Tolerance in TCR/Cognate Antigen Double-Transgenic Mice Mediated by Incomplete Thymic Deletion and Peripheral Receptor Downregulation

Influenza nucleoprotein (NP)-specific T-cell receptor transgenic mice (F5) were crossed with transgenic mice expressing the cognate antigenic protein under the control of the H- 2K^b promoter. Double-transgenic mice show negative selection of thymocytes at the CD4⁺8⁺TCR¹⁰ to CD4⁺8⁺TCR^hi transition stage. A few CD8 T cells, however, escape clonal deletion, and in the peripheral lymphoid organs of these mice, they exhibit low levels of the transgenic receptor and upregulated levels of the CD44 memory marker. Such cells do not proliferate upon exposure to antigen stimulation in vivo or ex vivo, however, they can develop low but detectable levels of antigen-specific cytotoxic function after stimulation in vitro in the presence of IL-2.     

In vitro study on cytotoxic effect of some chemicals on serum McCoy and serum-free McCoy-Plovdiv cell culture systems.

The cytotoxicity of test agents on serum-free McCoy cultures has not been studied at all. The cytotoxic effect of EDTA, methanol, DMSO, and cycloheximide on serum-free McCoy-Plovdiv cell culture (SF) was detected visually on inverted microscope and quantitatively by tests for viability (NR) and total protein (KBP). The IC50 values for the tested chemicals were calculated. SF showed the lowest IC50 values for cycloheximide, DMSO and EDTA and the highest for methanol according to both tests. EDTA, methanol, DMSO and cycloheximide had dose-effect relationship in the cell test systems after treatment. The data indicate that McCoy-Plovdiv cell line is a suitable serum-free cell system for in vitro cytotoxic studies.     

Immunopathological characteristics of cryptoglandular and Crohn's anorectal fistulas

Anorectal fistulas are quite common proctologic disorders. They can be either of cryptoglandular origin or can be associated with Crohn's disease and chronic ileocolitis. Mechanical obstruction and local infections are prime causes of this pathological condition. Genetic predisposition and inadequate immune response with overproduction of pro-inflammatory cytokines appear prominently in the course of Crohn's disease. Interferon-gamma, a Th1 type cytokine, reflecting the engagement of cellular immune mechanisms, is the first to be produced in the intestinal mucosa. The inflammatory process in the colon mucosa induced by the abundant microbial flora is sustained and turned chronic by the gradual elevation of the local TNF-a and regulatory cytokines levels (interleukin-10, transforming growth factor-beta). The number of activated local memory T cells CD4+CD45RBl0 increases significantly. The regulatory CD4+CD25+ T lymphocytes producing interleukin-10 increase also trying to counterbalance the cytokine reaction. The chronic inflammatory infiltrates of the colon mucosa are represented by lymphocytes, plasma cells, macrophages. The long-term activation of macrophages by the released interferon-gamma leads to tissue damage and potentiation of angiogenesis--a risk factor for carcinoma development. Management of anorectal abscesses and fistulas is complex aiming to alleviate the symptoms, prevent relapses, reduce the risk of sphincter damage and improve quality of life. The main approach (surgery) should be combined with antimicrobial infection control and immunomodulation by intravenous or local administration of anti-TNF-alpha antibodies.     

Quantitative system for diagnosis of vulvovaginal candidiasis

IntroductionVaginal infections are one of the most common reason for gynecological consultations. Many of them are the result of overgrowth of resident microorganisms. The clinical symptoms of vulvovaginal candidiasis are nonspecific and an accurate diagnosis is a problem that often leads to inadequate treatment or delays in treatment. The lack of an exact and practical diagnostic method is a common cause of misdiagnosis.AimTo create a complex, quantitative method for the diagnosis of vulvovaginal candidiasis which to enables differentiation from vaginal fungal colonization.Material and methodsA total of 2306 vaginal samples were examined. Clinical, microbiological, epidemiological methods and statistical models are used.Results and discussionThe proposed score system is a specific, sensitive and inexpensive method to routinely diagnose vulvovaginal candidiasis. Statistical processing of the obtained data shows the impact of the individual components on which the method is based: the presence of vaginal discharge, pruritus, direct microscopy and assessment of the fungal growth. The data analysis reveals good sensitivity (71%) and high specificity (98%) of the method. This allows accurate interpretation of the result of the clinical and microbiological examination of each patient.ConclusionThe system for diagnosing vulvovaginal candidiasis is complex and based on quantitative indicators. The method can be used to differentiate vulvovaginal candidiasis from vaginal fungal colonization (the cut-off value is 5.5 points) and to more accurately interpret a Candida positive result from quantitative real-time PCR in asymptomatic patients or in women with mixed vaginal infection.     

Inhibition of thymocyte negative selection by T cell receptor antagonist peptides

The T cell receptor (TCR) recognizes antigenic peptide presented by major histocompatibility complex (MHC) molecules. Analogs of antigenic peptides have been shown to inhibit antigen-specific T cell responses, a phenomenon described as TCR antagonism. We have examined the effect of a natural variant of an antigenic peptide and a synthetic peptide analog, on the responses of mature T cells and immature thymocytes from an αβ TCR-transgenic mouse (F5), the TCR of which recognizes a nonamer peptide from the nucleoprotein (NP) of influenza virus in the context of the H-2D^b MHC molecule. Both peptides were shown to antagonize specifically the T cell cytolytic response without being able directly to stimulate mature T cells from these transgenic mice. Furthermore, a negative selection assay in vitro was used to demonstrate for the first time that antagonistic peptides are capable of antagonizing thymocyte deletion induced by antigenic peptides. These data suggest that the final selection of a T cell could be the result of a balance between the positive and negative influences of endogenous peptide ligands.     

Increased Serum Immunoglobulin Responses to Gut Commensal Gram-Negative Bacteria in Unipolar Major Depression and Bipolar Disorder Type 1, Especially When Melancholia Is Present

Major depressive disorder (MDD) is accompanied by higher serum IgM/IgA responses to LPS of Gram-negative bacteria, suggesting increased bacterial translocation and gut dysbiosis while the latter may occur in bipolar disorder (BD). There are differences between MDD and BD type 1 (BP1) and 2 (BP2) in nitro-oxidative stress biomarkers associated with leaky gut. This study examines serum IgM/IgA responses directed to LPS of 6 Gram-negative bacteria as well as IgG responses to oxidized LDL (oxLDL) in 29 BP1, 37 BP2, 44 MDD, and 30 healthy individuals. Increased IgM/IgA responses to Pseudomonas aeruginosa significantly discriminated patients with affective disorders (MDD plus BD) from controls. BP1 patients showed higher IgM responses to Morganella morganii as compared with MDD and BP2 patients. Patients with melancholia showed higher IgA responses to Citrobacter koseri as compared to controls and non-melancholic depression. The total score on the Hamilton Depression Rating Scale was significantly associated with IgA responses to C. koseri. IgG to oxLDL was significantly associated with increased bacterial translocation. In conclusion, MDD, BP1, and BP2 are accompanied by an immune response due to the increased load of LPS while these aberrations in the gut-brain axis are most pronounced in BP1 and melancholia. Activated oxidative stress pathways and autoimmune responses to oxidative specific epitopes in mood disorders may be driven by a breakdown in gut paracellular, transcellular, and/or vascular pathways. If replicated, drugs that protect the integrity of the gut barrier may offer novel therapeutic opportunities for BP1 and MDD.