Identification of the novel HLA class I allele, HLA-A*31:215, identified by Next Generation Sequencing.

The novel HLA allele HLA-A*31:215 differs from HLA-A*31:01:02:01 by one nucleotide substitution c.G832A in exon 4.

[Extrapelvic endometriosis is a difficult diagnosis].

Extrapelvic endometriosis is a rare type of endometriosis. The diagnosis can be challenging, especially if the patient lacks characteristic endometriosis symptoms. This case report presents a 27-year-old woman diagnosed with both primary umbilical endometriosis and infiltrating endometriosis of the diaphragm. The woman presented with a painful bluish tumour in the umbilicus and cyclic pain in her upper right abdomen, but no lower abdominal pain. Endometriosis should be one of the differential diagnoses when symptoms like pain and/or swelling is cyclic and menstrual-related.

Cross-sectorial collaboration in return to work interventions: perspectives from patients, mental health care professionals and case managers in the social insurance sector.

PURPOSE: Common mental disorders (CMD) are major causes of sick leave and have huge negative impacts on psychosocial well-being and to overall society. The return to work (RTW) process involves various welfare sectors, but often lack collaboration across these sectors and may cause uneven RTW processes. This study reports from an intervention aimed to increase collaboration between the mental health care system (MHCS) and the social insurance sector (SIS). The intervention consisted of two phases: (a) a development phase building on a participatory approach and (b) an implementation phase. Interprofessional meetings, evaluation workshops and joint consultations with workers from SIS and MHCS, and patients, comprised the key components of the intervention. The study explores how the collaborative process is experienced during the implementation phase with a focus on how collaboration maintains and challenges professional identities. METHODS: A qualitative design was employed consisting of individual interviews (n20), focus group discussions (4), and observations of interactions between Patients with CMDs, case managers from SIS, and professionals working in MHCS. RESULTS: Patients with CMDs felt supported and did not feel pushed to RTW prematurely. SIS and MHCS professionals reported that they were not sufficiently prepared and it was not clear who was responsible for which tasks. MHCS professionals experienced that their professional identities were in conflict with the aim of the intervention whereas SIS professionals found that the intervention improved their poor reputation. CONCLUSIONS: The results suggest that a higher level of participation in the intervention design and implementation process can improve a clear and transparent role distribution between professionals in MHCS and SIS.IMPLICATIONS FOR REHABILITATIONCollaboration across sectors is key to successful Return-to-work after sickness-related absence.Professional identities are challenged in collaborative interventions as different agendas are at play.Collaboration is most likely to be successful when engaging involved professions to decide goals and methods which are compatible with local work procedures.

Perinatal exposure to the thyroperoxidase inhibitors methimazole and amitrole perturbs thyroid hormone system signaling and alters motor activity in rat offspring.

Disruption of the thyroid hormone system during development can impair brain development and cause irreversible damage. Some thyroid hormone system disruptors act by inhibiting the thyroperoxidase (TPO) enzyme, which is key to thyroid hormone synthesis. For the potent TPO-inhibiting drug propylthiouracil (PTU) this has been shown to result in thyroid hormone system disruption and altered brain development in animal studies. However, an outstanding question is which chemicals beside PTU can cause similar effects on brain development and to what degree thyroid hormone insufficiency must be induced to be able to measure adverse effects in rats and their offspring. To start answering these questions, we performed a perinatal exposure study in pregnant rats with two TPO-inhibitors: the drug methimazole (MMI) and the triazole herbicide amitrole. The study involved maternal exposure from gestational day 7 through to postnatal day 22, to MMI (8 and 16 mg/kg body weight/day) or amitrole (25 and 50 mg/kg body weight/day). Both MMI and amitrole reduced serum T4 concentrations in a dose-dependent manner in dams and offspring, with a strong activation of the hypothalamic-pituitary-thyroid axis. This reduction in serum T4 led to decreased thyroid hormone-mediated gene expression in the offspring's brains and caused adverse effects on brain function, seen as hyperactivity and decreased habituation in preweaning pups. These dose-dependent effects induced by MMI and amitrole are largely the same as those observed with PTU. This demonstrates that potent TPO-inhibitors can induce effects on brain development in rats and that these effects are driven by T4 deficiency. This knowledge will aid the identification of TPO-inhibiting thyroid hormone system disruptors in a regulatory context and can serve as a starting point in search of more sensitive markers of developmental thyroid hormone system disruption.

Voluntary wheel running can lead to modulation of immune checkpoint molecule expression.

BACKGROUND: Exercise and physical activity (PA) are associated with reduced tumor growth and enhanced intra-tumoral immune cell infiltration in mice. We aimed to investigate the role of PA achieved by voluntary wheel running in promoting the immunogenic profile across several murine tumor models, and to explore the potential of checkpoint blockade and PA in the form of voluntary wheel running as combination therapy. MATERIAL AND METHODS: The experiments were performed with C57BL/6 mice bearing subcutaneous tumors while having access to running wheels in their cages, where key immunoregulatory molecules expressed in the tumor tissue were measured by qPCR. Furthermore, we tested the hypothesis that wheel running combined with PD-L1 -or PD-1 inhibitor treatment could lead to an additive effect on tumor growth in mice bearing B16 melanoma tumors. RESULTS: Wheel running increased immune checkpoint expression (PD-1, PD-L1, PD-L2, CD28, B7.1 and B7.2) in B16 tumor-bearing mice, while induction of only PD-L2 was found in E0771 breast cancer and Lewis Lung Cancer. In studies combining voluntary wheel running with PD-1 -and PD-L1 inhibitors we found significant effects of wheel running on attenuating B16 melanoma tumor growth, in line with previous studies. We did, however, not find an additive effect of combining either of the two immunotherapeutic treatments with access to running wheels. CONCLUSION: B16 tumors displayed upregulated expression of immune regulatory molecules and decreased tumor growth in response to PA. However, combining PA with PD-1 or PD-L1 blockade did not lead to a further augmented inhibition of tumor growth.

Cost-Effectiveness of Insulin Degludec Versus Insulin Glargine U300 in the Netherlands: Evidence From a Randomised Controlled Trial.

INTRODUCTION: This study aimed to evaluate the short-term cost-effectiveness of insulin degludec 200 units/mL (degludec) versus insulin glargine 300 units/mL (glargine U300) from a Dutch societal perspective. METHODS: A previously published model estimated costs [2018 euros (EUR)] and effectiveness [quality-adjusted life years (QALYs)] with degludec compared with glargine U300 over a 1-year time horizon. The model captured hypoglycaemia rates and insulin dosing. Clinical outcomes were informed by CONCLUDE (NCT03078478), a head-to-head randomised controlled trial in insulin-experienced patients with type 2 diabetes. RESULTS: Treatment with degludec was associated with mean annual cost savings (EUR 24.71 per patient) relative to glargine U300, driven by a lower basal insulin dose and lower severe hypoglycaemia rate with degludec compared with glargine U300. Lower rates of non-severe nocturnal and severe hypoglycaemia resulted in improved effectiveness (+ 0.0045 QALYs) with degludec relative to glargine U300. In sensitivity analyses, changes to the vast majority of model parameters did not materially affect model outcomes. CONCLUSIONS: This short-term analysis, informed by the latest clinical trial evidence, demonstrated that degludec was a cost-effective treatment option relative to glargine U300. As such, our modelling analysis suggests that degludec would represent an efficient use of Dutch public healthcare resources in this patient population.

Exercise-Mediated Lowering of Glutamine Availability Suppresses Tumor Growth and Attenuates Muscle Wasting.

Glutamine is a central nutrient for many cancers, contributing to the generation of building blocks and energy-promoting signaling necessary for neoplastic proliferation. In this study, we hypothesized that lowering systemic glutamine levels by exercise may starve tumors, thereby contributing to the inhibitory effect of exercise on tumor growth. We demonstrate that limiting glutamine availability, either pharmacologically or physiologically by voluntary wheel running, significantly attenuated the growth of two syngeneic murine tumor models of breast cancer and lung cancer, respectively, and decreased markers of atrophic signaling in muscles from tumor-bearing mice. In continuation, wheel running completely abolished tumor-induced loss of weight and lean body mass, independently of the effect of wheel running on tumor growth. Moreover, wheel running abolished tumor-induced upregulation of muscular glutamine transporters and myostatin signaling. In conclusion, our data suggest that voluntary wheel running preserves muscle mass by counteracting muscular glutamine release and tumor-induced atrophic signaling.

Recurrent apical prolapse after high uterosacral ligament suspension - in a heterogenous cohort characterised by a high prevalence of previous pelvic operations.

BACKGROUND: The apical prolapse is probably the most complex form of pelvic organ prolapse (POP). Adequate apical support is essential in the treatment of POP, as it contributes to the support in all vaginal compartments. This study aimed to evaluate the rate of symptomatic recurrent apical prolapse after high uterosacral ligament suspension (HUSLS), in a cohort of women characterised by a high prevalence of previous pelvic operations and a significant degree of prolapse. METHODS: This is a retrospective chart review of 95 women who underwent HUSLS for symptomatic apical prolapse from 2002 to 2009 at Aarhus University Hospital, Denmark. Of these women, 97% attended a six-month clinical control. Recurrence was defined as symptomatic vaginal vault prolapse stage 2 or more (according to the International Continence Society (ICS) quantification system). Medical charts were reviewed for a mean period of 7.2 years. Any new contacts due to prolapse were noted. RESULTS: Before the operation, 73% of the women were hysterectomised, and 52% had previous prolapse surgery. Stage 2 apical prolapse was reported in 71% of the women, whereas 26% had stage 3 or 4. At six-month follow-up, 19% of the women had recurrent symptomatic apical prolapse, and 9% of the women had symptomatic recurrent prolapse in other compartments 6 months after operation. In all, 35% of the women had a renewed prolapse operation during the long-term follow-up period. Perioperative adverse events were seen in 7%. Two women were re-operated due to postoperative complications. CONCLUSIONS: This retrospective study of 95 women with a significant degree of prolapse and a high prevalence of previous pelvic operations demonstrates that the rate of recurrent prolapse associated with HUSLS might be higher than originally described. In conclusion, HUSLS may not be the optimal first choice of operation in this group of patients.

Every exercise bout matters: linking systemic exercise responses to breast cancer control.

Cumulative epidemiological evidence shows that regular exercise lowers the risk of developing breast cancer and decreases the risk of disease recurrence. The causality underlying this relation has not been fully established, and the exercise recommendations for breast cancer patients follow the general physical activity guidelines, prescribing 150 min of exercise per week. Thus, elucidations of the causal mechanisms are important to prescribe and implement the most optimal training regimen in breast cancer prevention and treatment. The prevailing hypothesis on the positive association within exercise oncology has focused on lowering of the basal systemic levels of cancer risk factors with exercise training. However, another rather overlooked systemic exercise response is the marked acute increases in several potential anti-cancer components during each acute exercise bout. Here, we review the evidence of the exercise-mediated changes in systemic components with the ability to influence breast cancer progression. In the first part, we focus on systemic risk factors for breast cancer, i.e., sex hormones, insulin, and inflammatory markers, and their adaptation to long-term training. In the second part, we describe the systemic factors induced acutely during exercise, including catecholamines and myokines. In conclusion, we propose that the transient increases in exercise factors during acute exercise appear to be mediating the positive effect of regular exercise on breast cancer progression.

Voluntary Running Suppresses Tumor Growth through Epinephrine- and IL-6-Dependent NK Cell Mobilization and Redistribution.

Regular exercise reduces the risk of cancer and disease recurrence. Yet the mechanisms behind this protection remain to be elucidated. In this study, tumor-bearing mice randomized to voluntary wheel running showed over 60% reduction in tumor incidence and growth across five different tumor models. Microarray analysis revealed training-induced upregulation of pathways associated with immune function. NK cell infiltration was significantly increased in tumors from running mice, whereas depletion of NK cells enhanced tumor growth and blunted the beneficial effects of exercise. Mechanistic analyses showed that NK cells were mobilized by epinephrine, and blockade of ß-adrenergic signaling blunted training-dependent tumor inhibition. Moreover, epinephrine induced a selective mobilization of IL-6-sensitive NK cells, and IL-6-blocking antibodies blunted training-induced tumor suppression, intratumoral NK cell infiltration, and NK cell activation. Together, these results link exercise, epinephrine, and IL-6 to NK cell mobilization and redistribution, and ultimately to control of tumor growth.

[Metformin and the obstetric patient]. / Metformin og obstetriske patienter

Polycystic ovary syndrome (PCOS) is associated with insulin resistance, infertility, obesity and gestational complications. Metformin is widely used in fertility treatment of women with PCOS, due to a suggested positive effect of continued metformin treatment beyond the first trimester on pregnancy complications. Larger randomized trials have failed to confirm this. Metformin treatment has not been found to be superior to insulin treatment in women with gestational diabetes and may be associated with long-term consequences in the children in the form of overweight and disturbed glucose metabolism.

Impaired oxidative capacity due to decreased CPT1b levels as a contributing factor to fat accumulation in obesity.

To characterize mechanisms responsible for fat accumulation we used a selectively bred obesity-prone (OP) and obesity-resistant (OR) rat model where the rats were fed a Western diet for 76 days. Body composition was assessed by magnetic resonance imaging scans, and as expected, the OP rats developed a higher degree of fat accumulation compared with OR rats. Indirect calorimetry showed that the OP rats had higher respiratory exchange ratio (RER) compared with OR rats, indicating an impaired ability to oxidize fat. The OP rats had lower expression of carnitine palmitoyltransferase 1b in intra-abdominal fat, and higher expression of stearoyl-CoA desaturase 1 in subcutaneous fat compared with OR rats, which could explain the higher fat accumulation and RER values. Basal metabolic parameters were also examined in juvenile OP and OR rats before and during the introduction of the Western diet. Juvenile OP rats likewise had higher RER values, indicating that this trait may be a primary and contributing factor to their obese phenotype. When the adult obese rats were exposed to the orexigenic and adipogenic hormone ghrelin, we observed increased RER values in both OP and OR rats, while OR rats were more sensitive to the orexigenic effects of ghrelin as well as ghrelin-induced attenuation of activity and energy expenditure. Thus increased fat accumulation characterizing obesity may be caused by impaired oxidative capacity due to decreased carnitine palmitoyltransferase 1b levels in the white adipose tissue, whereas ghrelin sensitivity did not seem to be a contributing factor.

Genetic variation and posttranslational modification of bovine κ-casein: effects on caseino-macropeptide release during renneting.

Chymosin-induced cleavage of κ-casein (κ-CN) occurs during the first enzymatic phase in milk coagulation during cheese manufacturing, where the hydrophilic C-terminal peptide of κ-CN, named caseino-macropeptide (CMP), is released into the whey. The CMP peptide is known for its rather heterogeneous composition with respect to both genetic variation and multiple posttranslational modifications, including phosphorylation and O-linked glycosylation. An approach of liquid chromatography coupled with mass spectrometry was used to investigate (1) the overall protein profile and (2) the release of various forms of CMP after addition of chymosin to individual cow milk samples from 2 breeds, Danish Jersey (DJ) and Danish Holstein-Friesian (DH). The cows were selected to represent distinct homo- and heterozygous types of the κ-CN genetic variants A, B, and E (i.e., genotypes AA, BB, AB, EE, and AE). Initially, investigation of the protein profile showed milk with κ-CN BB exhibited the highest relative content of κ-CN, whereas AE milk exhibited the lowest, and after 40min of renneting >90% of intact κ-CN was hydrolyzed by chymosin in milk representing all κ-CN genotype. By in-depth analysis of the CMP chromatographic profile, multiple CMP isoforms with 1 to 3 O-linked glycans (1-3 G) and 1 to 3 phosphate groups (1-3 P) were identified, as well as nonmodified CMP isoforms. The number of identified CMP isoforms varied to some extent between breeds (21CMP isoforms identified in DJ, 26CMP isoforms in DH) and between κ-CN genetic variants (CMP variant A being the most heterogeneous compared with CMP B and E), as well as between individual samples within each breed. The predominant forms of glycans attached to CMP were found to be the acidic tetrasaccharide {N-acetyl-neuraminic acid α(2-3)galactose ß(1-3)[N-acetyl-neuraminic acid α(2-6)]N-acetyl galactose} or trisaccharides {N-acetyl-neuraminic acid α(2-3)galactose ß(1-3)N-acetyl galactose and galactose ß(1-3)[N-acetyl-neuraminic acid (α2-6)]N-acetyl galactose}. The CMP release was calculated to follow first-order kinetics and was determined by the measurement of CMP content during renneting. The highest rate of release for all CMP isoforms occurred from 0 to 2min after chymosin addition. Concurring results from both breeds showed that CMP variant A with 1-2 P had the highest reaction rate of CMP release, followed by CMP B 1-2 P and then by CMP E 1-2 P (only in DH). All the identified glycosylated CMP isoforms had lower reaction rates of release compared with that of nonglycosylated CMP, thus glycan modifications seemed to negatively influence the reaction rate of chymosin-induced hydrolysis of κ-CN.

High sensitivity point-of-care device for direct virus diagnostics.

Influenza infections are associated with high morbidity and mortality, carry the risk of pandemics, and pose a considerable economic burden worldwide. To improve the management of the illness, it is essential with accurate and fast point-of-care diagnostic tools for use in the field or at the patient's bedside. Conventional diagnostic methods are time consuming, expensive and require specialized laboratory facilities. We present a highly sensitive, highly specific, and low cost platform to test for acute virus infections in less than 15 min, employing influenza A virus (H1N1) as an example of its usability. An all polymer microfluidic system with a functionalized conductive polymer (PEDOT-OH:TsO) microelectrode array was developed and exploited for label free and real time electrochemical detection of intact influenza A virus (H1N1) particles. DNA aptamers with affinity for influenza A virus (H1N1) were linked covalently to the conductive polymer microelectrodes in the microfluidic channel. Based on changes in the impedance when virions were captured by immobilized probes, we could detect clinically relevant concentrations of influenza A virus (H1N1) in saliva. This is a new, stable and very sensitive point-of-care platform for detection and diagnostics of intact virus particles.

Human cytomegalovirus chemokine receptor US28 induces migration of cells on a CX3CL1-presenting surface.

Human cytomegalovirus (HCMV)-encoded G protein-coupled-receptor US28 is believed to participate in virus dissemination through modulation of cell migration and immune evasion. US28 binds different CC chemokines and the CX3C chemokine CX3CL1. Membrane-anchored CX3CL1 is expressed by immune-activated endothelial cells, causing redirection of CX3CR1-expressing leukocytes in the blood to sites of infection. Here, we used stable transfected cell lines to examine how US28 expression affects cell migration on immobilized full-length CX3CL1, to model how HCMV-infected leukocytes interact with inflamed endothelium. We observed that US28-expressing cells migrated more than CX3CR1-expressing cells when adhering to immobilized CX3CL1. US28-induced migration was G protein-signalling dependent and was blocked by the phospholipase Cß inhibitor U73122 and the intracellular calcium chelator BAPTA-AM. In addition, migration was inhibited in a dose-dependent manner by competition from CCL2 and CCL5, whereas CCL3 had little effect. Instead of migrating, CX3CR1-expressing cells performed 'dancing-on-the-spot' movements, demonstrating that anchored CX3CL1 acts as a strong tether for these cells. At low receptor expression levels, however, no significant difference in migration potential was observed when comparing the migration of CX3CR1- and US28-expressing cells. Thus, these data showed that, in contrast to CX3CR1, which promotes efficient cell capture upon binding to anchored CX3CL1, US28 acts to increase the migration of cells upon binding to the same ligand. Overall, this indicates that infected cells probably move more than uninfected cells in inflamed tissues with high CX3CL1 expression, with soluble chemokines affecting the final migration.

Cell-Based Biosensors: Electrical Sensing in Microfluidic Devices.

Cell-based biosensors provide new horizons for medical diagnostics by adopting complex recognition elements such as mammalian cells in microfluidic devices that are simple, cost efficient and disposable. This combination renders possible a new range of applications in the fields of diagnostics and personalized medicine. The review looks at the most recent developments in cell-based biosensing microfluidic systems with electrical and electrochemical transduction, and relevance to medical diagnostics.

Polymer based biosensor for rapid electrochemical detection of virus infection of human cells.

The demand in the field of medical diagnostics for simple, cost efficient and disposable devices is growing. Here, we present a label free, all-polymer electrochemical biosensor for detection of acute viral disease. The dynamics of a viral infection in human cell culture was investigated in a micro fluidic system on conductive polymer PEDOT:TsO microelectrodes by electrochemical impedance spectroscopy and video time lapse microscopy. Employing this sensitive, real time electrochemical technique, we could measure the immediate cell response to cytomegalovirus, and detect an infection within 3h, which is several hours before the cytopathic effect is apparent with conventional imaging techniques. Atomic force microscopy and scanning ion conductance microscopy imaging consolidate the electrochemical measurements by demonstrating early virus induced changes in cell morphology of apparent programmed cell death.

Microfluidic device to study cell transmigration under physiological shear stress conditions.

The development of new drug therapies relies on studies of cell transmigration in in vitro systems. Migration has traditionally been studied using two methods, the Boyden chamber and a shear flow chamber assay. Though, commonly applied in cell transmigration studies, they are far from imitating a natural migration process. Here we describe a novel in vitro cell transmigration microfluidic assay, which mimicks physiological shear flow conditions in blood vessels. The device was designed to incorporate the principles of both the Boyden chamber and the shear flow chamber assay, i.e. migration through the membrane under flow conditions. The 3D environment of migrating cells is imitated by injecting cell adhesion proteins to coat the membrane in the device. We tested the developed device with Jurkat cells migration towards medium supplemented with serum, and with chemokine induced lymphocytes migration. The applied continuous flow of cell suspension and chemoattractant ensures that the concentration gradient is maintained in time and space. The cell adhesion proteins used to enhance cell migration in the device were fibronectin and VCAM-1. We successfully observed a multistep transmigration process by means of the developed microfluidic migration assay. The presented device is inexpensive, easy to fabricate and disposable, having a potential to be applied in basic research as well as in the drug development process.

Binding areas of urokinase-type plasminogen activator-plasminogen activator inhibitor-1 complex for endocytosis receptors of the low-density lipoprotein receptor family, determined by site-directed mutagenesis.

Some endocytosis receptors related to the low-density lipoprotein receptor, including low-density lipoprotein receptor-related protein-1A, very-low-density lipoprotein receptor, and sorting protein-related receptor, bind protease-inhibitor complexes, including urokinase-type plasminogen activator (uPA), plasminogen activator inhibitor-1 (PAI-1), and the uPA-PAI-1 complex. The unique capacity of these receptors for high-affinity binding of many structurally unrelated ligands renders mapping of receptor-binding surfaces of serpin and serine protease ligands a special challenge. We have mapped the receptor-binding area of the uPA-PAI-1 complex by site-directed mutagenesis. Substitution of a cluster of basic residues near the 37-loop and 60-loop of uPA reduced the receptor-binding affinity of the uPA-PAI-1 complex approximately twofold. Deletion of the N-terminal growth factor domain of uPA reduced the affinity 2-4-fold, depending on the receptor, and deletion of both the growth factor domain and the kringle reduced the affinity sevenfold. The binding affinity of the uPA-PAI-1 complex to the receptors was greatly reduced by substitution of basic and hydrophobic residues in alpha-helix D and alpha-helix E of PAI-1. The localization of the implicated residues in the 3D structures of uPA and PAI-1 shows that they form a continuous receptor-binding area spanning the serpin as well as the A-chain and the serine protease domain of uPA. Our results suggest that the 10-100-fold higher affinity of the uPA-PAI-1 complex compared with the free components depends on the bonus effect of bringing the binding areas on uPA and PAI-1 together on the same binding entity.