Low Expression of ICAM-1 in Blood Eosinophils in Patients With Active Eosinophilic Esophagitis.

BACKGROUND AND OBJECTIVE: Eosinophilic esophagitis (EoE) is a chronic and isolated inflammation of the esophagus characterized by a marked infiltration of eosinophilic leukocytes. Diagnosis and course of the disease are based exclusively on histopathology. Therefore, patients must undergo several esophageal biopsies, implying a risk associated with the procedure and considerable use of resources. Objective: The presence of active circulating eosinophils, which are quantifiable through the expression of specific cellular activation proteins in their membrane, could be consistent with histopathological findings, which are currently the only valid parameters in studies on EoE. METHODS: The activity of peripheral blood eosinophils from patients with EoE was analyzed by identifying 5 surface molecules (CD69, IL- 5Rα, CD44, ICAM-1, CD63), which are seen to be expressed by the active eosinophils in flow cytometry. The results were compared with the infiltrate of eosinophils present in patients' esophageal biopsies. RESULTS: ICAM-1 levels decreased significantly in patients with active EoE compared with nonactive EoE patients, allergic patients, and healthy controls. In patients with EoE, an inverse correlation was observed between the number of eosinophils in the esophageal biopsy and the percentage of ICAM-1 expression in peripheral blood eosinophils. No differences were observed for the remaining molecules studied. CONCLUSION: Expression of ICAM-1 in blood eosinophils could be a useful noninvasive marker for the diagnosis and assessment of patients with EoE.

Selective CD8 cell reduction by SARS-CoV-2 is associated with a worse prognosis and systemic inflammation in COVID-19 patients.

The lymphopenia exhibited in patients with COVID-19 has been associated with a worse prognosis in the development of the disease. To understand the factors associated with a worse evolution of COVID-19, we analyzed comorbidities, indicators of inflammation such as CRP and the ratio of neutrophils/lymphocytes, as well as the count of blood cells with T-lymphocyte subtypes in 172 hospitalized patients with COVID-19 pneumonia. Patients were grouped according to their needs for mechanical ventilation (ICU care) or not. Within the comorbidities studied, obesity was the only associated with greater severity and ICU admission. Both the percentage and the absolute number of neutrophils were higher in patients needing ICU care than non-ICU patients, whereas absolute lymphocyte count, and especially the percentage of lymphocytes, presented a deep decline in critical patients. There was no difference between the two groups of patients for CD4 T-lymphocytes, neither in percentage of lymphocyte nor in absolute number, however for CD8 T-cells the differences were significant for both parameters which were in decline in ICU patients. There was a firm correlation between the highest values of inflammation indicators with the decrease in percentage of CD8 T-lymphocytes. This effect was not seen with CD4 cells. Obesity together with lymphopenia, especially whether preferentially affects to CD8 T- lymphocytes, are factors that can predict a poor prognosis in patients with COVID-19.

Serological profile and clinical features of nucleolar antinuclear pattern in patients with systemic lupus erythematosus from southwestern Spain.

Nucleolar staining of antinuclear antibodies (ANAs) is not exclusive to patients suffering systemic sclerosis (SSc) since it can occur in other autoimmune diseases, such as systemic lupus erythematosus (SLE). The nucleolar ANA pattern presents a low incidence in patients with SLE, with less than 9% reported in some studies. The significance of nucleolar staining and antinucleolar antibodies (ANoA) in SLE is still unknown, as is its association with clinical manifestations. To address these issues, a case-control study was carried out. Twenty-eight cases of SLE with nucleolar staining were enrolled, as well as 73 controls with no nucleolar staining and different ANA patterns (homogeneous, speckled, and combined homogeneous and speckled). The homogeneous nucleolar pattern was the most frequent (27 out of 28), and in 75% was combined with other ANA patterns. The anti-double stranded DNA antibodies showed no differences between the two groups of patients, nor the auto-antibodies detected by line immunoassay (LIA). However, we have found an increased frequency of anti-PM-Scl antibodies with respect to the controls (p = 0.02), in addition to the association between Raynaud's phenomenon (RP) and anti-PM-Scl antibodies (OR = 20.72, 95% CI 1.33-323.19, p = 0.03). Moreover, the cases of SLE showed a 7.78-fold increase in the risk of developing cancer (95%, CI 1.85-32.75, p = 0.005) with respect to the control group. Taken together these findings suggest that nucleolar staining represents a comorbidity factor in patients with SLE, although its significance must still be determined.

Dopamine in human follicular fluid is associated with cellular uptake and metabolism-dependent generation of reactive oxygen species in granulosa cells: implications for physiology and pathology.

STUDY QUESTION: Is the neurotransmitter dopamine (DA) in the human ovary involved in the generation of reactive oxygen species (ROS)? SUMMARY ANSWER: Human ovarian follicular fluid contains DA, which causes the generation of ROS in cultured human granulosa cells (GCs), and alterations of DA levels in follicular fluid and DA uptake/metabolism in GCs in patients with polycystic ovary syndrome (PCOS) are linked to increased levels of ROS. WHAT IS KNOWN ALREADY: DA is an important neurotransmitter in the brain, and the metabolism of DA results in the generation of ROS. DA was detected in human ovarian homogenates, but whether it is present in follicular fluid and plays a role in the follicle is not known. STUDY DESIGN, SIZE AND DURATION: We used human follicular fluid from patients undergoing in vitro fertilization (IVF), GCs from patients with or without PCOS and also employed mathematical modeling to investigate the presence of DA and its effects on ROS. PARTICIPANTS/MATERIALS, SETTING AND METHODS: DA in follicular fluid and GCs was determined by enzyme-linked immunosorbent assay. GC viability, apoptosis and generation of ROS were monitored in GCs upon addition of DA. Inhibitors of DA uptake and metabolism, an antioxidant and DA receptor agonists, were used to study cellular uptake and the mechanism of DA-induced ROS generation. Human GCs were examined for the presence and abundance of transcripts of the DA transporter (DAT; SLC6A3), the DA-metabolizing enzymes monoamine oxidases A/B (MAO-A/B) and catechol-O-methyltransferase and the vesicular monoamine transporter. A computational model was developed to describe and predict DA-induced ROS generation in human GCs. MAIN RESULTS AND ROLE OF CHANCE: We found DA in follicular fluid of ovulatory follicles of the human ovary and in GCs. DAT and MAO-A/B, which are expressed by GCs, are prerequisites for a DA receptor-independent generation of ROS in GCs. Blockers of DAT and MAO-A/B, as well as an antioxidant, prevented the generation of ROS (P < 0.05). Agonists of DA receptors (D1 and D2) did not induce ROS. DA, in the concentration range found in follicular fluid, did not induce apoptosis of cultured GCs. Computational modeling suggested, however, that ROS levels in GCs depend on the concentrations of DA and on the cellular uptake and metabolism. In PCOS-derived follicular fluid, the levels of DA were higher (P < 0.05) in GCs, the transcript levels of DAT and MAO-A/B in GCs were 2-fold higher (P < 0.05) and the DA-induced ROS levels were found to be more than 4-fold increased (P < 0.05) compared with non-PCOS cells. Furthermore, DA at a high concentration induced apoptosis in PCOS-derived GCs. LIMITATIONS, REASONS FOR CAUTION: While the results in IVF-derived follicular fluid and in GCs reveal for the first time the presence of DA in the human follicular compartment, functions of DA could only be studied in IVF-derived GCs, which can be viewed as a cellular model for the periovulatory follicular phase. The full functional importance of DA-induced ROS in small follicles and other compartments of the ovary, especially in PCOS samples, remains to be shown. WIDER IMPLICATIONS OF THE FINDINGS: The results identify DA as a factor in the human ovary, which, via ROS generation, could play a role in ovarian physiology and pathology. The results obtained in samples from women with PCOS suggest the involvement of DA, acting via ROS, in this condition. STUDY FUNDING/COMPETING INTERESTS: This work was supported by a grant from DFG MA1080/17-3 and in part MA1080/19-1. There are no competing interests.

Immunotherapy reduces CD40L expression and modifies cytokine production in the CD4 cells of pollen allergy patients.

BACKGROUND: Allergen-specific immunotherapy (SIT) is the only intervention for IgE-mediated respiratory disorders. OBJECTIVE: The aim of the study was to investigate the immunological modifications induced by SIT in patients allergic to olive and/or grass pollen by attempting to establish an association between these modifications and clinical improvements. METHODS: We studied 29 patients who were allergic to olive and/or grass pollen. Patients were randomized to 2 groups: an active treatment group, comprising 19 allergic patients who received SIT, and a control group, formed by 10 allergic patients who received pharmacological treatment for their allergic symptoms but not immunotherapy. We used flow cytometry to analyze intracellular expression of the cytokines IL-4, IFN-gamma, IL-10, and TGF-beta1 in CD4+ T cells, as well as expression of Foxp3, the costimulatory CTLA-4 molecule, and the non-costimulatory CD40L molecule. To assess clinical changes, patients recorded their medication consumption, symptoms, and the limitation of daily activities using diary cards and quality of life questionnaires. RESULTS: Six months after initiation of SIT, we recorded a reduction in cell surface CD40L expression in the CD4+ T-cell population and a shift in the cytokine production profile (decrease in IL-4-producing CD4+ T cells and increase in IFN-gamma, IL-10, and TGF-beta1). These changes persisted after 12 months. Simultaneously, a clinical improvement was observed. CONCLUSIONS: SIT-induced clinical improvement is the result of immunological modifications such as a reduction in CD40L expression on CD4 cells and alteration in the cytokine production profile.

Differential expression of CD30 on CD3 T lymphocytes in patients with systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is an autoimmune systemic disease caused as a result of an imbalance of Th1-/Th2-type cytokines. The soluble form of CD30 (CD30s) released from peripheral blood cells has been described as a marker of active disease in Th2-type immune response as in SLE. However, the expression of CD30 on CD3 T lymphocytes from patients with SLE has not been studied yet. Therefore, we have addressed our study to attempt this issue, studying CD30 expression by flow cytometry on CD3 T lymphocytes and CD4/CD8 subsets in samples from SLE patients mainly with lupus nephritis. In parallel, we have determined the production of the cytokines IL-4 (Th2), IFNγ (Th1), IL-10 and TGFß by intracellular staining. Differences between positive CD30 T cells in healthy controls and patients with SLE were found, with a higher percentage of CD30-expressing T cells in patients with SLE (P = 0.001). In contrast to healthy controls, CD30 was mainly expressed on CD8 T cells from patients with SLE. The intracellular cytokine staining showed that TGFß is the main cytokine expressed in CD3 T cells from patients with SLE. In addition to this, we have found a positive correlation between CD30-expressing T cells and IL-4, IFNγ, and immunosuppressive cytokines (IL-10 and TGFß) (P < 0.05). These results suggest that CD30 could play a role in the pathogenesis of SLE and its expression on CD3 T lymphocytes is not restricted only to Th2-type response.

Partial recovery of SARS-CoV-2 immunity after booster vaccination in renal transplant recipients.

The pandemic caused by the SARS-CoV-2 coronavirus has been especially detrimental to patients with end-stage renal disease. History with other vaccines suggests that patients with renal disease may not respond adequately to the SARS-CoV-2 vaccine. The aim of this study is to evaluate the immunity to SARS-CoV-2 mRNA vaccines in renal patients. Post SARS-CoV-2 vaccination first, and after the booster dose, antibodies and cellular immunity were studied in patients on hemodialysis (N = 20), peritoneal dialysis (N = 10) and renal transplantation (N = 10). After the two doses of vaccine, there was an effective immunity in dialysis patients, with 100% seroconversion and 87% detection of cellular immunity (85% in hemodialysis and 90% in peritoneal dialysis). In contrast, in renal transplant recipients there was only 50% seroconversion and cellular immunity was detected in 30% of patients. After the booster dose, all dialysis patients achieved a cellular and antibody immunity, whereas in transplant patients, despite improvement, 20% did not produce antibodies and in 37.5% cellular immunity could not be detected. The mRNA vaccine plus booster performs excellently in dialysis patients, whereas in kidney transplant recipients, despite the booster, complete immunization is not achieved.

Basophil activation test as alternative method to CAP-inhibition in patients with double sensitization to vespid venoms.

Most patients with Hymenoptera venom allergy (HVA) to vespid venoms present double sensitization by specific IgE (sIgE)-mediated cross-reactivity. Thus, it is mandatory could discriminate between a true double and primary sensitization to implement an accurate venom-specific immunotherapy (VIT). To date, CAP-inhibition is the reference method in the diagnosis of cross-reactivity in double sensitized patients to vespid venoms, being the results obtained with the component resolved diagnostics (CRD) conflicting. For this, we have studied in a cohort of double sensitized patients to Vespula vulgaris (VV) and Polistes dominulus (PD) venoms (n = 40) the diagnostic accuracy of CRD using the CAP-inhibition as reference method, as well as to investigate whether basophil activation test (BAT) is an alternative method for inconclusive results obtained by CAP-inhibition. CAP-inhibition showed a sensitivity of 59.46 % in view of the indeterminate results; most patients had true double sensitization (54.5 %), followed by single sensitization to PD (27.27 %) and VV (18.18 %) venoms. CRD based on rVes v 5/rPol d 5 (or vice versa) ratio as well as whole extracts I3/I77 (or vice versa) ratio (specific IgE-I3 to VV/specific IgE-I77 to PD) showed a low diagnostic accuracy (AUC = 0.504, p = 0.974; AUC = 0.35, p = 0.235; respectively). BAT was determined in parallel with CAP-inhibition in 12 patients, presented higher sensitivity than CAP-inhibition (p = 0.021) and a positive agreement of 71.43 %. Likewise it was able to identify 100% of inconclusive results, showing a specificity of 83.3 %. Therefore, CRD is not a suitable method to distinguish monosensitization and BAT appears to be an appropriate method resolving indeterminate results from the gold standard method.

Oral rush desensitization to egg: efficacy and safety.

BACKGROUND: Current management of egg allergy relies on egg elimination from the diet. It does not protect patients from reactions after accidental ingestion of the food and it has a negative influence on quality of life. To solve these problems, some desensitization protocols have been described that are safe and effective, but only one study of a rush regimen for egg with a small patient sample has been published. OBJECTIVE: To evaluate the safety, efficacy and immunologic effects of an oral rush desensitization protocol for immediate egg allergy. METHODS: Subjects aged 5 years or older with symptomatic IgE-mediated allergy to hen's egg underwent a 5-day oral tolerance induction regimen and were subsequently maintained on a regular egg intake. The variables studied were the reactions that occurred during the induction regimen and follow-up and the duration of desensitization. Prick test weal size and egg white-specific IgE and IgG concentrations were monitored. RESULTS: Twenty-three patients between 5 and 17 years of age entered the protocol. Twenty (86.9%) achieved the daily intake of a whole cooked egg, 14 of them within the scheduled 5 days. One abandoned the protocol and two were changed to a slower regimen because of repeated reactions. Allergic reactions were frequent but in general were mild. No severe reactions occurred. During follow-up of at least 6 months, egg was well tolerated by all patients. Compared with baseline, skin prick test weal size and egg white-sIgE levels had fallen at 3 months, although the differences were only significant at 6 months. CONCLUSIONS AND CLINICAL RELEVANCE: The rush protocol described is useful and safe for achieving tolerance to egg within a few days but it should always be performed in a highly supervised setting. A high proportion of patients allergic to egg can effectively be desensitized using the described schedule, with the advantage of shortening the time to become protected from reactions after inadvertent ingestion of egg, with no increase in the risk compared with the earlier reported slower protocols.

Decreased Expression of the CD57 Molecule in T Lymphocytes of Patients with Chronic Fatigue Syndrome.

The chronic fatigue syndrome (CFS) is characterized by a prolonged incapacitating fatigue, headaches, sleep disturbances, and decreases in cognition, besides alterations in other physiological functions. At present, no specific biological markers have been described in this pathology. In the present study, we analyzed in lymphocytes the CD57 expression for the diagnosis of CFS, evaluating both the percentage of blood lymphocytes expressing CD57 and the average amount of the molecule expressed per cell. The study demonstrated a marked and significant decrease in the expression of CD57 in lymphocytes of CFS patients regarding healthy controls. In T lymphocytes, the decrease was significant both in the percentage of cells expressing CD57 (7.5 ± 1.2 vs 13.3 ± 1.6, p = 0.024) and in a more relevant way in the amount of CD57 molecule expressed per cell (331 ± 59 vs 1003 ± 104, p ≤ 0.0001). In non-T lymphocytes, the decrease was significant only in the amount of CD57 expressed per cell (379 ± 114 vs 691 ± 95, p = 0.007). The study of CD57 antigen in blood lymphocytes is a useful marker that could cooperate in the diagnosis of CFS patients. Its decrease in T lymphocytes provides most valuable results than the results in other lymphocyte subpopulations.

Analysis of the methodology to determine anaerobic toxicity: evaluation of main compounds present in wastewater treatment plants (WWTPs).

The influence of the concentration of biomass on the level of inhibition and anaerobic degradation kinetics in batch systems was studied with toxic compounds that can generate destabilization in the operation of sludge anaerobic digesters. The compounds were grouped in four families; long chain fatty acids, polycyclic aromatic hydrocarbons, linear alkylbenzene sulphonates and organic acids. For the organic acids, there is no effect due to the biomass concentration variation, therefore it is a competitive inhibition; but that doesn't happen with the remaining compounds, where there is a dependence on the complexity of their structure, becoming a non-competitive inhibition. In addition, it was observed that the degradation kinetics is affected, whether diminishing the methane production (polycyclic aromatic hydrocarbons, linear alkylbenzene sulphonates, organics acids) or increasing the initial latency time (long chain fatty acids) without this becoming an obstacle to obtain the maximum methane productions for the latter ones.

Low Expression of ICAM-1 in Blood Eosinophils in Patients With Active Eosinophilic Esophagitis

Background: Eosinophilic esophagitis (EoE) is a chronic and isolated inflammation of the esophagus characterized by a marked infiltration of eosinophilic leukocytes. Diagnosis and course of the disease are based exclusively on histopathology. Therefore, patients must undergo several esophageal biopsies, implying a risk associated with the procedure and considerable use of resources. Objective: The presence of active circulating eosinophils, which are quantifiable through the expression of specific cellular activation proteins in their membrane, could be consistent with histopathological findings, which are currently the only valid parameters in studies on EoE. Methods: The activity of peripheral blood eosinophils from patients with EoE was analyzed by identifying 5 surface molecules (CD69, IL- 5Rα, CD44, ICAM-1, CD63), which are seen to be expressed by the active eosinophils in flow cytometry. The results were compared with the infiltrate of eosinophils present in patients’ esophageal biopsies. Results: ICAM-1 levels decreased significantly in patients with active EoE compared with nonactive EoE patients, allergic patients, and healthy controls. In patients with EoE, an inverse correlation was observed between the number of eosinophils in the esophageal biopsy and the percentage of ICAM-1 expression in peripheral blood eosinophils. No differences were observed for the remaining molecules studied. Conclusion: Expression of ICAM-1 in blood eosinophils could be a useful noninvasive marker for the diagnosis and assessment of patients with EoE (AU)

[Introduction of an accreditation system for hospital informed consent forms]. / Introducción de un sistema de acreditación para los documentos de consentimiento informado hospitalario.

OBJECTIVE: To describe an accreditation system for informed consent forms (ICF) in a tertiary hospital, as an intervention to improve their quality, and to check the improvements achieved. MATERIAL AND METHODS: Following an external evaluation of the ICF quality in a public hospital in Murcia (Spain), an accreditation committee set the ICF requirements and associated procedures. Effectiveness is assessed by comparing two external evaluations carried out by the EMCA Program (2011 and 2013) and based on 19 criteria and a sample of 60 ICF for every public hospital in Murcia Region. RESULTS: To be accredited, every ICF must meet the 19 external criteria plus 5 based on legibility, readability and scientific and technical validity. A form to fill in the contents of every ICF was agreed, which would be reviewed, approved and validated for five years. Before the implementation, 8.2 defects/ICF were detected. The accreditation system obtained an 89% improvement (0.9 defects/ICF) and achieved significant improvements in 18 criteria, 16 of which are benchmarked. DISCUSSION: The accreditation system achieved a substantial improvement in the ICF (obtaining a better result in external evaluations) and guarantees their contents, legibility and readability. This system needs to be extended to other hospitals, since it is not clear whether common ICFs would be suitable. However, this improvement is structural and does not guarantee that the overall information/consent procedure is done properly, thus complementary strategies for measurement and improvement are required.

DNA analysis of HLA-DR4B1 subtypes in multiple sclerosis by specific oligonucleotide probes.

Conversely to the well-established association of DR2/Dw2 with multiple sclerosis (MS) susceptibility in Caucasoids, several studies have found an association of DR4 in populations from Mediterranean origin. We have studied the distribution of the different DR4B1 subtypes in Spanish MS patients. Oligonucleotide probes were selected in order to type samples amplified by polymerase chain reaction (PCR) from Spanish DR4+ MS patients (25) and controls (28). No DR4B1 subtypes were found to be increased in MS. MS susceptibility linked to DR4 may be due to the presence of shared functional epitopes common to the different HLA-DR4B1 subtypes.

Germline repertoire of T-cell receptor beta-chain genes in patients with insulin-dependent diabetes mellitus.

We have investigated the genotype and allelic distribution of germline restriction fragment length polymorphisms of the T-cell receptor beta chain, segment C beta, and two variable segments which are in linkage disequilibrium, V beta 8 and V beta 11, in 42 insulin-dependent diabetes mellitus (IDDM) patients and in 51 healthy blood donors used as controls. Recently, several works have reported contradictory results showing or not showing an association between polymorphic alleles of the C beta gene and diabetes type I. We found no significant differences in the allele, genotype, and haplotype distribution of the gene segments studied, between IDDM patients and control populations.

A chimeric anti-TNFalpha monoclonal antibody (cA2) in vivo removes TNFalpha-producing cells in Crohn's disease.

Tumor necrosis factor alpha (TNFalpha) has been described as a citokine involved in gastrointestinal mucosal inflammation in Crohn's disease. A single infusion of the chimeric mouse/human monoclonal antibody cA2 anti-TNFalpha has been established as a new therapeutical procedure. The aim of these study was to determine the effect of the monoclonal antibody cA2 on lymphocyte and monocyte TNFalpha-producing cells. Initially the patient, with severe Crohn's disease (Crohn's disease activity index CDAI > 350), presented a higher number of peripheral blood TNFalpha-producing cells than healthy controls. The patient received two cA2 treatments throughout one year due the severe activity of the disease. Before treatment the patient had a large number of TNFalpha producer cells. A dramatic reduction in lymphocyte and monocyte TNFalpha producing cells, together a clinical remission (CDAI < 150), was shown after the treatments. Four months after the first cA2 treatment, the patient had a clinical response associated with an important increment of TNFalpha-producing cells that extended increasing until the second cA2 treatment was averaged. These results suggest that the clinical activity of the Crohn's disease correlates with peripheral TNFalpha-expressing cells. The cA2 antibody, as well as of neutralize soluble TNFalpha, also removes TNFalpha-producer cells, which may collaborate with the anti-TNFalpha activity of the antibody treatment.

[Formal quality assessment of informed consent documents in 9 hospitals]. / Evaluación de la calidad formal de los documentos de consentimiento informado en 9 hospitales.

INTRODUCTION: Informed consent forms are very important in the process of medical information. The aim of this study is to design reliable formal quality criteria of these documents and their application in the evaluation of those used in the hospitals of a regional health service. MATERIAL AND METHODS: Criteria have been designed from the analysis of existing regulations, previous studies and consultation with key experts. The interobserver concordance was assessed using the kappa index. Criteria evaluation was performed on 1425 documents of 9 hospitals. RESULTS: A total of 19 criteria used in the evaluation of the quality of informed consent forms have been obtained. Kappa values were higher than 0,60 in 17 of them and higher than 0,52 in the other 2. The average number of defects per document was 7.6, with a high-low ratio among hospitals of 1.84. More than 90% of the documents had defects in the information on consequences and contraindications, and in about 90% it did not mention the copy to the patient. More than 60% did not comply with stating the purpose of the procedure, a statement of having understood and clarified doubts, and the treatment options. CONCLUSIONS: A tool has been obtained to reliably assess the formal quality of the informed consent forms. The documents assessed have a wide margin for improvement related to giving a copy to the patient, and some aspects of the specific information that patients should receive.