Berta Sánchez

CD4+ and CD8+ T lymphocyte regeneration after anti-retroviral therapy in HIV-1-infected children and adult patients

Previous studies have shown a slow recovery of naive CD4+ T cell counts after anti‐retroviral therapy in HIV‐1‐infected adults, which is in accordance with thymus atrophy after puberty. Here we investigate whether or not different patterns of naive CD4+ and CD8+ T cell repopulation are present in adult and child patients undergoing anti‐retroviral treatment. Thus, 25 adults under highly active anti‐retroviral therapy and 10 children under combined anti‐retroviral therapy were retrospectively analysed for T cell subpopulations at baseline (T0) and around week 12 (T1) and week 24 (T2) of anti‐retroviral treatment. Mean serum HIV‐1 RNA levels dropped in both groups. Recovery of T cells in adults was characterized by a heterogeneous response between patients, with only 44% of them increasing their naive CD4+ and CD8+ T cell counts at T1, and changes in mean total CD4+ T cells were mainly shaped by memory cells. Otherwise, children were characterized by an early increase in naive T cells. Thus, at T1, all children analysed had a strong rise in CD4+ (from 389 ± 116 to 569 ± 121 cells/μl; P < 0·01), and nine out of 10 also in naive CD8+ T cells (from 244 ± 58 to 473 ± 85 cells/μl; P < 0·05). However, no significant correlation between age and naive repopulation was observed (P= 0·22) in children. Thus, children had the earlier and greater increases in naive T cell subsets than adults, probably due to a more active thymus, with the potential for immune reconstitution when HIV‐1 replication is controlled.

Molecular structure of eight human autoreactive monoclonal antibodies

The heavy (H) and light (L) chain V‐region sequences of eight human autoreactive immunoglobulin M (IgM) monoclonal antibodies (mAbs: BY‐4, BY‐7, BY‐12, IRM‐3, IRM‐7, IRM‐8, IRM‐10 and CDC‐1) were determined at the cDNA level. All VH and VL families were identified. Four different VH families were represented, VH3 being the most common as five of the mAbs (BY‐7, BY‐12, IRM‐3, IRM‐8 and CDC‐1) used genetic elements of this family, whereas VH1, VH2 and VH4 were only present in IRM‐7, BY‐4 and IRM‐10, respectively. BY‐4, BY‐7, BY‐12, IRM‐7 and IRM‐10 reacted with a variety of self as well as non‐self antigens, thus exhibiting polyreactive behaviour. Comparison of the gene segments utilized by these mAbs with their germline counterparts revealed that the gene segments were close to germline configuration. The length of H‐CDR3 was found to be relatively long (27–60 nucleotides) among the polyreactive mAbs and the presence of Tyr and Trp residues in this region seems to be of vital importance for polyreactivity. We have analysed the utilization of gene elements and the presence of amino acid residues in regions particularly important for antigen binding, such as CDR. Common molecular features relating to the function of the mAbs are discussed.

Therapeutic effect of the acquisition of cytomegalovirus-specific immune response during preemptive treatment.

Background. It has been suggested that preemptive therapy against cytomegalovirus (CMV) infection after transplantation promotes a CMV-specific immune response. Our objective was to determine whether solid-organ transplant patients at high risk for CMV infection treated preemptively acquire a CMV-specific immune response and whether the acquired immune response confers immunity by controlling subsequent CMV replication episodes and by protecting from late-onset CMV disease. Methods. Patients were followed up for 18 months after transplantation. CMV viral load was determined using real-time polymerase chain reaction assays, and the T-cell immune response was characterized by intracellular cytokine staining. Results. The 21 patients studied developed CMV replication episodes at a median of 4 weeks (range 2–8 weeks) after transplantation and a CMV-specific T-cell response within a median of 12 weeks (range 10–20 weeks). The decline in the incidence of CMV replication episodes is inversely correlated with the acquisition of the CMV-specific T-cell response (linear regression r2=0.781, Pearson correlation=−0.883; P=0.001). There were no CMV replication episodes after week 47 of transplantation. In addition, after acquisition of the immune response, 42 replication episodes were cleared without treatment. The time taken for immune clearance of replication correlated with the peak viral load (P=0.01). No incidence of CMV early or late-onset disease was detected. Conclusions. Our results demonstrate that preemptive therapy is a safe and an effective strategy for the control of CMV infection in solid-organ transplant recipients at high risk for CMV infection. This is the first study that reports a therapeutic effect of the acquisition of CMV-specific immune response during preemptive treatment.

Psychological problems in the family members of gravely traumatised patients admitted into an intensive care unit.

The aim of these studies was the analysis of the psychological repercussions on the closest members of families of 76 gravely traumatised patients admitted into the Intensive Care Unit (ICU) of the Hospital Universitario de Rehabilitación y Traumatología “Virgen del Rocio”, Sevilla (Spain). An investigation based on social information and the Clinical Analysis Questionnaire was used. The sample of family members was composed of 42 women and 34 men, with an average age of 41.3 years (SD±12.8). Results showed that (a) more than 50% of the family members of gravely traumatised patients admitted into an ICU showed symptoms of depression, (b) the women scored more points in hypochondria, suicidal depression, anxious depression, low-energy depression, guilt-resentment, apathy-withdrawal, paranoia, schizophrenia, psychasthenia and psychological disadjustment, and (c) in general terms, the psychological characteristics of the families were far from the norm of the control group.

TAP polymorphism in patients with Behçet's disease.

OBJECTIVE--To determine if susceptibility to Behçets disease (BD) is associated with polymorphism of HLA-DRB1, HLA-DQB1, DQB1, and TAP1 and TAP2 genes. METHODS--Fifty eight Spanish BD patients and 116 ethnically matched unrelated healthy subjects were typed at the HLA-DRB1 and HLA-DQB1 loci using polymerase chain reaction/sequence specific oligotyping (PCR/SSO). TAP1 and TAP2 alleles were assigned using amplification refractory mutation system-PCR. RESULTS--TAP1C was absent in BD patients, but was found in 12.1% of control subjects (pcorr < 0.05; relative risk = 0.06). Additionally, a linkage disequilibrium between HLA-DQB1*0501 and TAP2B was observed in BD patients (delta = 0.095, pcorr < 0.02), but not in the control group (delta = -0.0031, p > 0.05). CONCLUSIONS--The complete absence of TAP1C alleles in BD patients may indicate that TAP1 polymorphism is not without some significance in the development of BD. Furthermore, the existence of a linkage disequilibrium between HLA-DQB1*0501 and TAP2B in our patients suggests that the gene conferring susceptibility for BD is inherited as an extended haplotype in the population studied.

Tonsillar lymphocyte subsets in recurrent acute tonsillitis and tonsillar hypertrophy

Recurrent acute tonsillitis is usually produced directly by micro-organisms, mainly beta-hemolytic streptococcus. Idiopathic tonsillar hypertrophy is presented without infection history and usually leads to obstructive sleep apnea. We have measured lymphocyte subsets in tonsillar cellular suspensions of infectious and obstructive tonsillar pathology by flow cytometry. Comparing with peripheral blood, the CD4+/CD8+ ratio for tonsillar pathology varies from 4.0 to 5.0 while in peripheral blood the ratio was 1.3. In tonsils the ratio of B lymphocytes/T lymphocytes is 1.6, being 0.3 in peripheral blood, cytotoxic T lymphocytes represent 8% in tonsils and 29% in peripheral blood, virgin or nonstimulated T lymphocytes (CD4+ CD62L+) consist of 3% in tonsils and 16% in peripheral blood. The immature B lymphocytes (CD20+ CD5+) represent 23% in tonsils and 12% in peripheral blood. In regards to NK cells (CD3- CD16+), 1% was found in tonsils and 11% in peripheral blood. In tonsils B lymphocytes and a low proportion of cytotoxic T lymphocytes predominate, in comparison to peripheral blood, with a CD4+/CD8+ ratio four times greater than tonsils. We have found in tonsils a significant increase of T cells (CD3+ and TCR alpha+ beta+) in infectious processes in comparison to obstructive pathology.

HUMAN POLYREACTIVE IGM MONOCLONAL ANTIBODIES WITH BLOCKING ACTIVITY AGAINST SELF-REACTIVE IGG

Natural IgM antibodies have been found to be involved in the control of IgG reactivity in normal serum. The authors investigated the blocking activity of four human IgM monoclonal antibodies (BY‐2, BY‐7, BY‐10 and IRM‐7) derived from B‐cells from blood samples of three renal dialysis patients, which had shown multispecific properties similar to those observed for natural polyreactive autoantibodies. To achieve this, competitive inhibition assays were performed with these MoAbs on the binding of IgG purified from a healthy control, three patients with SLE, and two patients with autoimmune thyroiditis, to histone, dsDNA, RNP and thyroglobulin. MoAbs inhibited binding of self‐reactive IgG to histone and dsDNA, but not to thyroglobulin or RNP, of natural and active or inactive phase disease‐associated autoreactive IgG. The inhibitory effect of the MoAbs was mediated by V‐region dependent interactions with autoreactive IgG, as shown by the ability of these MoAbs to block the binding of F(ab′)2 fragments of autoreactive IgG to antigens (histone and dsDNA). The blocking of autoantibody activity was dose‐dependent with maximal inhibition occurring at a specific molar ratio between the patients IgG and a given MoAb. In contrast, MoAbs did not inhibit binding of IgG alloantibodies present in the sera of four polytransfused renal dialysis patients to target antigens on the surface of different cells. These results support the concept of a functional idiotypic network regulating autoimmune responses, and suggest that the IgM MoAbs under study may be natural polyreactive antibodies belonging to the physiological network of autoantibodies with highly connected V‐regions, capable of binding and functionally neutralizing V‐regions of natural and pathologic autoantibodies.

Reduction of immune system activation in HIV-1-infected patients undergoing highly active antiretroviral therapy

Abstract The aim of this work was to analyze the effects of highly active antiretroviral therapy on the chronically activated immune system of 26 antiretroviral-naive HIV-1-infected patients. Samples from baseline to week 24 or 36 of treatment were tested for serum levels of β2-microglobulin, tumor necrosis factor α and soluble tumor necrosis factor α receptor type II, as well as for human leukocyte antigen-DR expression on T cells. After starting therapy, the mean HIV-1 RNA serum levels decreased and the mean CD4+ cell counts increased from baseline to week 36 (P<0.001). Mean levels of tumor necrosis factor α receptor type II, tumor necrosis factor α and β2-microglobulin as well as expression of human leukocyte antigen-DR were significantly reduced at the end of follow-up (P<0.01). Deactivation kinetics of these parameters was similar in patients with CD4+ counts>200 cells/μl at baseline versus those with CD4+ counts <200 cells/μl at baseline, despite higher activation at baseline in the group with <200 cells/μl. In summary, this study shows that highly active antiretroviral therapy is able to induce a strong deactivation of the immune system of HIV-1-infected patients.

Association of human beta-defensin-2 serum levels and sepsis in preterm neonates*.

Objectives: To determine human beta-defensin-2 levels in term and preterm neonates at birth and to evaluate its impact on sepsis. Design: Observational study. Setting: Single tertiary care hospital. Patients: Term neonates and preterm neonates were recruited and divided in groups according to important clinical events. Interventions: Cord blood samples were drawn from all newborns immediately after birth. Human beta-defensin-2 levels were determined using enzyme-linked immunosorbent assay technology. All neonates were followed clinically during the first 30 days of life. Measurements and Main Results: Forty-two term and 31 preterm neonates were enrolled. Human beta-defensin-2 levels in term neonates were higher compared with preterm infants (median, 1,882 vs 918 pg/mL; p = 0.003) and correlated with gestational age and birth weight. Of 31 preterm neonates, seven suffered from late-onset sepsis, and this was associated with lower human beta-defensin-2 levels (median, 513 vs 1,411 pg/mL; p = 0.006). Conclusion: Preterm neonates show lower human beta-defensin-2 levels in cord blood compared with term neonates. Low human beta-defensin-2 levels in preterm neonates might be associated with an increased risk of late-onset sepsis.

Cellular immune responses and occult infection in seronegative heterosexual partners of chronic hepatitis C patients

Summary.  It is unknown whether hepatitis C virus (HCV)‐specific cellular immune responses can develop in seronegative sexual partners of chronically HCV‐infected patients and whether they have occult infection. Thirty‐one heterosexual partners of patients with chronic HCV were studied, fifteen of them with HCV transmission risks. Ten healthy individuals and 17 anti‐HCV seropositive patients, without viremia, were used as controls. Virus‐specific CD4+ and CD8+ T‐cell responses were measured by flow cytometry against six HCV peptides, situated within the nonstructural (NS) proteins NS3, NS4 and NS5, through intracellular detection of gamma interferon (IFN‐γ) or interleukin 4 (IL‐4) production and CD69 expression. Sexual partners had a higher production of IFN‐γ and IL‐4 by CD4+ cells against NS3‐p124 (P = 0.003), NS5b‐p257 (P = 0.005) and NS5b‐p294 (P = 0.012), and CD8+ cells against NS3‐p124 (P = 0.002), NS4b‐p177 (P = 0.001) and NS3‐p294 (P = 0.004) as compared with healthy controls. We observed elevated IFN‐γ production by CD4+ T cells against NS5b‐p257 (P = 0.042) and NS5b‐p294 (P = 0.009) in the sexual partners with HCV transmission risks (sexual, professional and familial altogether) than in those without risks. RNA was extracted from peripheral blood mononuclear cells (PBMC), and detection of HCV‐RNA positive and replicative (negative) strands was performed by strand‐specific real‐time PCR. In four sexual partners, the presence of positive and negative HCV‐ RNA strands in PBMC was confirmed. Hence, we found an HCV‐specific cellular immune response as well as occult HCV infection in seronegative and aviremic sexual partners of chronically HCV‐infected patients.