we found an increased IL-10 production in clinically active MS patients (P = 0.03). Treatment with IFN-beta of active MS patients showed lower levels of cytokines when compared with untreated MS patients. This methodological approach could help in the follow up and therapeutic monitoring of MS patients.

Progressive decrease of CD8high+ CD28+ CD57- cells with ageing.

An age-dependent decrease in T cell responsiveness to CD28 costimulation has been described. In order to test the hypothesis that an age-related decrease in CD28 expression by CD8+ T lymphocytes might be involved, we analysed 67 healthy donors ranging in age from 15 to 69 years for their CD8+ T cell expression of CD28 and CD57. We found a statistically significant decrease of CD28 expression through ageing and a significant increase of CD57 expression, both markers being mutually exclusive. Given that cytomegalovirus (CMV) is reported to induce CD57 expression, and since the carrier status for this ubiquitous virus increases with age in the general population, it seemed essential to evaluate whether the phenotypic age-related changes described in CD8high+ cells were not influenced by the CMV carrier status of the individuals. Accordingly, we performed a multivariate analysis to assess the independent association of age and CMV carrier status with CD28 and CD57 expression in CD8high+ cells. Results showed that the progressive decrease in CD8high+ CD28+ CD57- cells was associated only with age, while the expansion of the CD8high+ CD28- CD57+ subset depended both on age and CMV, although mainly on age. We conclude that ageing is accompanied by a progressive loss of CD28 expression in CD8+ T cells and a reciprocal enhancement of CD57 expression, both facts being probably related to the repeated antigenic stimulation occurring throughout life.

[Oxidizing agents and free radicals in biomedicine]. / Oxidantes y radicales libres en biomedicina.

The reactions which involve oxidants and free radicals species have played an essential role at the beginning of aerobic life, and they are an intrinsic part in the regulation of the cellular processes. However, as a result of the derived toxic effects of these oxidative processes, antioxidants molecules appeared in the very early stages of the evolution and they are able to control the production of these reactants and their dangerous effects. Oxidants and antioxidants have a clear function in the cellular physiology. When this delicate balance change many biochemical and cellular reactions are altered, and that may cause different pathologic diseases. In addition, free radicals of oxygen are thought to play a growing role in the biological process of ageing (1). It is not unusual to find papers about these reactants in every topic of the biomedicine. The main oxidants and free radicals in the organisms are oxygen-related agents, which are globally called reactive oxygen intermediates (ROI). These unavoidable, useful and dangerous biological oxidants are well characterized, although the recent implications they received in some pathologies deserve, in our opinion, a general review.