Interleukin-23 is critical for full-blown expression of a non-autoimmune destructive arthritis and regulates interleukin-17A and RORγt in γδ T cells

Introduction: Interleukin (IL)-23 is essential for the development of various experimental autoimmune models. However, the role of IL-23 in non-autoimmune experimental arthritis remains unclear. Here, we examined the role of IL-23 in the non-autoimmune antigen-induced arthritis (AIA) model. In addition, the regulatory potential of IL-23 in IL-17A and retinoic acid-related orphan receptor gamma t (RORγt) expression in CD4+and TCRγδ+T cells was evaluated systemically as well as at the site of inflammation.Methods: Antigen-induced arthritis was induced in wild-type, IL-23p19-deficient and IL-17 Receptor A - knockout mice. At differe

Intracellular signaling in key pathways is induced by treatment with ultrasound and microbubbles in a leukemia cell line, but not in healthy peripheral blood mononuclear cells

Treatment with ultrasound and microbubbles (sonoporation) to enhance therapeutic efficacy in cancer therapy is rapidly expanding, but there is still very little consensus as to why it works. Despite the original assumption that pore formation in the cell membrane is responsible for increased uptake of drugs, the molecular mechanisms behind this phenomenon are largely unknown. We treated cancer cells (MOLM-13) and healthy peripheral blood mononuclear cells (PBMCs) with ultrasound at three acoustic intensities (74, 501, 2079 mW/cm2) ± microbubbles. We subsequently monitored the intracellular response of a number of key signaling pathways using flow cytometry or western blotting 5 min, 30 min and 2 h post-treatment. This was complemented by studies on uptake of a cell impermeable dye (calcein) and investigations of cell viability (cell count, Hoechst staining and colony forming assay). Ultrasound + microbubbles resulted in both early changes (p38 (Arcsinh ratio at high ultrasound + microbubbles: +0.5), ERK1/2 (+0.7), CREB (+1.3), STAT3 (+0.7) and AKT (+0.5)) and late changes (ribosomal protein S6 (Arcsinh ratio at low ultrasound: +0.6) and eIF2α in protein phosphorylation). Observed changes in protein phosphorylation corresponded to changes in sonoporation efficiency and in viability, predominantly in cancer cells. Sonoporation induced protein phosphorylation in healthy cells was pronounced (p38 (+0.03), ERK1/2 (−0.03), CREB (+0.0), STAT3 (−0.1) and AKT (+0.04) and S6 (+0.2)). This supports the hypothesis that sonoporation may enhance therapeutic efficacy of cancer treatment, without causing damage to healthy cells.publishedVersio

Humanized Ovarian Cancer Patient-Derived Xenografts for Improved Preclinical Evaluation of Immunotherapies

High-grade serous ovarian cancer (HGSOC) has poor prognosis and new treatment modalities are needed. Immunotherapy, with checkpoint inhibitors, have demonstrated limited impact. To evaluate the suitability for immunotherapeutics, contextualized preclinical models are required to secure meaningful clinical translation. Therefore, we developed and characterized humanized patient-derived xenograft (hu PDX) murine models of HGSOC, which were established by orthotopic implantation of tumor cell suspensions and intravenous injection of CD34+ cells isolated from umbilical cord blood samples. The developing human immune system in NSG and NSGS mice was followed longitudinally by flow cytometry and characterized by mass cytometry with a panel of 34 surface markers. Molecular imaging of tumor burden, survival analysis, and characterization of tumor-infiltrating immune cells was performed to assess the treatment response to anti-PD-1 (nivolumab) monotherapy. Successful generation of hu PDX models was achieved. Mice treated with nivolumab showed a decrease in tumor burden, however no significant survival benefit was identified when compared to untreated controls. No correlation was seen between PD-L1 expression and CD8 T cell infiltration and response parameters. As the characterization showed an immune infiltration of predominantly myeloid cells, similar to what is observed in HGSOC patients, the models may have the potential to evaluate the importance of myeloid cell immunomodulation as well.publishedVersio

Tyrosine kinase inhibitors and interferon‐α increase tunneling nanotube (TNT) formation and cell adhesion in chronic myeloid leukemia (CML) cell lines (

Chronic myeloid leukemia (CML) is a stem cell disease of the bone marrow where mechanisms of inter‐leukemic communication and cell‐to‐cell interactions are proposed to be important for optimal therapy response. Tunneling nanotubes (TNTs) are novel intercellular communication structures transporting different cargos with potential implications in therapy resistance. Here, we have investigated TNTs in CML cells and following treatment with the highly effective CML therapeutics tyrosine kinase inhibitors (TKIs) and interferon‐α (IFNα). CML cells from chronic phase CML patients as well as the blast crisis phase cell lines, Kcl‐22 and K562, formed few or no TNTs. Treatment with imatinib increased TNT formation in both Kcl‐22 and K562 cells, while nilotinib or IFNα increased TNTs in Kcl‐22 cells only where the TNT increase was associated with adherence to fibronectin‐coated surfaces, altered morphology, and reduced movement involving β1integrin. Ex vivo treated cells from chronic phase CML patients showed limited changes in TNT formation similarly to bone marrow cells from healthy individuals. Interestingly, in vivo nilotinib treatment in a Kcl‐22 subcutaneous mouse model resulted in morphological changes and TNT‐like structures in the tumor‐derived Kcl‐22 cells. Our results demonstrate that CML cells express low levels of TNTs, but CML therapeutics increase TNT formation in designated cell models indicating TNT functionality in bone marrow derived malignancies and their microenvironment.publishedVersio

Clinical categories of patients and encounter rates in primary health care – a three-year study in defined populations

BACKGROUND: The objective was to estimate the proportion of inhabitants with a diagnosis-registered encounter with a general practitioner, and to elucidate annual variations of clinical categories of patients in terms of their individual comorbidity. METHODS: A three-year retrospective study of encounter data from electronic patient records, with an annual-based application of the Johns Hopkins Adjusted Clinical Groups (ACG) system. Data were retrieved from every patient with a diagnosis-registered encounter with a GP during the period 2001–2003 at 13 publicly managed primary health care centres in Blekinge county, southeastern Sweden, with about 150000 inhabitants. Main outcome measures: Proportions of inhabitants with a diagnosis-registered encounter, and ranges of the annual proportions of categories of patients according to ACGs. RESULTS: The proportion of inhabitants with a diagnosis-registered encounter ranged from about 64.0% to 90.6% for the primary health care centres, and averaged about 76.5% for all inhabitants. In a three-year perspective the average range of categories of patients was about 0.4% on the county level, and about 0.9% on the primary health care centre level. About one third of the patients each year had a constellation of two or more types of morbidity. CONCLUSION: About three fourths of all inhabitants had one or more diagnosis-registered encounters with a general practitioner during the three-year period. The annual variation of categories of patients according to ACGs was small on both the county and the primary health care centre level. The ACG system seems useful for demonstrating and predicting various aspects of clinical categories of patients in Swedish primary health care

Coupling of alpha(1)-Adrenoceptors to ERK1/2 in the Human Prostate

Introduction: alpha(1)-Adrenoceptors are considered critical for the regulation of prostatic smooth muscle tone. However, previous studies suggested further alpha(1)-adrenoceptor functions besides contraction. Here, we investigated whether alpha(1)-adrenoceptors in the human prostate may activate extracellular signal-regulated kinases (ERK1/2). Methods: Prostate tissues from patients undergoing radical prostatectomy were stimulated in vitro. Activation of ERK1/2 was assessed by Western blot analysis. Expression of ERK1/2 was studied by immunohistochemistry. The effect of ERK1/2 inhibition by U0126 on phenylephrine-induced contraction was studied in organ-bath experiments. Results: Stimulation of human prostate tissue with noradrenaline (30 mu M) or phenylephrine (10 mu M) resulted in ERK activation. This was reflected by increased levels of phosphorylated ERK1/2. Expression of ERK1/2 in the prostate was observed in smooth muscle cells. Incubation of prostate tissue with U0126 (30 mu M) resulted in ERK1/2 inhibition. Dose-dependent phenylephrine-induced contraction of prostate tissue was not modulated by U0126. Conclusions: alpha(1)-Adrenoceptors in the human prostate are coupled to ERK1/2. This may partially explain previous observations suggesting a role of alpha(1)-adrenoceptors in the regulation of prostate growth. Copyright (C) 2011 S. Karger AG, Base

Locomotor adaptability in persons with unilateral transtibial amputation

Background Locomotor adaptation enables walkers to modify strategies when faced with challenging walking conditions. While a variety of neurological injuries can impair locomotor adaptability, the effect of a lower extremity amputation on adaptability is poorly understood. Objective Determine if locomotor adaptability is impaired in persons with unilateral transtibial amputation (TTA). Methods The locomotor adaptability of 10 persons with a TTA and 8 persons without an amputation was tested while walking on a split-belt treadmill with the parallel belts running at the same (tied) or different (split) speeds. In the split condition, participants walked for 15 minutes with the respective belts moving at 0.5 m/s and 1.5 m/s. Temporal spatial symmetry measures were used to evaluate reactive accommodations to the perturbation, and the adaptive/de-adaptive response. Results Persons with TTA and the reference group of persons without amputation both demonstrated highly symmetric walking at baseline. During the split adaptation and tied post-adaptation walking both groups responded with the expected reactive accommodations. Likewise, adaptive and de-adaptive responses were observed. The magnitude and rate of change in the adaptive and de-adaptive responses were similar for persons with TTA and those without an amputation. Furthermore, adaptability was no different based on belt assignment for the prosthetic limb during split adaptation walking. Conclusions Reactive changes and locomotor adaptation in response to a challenging and novel walking condition were similar in persons with TTA to those without an amputation. Results suggest persons with TTA have the capacity to modify locomotor strategies to meet the demands of most walking conditions despite challenges imposed by an amputation and use of a prosthetic limb