Loss of lineage antigens is a common feature of apoptotic lymphocytes

The analysis of apoptosis in cell populations involves the detection of their specific lineage antigen (LAg) expression. This experimental approach relies on their assumed constant expression, but it is unclear whether such expression is actually maintained during cell death. We examined whether the loss of LAgs is a common feature of apoptotic lymphocytes and whether some might completely lose their LAgs. The changes in the expression of CD3, CD5, CD8, CD4, CD28, CD56, and CD19 were monitored in highly purified lymphocyte populations obtained by negative selection in a fluorescence-activated cell sorter. These were cultured for 24 h with or without phytohemagglutinin or staurosporin. For each LAg-positive subset studied, apoptosis was consistently more common among cells showing partial or total loss of LAg expression compared with cells maintaining their initial LAg levels. The kinetics of expression loss was rapid for CD8, CD56, and CD28, and more than 80% of initial expression was lost in the early stages of apoptosis but was slower for CD3, CD5, and CD4. For CD3 and CD5, expression was dependent on the apoptotic stimulus used. It is interesting that loss of antigen expression was independent of cell size. This phenomenon was also found in nonmanipulated, highly pure CD19 B lymphocytes of peripheral blood mononuclear cells from B chronic lymphocytic leukemia patients. Loss of LAg expression appeared to be a common feature of apoptotic lymphocytes under all the conditions assayed. The different kinetic patterns of LAg loss suggest apoptotic cells might actively regulate this process.     

Cell surface CD28 levels define four CD4+ T cell subsets: abnormal expression in rheumatoid arthritis

CD28 is a costimulatory receptor expressed in most CD4(+) T cells. Despite the long-standing evidence for up- and downregulation of surface CD28 expression in vitro, and the key regulatory role assigned to the upregulation of CD28 counterreceptor [the CD152 (CTLA-4) molecule], in vivo CD28 induction has attracted little attention. We studied CD28 and CD152 expression and function in 33 rheumatoid arthritis (RA) patients, 20 clinically active and 13 inactive, and in 24 healthy donors. Four subsets of CD28(-), CD28(low), CD28(int), and CD28(high) peripheral blood human CD4(+) T cells were defined using three-color flow cytometry. The three CD28(+) subsets displayed a one-, two-, or threefold quantitative difference in their relative number of CD28 antibody binding sites, respectively (P < 0.01). RA patients, whether active or inactive, showed a distinct phenotype when compared to healthy donors: (i) the percentage of CD4(+)CD28(high) cells was increased twofold and the CD4(+)CD28(low) subset was reduced twofold (P < 0.01) and (ii) the CD4(+)CD28(high) cells from RA patients showed an in vivo activated phenotype, CD45RO(+)CD5(high)IL-2Ralpha(+) (P < 0.01). Active RA patients were different from inactive patients. They showed a twofold increase in mean CD28 expression (P < 0.05), whereas each of the CD28(+) subsets in the inactive RA patients showed reduced expression when compared to healthy donors. Notably, both active and inactive RA patients showed abnormal CD28 upregulation when T cells were activated in vitro with CD3 antibodies, but only inactive RA patients showed a hypoproliferative response to TCR/CD3 triggering when compared to healthy donors (P < 0.01). This defective proliferation was normalized by concurrent crosslinking with CD28 antibody. No differences were noted in the expression of CD152 or CD80, a CD28 and CD152 shared ligand. The disregulated in vivo expression of CD28 was related to the RA patients' disease activity and suggests that modulation of CD28 surface levels may be an additional mechanism to finely tune the delicate responsiveness/tolerance balance.     

Defective functional response to membrane stimuli in lymphocytes from patients with benign prostatic hyperplasia.

Benign prostatic hyperplasia (BPH) is a local disturbance in the prostate that may involve an inflammatory infiltrate predominantly composed of activated lymphocytes and macrophages. The activation and proliferative response of T lymphocytes to different mitogenic signals has been analysed in peripheral blood mononuclear cells (PBMC) from 45 patients with BPH and 55 healthy controls. The PBMC obtained from the patients showed a significant specific impairment in proliferation, CD25 expression and IL-2 production in response to stimulation with lectins (phytohaemagglutinin (PHA), concanavalin A (Con A)), that was not corrected by the addition of IL-2 or of phorbol esters (phorbol myristate acetate (PMA)). Also, the CD28 response was defective in patient PBMC. Activation with anti-CD3 or anti-CD2 MoAbs was normal, but the addition of PMA to these stimuli provoked a significant defective response. Only the use of transmembrane stimuli (PMA and ionomycin) elicited responses similar to those found in the control group. The results indicate that peripheral T lymphocytes from BPH patients show a functional impairment that is mainly explained by an alteration of membrane signals (PHA, CD28) and is distal to protein kinase C (PKC) activation.     

The B-cell activation pathway in human systemic lupus erythematosus: Imbalancedin vitro production of lymphokines and association with serum analytical findings

Recent knowledge of B-lymphocyte physiology has clarified the role of T cell-derived lymphokines in clonal proliferation and differentiation of B-cell responses. Lymphokine production was analyzed in 19 systemic lupus erythematosus (SLE) patients and sex- and age-matched controls in relation to clinical activity and steroid treatment. Whenin vitro production of interleukin-2 (IL-2) and B-cell growth factor (BCGF) was tested, both activities were found to be diminished in the group of patients (P<0.01), while B-cell differentiation factor (BCDF) activity was higher in this group with respect to normal controls (P<0.01). Interestingly enough, thisin vitro BCDF synthesis was positively correlated with clinical activity regardless of low-dose steroid treatment. A correlation was also found between BCDF production and the levels of IgG (r=0.64,P<0.01), anti-DNA antibodies (r=0.52,P<0.05), and the IgG/IgM ratio (r=0.7,P<0.01) in serum. Implications of these abnormal T-lymphocyte functions in SLE with respect toin vivo B-cell function are discussed.     

Immunohistochemical classification of non-BRCA1/2 tumors identifies different groups that demonstrate the heterogeneity of BRCAX families.

Around 25% of hereditary breast and ovarian cancer families have mutations in the BRCA1 and BRCA2 genes. The search for other genes has until now failed, probably because there is not one single BRCAX gene, but rather various genes that may each be responsible for a small number of breast cancer families and/or may interact according to a polygenic model. We have studied 50 tumors from probands belonging to non-BRCA1/2 breast cancer families (BRCAX), using 25 immunohistochemical markers. The objective was to classify these tumors and confirm that they are heterogeneous. Unsupervised cluster analysis showed the existence of the following two main groups of tumors: high-grade and estrogen receptor (ER)-negative tumors (50%), and low-grade and ER-positive tumors (50%). In addition we identified five subgroups, three among the high-grade and two among the low-grade groups; one overexpressing HER-2 (18%); one with a basal-like phenotype (14%); one with a normal breast-like phenotype (18%); a luminal A subgroup (36%), and a luminal B subgroup (14%). Hypermethylation of the BRCA1 gene was observed in 42% of the cases, spread across all five subgroups, but only 37% of those had loss of heterozygosity as well. These latter cases were all clustered in the high-grade group and the majority of them in the basal-like subgroup. Our results show that familial non-BRCA1/2 tumors are heterogeneous and suggest a polygenic model for explaining the majority of BRCAX families. In addition we have defined a subset of them that have somatic inactivation of the BRCA1 gene.     

Decreased TcR-CD3+ T cell numbers in healthy aged humans. Evidence that T cell defects are masked by a reciprocal increase of TcR-CD3- CD2+ natural killer cells

While there is accumulating evidence to indicate the presence of functional abnormalities in T cells from aged healthy humans, their cellular basis remains unclear. By using two-color immunofluorescence and multiparameter flow cytometry we show that (a) the number of peripheral blood antigen receptor-positive (TcR-CD3+) T cells is significantly lower in aged than in young adults; (b) the numbers of E-rosette-forming (CD2+) cells are maintained in the elderly due to a reciprocal increase in the frequency of TcR-CD3- cells, which constitute only a minor lymphocyte subpopulation in young adults, and (c) TcR-CD3-CD2+5- lymphocytes exhibit the phenotypic features of natural killer (NK) cells. By using functional assays we show the TcR-CD3-CD2+16+ lymphocytes are indeed NK cells because they are activated by and lyse NK targets. In contrast, they are unresponsive to either phytohemagglutinin or mitogenic CD2 monoclonal antibody stimulation, which in turn activates TcR-CD3+CD2+16- T cells. We conclude that the increase in TcR-CD3-CD2+ NK cells masks the T cell reduction in aged humans by normalizing CD2+ cell frequencies. However, NK cells cannot functionally substitute the thymus-derived lymphocytes they replace.     

Resumen ejecutivo del Documento de Consenso de la Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica (SEIMC) y de la Asociación Española de Cirujanos (AEC) en profilaxis antibiótica en cirugía

Antibiotic prophylaxis in surgery is one of the most effective measures for preventing surgical site infection, although its use is frequently inadequate and may even increase the risk of infection, toxicities and antimicrobial resistance. As a result of advances in surgical techniques and the emergence of multidrug-resistant organisms, the current guidelines for prophylaxis need to be revised.The Sociedad Española de Enfermedades Infecciosas (Spanish Society of Infectious Diseases and Clinical Microbiology) (SEIMC) together with the Asociación Española de Cirujanos (Spanish Association of Surgeons) (AEC) have revised and updated the recommendations for antibiotic prophylaxis in surgery to adapt them to any type of surgical intervention and to current epidemiology. This document gathers together the recommendations on antimicrobial prophylaxis in the various procedures, with doses, duration, prophylaxis in special patient groups, and in epidemiological settings of multidrug resistance to facilitate standardized management and the safe, effective and rational use of antibiotics in elective surgery.     

Overview and update on molecular testing in non-small cell lung carcinoma utilizing endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) samples

Lung carcinoma remains one of the most frequent and aggressive human neoplasms. Fortunately, in the last decades, the increasing knowledge of the molecular mechanisms leading to cancer development has allowed the use of targeted therapies with improvement of prognosis in many patients. Clinical management has also changed after the introduction of endobronchialultrasonographic bronchoscopy that allows a conservative staging of lung tumors, avoiding the need of mediastinoscopy for lymph node staging. Lung pathologists and cytopathologists are facing the challenge of giving the more comprehensive prognostic and predictive information with ever smaller tissue or cytological samples. The aim of this review is to summarize the molecular testing for non-small cell lung carcinoma and how pathologists can contribute to the patient's outcome with a conscious management of biological samples.     

Chagasic heart disease in Mexico. 1st case diagnosed by positive xenodiagnosis

We describe the first case of chronic Chagas heart disease in México, in whom the illness was corroborated by means of the "xenodiagnosis". The patient had changing bundle branch block and also A-V block that required insertion of a pacemaker. This disease is seldom seen in Mexico.