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This paper presents a framework that integrates digital twin and virtual reality (VR) technologies to improve the efficiency and safety of human–robot collaborative systems in the disassembly domain. With the increasing complexity of the handling of end-of-life electronic products and as the related disassembly tasks are characterized by variabilities such as rust, deformation, and diverse part geometries, traditional industrial robots face significant challenges in this domain. These challenges require adaptable and flexible automation solutions that can work safely alongside human workers. We developed an architecture to address these challenges and support system configuration, training, and operational monitoring. Our framework incorporates a digital twin to provide a real-time virtual representation of the physical disassembly process, allowing for immediate feedback and dynamic adjustment of operations. In addition, VR is used to simulate and optimize the workspace layout, improve human–robot interaction, and facilitate safe and effective training scenarios without the need for physical prototypes. A unique case study is presented, where the collaborative system is specifically applied to the disassembly of antenna amplifiers, illustrating the potential of our comprehensive approach to facilitate engineering processes and enhance collaborative safety.

cDC2s occur abundantly in peripheral tissues and arise from circulating blood cDC2s. However, the factors governing cDC2 differentiation in tissues, especially under inflammatory conditions, remained poorly defined. We here found that psoriatic cDC2s express the efferocytosis receptor Axl and exhibit a bone morphogenetic protein (BMP) and p38MAPK signaling signature. BMP7, strongly expressed within the lesional psoriatic epidermis, cooperates with canonical TGF-β1 signaling for inducing Axl+cDC2s from blood cDC2s in vitro. Moreover, downstream induced p38MAPK promotes Axl+cDC2s at the expense of Axl+CD207+ Langerhans cell differentiation from blood cDC2s. BMP7 supplementation allowed to model cDC2 generation and their further differentiation into LCs from CD34+ hematopoietic progenitor cells in defined serum-free medium. Additionally, p38MAPK promoted the generation of another cDC2 subset lacking Axl but expressing the non-classical NFkB transcription factor RelB in vitro. Such RelB+cDC2s occurred predominantly at dermal sites in the inflamed skin. Finally, we found that cDC2s can be induced to acquire high levels of the monocyte lineage identity factor kruppel-like-factor-4 (KLF4) along with monocyte-derived DC and macrophage phenotypic characteristics in vitro. In conclusion, inflammatory and psoriatic epidermal signals instruct blood cDC2s to acquire phenotypic characteristics of several tissue-resident cell subsets.

Abstract Toll‐like receptor (TLR) stimulation induces innate immune responses involved in many inflammatory disorders including psoriasis. Although activation of the AP‐1 transcription factor complex is common in TLR signaling, the specific involvement and induced targets remain poorly understood. Here, we investigated the role of c‐Jun/AP‐1 protein in skin inflammation following TLR7 activation using human psoriatic skin, dendritic cells (DC), and genetically engineered mouse models. We show that c‐Jun regulates CCL2 production in DCs leading to impaired recruitment of plasmacytoid DCs to inflamed skin after treatment with the TLR7/8 agonist Imiquimod. Furthermore, deletion of c‐Jun in DCs or chemical blockade of JNK/c‐Jun signaling ameliorates psoriasis‐like skin inflammation by reducing IL‐23 production in DCs. Importantly, the control of IL‐23 and CCL2 by c‐Jun is most pronounced in murine type‐2 DCs. CCL2 and IL‐23 expression co‐localize with c‐Jun in type‐2/inflammatory DCs in human psoriatic skin and JNK‐AP‐1 inhibition reduces the expression of these targets in TLR7/8‐stimulated human DCs. Therefore, c‐Jun/AP‐1 is a central driver of TLR7‐induced immune responses by DCs and JNK/c‐Jun a potential therapeutic target in psoriasis.

Oxysterols are oxidized cholesterol derivatives whose systemic levels are found elevated in pregnancy disorders such as gestational diabetes mellitus (GDM). Oxysterols act through various cellular receptors and serve as a key metabolic signal, coordinating inflammation. GDM is a condition of low-grade chronic inflammation accompanied by altered inflammatory profiles in the mother, placenta and fetus. Higher levels of two oxysterols, namely 7-ketocholesterol (7-ketoC) and 7β-hydroxycholesterol (7β-OHC), were observed in fetoplacental endothelial cells (fpEC) and cord blood of GDM offspring. In this study, we tested the effects of 7-ketoC and 7β-OHC on inflammation and investigated the underlying mechanisms involved. Primary fpEC in culture treated with 7-ketoC or 7β-OHC, induced the activation of mitogen-activated protein kinase (MAPK) and nuclear factor kappa B (NFκB) signaling, which resulted in the expression of pro-inflammatory cytokines (IL-6, IL-8) and intercellular cell adhesion molecule-1 (ICAM-1). Liver-X receptor (LXR) activation is known to repress inflammation. Treatment with LXR synthetic agonist T0901317 dampened oxysterol-induced inflammatory responses. Probucol, an inhibitor of LXR target gene ATP-binding cassette transporter A-1 (ABCA-1), antagonized the protective effects of T0901317, suggesting a potential involvement of ABCA-1 in LXR-mediated repression of inflammatory signaling in fpEC. TLR-4 inhibitor Tak-242 attenuated pro-inflammatory signaling induced by oxysterols downstream of the TLR-4 inflammatory signaling cascade. Taken together, our findings suggest that 7-ketoC and 7β-OHC contribute to placental inflammation through the activation of TLR-4. Pharmacologic activation of LXR in fpEC decelerates its shift to a pro-inflammatory phenotype in the presence of oxysterols.

MiR-451a is best known for its role in erythropoiesis and for its tumour suppressor features. Here we show a role for miR-451a in neuronal differentiation through analysis of endogenous and ectopically expressed or silenced miR-451a in Ntera2/D1 cells during neuronal differentiation. Furthermore, we compared neuronal differentiation in the dentate gyrus of hippocampus of miR-451a-/- and wild type mice. MiR-451a overexpression in lentiviral transduced Ntera2/D1 cells was associated with a significant shifting of mRNA expression of the developmental markers Nestin, βIII Tubulin, NF200, DCX and MAP2 to earlier developmental time points, compared to control vector transduced cells. In line with this, accelerated neuronal network formation in AB.G.miR-451a transduced cells, as well as an increase in neurite outgrowth both in number and length was observed. MiR-451a targets genes MIF, AKT1, CAB39, YWHAZ, RAB14, TSC1, OSR1, POU3F2, TNS4, PSMB8, CXCL16, CDKN2D and IL6R were, moreover, either constantly downregulated or exhibited shifted expression profiles in AB.G.miR-451a transduced cells. Lentiviral knockdown of endogenous miR-451a expression in Ntera2/D1 cells resulted in decelerated differentiation. Endogenous miR-451a expression was upregulated during development in the hippocampus of wildtype mice. In situ hybridization revealed intensively stained single cells in the subgranular zone and the hilus of the dentate gyrus of wild type mice, while genetic ablation of miR-451a was observed to promote an imbalance between proliferation and neuronal differentiation in neurogenic brain regions, suggested by Ki67 and DCX staining. Taken together, these results provide strong support for a role of miR-451a in neuronal maturation processes in vitro and in vivo.

BACKGROUND:Chronic inflammation is frequently observed on histological analysis of malignant and non-malignant prostate specimens. It is a suspected supporting factor for prostate diseases and their progression and a main cause of false positive PSA tests in cancer screening. We hypothesized that inflammation induces autoantibodies, which may be useful biomarkers. We aimed to identify and validate prostate inflammation associated serum autoantibodies in prostate cancer patients and evaluate the expression of corresponding autoantigens. METHODS:Radical prostatectomy specimens of prostate cancer patients (N = 70) were classified into high and low inflammation groups according to the amount of tissue infiltrating lymphocytes. The corresponding pre-surgery blood serum samples were scrutinized for autoantibodies using a low-density protein array. Selected autoantigens were identified in prostate tissue and their expression pattern analyzed by immunohistochemistry and qPCR. The identified autoantibody profile was cross-checked in an independent sample set (N = 63) using the Luminex-bead protein array technology. RESULTS:Protein array screening identified 165 autoantibodies differentially abundant in the serum of high compared to low inflammation patients. The expression pattern of three corresponding antigens were established in benign and cancer tissue by immunohistochemistry and qPCR: SPAST (Spastin), STX18 (Syntaxin 18) and SPOP (speckle-type POZ protein). Of these, SPAST was significantly increased in prostate tissue with high inflammation. All three autoantigens were differentially expressed in primary and/or castration resistant prostate tumors when analyzed in an inflammation-independent tissue microarray. Cross-validation of the inflammation autoantibody profile on an independent sample set using a Luminex-bead protein array, retrieved 51 of the significantly discriminating autoantibodies. Three autoantibodies were significantly upregulated in both screens, MUT, RAB11B and CSRP2 (p>0.05), two, SPOP and ZNF671, close to statistical significance (p = 0.051 and 0.076). CONCLUSIONS:We provide evidence of an inflammation-specific autoantibody profile and confirm the expression of corresponding autoantigens in prostate tissue. This supports evaluation of autoantibodies as non-invasive markers for prostate inflammation.

We describe a novel procedure for determination and validation of off-target activities of anti-cluster designation antigen identity 44 (CD44) variant 6 recombinant antibodies by combining two complementary technology platforms; namely UNIchip AV-400, containing a printed serial dilution of CD44 variant 6 and approximately 400 different human proteins, and TISSOMICS, enabling human tissue microarray analysis in high-throughput mode. We have analyzed the performance of two human monoclonal recombinant antibodies directed against CD44 variant 6 protein, BMS 116 and BMS 125, in a blinded study. The antibodies exhibit a clear differentiation with regard to their binding profiles in the two systems. BMS 116 shows a low degree of specificity in the normal human Food and Drug Administration (FDA) tissue panel, which was confirmed by binding to more than 206 proteins on the protein biochip. In contrast, BMS 125 gives a highly selective membranous staining on selected human epithelial tissue components with no off-target activities observed on the protein biochip. Additionally, antibody BMS 125 shows a higher sensitivity to its antigen CD44 variant 6 than antibody BMS 116 as determined by the protein biochip.

Bone morphogenetic proteins (BMPs) are members of the TGF-β family that signal via the BMP receptor (BMPR) signaling cascade, distinct from canonical TGF-β signaling. BMP downstream signaling is strongly induced within epidermal keratinocytes in cutaneous psoriatic lesions, and BMP7 instructs monocytic cells to acquire characteristics of psoriasis-associated Langerhans dendritic cells (DCs). Regulatory T (Treg)-cell numbers strongly increase during psoriatic skin inflammation and were recently shown to limit psoriatic skin inflammation. However, the factors mediating Treg-cell accumulation in psoriatic skin currently remain unknown. We sought to investigate the role of BMP signaling in Treg-cell accumulation in psoriasis. The following methods were used: immunohistology of patients and healthy controls; ex vivo models of Treg-cell generation in the presence or absence of Langerhans cells; analysis of BMP versus canonical TGF-β signaling in DCs and Treg cells; and modeling of psoriatic skin inflammation in mice lacking the BMPR type 1a in CD11c+ cells. We here demonstrated a positive correlation between Treg-cell numbers and epidermal BMP7 expression in cutaneous psoriatic lesions and show that unlike Treg cells from healthy skin, a portion of inflammation-associated Treg cells exhibit constitutive-active BMP signaling. We further found that BMPR signaling licenses inflammation-associated Langerhans cell/DC to gain an enhanced capacity to promote Treg cells via BMPR-mediated CD25 induction and that this effect is associated with reduced skin inflammation. Psoriatic lesions are marked by constitutive high BMP7/BMPR signaling in keratinocytes, which instructs inflammatory DCs to gain enhanced Treg-cell–stimulatory activity. Locally secreted BMP7 can directly promote Treg-cell generation through the BMP signaling cascade. [Display omitted] [ABSTRACT FROM AUTHOR]

Epidermal hyperplasia represents a morphologic hallmark of psoriatic skin lesions. Langerhans cells (LCs) in the psoriatic epidermis engage with keratinocytes (KCs) in tight physical interactions; moreover, they induce T-cell-mediated immune responses critical to psoriasis. This study sought to improve the understanding of epidermal factors in psoriasis pathogenesis. BMP7-LCs versus TGF-β1-LCs were phenotypically characterized and their functional properties were analyzed using flow cytometry, cell kinetic studies, co-culture with CD4 T cells, and cytokine measurements. Furthermore, immunohistology of healthy and psoriatic skin was performed. Additionally, in vivo experiments with Jun f /f JunB f/f K5cre-ER T mice were carried out to assess the role of bone morphogenetic protein (BMP) signaling in psoriatic skin inflammation. This study identified a KC-derived signal (ie, BMP signaling) to promote epidermal changes in psoriasis. Whereas BMP7 is strictly confined to the basal KC layer in the healthy skin, it is expressed at high levels throughout the lesional psoriatic epidermis. BMP7 instructs precursor cells to differentiate into LCs that phenotypically resemble psoriatic LCs. These BMP7-LCs exhibit proliferative activity and increased sensitivity to bacterial stimulation. Moreover, aberrant high BMP signaling in the lesional epidermis is mediated by a KC intrinsic mechanism, as suggested from murine data and clinical outcome after topical antipsoriatic treatment in human patients. These data indicate that available TGF-β family members within the lesional psoriatic epidermis preferentially signal through the canonical BMP signaling cascade to instruct inflammatory-type LCs and to promote psoriatic epidermal changes. Targeting BMP signaling might allow to therapeutically interfere with cutaneous psoriatic manifestations. [ABSTRACT FROM AUTHOR]

Background: Chronic inflammation is frequently observed on histological analysis of malignant and non-malignant prostate specimens. It is a suspected supporting factor for prostate diseases and their progression and a main cause of false positive PSA tests in cancer screening. We hypothesized that inflammation induces autoantibodies, which may be useful biomarkers. We aimed to identify and validate prostate inflammation associated serum autoantibodies in prostate cancer patients and evaluate the expression of corresponding autoantigens., Methods: Radical prostatectomy specimens of prostate cancer patients (N = 70) were classified into high and low inflammation groups according to the amount of tissue infiltrating lymphocytes. The corresponding pre-surgery blood serum samples were scrutinized for autoantibodies using a low-density protein array. Selected autoantigens were identified in prostate tissue and their expression pattern analyzed by immunohistochemistry and qPCR. The identified autoantibody profile was cross-checked in an independent sample set (N = 63) using the Luminex-bead protein array technology., Results: Protein array screening identified 165 autoantibodies differentially abundant in the serum of high compared to low inflammation patients. The expression pattern of three corresponding antigens were established in benign and cancer tissue by immunohistochemistry and qPCR: SPAST (Spastin), STX18 (Syntaxin 18) and SPOP (speckle-type POZ protein). Of these, SPAST was significantly increased in prostate tissue with high inflammation. All three autoantigens were differentially expressed in primary and/or castration resistant prostate tumors when analyzed in an inflammation-independent tissue microarray. Cross-validation of the inflammation autoantibody profile on an independent sample set using a Luminex-bead protein array, retrieved 51 of the significantly discriminating autoantibodies. Three autoantibodies were significantly upregulated in both screens, MUT, RAB11B and CSRP2 (p>0.05), two, SPOP and ZNF671, close to statistical significance (p = 0.051 and 0.076)., Conclusions: We provide evidence of an inflammation-specific autoantibody profile and confirm the expression of corresponding autoantigens in prostate tissue. This supports evaluation of autoantibodies as non-invasive markers for prostate inflammation.

Background: The currently used prostate cancer serum marker has a low cancer specificity and improved diagnostics are needed. Here we evaluated whether autoantibodies are present in sera of prostate cancer patients and whether they are useful diagnostic markers for prostate cancer., Methods: Sera from 20 prostate cancer patients and 20 healthy controls were incubated on expression clone arrays containing more than 37,000 recombinant human proteins. Functional annotation clustering of the identified autoantigens was performed using the DAVID database. Autoantigens identified in the prostate cancer group were validated on microarrays using sera of 40 prostate cancer patients, 40 patients with elevated PSA levels but prostate cancer negative biopsies (benign disease), and 40 healthy controls., Results: We detected autoantibodies against 408 different antigens in sera of prostate cancer patients. One hundred seventy-four of these were exclusively detected in the cancer group compared to the healthy control group. Functional annotation clustering revealed an enrichment of RNA-associated, cytoskeleton, and nuclear proteins. The autoantibody panel was validated in serum samples of independent prostate cancer patients. Autoantibody profiles discriminated between prostate cancer patients and benign disease patients with an ROC curve AUC of 0.71. TTLL12, a protein recently described to be over-expressed in prostate cancer, was the highest ranked discrimination autoantigen., Conclusion: A variety of autoantibodies were identified in sera of prostate cancer patients and provide a first step towards autoantibody diagnostics. Serum autoantibodies reflect the disease and represent valuable tools not only for prostate cancer, but also for other diseases affecting the immune response., (Copyright © 2011 Wiley Periodicals, Inc.)

Background: Several studies have demonstrated the value of DNA methylation in urine-based assays for prostate cancer diagnosis. However, a multicenter validation with a clinical prototype has not been published., Methods: We developed a multiplexed, quantitative methylation-specific polymerase chain reaction (MSP) assay consisting of 3 methylation markers, GSTP1, RARB, and APC, and an endogenous control, ACTB, in a closed-tube, homogeneous assay format. We tested this format with urine samples collected after digital rectal examination from 234 patients with prostate-specific antigen (PSA) concentrations > or =2.5 microg/L in 2 independent patient cohorts from 9 clinical sites., Results: In the first cohort of 121 patients, we demonstrated 55% sensitivity and 80% specificity, with area under the curve (AUC) 0.69. In the second independent cohort of 113 patients, we found a comparable sensitivity of 53% and specificity of 76% (AUC 0.65). In the first cohort, as well as in a combined cohort, the MSP assay in conjunction with total PSA, digital rectal examination status, and age improved the AUC without MSP, although the difference was not statistically significant. Importantly, the GSTP1 cycle threshold value demonstrated a good correlation (R = 0.84) with the number of cores found to contain prostate cancer or premalignant lesions on biopsy. Moreover, samples that exhibited methylation for either GSTP1 or RARB typically contained higher tumor volumes at prostatectomy than those samples that did not exhibit methylation., Conclusions: These data confirm and extend previously reported studies and demonstrate the performance of a clinical prototype assay that should aid urologists in identifying men who should undergo biopsy.

Understanding the antibody response in botulinum intoxication is important for vaccine design and passive prophylaxis. To investigate this activity, we have studied the immune response to BoNT/A (botulinum neurotoxin serotype A) binding domain (HC) at the molecular level using phage display. The scFv antibodies were isolated from V-gene repertoires prepared from (a) human volunteer immunized with pentavalent botulinum toxoid and (b) non-immune human peripheral blood lymphocytes and spleenocytes. A large panel of serotype specific phage expressing botulinum binding scFv could be selected from both libraries. Epitope mapping of immune scFv binders towards BoNT/A HC revealed surprisingly a limited number of scFv recognizing conformational epitopes that corresponded to two distinct groups, clusters I and II. Only scFv from cluster I exhibited neutralizing activity in the mouse hemidiaphragm assay. Anti- BoNT/A HC clones derived from a non-immune library could be conveniently grouped into clusters III-XI and appeared to share no overlapping epitopes with cluster I or II. In addition they showed no neutralization of toxin at biologically significant concentrations. We therefore suggest that a vaccine based on the pentavalent botulinum toxoid directs the humoral immune response to a limited number of immunodominant epitopes exposed on the binding domain HC.

This investigation was pursued to test the use of intracellular antibodies (intrabodies) as a means of blocking the pathogenesis of Huntington's disease (HD). HD is characterized by abnormally elongated polyglutamine near the N terminus of the huntingtin protein, which induces pathological protein-protein interactions and aggregate formation by huntingtin or its exon 1-containing fragments. Selection from a large human phage display library yielded a single-chain Fv (sFv) antibody specific for the 17 N-terminal residues of huntingtin, adjacent to the polyglutamine in HD exon 1. This anti-huntingtin sFv intrabody was tested in a cellular model of the disease in which huntingtin exon 1 had been fused to green fluorescent protein (GFP). Expression of expanded repeat HD-polyQ-GFP in transfected cells shows perinuclear aggregation similar to human HD pathology, which worsens with increasing polyglutamine length; the number of aggregates in these transfected cells provided a quantifiable model of HD for this study. Coexpression of anti-huntingtin sFv intrabodies with the abnormal huntingtin-GFP fusion protein dramatically reduced the number of aggregates, compared with controls lacking the intrabody. Anti-huntingtin sFv fused with a nuclear localization signal retargeted huntingtin analogues to cell nuclei, providing further evidence of the anti-huntingtin sFv specificity and of its capacity to redirect the subcellular localization of exon 1. This study suggests that intrabody-mediated modulation of abnormal neuronal proteins may contribute to the treatment of neurodegenerative diseases such as HD, Alzheimer's, Parkinson's, prion disease, and the spinocerebellar ataxias.

A large library of phage-displayed human single-chain Fv antibodies (scFv), containing 6.7 x 10(9) members, was generated by improving the steps of library construction. Fourteen different protein antigens were used to affinity select antibodies from this library. A panel of specific antibodies was isolated with each antigen, and each panel contained an average of 8.7 different scFv. Measurements of antibody-antigen interactions revealed several affinities below 1 nM, comparable to affinities observed during the secondary murine immune response. In particular, four different scFv recognizing the ErbB2 protein had affinities ranging from 220 pM to 4 nM. Antibodies derived from the library proved to be useful reagents for immunoassays. For example, antibodies generated to the Chlamydia trachomatis elementary bodies stained Chlamydia-infected cells, but not uninfected cells. These results demonstrate that phage antibody libraries are ideally suited for the rapid production of panels of high-affinity mAbs to a wide variety of protein antigens. Such libraries should prove especially useful for generating reagents to study the function of gene products identified by genome projects.