Polymyositis And Dermatomyositis - Inflammation, Muscle Structure & Immunosuppressive Treatment

Polymyositis and dermatomyositis are chronic, inflammatory disorders characterized by muscle weakness, low muscle endurance and by inflammation in skeletal muscle tissues. The pathogenesis is not completely understood and several mechanisms are believed to be involved. Conventional immunosuppressive treatment for patients is primarily with glucocorticoids in combination with another immune-modulating drug. Although most patients respond to the treatment to some degree, persisting muscle weakness as well as continued presence of muscle tissue inflammation is often evident. The main aim was to investigate different pathways that may play a role in the disease pathogenesis and contribute to the muscle weakness in patients with polymyositis and dermatomyositis. We wanted to determine whether these mechanisms were associated with clinical outcome and if immunosuppressive treatment could alter the processes. Studies were thus performed by characterizing muscle tissue samples from patients before and after immunosuppressive treatment, and through comparison with samples from healthy individuals. We also collected in vivo samples from quadriceps muscles by microdialysis before and after exercise in both patients and healthy individuals. Some novel observations were made. Firstly, we determined that the pro-inflammatory cytokine interleukin (IL) 15 and the IL-15 receptor alpha (IL-15Rα) were over-expressed in muscle tissues from the patients compared to in healthy individuals. Immunosuppressive treatment had no effect on IL-15 expression in 1/3 of the patients along with a poorer functional outcome. Secondly, we investigated the prostaglandin (PG) E2 and leukotriene (LT) B4 pathways. Here we found that PGE2 pathway is up-regulated in muscle tissue from patients with polymyositis or dermatomyositis compared to healthy individuals. A higher expression of cyclooxygenase (COX) 1, COX-2 and microsomal PGE synthase (mPGES)-1 in the patients was apparent and while COX-2 was down-regulated by treatment, COX-1 and mPGES-1 levels remained high. LTB4 pathway was also found to be active in patients and the expression of both 5-lipoxygenase (5-LO) and the LTB4 receptor 1 (BLT1) were higher compared to in healthy individuals. 5-LO activating protein (FLAP), was highly expressed in a subgroup of patients who did not respond to immunosuppressive treatment. These results may suggest an involvement of PGE2 and LTB4 in vascular permeability and chemotaxis for infiltrating immune cells, but perhaps also to serve a purpose in skeletal muscle regeneration in patients with polymyositis and dermatomyositis. Lastly, we could report that patients with newly diagnosed polymyositis or dermatomyositis did not display a low proportion of oxidative type I muscle fibers typically seen in muscle tissue of patients in a chronic state of disease. This finding implies that the proportion of oxidative fibers does not contribute to the low muscle endurance, at least not in early polymyositis and dermatomyositis. The conclusions drawn from this thesis are that several mechanisms may contribute to the muscle weakness in different subgroups of polymyositis and dermatomyositis patients. New pathways discovered in this thesis include IL-15 and the eicosanoids PGE2 and LTB4, but the fiber type composition in the muscle tissue might play a lesser role, at least in early disease. The mechanisms identified could provide a basis for further studies and possibly new therapies

A test of the pioneer factor hypothesis using ectopic liver gene activation

The pioneer factor hypothesis (PFH) states that pioneer factors (PFs) are a subclass of transcription factors (TFs) that bind to and open inaccessible sites and then recruit non-pioneer factors (non-PFs) that activate batteries of silent genes. The PFH predicts that ectopic gene activation requires the sequential activity of qualitatively different TFs. We tested the PFH by expressing the endodermal PF FOXA1 and non-PF HNF4A in K562 lymphoblast cells. While co-expression of FOXA1 and HNF4A activated a burst of endoderm-specific gene expression, we found no evidence for a functional distinction between these two TFs. When expressed independently, both TFs bound and opened inaccessible sites, activated endodermal genes, and \u27pioneered\u27 for each other, although FOXA1 required fewer copies of its motif for binding. A subset of targets required both TFs, but the predominant mode of action at these targets did not conform to the sequential activity predicted by the PFH. From these results, we hypothesize an alternative to the PFH where \u27pioneer activity\u27 depends not on categorically different TFs but rather on the affinity of interaction between TF and DNA

Activation domains can decouple the mean and noise of gene expression

Regulatory mechanisms set a gene\u27s average level of expression, but a gene\u27s expression constantly fluctuates around that average. These stochastic fluctuations, or expression noise, play a role in cell-fate transitions, bet hedging in microbes, and the development of chemotherapeutic resistance in cancer. An outstanding question is what regulatory mechanisms contribute to noise. Here, we demonstrate that, for a fixed mean level of expression, strong activation domains (ADs) at low abundance produce high expression noise, while weak ADs at high abundance generate lower expression noise. We conclude that differences in noise can be explained by the interplay between a TF\u27s nuclear concentration and the strength of its AD\u27s effect on mean expression, without invoking differences between classes of ADs. These results raise the possibility of engineering gene expression noise independently of mean levels in synthetic biology contexts and provide a potential mechanism for natural selection to tune the noisiness of gene expression

Transcription factor interactions explain the context-dependent activity of CRX binding sites

The effects of transcription factor binding sites (TFBSs) on the activity of a cis-regulatory element (CRE) depend on the local sequence context. In rod photoreceptors, binding sites for the transcription factor (TF) Cone-rod homeobox (CRX) occur in both enhancers and silencers, but the sequence context that determines whether CRX binding sites contribute to activation or repression of transcription is not understood. To investigate the context-dependent activity of CRX sites, we fit neural network-based models to the activities of synthetic CREs composed of photoreceptor TFBSs. The models revealed that CRX binding sites consistently make positive, independent contributions to CRE activity, while negative homotypic interactions between sites cause CREs composed of multiple CRX sites to function as silencers. The effects of negative homotypic interactions can be overcome by the presence of other TFBSs that either interact cooperatively with CRX sites or make independent positive contributions to activity. The context-dependent activity of CRX sites is thus determined by the balance between positive heterotypic interactions, independent contributions of TFBSs, and negative homotypic interactions. Our findings explain observed patterns of activity among genomic CRX-bound enhancers and silencers, and suggest that enhancers may require diverse TFBSs to overcome negative homotypic interactions between TFBSs

Effects on muscle tissue remodeling and lipid metabolism in muscle tissue from adult patients with polymyositis or dermatomyositis treated with immunosuppressive agents.

BACKGROUND: Polymyositis (PM) and dermatomyositis (DM) are autoimmune muscle diseases, conventionally treated with high doses of glucocorticoids in combination with immunosuppressive drugs. Treatment is often dissatisfying, with persisting muscle impairment. We aimed to investigate molecular mechanisms that might contribute to the persisting muscle impairment despite immunosuppressive treatment in adult patients with PM or DM using gene expression profiling of repeated muscle biopsies. METHODS: Paired skeletal muscle biopsies from six newly diagnosed adult patients with DM or PM taken before and after conventional immunosuppressive treatment were examined by gene expression microarray analysis. Selected genes that displayed changes in expression were analyzed by Western blot. Muscle biopsy sections were evaluated for inflammation, T lymphocytes (CD3), macrophages (CD68), major histocompatibility complex (MHC) class I expression and fiber type composition. RESULTS: After treatment, genes related to immune response and inflammation, including inflammasome pathways and interferon, were downregulated. This was confirmed at the protein level for AIM-2 and caspase-1 in the inflammasome pathway. Changes in genes involved in muscle tissue remodeling suggested a negative effect on muscle regeneration and growth. Gene markers for fast type II fibers were upregulated and fiber composition was switched towards type II fibers in response to treatment. The expression of genes involved in lipid metabolism was altered, suggesting a potential lipotoxic effect on muscles of the immunosuppressive treatment. CONCLUSION: The anti-inflammatory effect of immunosuppressive treatment was combined with negative effects on genes involved in muscle tissue remodeling and lipid metabolism, suggesting a negative effect on recovery of muscle performance which may contribute to persisting muscle impairment in adult patients with DM and PM

Helicobacter pylori patient isolates from South Africa and Nigeria differ in virulence factor pathogenicity profile and associated gastric disease outcome

Helicobacter pylori is a gram-negative, spiral-shaped bacterial pathogen and the causative agent for gastritis, peptic ulcer disease and classified as a WHO class I carcinogen. While the prevalence of H. pylori infections in Africa is among the highest in the world, the incidence of gastric cancer is comparably low. Little is known about other symptoms related to the H. pylori infection in Africa and the association with certain phenotypes of bacterial virulence. We established a network of study sites in Nigeria (NG) and South Africa (ZA) to gain an overview on the epidemiological situation. In total 220 isolates from 114 patients were analyzed and 118 different patient isolates examined for the presence of the virulence factors cagA, vacA, dupA, their phylogenetic origin and their resistance against the commonly used antibiotics amoxicillin, clarithromycin, metronidazole and tetracycline. We report that H. pylori isolates from Nigeria and South Africa differ significantly in their phylogenetic profiles and in their expression of virulence factors. VacA mosaicism is intensive, resulting in m1-m2 vacA chimeras and frequent s1m1 and s1m2 vacA subtypes in hpAfrica2 strains. Gastric lesions were diagnosed more frequent in Nigerian versus South African patients and H. pylori isolates that are resistant against one or multiple antibiotics occur frequently in both countries

Helicobacter pylori cag-Pathogenicity Island-Dependent Early Immunological Response Triggers Later Precancerous Gastric Changes in Mongolian Gerbils

Infection with Helicobacter pylori, carrying a functional cag type IV secretion system (cag-T4SS) to inject the Cytotoxin associated antigen (CagA) into gastric cells, is associated with an increased risk for severe gastric diseases in humans. Here we studied the pathomechanism of H. pylori and the role of the cag-pathogenicity island (cag-PAI) for the induction of gastric ulcer and precancerous conditions over time (2–64 weeks) using the Mongolian gerbil model. Animals were challenged with H. pylori B128 (WT), or an isogenic B128ΔcagY mutant-strain that produces CagA, but is unable to translocate it into gastric cells. H. pylori colonization density was quantified in antrum and corpus mucosa separately. Paraffin sections were graded for inflammation and histological changes verified by immunohistochemistry. Physiological and inflammatory markers were quantitated by RIA and RT-PCR, respectively. An early cag-T4SS-dependent inflammation of the corpus mucosa (4–8 weeks) occurred only in WT-infected animals, resulting in a severe active and chronic gastritis with a significant increase of proinflammatory cytokines, mucous gland metaplasia, and atrophy of the parietal cells. At late time points only WT-infected animals developed hypochlorhydria and hypergastrinemia in parallel to gastric ulcers, gastritis cystica profunda, and focal dysplasia. The early cag-PAI-dependent immunological response triggers later physiological and histopathological alterations towards gastric malignancies

Helicobacter pylori strains from a Nigerian cohort show divergent antibiotic resistance rates and a uniform pathogenicity profile

Antibiotic resistance in Helicobacter pylori is a factor preventing its successful eradication. Particularly in developing countries, resistance against commonly used antibiotics is widespread. Here, we present an epidemiological study from Nigeria with 111 isolates. We analyzed the associated disease outcome, and performed a detailed characterization of these isolated strains with respect to their antibiotic susceptibility and their virulence characteristics. Furthermore, statistical analysis was performed on microbiological data as well as patient information and the results of the gastroenterological examination. We found that the variability concerning the production of virulence factors between strains was minimal, with 96.4% of isolates being CagA-positive and 92.8% producing detectable VacA levels. In addition, high frequency of bacterial resistance was observed for metronidazole (99.1%), followed by amoxicillin (33.3%), clarithromycin (14.4%) and tetracycline (4.5%). In conclusion, this study indicated that the infection rate of H. pylori infection within the cohort in the present study was surprisingly low (36.6%). Furthermore, an average gastric pathology was observed by histological grading and bacterial isolates showed a uniform pathogenicity profile while indicating divergent antibiotic resistance rates