Anti-Anisakis antibodies in colon cancer patients and their relationship with γδ T-cells.

Many pathogens are related to carcinogenesis. Chronic inflammation, as a result of persistent infection, leads to DNA damage, higher expression of oncogenes, decreased apoptosis and immunosuppression, which are some of the reasons for cancer induction. Among parasites, Schistosoma, Opistorchis and Clonorchis are recognised as infectious agents which contribute to cancer. A relationship between Anisakis and cancer was hypothesised because cellular responses to Anisakis products could result in inflammation and DNA damage. Previous research has shown a decrease in CD8+ γδ T-cells and an increase in αß and γδ T-cell apoptosis in colon cancer (CC) samples. Ninety-two CC patients and 60 healthy subjects were recruited. γδ and αß T-cells were analysed, and their apoptosis was evaluated. Anti-Anisakis antibodies were tested in sera from CC patients and controls. Anti-Anisakis IgG, IgM, IgA and IgE antibodies were significantly higher in CC patients. A significant increase in anti-Anisakis IgA levels was observed in patients with angiolymphatic invasion. The number of all γδ T-cells, as well as CD3+ CD4+ αß T-cells, was significantly lower in CC patients. The apoptosis of all T-cells was significantly increased in patients with CC. We observed a significantly higher percentage of anti-Anisakis IgE positive patients having a deficit of CD3+ γδ T-cells. Our results suggest a relationship between Anisakis and CC.

Expansion of T regulatory lymphocytes by murine bone marrow dendritic cells previously stimulated with Anisakis simplex larval antigens.

BACKGROUND: Anisakis simplex antigens present immunomodulatory properties by the induction of tolerogenic dendritic cells (DCs) in mice. OBJECTIVES: To study the capacity of DCs stimulated with A. simplex excretory-secretory (ES) or crude extract (CE) to generate Tregs. To investigate in vitro effects of antigens on the metabolic activity of splenocytes induced by LPS or CpG. METHODS: Phenotypic and functional characterization of T cells co-cultured with A. simplex-pulsed DCs was performed by flow cytometry. Lymphocyte mitochondrial respiratory activity was estimated by the Alamar Blue® Assay. FINDINGS: In C57BL/6J, CD4+CD25-Foxp3+ and CD8+CD25-Foxp3+ populations increased by CE-stimulated-DCs. In BALB/c, CE-stimulated-DCs caused the expansion of CD4+CD25+Foxp3+IL-10+ and CD8+CD25+Foxp3+IL-10+. IFN-γ expression raised in BALB/c CD4+CD25+ and CD4+CD25- for CE and ES, respectively. ES-stimulated-DCs increased CD4+CD25+ Foxp3+ and CD8+CD25- Foxp3+ expression in T cells. The association of ES or CE with LPS produced the increase in splenocyte activity in C57BL/6J. The association of CE with CpG decreased the proliferation caused by CpG in C57BL/6J. MAIN CONCLUSIONS: A. simplex increase the frequency of Tregs, which in turn produce IL-10 and IFN-γ. The host genetic base is essential in the development of anti-Anisakis immune responses (Th2, Th1, Treg).

Corticosteroids for COVID-19 patients requiring oxygen support? Yes, but not for everyone: Effect of corticosteroids on mortality and intensive care unit admission in patients with COVID-19 according to patients' oxygen requirements.

Corticosteroids reduce mortality in hospitalized patients with coronavirus disease 2019 (COVID-19), but the response seems to vary according to the level of respiratory support needed. This retrospective cohort study included COVID-19 patients with oxygen saturation (SatO2 ) in room air <92% admitted between March 3 and April 30, 2020. Following the interim protocol, patients could receive dexamethasone or methylprednisolone, and were classified according to oxygen requirements. The primary endpoint was admission to the intensive care unit (ICU) or mortality. Kaplan-Meier and Cox hazards analyses were used. Of the 115 patients included, 38 received corticosteroids. Among requiring high-flow, noninvasive ventilation (NIV) or fraction of inspired oxygen (FiO2 ) > 0.40, the hazard ratio (HR) for death or ICU admission, between the corticosteroids and non-corticosteroids group, was 0.07 (95% CI 0.01-0.4), p = .002, and for patients requiring low-flow oxygen, the HR was 0.70 (95% CI 0.13-3.8), p = .68. Significant differences were also observed when all patients were analyzed together. A significant reduction in mortality and ICU admission frequency was observed among patients requiring high-flow oxygen or NIV, but not among those requiring low-flow oxygen. Better targeting of COVID-19 patients is needed for the beneficial use of corticosteroids.

Expansion of T regulatory lymphocytes by murine bone marrow dendritic cells previously stimulated with Anisakis simplex larval antigens

BACKGROUND Anisakis simplex antigens present immunomodulatory properties by the induction of tolerogenic dendritic cells (DCs) in mice. OBJECTIVES To study the capacity of DCs stimulated with A. simplex excretory-secretory (ES) or crude extract (CE) to generate Tregs. To investigate in vitro effects of antigens on the metabolic activity of splenocytes induced by LPS or CpG. METHODS Phenotypic and functional characterization of T cells co-cultured with A. simplex-pulsed DCs was performed by flow cytometry. Lymphocyte mitochondrial respiratory activity was estimated by the Alamar Blue® Assay. FINDINGS In C57BL/6J, CD4+CD25-Foxp3+ and CD8+CD25-Foxp3+ populations increased by CE-stimulated-DCs. In BALB/c, CE-stimulated-DCs caused the expansion of CD4+CD25+Foxp3+IL-10+ and CD8+CD25+Foxp3+IL-10+. IFN-γ expression raised in BALB/c CD4+CD25+ and CD4+CD25- for CE and ES, respectively. ES-stimulated-DCs increased CD4+CD25+ Foxp3+ and CD8+CD25- Foxp3+ expression in T cells. The association of ES or CE with LPS produced the increase in splenocyte activity in C57BL/6J. The association of CE with CpG decreased the proliferation caused by CpG in C57BL/6J. MAIN CONCLUSIONS A. simplex increase the frequency of Tregs, which in turn produce IL-10 and IFN-γ. The host genetic base is essential in the development of anti-Anisakis immune responses (Th2, Th1, Treg).

Anti-Anisakis sp. antibodies in serum of patients with sepsis and their relationship with γδ T cells and disease severity.

Immunosuppression in sepsis reduces both αß and γδ T cell subsets. Anisakis sp. is a parasitic nematode with a high prevalence in Spain. Previous contact with the parasite is related to a decrease in γδ T cells. Anti-Anisakis antibodies were measured and related to αß and γδ T cells in 114 septic patients versus 97 healthy controls. Significant differences were seen with respect to the groups with severe sepsis and septic shock where lower anti-Anisakis levels were observed. A similar decrease appeared in the case of specific IgM with significant differences between the groups of control/uncomplicated sepsis versus severe sepsis and septic shock. These differences were also apparent in the case of specific IgA. The lowest IgE levels were detected in the septic shock group. Anti-Anisakis IgG levels significantly increased in septic shock groups compared with the controls. We observed positive correlations among anti-Anisakis IgA levels and all γδ T cell subsets. There were negative correlations among IgA levels and APACHE and SOFA indices. Greater contact with the parasite (IgG) was directly related with septic shock, inflammation and markers of sepsis severity. A lack of protection in the mucosa (IgA and γδ T cells) was associated with the disease severity.

Anti-Anisakis sp. antibodies in serum of healthy subjects. Relationship with αß and γδ T cells.

Anisakiosis is nowadays one of the nematodoses more prevalent in Spain, with rates that oscillate between 0.43% in Galicia (N.W. Spain), and 15.7% and 22.1% in inland and southern regions, respectively. Likewise, it has been proved that Anisakis larvae have developed mechanisms to modulate the dichotomy of the host immune response for their own benefit. The experimental hypothesis of the present study was that Anisakis sp. larval products can be mediators of immune suppression and induce changes on the populations of αß+ and γδ+ T cells. In the present study we determined the levels of anti-Anisakis antibodies in the serum of healthy people, and their relationship with the B and T cell subsets. Levels of anti-Anisakis antibodies (Ig's, IgG, IgM, IgA and IgE) were measured by ELISA, while B and T cell subsets were studied by flow cytometry. Cells were labelled with monoclonal antibodies against CD45, CD4, CD8, CD56, CD3, CD19, TCRαß and TCRγδ. All the specific isotypes studied were negatively correlated with NKT cell rates with the exception of IgG. A previous contact with Anisakis was related to a decrease in CD56+αß+ and all γδ+ T cell subsets. The CD3+γδ+ population was lower in the group of subjects that showed IgA anti-Anisakis. We observed an inverse correlation among αß-γδ NKT cells and anti-Anisakis sp. antibodies. CD3+CD56+ cells showed a significant decrease in the group of anti-Anisakis positive subjects. This fact was especially significant with CD3+CD56+γδ+ cells in the case of the anti-Anisakis IgA positive group.

Epstein-Barr virus is related with 5-aminosalicylic acid, tonsillectomy, and CD19(+) cells in Crohn's disease.

AIM: To study anti-Epstein-Barr virus (EBV) IgG antibodies in Crohn's disease in relation to treatment, immune cells, and prior tonsillectomy/appendectomy. METHODS: This study included 36 CD patients and 36 healthy individuals (controls), and evaluated different clinical scenarios (new patient, remission and active disease), previous mucosa-associated lymphoid tissue removal (tonsillectomy and appendectomy) and therapeutic regimens (5-aminosalicylic acid, azathioprine, anti-tumor necrosis factor, antibiotics, and corticosteroids). T and B cells subsets in peripheral blood were analyzed by flow cytometry (markers included: CD45, CD4, CD8, CD3, CD19, CD56, CD2, CD3, TCRαß and TCRγδ) to relate with the levels of anti-EBV IgG antibodies, determined by enzyme-linked immunosorbent assay. RESULTS: The lowest anti-EBV IgG levels were observed in the group of patients that were not in a specific treatment (95.4 ± 53.9 U/mL vs 131.5 ± 46.2 U/mL, P = 0.038). The patients that were treated with 5-aminosalicylic acid showed the highest anti-EBV IgG values (144.3 U/mL vs 102.6 U/mL, P = 0.045). CD19(+) cells had the largest decrease in the group of CD patients that received treatment (138.6 vs 223.9, P = 0.022). The analysis of anti-EBV IgG with respect to the presence or absence of tonsillectomy showed the highest values in the tonsillectomy group of CD patients (169.2 ± 20.7 U/mL vs 106.1 ± 50.3 U/mL, P = 0.002). However, in the group of healthy controls, no differences were seen between those who had been tonsillectomized and subjects who had not been operated on (134.0 ± 52.5 U/mL vs 127.7 ± 48.1 U/mL, P = 0.523). CONCLUSION: High anti-EBV IgG levels in CD are associated with 5-aminosalicylic acid treatment, tonsillectomy, and decrease of CD19(+) cells.

Deficit of interleukin 7 in septic patients.

We recently demonstrated an overall decrease of all αß and specially γδ T cell subsets in patients with sepsis compared with healthy subjects. IL-7 is a crucial factor for development of γδ T cells and survival in sepsis but its association with sepsis severity, evolution of organ failure and death still has not been investigated. Sera from 78 patients who met criteria for sepsis were analyzed vs control group. Septic patients showed the lowest levels of IL-7. Patients with severe sepsis reached levels of IL-7 higher than those observed in the groups of uncomplicated sepsis and septic shock. The frequency of γδ T cells at admission was lower in septic patients vs control group. At the time of admission, the frequency of γδ T cells in septic patients who subsequently died was lower than the observed in the group of patients that instead survived.

Microsporidia and its relation to Crohn's disease. A retrospective study.

BACKGROUND: The cause of Crohn's Disease (CD) remains unknown. Recently a decrease in the global lymphocyte population in the peripheral blood of CD patients has been reported. This decrease was more evident in γδ T lymphocytes, especially γδ CD8+T subsets. Furthermore, a decrease of IL-7 was also observed in these patients. We propose the hypothesis that microsporidia, an obligate intracellular opportunistic parasite recently related to fungi, in CD patients can take advantage of the lymphocytes and IL-7 deficits to proliferate and to contribute to the pathophysiology of this disease. METHODS AND FINDINGS: In this case-control study, serum samples were collected from 36 CD patients and from 36 healthy individuals (controls), IgE and IgG anti-Encephalitozoon antibodies were determined by ELISA; and forty-four intestinal tissue samples were analyzed through real time Polymerase Chain Reaction (PCR), twenty CD patients, nine with others diseases and 15 healthy subjects. We observed that IgE anti-Encephalitozoon levels were significantly higher in patients with CD: 0.386(±0.256) vs control group, 0.201(±0.147), P<0.001. However, IgG anti-Encephalitozoon values were significantly lower in CD patients: 0.361(±0.256) vs control group, 0.876(±0.380), P<0.001. In the group of CD patients, 6/20 (30%) were positive by real time PCR for microsporidia and, all the patients of the control group were negative by real time PCR. CONCLUSIONS: These results suggest that CD patients are a group at risk for microsporidiasis and, moreover that microsporidia may be involved as a possible etiologic factor of CD.

Association of γδ T cells with disease severity and mortality in septic patients.

Gamma-delta T cells are the most abundant of all epithelial-resident lymphocytes and are considered a first line of defense against pathogens in the mucosa. Our objective was to confirm the reduction in γδ T cell subsets and its relationship with mortality in patients with sepsis. We studied 135 patients with sepsis attended in the emergency department and intensive care unit of two hospitals and compared them with a similar control group of healthy subjects. The αß and γδ T cell subsets were determined via flow cytometry according to the stage of the sepsis and its relationship with mortality. All the lymphocyte subsets were reduced with respect to the corresponding subsets in the control group. All the γδ T cell populations decreased significantly as the septic picture worsened. Furthermore, γδ T cells showed decreases at days 2, 3, and 4 from the start of sepsis. Twenty-six patients with sepsis died (19.3%). The γδ T cells, specifically, the CD3(+) CD56(+) γδ T cells, were significantly reduced in those septic patients who died. Our results indicate that, during sepsis, γδ T cells show the largest decrease and this reduction becomes more intense when the septic process becomes more severe. Mortality was associated with a significant decrease in γδ T cells.