Dynamic weight bearing as a non-reflexive method for the measurement of abdominal pain in mice.

BACKGROUND: Chronic pelvic pain (CPP) is a high burden for patients and society. It affects 15-24% of women in reproductive age and is an area of high unmet medical need. CPP can be caused by a wide range of visceral diseases such as abdominal infections, gastrointestinal or gynaecological diseases like endometriosis. Despite the high medical need for this condition, pharmacological approaches are hampered by the limited number of available methods for the behavioural evaluation of pain in inflammation-driven animal models of pelvic pain. METHODS: The dynamic weight bearing (DWB) system was used for the evaluation of spontaneous behaviour changes in the zymosan-induced peritonitis mouse model. Inflammatory mediator levels were evaluated in peritoneal lavage and their correlation with the behavioural endpoints was assessed. We evaluated the effect on behavioural endpoints of the selective cyclooxygenase-2 (COX-2) inhibitor celecoxib and the Nav 1.8 blocker A-803467. RESULTS: The presence of a relief posture, characterized by a significantly increased weight distribution towards the front paws, was observed following intraperitoneal injection of zymosan. A positive correlation was detected between PGE2 levels in the peritoneal lavage and DWB endpoints. In addition, zymosan-induced weight bearing changes were reverted by celecoxib and A-803467. CONCLUSIONS: This study described for the first time the use of DWB as a non-subjective and non-reflexive method for the evaluation of inflammatory-driven abdominal pain in a mouse model.

The TCR-delta repertoire in normal human skin is restricted and distinct from the TCR-delta repertoire in the peripheral blood.

The skin and the intestinal mucosa form surfaces to external environments and share similarities in anatomic structure and immunologic defense. In healthy humans, intestinal gamma/delta T cells express a highly restricted gamma/delta T cell receptor repertoire whereas gamma/delta T cells of the skin were thought to express a polyclonal repertoire. Herein we report, using complementarity-determining region 3 size spectratyping and nucleotide sequencing of T cell receptor DV1 and DV2 rearrangements, that the human skin is also composed of clonally expanded gamma/delta T cells that are widely distributed. Identical complementarity-determining region 3 profiles and T cell receptor delta rearrangements were found in two separate skin samples that were obtained as far as 2-10 cm apart. Furthermore, analysis of peripheral blood mononuclear cells of these subjects clearly demonstrated that the skin harbors a unique population of gamma/delta T cells that is distinct from that in the peripheral blood. In addition comparable data were obtained irrespective of whether DNA or RNA was analyzed, indicating that the observed oligoclonality is not secondary to the expression of large amounts of mRNA from a few activated cells. Thus, gamma/delta T cells of the skin and the intestine both express an oligoclonal repertoire that enables them to respond to a variety of deleterious antigens without the need for diverse T cell receptors, possibly by recognition of stress-induced self-antigens or of conserved foreign antigens.

T-cell-receptor repertoire in chronic plaque-stage psoriasis is restricted and lacks enrichment of superantigen-associated V beta regions.

Preferential usage of certain T-cell receptors by the lymphocytic infiltrate in psoriasis might indicate the involvement of an antigen in the pathogenesis of this disease. However, to date there are no data on the complete T-cell-receptor V alpha and V beta repertoire in psoriatic patients. We therefore compared the usage of T-cell-receptor variable regions in blood and skin of 10 patients with chronic plaque-stage psoriasis by means of semiquantitative polymerase chain reaction. Additionally, HLA class II alleles were analyzed by means of sequence-specific oligonucleotide typing. A considerable restriction of the T-cell-receptor repertoire was observed in the skin, where up to 20% of the variable regions present in the blood were not detectable. This was true for both alpha- and beta-chains. However, no interindividually constant pattern of T-cell-receptor restriction was deducible. Inconsistently, a certain preferential usage of some beta chains occurred within the cutaneous compartment. This report on the complete T-cell-receptor V alpha and V beta repertoire in psoriasis documents the restricted receptor repertoire of infiltrating T cells and a lack of enrichment of superantigen-associated V beta regions. Thus superantigens seem not to play a pathogenetically relevant role in chronic plaque-stage psoriasis.

Redox-modulated pathways in inflammatory skin diseases.

Inflammatory skin diseases account for a large proportion of all skin disorders and constitute a major health problem worldwide. Contact dermatitis, atopic dermatitis, and psoriasis represent the most prevalent inflammatory skin disorders and share a common efferent T-lymphocyte mediated response. Oxidative stress and inflammation have recently been linked to cutaneous damage in T-lymphocyte mediated skin diseases, particularly in contact dermatitis. Insights into the pathophysiology responsible for contact dermatitis can be used to better understand the mechanism of other T-lymphocyte mediated inflammatory skin diseases, and may help to develop novel therapeutic approaches. This review focuses on redox sensitive events in the inflammatory scenario of contact dermatitis, which comprise for example, several kinases, transcription factors, cytokines, adhesion molecules, dendritic cell surface markers, the T-lymphocyte receptor, and the cutaneous lymphocyte-associated antigen (CLA). In vitro and animal studies clearly point to a central role of several distinct but interconnected redox-sensitive pathways in the pathogenesis of contact dermatitis. However, clinical evidence that modulation of the skin's redox state can be used therapeutically to modulate the inflammatory response in contact dermatitis is presently not convincing. The rational for this discrepancy seems to be multi-faceted and complex and will be discussed.

Activation of human monocyte/macrophage cytotoxicity by IL-2/IFN gamma is linked to increased expression of an antitumor receptor with specificity for acetylated mannose.

Spontaneous cytotoxicity of human monocytes (purity: 92-95%) against K562 tumor cells was only observed in 31% healthy donors but, in the presence of rhamnogalacturonan (500 ng/ml), enhanced cytotoxicity was recorded for 79% (n = 14) of the donors. Monocytes activated by culturing with interleukin-2 and/or IFN gamma showed increased antitumor cytotoxicity against K562 tumor cells in 86% (n = 21) of the donors exhibiting additional increases in specific cytotoxicity when the cytotoxicity assays were carried out in the presence of rhamnogalacturonan. Increases of monocyte cytotoxicity achieved by activation with cytokines coincided with increased formation of monocyte/tumor cell conjugates. Similarly, increased monocyte cytotoxicity mediated by rhamnogalacturonan also correlated with increased monocyte/tumor cell conjugate formation most likely due to effector cell/target cell bridging as was originally described for rhamnogalacturonan interacting with CD56+ natural killer or lymphokine-activated killer cells and tumor cells. The chemospecificity of the monocyte-based receptors responsible for cytotoxicity and for monocyte/tumor cell conjugate formation, as well as for their rhamnogalacturonan-mediated enhancements, appears to be identical since all these effects could be inhibited in a dose-dependent manner by partially deacetylated (60%) mannose pentaacetate.

The superantigen exfoliative toxin induces cutaneous lymphocyte-associated antigen expression in peripheral human T lymphocytes.

Several immune-mediated dermatoses including psoriasis and atopic dermatitis can be exacerbated by bacterial infections. Superantigen producing bacteria can be isolated from skin lesions of these dermatoses. Consistent with superantigen effects, skewed T cell receptor variable gene usage has been demonstrated within these lesions. Therefore, the question arises whether superantigen induce a skin-seeking phenotype within peripheral T cells. In this study, we investigated the in vitro influence of the V beta 2-selective superantigen exfoliative toxin from Staphylococcus aureus on the expression of the cutaneous lymphocyte-associated antigen on peripheral T lymphocytes of healthy donors. We demonstrate that exfoliative toxin dramatically upregulates cutaneous lymphocyte-associated antigen expression on T cell receptor V beta 2+ lymphocytes. Up to 69% of V beta 2+ lymphocytes expressed cutaneous lymphocyte-associated antigen after 5 days of in vitro culture. Additionally, exfoliative toxin also increased cutaneous lymphocyte-associated antigen expression in CD3+ T cell receptor V beta 2- lymphocytes indicating a different effect as caused by the superantigen-T cell receptor V beta 2 interaction. Our findings suggest influence of bacterial superantigens on T lymphocyte skin homing in vivo.

Enhancement of MHC-unrestricted cytotoxic activity of human CD56+ CD3- natural killer (NK) cells and CD3+ T cells by rhamnogalacturonan: target cell specificity and activity against NK-insensitive targets.

Rhamnogalacturonan-mediated enhancement of MHC-unrestricted cytotoxicity was studied with freshly isolated CD56+CD3- natural killer (NK) cells, interleukin-2 (IL-2)-activated CD56+ lymphokine-activated killer (LAK) cells und IL-2/anti-CD3-activated T cells as effector cells using NK-sensitive and NK-insensitive tumor cells as targets. The rhamnogalacturonan fractions IM, IP, and IQ were prepared from commercially available extracts of Viscum album. The dose/response relation of IM, IP, and IQ demonstrated the presence of various concentrations of cytotoxicity-enhancing compounds in all three fractions that were identified as rhamnogalacturonans by degradation studies with poly-alpha-D-galacturonidase (EC 3.2.1.15) and alpha-1,6-rhamnosidase (EC 3.2.1.40). Specific cytotoxicity of all three effector cell populations as well as the respective rhamnoagalacturonan-mediated cytotoxicity enhancement was readily inhibited in a dose-dependent manner by 60%-deacetylated mannose pentaacetate. Rhamnogalacturonan-mediated enhancement of cytotoxicity of fresh CD56+ NK cells was also observed with four of five NK-insensitive tumor cells as targets, indicating that the effector-cell/tumor-cell bridging activity of rhamnogalacturonans renders NK-insensitive targets susceptible to NK-mediated lysis. Moreover, the rhamnogalacturonan-mediated cytotoxicity enhancement became even more prominent when lymphokine-activated CD56+ LAK and CD3+ T cells were assayed with the NK-insensitive tumor cell targets.

Mycophenolate mofetil is effective in the treatment of atopic dermatitis.

OBJECTIVE: To evaluate whether mycophenolate mofetil, a new immunosuppressive agent, is effective for treating moderate-severe atopic dermatitis (AD). DESIGN: In an open-label pilot study, mycophenolate mofetil, 1 g, was given orally twice daily for 4 weeks. At week 5, the dosage was reduced to 500 mg twice daily until study end (week 8). Patients were followed up for 20 weeks. SETTING: University hospital dermatology department. PATIENTS: Ten consecutive patients with moderate-severe AD nonresponsive to standard therapy. MAIN OUTCOME MEASURE: Severity of AD as measured using the subjective SCORAD [SCORing Atopic Dermatitis] index. RESULTS: Clinical efficacy was measured every 2 weeks using the subjective SCORAD index. Treatment with mycophenolate notably reduced the severity of AD within 4 weeks in all patients (P<.05), and after 8 weeks the mean +/- SD SCORAD index dropped from the pretreatment value of 49.2 +/- 13.8 to 21.9 +/- 26.5 (P<.01). One patient had to discontinue mycophenolate therapy after 4 weeks because of the development of herpes retinitis. Except for this event, mycophenolate was tolerated well in all patients. Six of 7 patients who had responded to mycophenolate monotherapy had no relapse of disease during 20-week follow-up. In the 7 patients who finished the study, the SCORAD index was reduced by 74%, from 44.0 +/- 7.8 before treatment to 11.4 +/- 5.9 at 20-week follow-up. CONCLUSIONS: Mycophenolate is a highly effective drug for treating moderate-severe AD, with no serious adverse effects occurring in any patients. Thus, mycophenolate might develop into a promising alternative in the therapy of moderate-severe AD.

Induction of NK-like activity in T cells by IL-2/anti-CD3 is linked to expression of a new antitumour receptor with specificity for acetylated mannose.

The generation of MHC-unrestricted cytotoxicity of highly enriched human CD3+ T cells (95-99%) by treatment with IL-2 and/or anti-CD3 antibodies was studied. T cells obtained by positive immunomagnetic sorting (anti-CD3) developed comparable specific cytotoxicities against K562 and Daudi cells when cultured with IL-2 and anti-CD3 for 96 h (80% of donors; n = 25). This increase of MHC-unrestricted cytotoxicity correlated fairly well with an increased formation of T cell/tumour cell conjugates. Moreover, simultaneous expression of a rhamnogalacturonan-binding receptor on activated T cells could be demonstrated. Rhamnogalacturonan was reported to enhance cytotoxicity of CD56+ NK cells by effector cell/target cell bridging. Untreated CD3+ cells hardly reacted with rhamnogalacturonan and IL-2-activated T cells showed only a moderate enhancement of cytotoxicity in the presence of rhamnogalacturonan. However, when CD3+ T cells had interacted with anti-CD3 antibodies during cell-sorting or during subsequent culturing with IL-2, enhancement in cytotoxicity and increased formation of lytic effector cell/tumour cell conjugates in the presence of rhamnogalacturonan could be readily demonstrated, indicating a bridging effect analogous to CD56+ NK cells. The conjugate formation of activated T cells with tumour cells as well as the additional rhamnogalacturonan-mediated bridging must be based on the expression of receptors with acetyl mannose specificity, since enhancements of MHC-unrestricted T cell cytotoxicity and conjugate formation were inhibited in a dose-dependent manner when acetylated mannose was present in the assays.

Differential expression of heat shock protein 70 (HSP70) and heat shock cognate protein 70 (HSC70) in human epidermis.

Autoimmunity and microbial agents have been suggested as playing a pathogenetic role in psoriasis. Since immune responses to microbial infections are often directed towards heat shock proteins (HSP), we investigated the expression of three HSP families in normal and inflamed human skin. Specimens from ten patients with psoriasis and three patients with positive patch tests for nickel and from five healthy volunteers were analysed by means of immunohistochemistry. The patterns observed were qualitatively similar in these conditions showing only minor quantitative differences. Psoriatic epidermis exhibited the highest level of expression. HSP27, HSP70 and heat shock cognate protein 70 (HSC70) were readily detectable. HSP27 was homogeneously distributed throughout the epidermis, whereas HSP70 was restricted to the basal layer and HSC70 primarily to the suprabasal layers. Other HSPs were detected to a lesser degree and showed a more irregular pattern. Thus, the qualitative expression pattern of HSPs seems to be constant between different skin conditions, but the expression of constitutive and inducible HSP70 depends on the differentiation state of keratinocytes.

Characterization of ZK 245186, a novel, selective glucocorticoid receptor agonist for the topical treatment of inflammatory skin diseases.

BACKGROUND AND PURPOSE: Glucocorticoids are highly effective in the therapy of inflammatory diseases. Their value, however, is limited by side effects. The discovery of the molecular mechanisms of the glucocorticoid receptor and the recognition that activation and repression of gene expression could be addressed separately opened the possibility of achieving improved safety profiles by the identification of ligands that predominantly induce repression. Here we report on ZK 245186, a novel, non-steroidal, low-molecular-weight, glucocorticoid receptor-selective agonist for the topical treatment of inflammatory dermatoses. EXPERIMENTAL APPROACH: Pharmacological properties of ZK 245186 and reference compounds were studied in terms of their potential anti-inflammatory and side effects in functional bioassays in vitro and in rodent models in vivo. KEY RESULTS: Anti-inflammatory activity of ZK 245186 was demonstrated in in vitro assays for inhibition of cytokine secretion and T cell proliferation. In vivo, using irritant contact dermatitis and T cell-mediated contact allergy models in mice and rats, ZK 245186 showed anti-inflammatory efficacy after topical application similar to the classical glucocorticoids, mometasone furoate and methylprednisolone aceponate. ZK 245186, however, exhibits a better safety profile with regard to growth inhibition and induction of skin atrophy after long-term topical application, thymocyte apoptosis, hyperglycaemia and hepatic tyrosine aminotransferase activity. CONCLUSIONS AND IMPLICATIONS: ZK 245186 is a potent anti-inflammatory compound with a lower potential for side effects, compared with classical glucocorticoids. It represents a promising drug candidate and is currently in clinical trials.

Psoriasis: a possible risk factor for development of coronary artery calcification.

BACKGROUND: Psoriasis is a chronic inflammatory skin disorder affecting about 2% of white-skinned individuals. Epidemiological data on the prevalence and degree of coronary artery calcification (CAC) as an indicator for cardiovascular diseases in patients with psoriasis are contradictory. OBJECTIVES: To study the prevalence and degree of CAC as an indicator for cardiovascular diseases in 32 patients with psoriasis matched for age, sex and risk factors to an equally sized control population. METHODS: Noncontrast-enhanced 16-row spiral computed tomography was performed in patients and controls. RESULTS: We found a significantly increased prevalence (59.4% vs. 28.1%, P = 0.015) and severity (CAC score according to Agatston 3.7 vs. 0.0, P = 0.019) of CAC in patients with psoriasis. Multiple linear regression calculations identified psoriasis as a likely independent risk factor for CAC. CONCLUSIONS: Our results point towards the potentially systemic nature of the inflammatory processes underlying the pathogenesis of psoriasis, which may therefore be considered a potentially severe systemic disease.

[The role of bacterial superantigens in pathophysiology of the skin]. / Die Rolle bakterieller Superantigene in der Pathophysiologie der Haut.

In contrast to conventional antigens, bacterial superantigens activate a large percentage of an organism's total T-cell repertoire. This has clinical implications particularly in the case of infections with Gram-positive bacteria. Unravelling the molecular mechanisms of superantigen-mediated T-cell activation has yielded new insights in the pathogenesis of a number of diseases, in the field of dermatology namely atopic dermatitis and psoriasis. Moreover, therapeutic applications of superantigens have begun to emerge.

Mycetoma due to Exophiala jeanselmei and Mycobacterium chelonae in a 73-year-old man with idiopathic CD4+ T lymphocytopenia.

Exophiala jeanselmei and Mycobacterium chelonae were isolated from cutaneous nodules in a 73-year-old man with mycetoma of the right lower leg. Further evaluation revealed CD4+ lymphocytopenia without evidence of HIV infection. Antibodies to HIV 1/2, p24 antigen and HIV 1/2 (PCR) and reverse transcriptase activity were not detectable. The patient was not a member of any HIV risk group. He had not previously undergone therapy or suffered from immunodeficiency. This case clearly demonstrates that infections with opportunistic moulds and/or atypical mycobacteria should be taken into consideration not only in patients with classical immundeficiency diseases but also in apparently healthy patients because infection with these agents can be the first sign of underlying immunodeficiency.

Predominant usage of distinct T-cell receptor V beta regions by epidermotropic T cells in psoriasis.

To investigate the role of T cells in psoriasis, we analyzed the T-cell receptor (TCR) repertoire in peripheral blood lymphocytes (PBLs) and in skin lesions of 9 patients with chronic plaque-stage psoriasis by means of immunohistochemistry. An extended panel of monoclonal antibodies detecting 11 different V beta families was employed in this study. In the peripheral blood, no predominant TCR V beta expression was detected and interindividual differences were small. Skin-infiltrating T cells showed only marginal differences in the TCR V beta usage when compared with PBLs. However, in 3 patients the intraepidermal infiltrate comprised up to 40% of T cells expressing a TCR V beta region which was virtually absent in the peripheral blood and the dermal infiltrate. Thus, although the TCR V beta usage of skin-infiltrating T cells showed little difference compared to PBLs, several cases exhibited a significant increase of epidermotrophic T cells using one particular V beta region.

[Nail polish allergy. An important differential diagnosis in contact dermatitis]. / Nagellack-Allergie. Eine wichtige Differentialdiagnose bei Kontaktdermatitis.

HISTORY AND CLINICAL FINDINGS: Case 1. A 34-year-old woman presented with a recurrent, itching and erythematous plaque on her right cheek. Case 2. A 27-year-old woman reported itching erythema on her fingertips from time to time recently. Inspection revealed dyshidrosiform blisters. Both patients had been using nail varnish and varnish remover. Case 3. A 49-year-old woman had, for about 5 years, repeatedly experienced itching erythema at the perionychium of several fingers on which she had placed artificial nails. Marked oedema and erythema was noted. INVESTIGATION: Extensive epicutaneous tests were performed on these patients. They showed sensitisation against important allergens in nail varnish (toluolsulphonamide-formaldehyde resin), nail varnish remover (benzophenone-2) and artificial nails (ethylacrylate), respectively. The three patients also showed type IV sensitisation against other substances. DIAGNOSIS, TREATMENT AND COURSE: Epicutaneous tests documented type IV allergies to important constituents of nail varnish, nail varnish remover and artificial nails. Symptoms and skin changes disappeared when these three items were no longer used. CONCLUSION: Allergies against nail varnish and its remover and against artificial nails should be included in the differential diagnosis of skin allergies, even when patients have additional type IV sensitizations against common allergens. As the causative allergen can be easily avoided, knowledge of the particular problems associated with these allergies is of a great practical significance.