Indicators of active disease and steroid dependency in patients with inflammatory bowel diseases not treated with biologics in a German real-world-setting.

BACKGROUND AND AIMS: While a minority of inflammatory bowel disease (IBD) patients receives biologics in Germany, little is known about therapeutic needs of patients receiving non-biologic therapies. This study aimed to identify indicators of active disease/steroid dependency in patients with moderate to severe Crohn's disease (CD) and ulcerative colitis (UC) treated with conventional therapies and to describe health care resource use (HCRU)/cost. METHODS: CD/UC patients treated with immunosuppressants (IS) and/or systemic or locally acting oral corticosteroids (CS) were identified in German claims data (2013-2017) and followed for 12 months post-therapy start. Indicators of active disease/steroid dependency during follow-up period were (i) ≥ 2 prescriptions of CS (sensitivity ≥ 4) or (ii) ≥ 1 IBD-related surgery or (iii) > 7 days IBD-related hospitalization(s). RESULTS: Of 9871 included IBD patients (5170 CD, 4701 UC), 25.7%/19.9% (CD/UC) received ≥ 2 prescriptions of CS (sensitivity, 17.4%/15.7%) (i), 3.2% experienced IBD-related surgeries (ii), and 2.5% > 7 days of hospitalizations (iii). Altogether, 44.4% had indicators of active disease/steroid dependency (sensitivity, 23.9%). Among patients with active disease/steroid dependency, 78.0% received CS monotherapy at baseline. Of these, 89.6% received a CS monotherapy in the follow-up period, too. Proportionally, fewer patients with CS monotherapy (57.4%) than IS therapy (91.0%) visited a specialist. HCRU/cost per patient year was significantly higher in patients with than without active disease/steroid dependency. CONCLUSIONS: A substantial percentage of biologic-naïve IBD patients suffers from active disease/steroid dependency. The majority receives a monotherapy with systemic CS. Referral to gastroenterologists for treatment optimization is recommended, also because active disease/steroid dependency is associated with increased HCRU/cost.

Alcohol and delirium tremens: effects of average number of drinks per day and beverage type.

OBJECTIVE: Associations of amount of alcohol intake and beverage type with the risk of delirium tremens (DT) have not been studied. This longitudinal study investigated if the average number of drinks per day and beverage type predict DT. METHODS: A cohort of 3 582 alcohol-dependent men and women aged 19-82 without previous DT were interviewed about alcohol intake and beverage type at baseline in 1994-2005 and followed through record linkage in Danish nationwide registers to identify incident DT. Data were analyzed by means of Cox regression models. RESULTS: An average number of drinks per day of 20-30 or >30 was associated with hazard ratios (HRs) of 1.38 (95% CI 1.03-1.84) and 1.64 (95% CI 1.19-2.27) relative to the reference category (1-9 drinks). Independently of amount consumed and covariates (age, gender, civil status and work status), beverage type (spirits vs. mixed alcohol) was associated with a HR of 1.63 (95% CI 1.08-2.46). Male gender was robustly associated with increased risk (HR = 1.62 (95% CI 1.25-2.08). CONCLUSIONS: In alcohol-dependent men and women, daily alcohol intake above a threshold of 20 beverages or 240 g alcohol and a preference for spirits increase the risk of developing DT.

Induction of interleukin 10-producing, nonproliferating CD4(+) T cells with regulatory properties by repetitive stimulation with allogeneic immature human dendritic cells.

The functional properties of dendritic cells (DCs) are strictly dependent on their maturational state. To analyze the influence of the maturational state of DCs on priming and differentiation of T cells, immature CD83(-) and mature CD83(+) human DCs were used for stimulation of naive, allogeneic CD4(+) T cells. Repetitive stimulation with mature DCs resulted in a strong expansion of alloreactive T cells and the exclusive development of T helper type 1 (Th1) cells. In contrast, after repetitive stimulation with immature DCs the alloreactive T cells showed an irreversibly inhibited proliferation that could not be restored by restimulation with mature DCs or peripheral blood mononuclear cells, or by the addition of interleukin (IL)-2. Only stimulation of T cells with mature DCs resulted in an upregulation of CD154, CD69, and CD70, whereas T cells activated with immature DCs showed an early upregulation of the negative regulator cytotoxic T lymphocyte-associated molecule 4 (CTLA-4). These T cells lost their ability to produce interferon gamma, IL-2, or IL-4 after several stimulations with immature DCs and differentiated into nonproliferating, IL-10-producing T cells. Furthermore, in coculture experiments these T cells inhibited the antigen-driven proliferation of Th1 cells in a contact- and dose-dependent, but antigen-nonspecific manner. These data show that immature and mature DCs induce different types of T cell responses: inflammatory Th1 cells are induced by mature DCs, and IL-10-producing T cell regulatory 1-like cells by immature DCs.

Induction of hapten-specific tolerance by interleukin 10 in vivo.

Interleukin 10 (IL-10) is released during the induction phase of contact sensitivity and was shown in prior functional studies to convert epidermal Langerhans cells (LC) from potent inducers of primary immune responses to specifically tolerizing cells in vitro. To investigate whether IL-10 also subserves the function of a tolerizing agent in vivo ears of BALB/c or C3H mice were injected intradermally with 1-2 micrograms of recombinant mouse (rm)IL-10 8 h before epicutaneous application of 3% trinitrochlorobenzene (TNCB; a contact allergen). As a control, mice were injected with phosphate-buffered saline or IL-10 plus neutralizing amounts of anti-IL-10 mAb. 5 d later, mice were challenged with 1% TNCB on contralateral ears and ear swelling response was measured 24 h later. Whereas control-treated mice showed a normal ear swelling response to epicutaneous challenge (delta mm-2 = 25 +/- 5), ear swelling response of IL-10-treated animals was significantly inhibited (delta mm-2 = 3 +/- 2). Coinjection of IL-10-specific mAb together with rmIL-10 completely abrogated this effect. To differentiate between a state of nonresponsiveness and induction of tolerance by IL-10, mice initially treated with IL-10 and TNCB were resensitized with 3% TNCB in the absence of any treatment after 14 d of rest (group 1). Again mice were challenged 5 d later and ear swelling responses were tested. Whereas control mice treated with allergen alone (group 2) showed a good swelling response (delta mm-2 = 28 +/- 6), IL-10-treated mice (group 1) showed a minimal response towards application of allergen (delta mm-2 = 4 +/- 2). To show that anergy induction by IL-10 was antigen-specific, mice initially treated with IL-10 plus TNCB were exposed to 0.5% dinitrofluorobenzene (DNFB) 14 d later (group 1). After challenge with 0.1% DNFB, IL-10-treated mice showed an ear swelling response (delta mm-2 = 13 +/- 3; group 1) similar to that of control mice only sensitized with DNFB (delta mm-2 = 14 +/- 3; group 3). In an attempt to show the induction of antigen-specific tolerance in these mice in vitro, regional lymph nodes of mice initially treated with TNCB plus IL-10 (group 1) and control-treated mice (groups 2 and 3) were prepared and cultured in the presence of TNBS, dinitrobenzene sulfonate (DNBS), or medium to measure antigen-specific proliferation.(ABSTRACT TRUNCATED AT 400 WORDS)

Identification and functional characterization of human CD4(+)CD25(+) T cells with regulatory properties isolated from peripheral blood.

A subpopulation of peripheral human CD4(+)CD25(+) T cells that expresses CD45RO, histocompatibility leukocyte antigen DR, and intracellular cytotoxic T lymphocyte-associated antigen (CTLA) 4 does not expand after stimulation and markedly suppresses the expansion of conventional T cells in a contact-dependent manner. After activation, CD4(+)CD25(+) T cells express CTLA-4 on the surface detectable for several weeks. These cells show a G1/G0 cell cycle arrest and no production of interleukin (IL)-2, IL-4, or interferon (IFN)-gamma on either protein or mRNA levels. The anergic state of CD4(+)CD25(+) T cells is not reversible by the addition of anti-CD28, anti-CTLA-4, anti-transforming growth factor beta, or anti-IL-10 antibody. However, the refractory state of CD4(+)CD25(+) T cells was partially reversible by the addition of IL-2 or IL-4. These data demonstrate that human blood contains a resident T cell population with potent regulatory properties.

Identification and induction of human keratinocyte-derived IL-12.

Interleukin 12 is a heterodimeric molecule that serves as a potent co-stimulator enhancing the development of Th1 cells. As one of the classical Th1 cell-mediated responses is contact sensitivity in skin, we wondered whether IL-12 might be produced by epidermal cells and serve as a mediator of this immune response. Using a sensitive, quantitative PCR technique we demonstrate that p35 chain mRNA of IL-12 is produced constitutively by human epidermal cells, whereas p40 chain mRNA can only be detected in epidermis treated with contact allergen, but not epidermis exposed to irritants or tolerogens. Time course studies showed a dramatic induction of IL-12 p40 mRNA 4 h after in vivo allergen treatment reaching peak strength after 6 h. In cell depletion assays we show that epidermal keratinocytes are the major source of this cytokine in the epidermis. This was further supported by analysis of mRNA derived from the human keratinocyte cell line HaCat expressing IL-12 p35 and p40 mRNA upon stimulation. The presence of bioactive IL-12 in supernatants derived from allergen-stimulated epidermal cells was demonstrated by IL-12-specific bioassay. Additional evidence for the functional importance of IL-12 in primary immune reactions in skin was obtained in allogeneic proliferation assays using human haptenated epidermal cells containing Langerhans cells as APC and allogeneic CD4+ T cells as responders. Anti-IL-12 mAb inhibited the proliferation of T cells by approximately 50%. In aggregate our data demonstrate that nonlymphoid keratinocytes are capable of producing functional IL-12 and provide evidence for the functional significance of IL-12 in primary immune responses in skin.

Depletion of CD25(+) CD4(+) T cells and treatment with tyrosinase-related protein 2-transduced dendritic cells enhance the interferon alpha-induced, CD8(+) T-cell-dependent immune defense of B16 melanoma.

Transduction of B16 melanoma cells with IFN alpha (B16-IFN alpha) enhances CD8(+) T-cell-dependent tumor immunity in mice, resulting in delayed outgrowth in vivo. Here we provide evidence that CD4(+) T cells down-regulate the IFN alpha-induced tumor immune defense. Importantly, depletion of regulatory CD25(+) CD4(+) T cells prevented growth of B16-IFN alpha in most mice and promoted long-lasting protective tumor immunity. Rejection of B16-IFN alpha could also be achieved with therapeutic injections of dendritic cells genetically engineered to express the melanoma antigen tyrosinase-related protein 2. These results support the development of novel strategies for the immunotherapy of melanoma using IFN alpha in combination with elimination of regulatory T cells or antigen-specific immunization.

A prospective study of young men at high risk for alcoholism. Social and psychological characteristics.

In a prospective longitudinal study of alcoholism, we applied the high-risk method using a Danish birth cohort (9125 consecutive deliveries, 1959 to 1961). From the cohort, 134 sons of alcoholic fathers (high-risk group) and 70 matched controls without parental alcoholism were selected for study. Extensive data were collected in a multidisciplinary etiologic approach. We report the social and psychological characteristics from a "premorbid" assessment when the subjects were 19 to 20 years old. The high-risk group reported more disrupted familial conditions during childhood than the control group. Both groups had a drinking pattern similar to that of the general Danish population at the same age. No alcoholic subjects were found. The high-risk group was characterized by poor verbal ability and impulsive behavior. We plan a follow-up examination of the sample.

Psychopathology in adopted and nonadopted daughters of alcoholics.

This report completes a series of studies conducted in Denmark comparing drinking patterns and psychopathology in adopted and nonadopted children of alcoholics. Sons of alcoholics had higher rates of alcoholism than controls, whether raised by their alcoholic parents or by foster parents. They did not have more psychopathology otherwise. Daughters of alcoholics, adopted and nonadopted, had a higher rate of alcoholism than was the expected frequency in the general population, but so did controls in the adopted group; neither group had higher rates of other psychopathology, eg, depression. However, daughters of alcoholics raised by their biological parents had significantly more depression.

Alcoholism and depression in adopted-out daughters of alcoholics.

Forty-nine daughters of alcoholics were compared to 47 daughters of nonalcholics; both groups of women (average age, 35 years) had been adopted by nonrelatives early in life. Two women in each group were alcoholic or problem drinkers. Although this is above the expected rate of alcoholism among women, the numbers are too small to draw definite conclusions. Almost all were light drinkers. Daughters of alcoholics had no more depression than controls, indicating that alcoholism in the biological parents did not increase the risk of depression in daughters raised by foster parents. Environmental factors may be important in both alcoholism and depression in women, since both tended to be correlated with psychopathology in the foster parents.

The EEG after alcohol administration in men at risk for alcoholism.

The biologic sons of alcoholics constitute a group at high risk (HR) for alcoholism. A 0.5-g/kg dose of alcohol was administered to HR and control subjects aged 19 to 21 years. Blood alcohol concentration measurements failed to distinguish HR from control subjects, but quantitative measurements of EEG alpha activity differentiated them. The HR subjects exhibited greater increases in slow alpha energy and greater decreases of fast alpha energy after alcohol administration than controls; the HR subjects also showed greater decreases in mean alpha frequency after alcohol administration. These EEG findings suggest that subjects at high risk for alcoholism are physiologically more sensitive to alcohol than control subjects.

The electroencephalogram after alcohol administration in high-risk men and the development of alcohol use disorders 10 years later.

BACKGROUND: In 1979 through 1980, electroencephalographic (EEG) responses to an alcohol challenge in 19 year-old sons of alcoholics as well as in sons of nonalcoholic control subjects were examined. The familial risk status of the subjects and greater EEG sensitivity to alcohol were hypothesized to predict the development of alcoholism 10 years later. METHODS: In 1990 through 1992, diagnostic interviews were completed to ascertain alcohol and other substance use disorders in these subjects and to update their family history. RESULTS: Updated family history of alcoholism predicted the development of substance dependence. Density of alcoholic relatives (the number of alcoholic relatives divided by the number of known relatives) was positively related to the severity of alcohol use disorders in the probands. Contrary to expectation, a greater EEG response at age 19 years was not related to the later development of alcohol dependence. Instead, the opposite was observed: a smaller EEG alpha frequency response to alcohol at age 19 years was related to the development of alcohol dependence and high quantity and frequency of alcohol consumption 10 years later. CONCLUSIONS: Lower EEG response to a small dose of alcohol may be associated with the later development of alcohol dependence. This result is based on a small number of subjects and should be interpreted with caution. Although this result is opposite to our 1980 hypothesis, it is consistent with much of the recent literature.

Immunologic effects of interferon.

Interferons can be defined as a family of induced proteins sharing the capacity to exert pleiotropic effects on cell functions and to render cells resistant to virus infection. They are activating genes coding for a number of enzymes, most of which have not yet been characterized, and also by enhancing the synthesis of cell surface components. This enables interferons to modulate the immune response at different levels. This article will focus on the effects of interferon on antigen presentation, regulation of the immune response, activation of macrophage functions, and on its role in the pathogenesis of some diseases.

Different cellular reaction patterns of epidermal Langerhans cells after application of contact sensitizing, toxic, and tolerogenic compounds. A comparative ultrastructural and morphometric time-course analysis.

BALB/c mice were treated with the irritants croton oil (0.5%, 20%), sodium lauryl sulfate (15%), and benzalkonium chloride (25%), the contact sensitizers 2,4-dinitrofluorobenzene (DNFB, 0.3%) and picryl chloride (PCl, 1%), and the tolerogen 2,4-dinitrothiocyanatebenzene (DNTB, 2%). All irritants used produced degenerative alterations of Langerhans cells (LCs). After application of 0.5% croton oil, however, this degeneration was preceded by an activation of the cells with increased number of mitochondria and enlargement of nuclei. The DNFB and PCl application in sensitizing doses to nonsensitized animals resulted in a cellular activation similar to that observed for 0.5% croton oil. In addition, these LCs showed enhanced adsorptive endocytosis as demonstrated by increased numbers of Birbeck granules and coated vesicles. The endocytotic activity was more pronounced in DNFB-sensitized animals. The DNTB at a concentration that induced tolerance to DNFB did not cause either cellular or endocytotic activation of LCs. These results demonstrate that the contact sensitizers DNFB and PCl induce characteristic cellular reaction patterns of LCs, which may be related to their sensitizing property.

Ultrastructural localization of 2,4-dinitrophenyl groups in mouse epidermis following skin painting with 2,4-dinitrofluorobenzene and 2,4-dinitrothiocyanatebenzene: an immunoelectron microscopical study.

The ultrastructural localization of 2,4-dinitrophenyl (DNP) groups in mouse epidermis after epicutaneous application of the contact sensitizer 2,4-dinitrofluorobenzene (DNFB) and the tolerogen 2,4-dinitrothiocyanatebenzene (DNTB) was investigated at varying times using immunoelectron microscopy with the protein A - gold technique. After application of DNFB, there was a homogenous cytoplasmic labeling of all epidermal cells. The intracellular localization of the DNP groups was not restricted to cytoplasmic organelles belonging to the endocytotic - lysosomal system. The numerous endocytotic organelles, Birbeck granules, and lysosomes of the Langerhans cells (LCs) typically observed after application of contact sensitizers also did not show an increased number of gold particles. Skin painting with DNTB resulted in a similar distribution and time-course of immunolabeling, but this compound did not induce cellular and the endocytotic activation of LCs as seen after DNFB application. These results demonstrate that contact sensitizers do not require specific cellular uptake and intracellular processing by the endocytotic - lysosomal compartment of the LCs before membrane presentation. However, a cellular and endocytotic activation of the LCs by haptens may be an important mechanism for T effector cell activation.

Modulation of suppressor mechanisms in allergic contact dermatitis: 2. Inhibition of suppressor T-lymphocytes by Corynebacterium parvum.

Pretreatment of BALB/c mice with Corynebacterium parvum inhibited the induction of tolerance to the contact sensitizing agent 2,4-dinitrofluorobenzene induced by intravenous injection of DNBSO3. The suppressive effect on tolerance induction has further been analyzed by adoptive transfer experiments. Injection of C. parvum intraperitoneally (0.7-2.8 mg/mouse) before injection of the tolergen inhibited the generation of T-suppressor cells as shown by transfer of spleen cells from the tolerized donor to naive recipients. Pretreatment of the recipients of the suppressor T-cells from tolerized animals with C. parvum also inhibited the function of these cells in the recipient animals. Time-kinetic experiments suggested that more than one mechanism appeared to be responsible for the tolerance induced by DNBSO3; C. parvum (probably via activated macrophages) suppressed tolerance which is mediated by T-suppressor lymphocytes. These results suggest that T-suppressor lymphocytes may--similarly as T-helper cells--be modulated by an activated monocytic-phagocytic system.

Modulation of suppressor mechanisms in allergic contact dermatitis: 1. Effect of C. parvum on the induction phase of contact allergy.

The effect of a pretreatment with corynebacterium parvum (C. parvum) on contact allergy in BALB/c mice was studied. Mice sensitized with 50 microliter (supraoptimal dose) 2.4-dinitrofluorobenzene (DNFB, 0.5%) showed a suppressed response as measured by ear swelling after painting the right ear with 0.3% DNFB in comparison to an allergic response obtained with an optimal sensitization dose (15 microliter DNFB 0.5%). By transfer of spleen cells from donors sensitized with a supraoptimal or an optimal dose to recipients either challenged ro sensitized shortly afterwards with DNFB it could be shown that less functionally active immune T-lymphocytes of the delayed hypersensitivity type and significantly more suppressor T-cells were induced in supraoptimally sensitized mice in comparison to the optimally sensitized animals. Intraperitoneal injection of C. parvum (2.8 mg/mouse) one week before sensitization enhanced the contact allergic response in mice sensitized with a supraoptimal dose of DNFB, with little effect on the response in optimally sensitized animals. Further analysis of this enhancement in transfer experiments showed that C, parvum selectively suppressed the generation and/or functional expression of T-suppressor cells and, probably by this mechanism, increased the number of functionally active T-immune lymphocytes. It is proposed that possibly by activation of the immune system C. parvum will suppress suppressor cells in contact allergy and by this mechanism might facilitate sensitization to the contact allergen.

Modulation of suppressor mechanism in allergic contact dermatitis. IV. Selective inhibition of suppressor T-lymphocytes by serum obtained from Corynebacterium parvum treated mice.

We have previously shown that Corynebacterium parvum treatment reduced tolerance induced by intravenous injection of 2,4-dinitrobenzene sulfonic acid (DNBSO3) or by sensitizing BALB/c mice with a supraoptimal dose of 2,4-dinitrofluorobenzene (DNFB). Further analysis by transfer experiments revealed that the generation and/or functional expression of suppressor T-lymphocytes (Ts-cells) was inhibited in both tolerance models. Inhibition of Ts-cells was not entirely correlated with suppression of mitogen induced lymphocyte proliferation mediated by C. parvum activated macrophages. In this investigation we studied the effect of serum obtained at various times after C. parvum injection on tolerance and Ts-cells in DNFB contact allergy. Serum obtained 6 and 24 hr, however, not 72 hr after C.parvum injection inhibited Ts-cells, induced by DNBSO3 and tested by transfer to naive recipients, as efficiently as C. parvum itself. The serum had following characteristics: (1) It inhibited the functional expression of Ts in the recipient animal. (2) It inhibited tolerance induction by an epicutaneous supra-optimal dose of DNFB. (3) It did not inhibit the induction and functional expression of T-effector cells of delayed hypersensitivity (TDH-cells) as shown by transfer experiments. (4) The Ts-inhibitory factor was heat resistant (56 degrees C), not destroyed or lost by dialysis against tris-glycine buffer pH 2 and could not be detected in the serum of NMRI-mice injected 24 hr before with C parvum. (5) The C parvum serum did not significantly increase the spleen weight or suppress mitogen-induced spleen lymphocyte proliferation. The nature of this Ts-cells inhibitory factor is unknown, although the results suggest that cortisone or transfer of bacteria are not responsible. Factors mediating selective inhibition of Ts-cells may be of important regulatory function in delayed type hypersensitivity.

A novel monoclonal antibody to a distinct subset of cutaneous dendritic cells.

A monoclonal antibody was generated by immunizing rats with Langerhans cell (LC)-enriched epidermal cells obtained from BALB/c mouse earskin after epicutaneous application of the contact sensitizer 2,4-dinitrofluorobenzene (DNFB). The antibody 4F7 detects in normal mouse skin, few dermal cells showing the morphologic, phenotypic, and functional properties of accessory dendritic cells, but lacking Birbeck granules. The capacity to stimulate allogenic T cells in the mixed leucocyte reaction resembles that of freshly isolated LCs. After DNFB application, an increased number of 4F7+ dendritic cells are found in the dermis and, in addition, some labeled dendritic cells occur in the epidermis. Some of the latter cells exhibit cytoplasmic Birbeck granules. Remarkably, there is no increase of the 4F7+ cells in the regional lymph nodes after DNFB treatment. These data suggest that the 4F7 antibody labels distinct dendritic cells of the mouse skin that are involved in the mediation of contact sensitization and probably represent immature LCs.

Increased level of intracellular MHC class II molecules in murine Langerhans cells following in vivo and in vitro administration of contact allergens.

Treatment of murine Langerhans cells (LC) with contact allergens results in increased internalization of cell membrane constituents and therefore in depressed cell-surface expression of major histocompatibility complex (MHC) class II molecules during the first hours after haptenization. In this presentation we show that this downregulation of cell-surface-expressed Ia-antigens is accompanied by an augmentation of the intracellular pool of MHC class II molecules. Rat MoAb 2G9 was developed, which recognizes IA and IE molecules of the d-haplotype. This MoAb competes with the murine MoAb MK-D6 for binding sites to IAd-molecules. After blocking the cell-surface-expressed molecules with 2G9 and permeabilizing the cell membranes this allowed us to measure selectively the intracellular amount of IA molecules by double immunofluorescence staining and flow cytometric analysis. Cell-surface expression of IA molecules was found to be depressed but their internal pool was significantly elevated following in vivo treatment with the contact allergens DNFB, DNCB, oxazolone, and K2Cr2O7 for 3 h. In vitro culture of freshly prepared LC in the presence of 1 microgram/ml DNFB yielded similar results. Blocking of protein biosynthesis with cycloheximide did not prevent this intracellular class II accumulation. An augmented representation of internalized class II molecules in haptenized LC might play an important role in the presentation of contact allergens.