Lichen planus (LP) is a common, chronic relapsing inflammatory disorder of the skin and mucous membranes which often poses a major therapeutic challenge due to its refractory course. Novel ...pathogenesis-based therapies are urgently needed. As several studies have shown that IL-17 may contribute to LP pathogenesis, we investigated whether therapeutic targeting of IL-17
T cells leads to clinical improvement of mucosal and cutaneous LP lesions. A total of five patients with lichen planus were treated in a compassionate use trial with either secukinumab (anti-IL-17; 3 patients with acute and chronic recalcitrant muco-cutaneous LP), ustekinumab (anti-IL-12/IL-23; 1 patient with recalcitrant oral LP) or guselkumab (anti-IL-23; 1 patient with recalcitrant oral LP). The clinical course of the patients was assessed by the Autoimmune Bullous Skin Disorder Intensity Score (ABSIS) reflecting both extent and severity of disease and functional sequelae of oral involvement for at least 12 weeks. The inflammatory infiltrate in lesional and post-lesional skin was analyzed by immunohistochemistry before and after treatment. Furthermore, the cytokine profile of peripheral blood T cells from the treated patients was assessed by flow cytometry and/or ELISpot assay. Treatment with secukinumab induced rapid and prolonged clinical amelioration of muco-cutaneous LP. Clinical improvement was accompanied by a strong reduction of the Th1 and Th17/Tc17 cellular mucosal and cutaneous infiltrate. Moreover, long-term treatment of one patient with recalcitrant oral LP with ustekinumab led to healing of the ulcerative oral lesions and a reduction of peripheral blood and lesional IL-17
T cells. Finally, treatment with guselkumab led to a marked clinical improvement in a patient with recalcitrant erosive oral LP. These findings show for the first time that therapeutic targeting of Th17/Tc17 cells leads to a pronounced clinical amelioration of mucosal and cutaneous LP and strongly suggests that IL-17-producing T cells are central to disease pathogenesis. Thus, therapeutic targeting of Th17/Tc17 cells opens new therapeutic avenues in the treatment of recalcitrant LP.
Paraneoplastic autoimmune multi-organ syndrome (PAMS) is a rare clinical condition characterized by variable and heterogeneous clinical phenotypes in the presence of neoplasias which largely depend ...on the activation of humoral and cellular immune responses. Clinically, these patients present with a spectrum of antibody-driven pemphigus-like lesions to graft-vs.-host-disease-like exanthemas with a lichenoid inflammatory infiltrate in the skin. PAMS is occasionally associated with thymoma, in which altered immune surveillance eventually leads to multiorgan autoimmunity which often includes variable cutaneous symptoms. This disorder is associated with a profound disturbance of peripheral immune tolerance against human autoantigens.
We here present a patient with relapsing thymoma who developed PAMS with several cutaneous and extracutaneous autoimmune disorders.
Peripheral blood mononuclear cells (PBMC), sera, and lesional skin biopsies were obtained at different clinical disease stages. Peripheral T cell subsets were characterized phenotypically and the cytokine profile of the peripheral blood T cellular response against distinct epidermal and dermal autoantigens of the skin was analyzed by ELISpot assay. Serological screening was performed by ELISA and immunoblot analysis. Skin biopsies were subjected to immunohistochemical analysis of distinct T cell subsets. Thymoma tissue was analyzed for the presence of T regulatory cells and compared with adult thymus and indolent thymoma.
In the present case, thymoma was the cause of the observed multi-organ autoimmune syndromes as its recurrence and surgical removal was associated with the relapse and regression of the cutaneous symptoms, respectively. Initially, the patient presented with two autoimmune disorders with Th2/Th1 imbalance, myasthenia gravis (MG) and pemphigus foliaceus (PF), which regressed upon immunosuppressive treatment. Months later, the patient developed a lichenoid exanthema with a Th1-dominated skin infiltrate. Further clinical evaluation revealed the recurrence of the thymoma and the lichenoid exanthema gradually regressed upon thymectomy. Our contention that T cell recognition against distinct cutaneous autoantigens, such as desmoglein 1 (Dsg1), shifted from a Th2 to a Th1-dominated immune response could not be fully substantiated as the patient was on a stringent immunosuppressive treatment regimen. We could only observe a decrease of the initially present serum IgG autoantibodies against Dsg1. Phenotypic analysis of the associated thymoma showed a lower number of T regulatory cells compared to adult thymus and indolent thymoma, suggesting that impaired thymus-derived immune surveillance had a direct impact on the outcome of the observed cutaneous autoimmune disorders.
Inhibitors of protein deacetylases represent a novel therapeutic option for cancer diseases due to their effects on transcriptional regulation by interfering with histones acetylation and on several ...other cellular pathways. Recently, their ability to modulate several transcription factors and, interestingly, also co-factors, which actively participate in formation and modulation of transcription complexes was shown. We here investigate whether HMGA2 (High Mobility Group AT-2 hook), a nuclear non-histone transcriptional co-factor with known oncogenic properties, can be influenced by the novel pan-deacetylase inhibitor panobinostat (LBH589) in human hepatocellular carcinoma models. Panobinostat strongly downregulated HMGA2 in HepG2 and Hep3B cells; this effect was mediated by transcriptional upregulation and promotion of the maturation of the tumorsuppressor miRNA hsa-let-7b, which could inhibit HMGA2 expression via RNA interference pathways. siRNA knockdown of HMGA2 or transfection of hsa-let-7b mimicking oligonucleotides confirmed the role of HMGA2 in regulating cell proliferation and apoptosis in liver cancer cell lines. Co-incubation with panobinostat showed an additive effect on inhibition of cell proliferation using an impedance-based real-time cell analyzer. Treatment of HepG2 xenografts with panobinostat also led to a downregulation of HMGA2 in vivo. These findings show that pan-deacetylase inhibitors also modulate other signaling pathways and networks than histone modifications to influence cell fate.
► Panobinostat for the treatment of liver cancer. ► Panobinostat meddles with miRNAs-dependent transcriptional and translational control. ► Tumorsuppressor miRNA hsa-let-7b upregulation. ► HMGA2 is downregulated via RNA interference pathways mediated by hsa-let-7b. ► Panobinostat determines inhibition of proliferation via the axis hsa-let-7b – HMGA2.
Nitric oxide is a relatively stable free radical and an important signal molecule in plants, animals, and humans with high relevance for biological processes involving inflammatory processes, e.g. ...wound healing or cancer. The molecule can be detected in the gas phase of non-thermal plasma jets making it a valuable tool for clinical intervention, but transport efficiency from the gas phase into the liquid phase or tissue remains to be clarified. To elucidate this fact, the nitric oxide concentration in buffered solutions is determined using electron paramagnetic resonance spectroscopy. The origin of the nitric oxide in the liquid could be excluded, therefore, potential precursors such as hydroxyl radicals, superoxide anions, atomic hydrogen and stable species (nitrite, nitrate and hydrogen peroxide) were detected and the potential formation pathway as well as ways of enhancing the production of nitric oxide by alteration of the feed gas and the surrounding gas composition during plasma treatment of the liquid have been pointed out.
Atmospheric pressure plasma jets for biomedical applications are often sustained in He with small amounts of, for example, O2 impurities and typically propagate into ambient air. The resulting poorly ...controlled generation of reactive species has motivated the use of gas shields to control the interaction of the plasma plume with the ambient gas. The use of different gases in the shield yields different behavior in the plasma plume. In this paper, we discuss results from experimental and computational investigations of He plasma jets having attaching and non-attaching gas shields. We found that negative ion formation in the He-air mixing region significantly affects the ionization wave dynamics and promotes the propagation of negative guided streamers through an electrostatic focusing mechanism. Results from standard and phase resolved optical emission spectroscopy ratios of emission from states of N2 and He imply different electric fields in the plasma plume depending on the composition of the shielding gas. These effects are attributed to the conductivity in the transition region between the plasma plume and the shield gas, and the immobile charge represented by negative ions. The lower conductivity in the attaching mixtures enables more extended penetration of the electric field whereas the negative ions aid in focusing the electrons towards the axis.
Autoantibodies against desmoglein (Dsg) 1 and Dsg3 primarily cause blister formation in the autoimmune disease pemphigus vulgaris (PV). Src was proposed to contribute to loss of keratinocyte ...cohesion. However, the role and underlying mechanisms are unclear and were studied here. In keratinocytes, cell cohesion in response to autoantibodies was reduced in Src-dependent manner by two patient-derived PV-IgG fractions as well as by AK23 but not by a third PV-IgG fraction, although Src was activated by all autoantibodies. Loss of cell cohesion was progredient in a timeframe of 24 h and AK23, similar to PV-IgG, interfered with reconstitution of cell cohesion after Ca
-switch, indicating that the autoantibodies also interfered with desmosome assembly. Dsg3 co-localized along cell contacts and interacted with the Src substrate cortactin. In keratinocytes isolated from cortactin-deficient mice, cell adhesion was impaired and Src-mediated inhibition of AK23-induced loss of cell cohesion for 24 h was significantly reduced compared to wild-type (wt) cells. Similarly, AK23 impaired reconstitution of cell adhesion was Src-dependent only in the presence of cortactin. Likewise, Src inhibition significantly reduced AK23-induced skin blistering in wt but not cortactin-deficient mice. These data suggest that the Src-mediated long-term effects of AK23 on loss of cell cohesion and skin blistering are dependent on cortactin-mediated desmosome assembly. However, in human epidermis PV-IgG-induced skin blistering and ultrastructural alterations of desmosomes were not affected by Src inhibition, indicating that Src may not be critical for skin blistering in intact human skin, at least when high levels of autoantibodies targeting Dsg1 are present.
Finding a solution for air species contamination of atmospheric pressure plasmas in plasma medical treatment is a major task for the new field of plasma medicine. Several approaches use complex ...climate chambers to control the surrounding atmosphere. In this paper, ambient species are excluded in plasma-human-skin-cell treatment by ensheathing the plasma jet effluent with a shielding gas. Not only does this gas curtain protect the plasma jet effluent from inflow of air species but it also, more importantly, allows controlling the effluent reactive species composition by adjusting the mixture of the shielding gas. In the present investigations, the mixture of nitrogen to oxygen within the gas curtain around an argon atmospheric pressure plasma jet (kinpen) is varied. The resulting reactive plasma components produced in the jet effluent are thus either oxygen or nitrogen dominated. With this gas curtain, the effect of reactive oxygen species (ROS) and reactive nitrogen species (RNS) on the cell viability of indirectly plasma-treated HaCaT skin cells is studied. This human keratinocyte cell line is an established standard for a skin model system. The cell viability is determined by a fluorometric assay, where metabolically active cells transform nonfluorescent resazurin to the highly fluorescent resorufin. Plasma jet and gas curtain are characterized by numerical flow simulation as well as by optical emission spectroscopy. The generation of nitrite within the used standard cell culture medium serves as a measure for generated RNS. Measurements with the leukodye dichlorodihydrofluorescein diacetate show that, despite a variation of the shielding gas mixture, the total amount of generated reactive oxygen plus nitrogen species is constant. It is shown that a plasma dominated by RNS disrupts cellular growth less than a ROS-dominated plasma.
The influence of ambient air species is an ever-present problem for atmospheric pressure plasma jet applications. In particular, applications where the plasma-induced effects are extremely sensitive ...to specific types of ambient species (oxygen, nitrogen, humidity) - as, for example, in plasma medicine - require concepts to exclude or to control ambient species flux into the jet effluent that go beyond an environmental control via process chambers or even vacuum systems. In this paper, we demonstrate how to eliminate ambient species influence on effluent chemistry by ensheathing the effluent. With a designed shielding gas composition, we control the species flowing into the jet effluent and thus control the effluent chemistry. The proposed approach can be applied to the majority of possible jet plasma sources. Flow simulations as well as VUV-absorption spectroscopy measurements prove the gas curtain to be effective in shielding the jet gas from ambient species and show that a control of reactive species within the jet effluent is possible. On the example of plasma treatment of a NaCl solution, we demonstrate that, by adjusting the shielding gas composition, the generation of nitrite and nitrate in the solution can be finely controlled.
The influence of ambient air species especially humidity is an ever-present challenge for atmospheric pressure plasma jet applications. Especially, where the plasma-induced effects are extremely ...sensitive to humidity, such as in the field of plasma medicine, an understanding of the influence of ambient species diffusion on plasma chemistry and on reactive component composition is crucial. In this paper, we investigate the influence of ambient humidity versus feed gas humidity on the production of reactive components by atmospheric pressure plasma jets. By the use of a shielding gas curtain, we control the surrounding atmosphere around the active effluent region of the investigated argon RF-plasma jet (kinpen) and control the gas humidity of the ambient gas. By quantum cascade laser absorption spectroscopy and by Fourier transformed infrared (IR) absorption spectroscopy, the effect of diffusing surrounding molecular species on the chemistry of the long-living reactive oxygen species is investigated. Mechanisms of H 2 O 2 and O 3 production are studied. In this paper, we have quantified the influence that ambient species, namely, water molecules, have on the reactive species' generation in the gas phase. It is shown that the effect of ambient humidity is important for the long-living species production, feed gas humidity, however, has the much stronger effect. Finally, with the focus of applications in plasma medicine, the cell viability of human skin cells (HaCaT keratinocytes) as a function of feed gas and ambient gas humidity is compared.
Tumor-cell infiltration is a major obstacle to successful therapy for brain tumors. Membrane-type matrix metalloproteinases (MT-MMPs), a metzincin subfamily of six proteases, are important mediators ...of infiltration. The cellular source of MT-MMPs and their role in glioma biology, however, remain controversial. Thus, we comprehensively analyzed the expression of MT-MMPs in primary brain tumors. All MT-MMPs were differentially expressed in primary brain tumors. In diffuse gliomas, MT-MMP1, -3, and -4 were predominantly expressed by IDH1mutated tumor cells, while macrophages/microglia contributed significantly less to MT-MMP expression. For functional analyses, individual MT-MMPs were expressed in primary mouse p53−/− astrocytes. Invasion and migration potential of MT-MMP-transduced astrocytes was determined via scratch, matrigel invasion, and novel organotypic porcine spinal slice migration (OPoSSM) and invasion assays. Overall, MT-MMP-transduced astrocytes showed enhanced migration compared to controls. MMP14 was the strongest mediator of migration in scratch assays. However, in the OPoSSM assays, the glycosylphosphatidylinositol (GPI)-anchored MT-MMPs MMP17 and MMP25, not MMP14, mediated the highest infiltration rates of astrocytes. Our data unequivocally demonstrate for the first time that glioma cells, not microglia, are the predominant producers of MT-MMPs in glioma and can act as potent mediators of tumor-cell infiltration into CNS tissue. These proteases are therefore promising targets for therapeutic interventions.
PDF
