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G protein-coupled receptors (GPCRs) comprise an expanded superfamily of receptors in the human genome. Adhesion class G protein-coupled receptors (adhesion-GPCRs) form the second largest class of GPCRs. Despite the abundance, size, molecular structure, and functions in facilitating cell and matrix contacts in a variety of organ systems, adhesion-GPCRs are by far the most poorly understood GPCR class. Adhesion-GPCRs possess a unique molecular structure, with extended N-termini containing various adhesion domains. In addition, many adhesion-GPCRs are autoproteolytically cleaved into an N-terminal fragment (NTF, NT, α-subunit) and C-terminal fragment (CTF, CT, β-subunit) at a conserved GPCR autoproteolysis-inducing (GAIN) domain that contains a GPCR proteolysis site (GPS). These two features distinguish adhesion-GPCRs from other GPCR classes. Though active research on adhesion-GPCRs in diverse areas, such as immunity, neuroscience, and development and tumor biology has been intensified in the recent years, the general biological and pharmacological properties of adhesion-GPCRs are not well known, and they have not yet been used for biomedical purposes. The '6th International Adhesion-GPCR Workshop,' held at the Institute of Physiology of the University of Würzburg on September 6-8, 2012, assembled a majority of the investigators currently actively pursuing research on adhesion-GPCRs, including scientists from laboratories in Europe, the United States, and Asia. The meeting featured the nascent mechanistic understanding of the molecular events driving the signal transduction of adhesion-GPCRs, novel models to evaluate their functions, and evidence for their involvement in human disease. [ABSTRACT FROM AUTHOR]

Abstract: Background and aims: To understand biomechanics of ligaments and tendons data on their material properties are necessary. The iliotibial tract is a suitable model for virtual pelvic or lower extremity ligaments due to its parallel fibers, which facilitates biomechanical testing. Here, we determined Young''s modulus (YM) as secant stiffness between defined limits of the iliotibial tract and correlated the data to ultimate stress (US) of the specimens and to age, gender and body weight of the body donors. Materials and methods: Thirty eight specimens from 12 iliotibial tracts of 10 young donors (mean age 31.2±9.1 years) were investigated biomechanically. After preconditioning, YM were determined in the ranges of 0–4 and 4–11N/mm² of applied stress and from 4N/mm² of applied stress to US. Results: YM of the specimens were 84.7±30.2 (0–4N/mm²), 335.4±101.9 (4–11N/mm²), and 369.1±191.5 (4N/mm² to US) N/mm², respectively. The mean US was 35.8±16.4N/mm². YM and US correlated closely in the ranges of 4–11N/mm² (r=0.95) and 4N/mm² to US (r=0.91). YM did not correlate to age, body weight or gender within these young donors. Concerning tissue behavior a decrease of YM, i.e. weakening, is more common than an increase of YM, i.e. stiffening, before specimen failure. Overall, YM of specimens from young donors were significantly lower compared to those of old donors. Discussion and conclusions: This is the first study providing age-dependent nonlinear stiffness properties of the iliotibial tract. YM is significantly lower in young than in old donors and is thus a subject of alteration during life time. [Copyright &y& Elsevier]

Background The 2-chloro-N6-cyclopentyladenosine (CCPA) was proven to be a protective factor in ischemic reperfusion injury in myocardium and to reduce the infarct size in the heart. The purpose of this study was to determine whether flap necrosis could be reduced by intravenous administration of CCPA. Methods Fifty-six male Wistar rats were divided into 4 experimental groups. An epigastric adipocutaneous flap was raised, and the area of flap necrosis was assessed for all groups on the fifth postoperative day with planimetry software. Results The control group had a significantly lower rate of flap necrosis than the ischemic control group ( p < .05). The nonischemic CCPA group had a significantly lower rate of flap necrosis than the nonischemic control group ( p < .05). The ischemic CCPA group had a highly significant ( p < .0001) rate of lower flap necrosis than the ischemic control group. Conclusion Our data show that reduction of flap necrosis can be achieved both with and without ischemic periods by intravenous administration of CCPA. © 2011 Wiley Periodicals, Inc. Head Neck, 2011 [ABSTRACT FROM AUTHOR]

Evaluates the presence and number of eosinophils at varying stages in the human corpus luteum from the ovaries of women at reproductive age. Numbers of extravasated eosinophils at the granulosa layer, thecal layer and hemorrhages in the former antrum; Reduction of eosinophils during secretion and regression stages; Expression and regulation of eosinophil-attracting chemokines RANTES and eotaxin.

IA-2, a member of the protein tyrosine phosphatase family, represents a major target autoantigen in type 1 diabetes. To study the regulation of IA-2 gene expression, we used INS-1 insulinoma cells to analyze beta-cell signal transduction pathways as well as the effect of metabolic and hormonal factors involved in the regulation of the insulin secretory pathway. Quantitative competitive reverse transcriptase-polymerase chain reaction revealed that an increase of cellular cAMP mediated by forskolin (10 micromol/l, 24 h) or 3-isobutyl-1-methylxanthine (100 micromol/l, 24 h) induced maximal stimulation of IA-2 mRNA levels (451 +/- 85 and 338 +/- 86% compared with basal conditions; P < 0.001). In contrast, activation of protein kinase C (PKC) by short-term treatment with phorbol 12-myristate 13-acetate (PMA) (1 micromol/l, 6 h) did not alter IA-2 expression, whereas depletion of PKC by prolonged culturing (24 h) exerted a significant inhibition (57 +/- 24%; P < 0.05). cAMP-dependent upregulation was confirmed by the findings that glucagon (10 micromol/l, 24-48 h) increased levels of IA-2 mRNA (190 +/- 35%; P < 0.05), whereas short-term incubation with high glucose concentration showed no effect. However, prolonged incubation in high glucose (21 mmol/l) induced a time- and dose-dependent increase of IA-2 mRNA expression, reaching maximal values after 144 h (285 +/- 68%; P < 0.05). These studies demonstrate that stimuli of insulin secretion that operate by activation of adenylate cyclase generating cAMP significantly increase IA-2 gene expression. In contrast, activation of PKC by high glucose concentration or PMA exerted no effect, suggesting that IA-2 gene expression is not simply coupled to insulin secretion, but may be involved in the fine regulation of beta-cell function. These findings may be important to clarify the function of IA-2 in beta-cells and elucidate mechanisms involved in the induction of autoimmunity to IA-2. [ABSTRACT FROM AUTHOR]

Trauma causes the breakdown of membrane phospholipids and the subsequent degradation of the released polyunsaturated fatty acids (PUFAs) to partially bioactive oxylipins. Here, we screened for circulating PUFAs and oxylipins in patients (n = 34) differing from those of uninjured controls (n = 25) and analyzed their diagnostic potential. Patients were followed up for 1 to 240 h after minor/moderate, severe, and very severe injuries. Of the targeted oxylipins, 13 out of 80 (13/80) were detected in almost all patients and controls. Injury caused a long-term decrease in 9- and 13-hydroxyoctadecadienoic acids and in several dihydroxyeicosatetraenoic acids, the stable derivatives of bioactive anti-inflammatory epoxyeicosatrienoic acids, compared to controls. Frequently, these oxylipins correlated inversely to injury severity, days in the intensive care unit and hospital, and/or procalcitonin and pro-inflammatory cytokine levels 48 up to 240 h after trauma. Notably, 20/80 oxylipins were detected in some patients but not or less often in controls. Many of these oxylipins increased transiently immediately after injury. Their level is partly correlated with adverse clinical parameters at this early time point. The circulating oxylipidome was markedly affected by trauma. Several oxylipins showed injury-dependent alterations at different time points in the post-traumatic course. [ABSTRACT FROM AUTHOR]

Strain-dependent transmission data of nine iliotibial tract specimens are determined using a custom-built optical setup with a halogen light source and an industrial norm material testing machine. Polarized light microscopy and hematoxylin-eosin staining indicated that lateral contraction of collagen structures is responsible for total intensity variations during a 20-cycle preconditioning and a 5-cycle tensile test. Tensile force progress is opposite to total transmission progress. Due to dehydration, wavelength-specific radiation intensity shifting is determined during the test, primarily noticeable in a water absorption band between 1400 and 1500 nm. The results show the capability of integrating spectrophotometry technology into biomechanics for determining structural alterations of human collagen due to applied strain. Being more sensitive to drying, spectrophotometry may likely serve as a quality control in stress-strain testing of biological structures. [ABSTRACT FROM AUTHOR]

Background: Anti-tumour immune competence has an impact in hepatocarcinogenesis and success of anti-cancer therapies. Tumour-infiltrating lymphocytes (TILs) and monocytes/macrophages (TAMs) are proposed to have significance in cancer. However, there is only limited data concerning their impact on patient outcome and survival in hepatocellular carcinoma (HCC). Methods: Frequencies of CD68+, CD163+ M2-polarized TAMs and TILs were measured in de novo HCC tumours in non-cirrhosis (n = 58) using immunohistology and correlated to patients' clinicopathological characteristics and survival rates. Results: Patients with tumours marked by appearance of TILs and CD68+ TAMs showed an improved 1-, 3- and 5-year recurrence-free survival (all p ≤ 0.05). CD68+ TAMs were associated with reduced incidence of recurrent and multifocal disease. Conversely, CD163+ TAMs were associated with multifocal HCC and lymphangiosis carcinomatosa (all p ≤ 0.05). Conclusions: TILs and CD68+ TAMs are associated with multiple tumour characteristics and patient survival in HCC. However, there is only scarce data about the biology underlying their mechanistic involvement in human tumour progression. Thus, experimental data on functional links might help develop novel immunologic checkpoint inhibitor targets for liver cancer. [ABSTRACT FROM AUTHOR]

Background: Chemerin is an adipokine which plays a crucial role in atherosclerosis. Here, we examined whether circulating chemerin is enhanced in patients with advanced carotid stenosis.Methods: Chemerin was quantified in 178 patients prior to carotid end arterectomy (CEA) and in age- and gender-matched controls (n = 163). Chemerin levels were related to anthropometric, clinical and metabolic characteristics of the patients.Results: Chemerin levels were higher in patients compared to controls (p <  0.001). Chemerin correlated to parameters associated with inflammation such as C-reactive protein (CRP, p <  0.001), leukocyte blood count (p <  0.001) and circulating TNF-α (p = 0.004) in the patients. Chemerin levels did not differ between asymptomatic (n = 93) and symptomatic patients who experienced an ischemic event within 6 months prior to CEA (n = 85). However, in the case of high-grade carotid stenosis (≥ 90%), chemerin levels were higher in symptomatic (n = 44) compared to asymptomatic patients (n = 41, p = 0.014). Chemerin was increased in patients with (n = 50) compared to patients without (n = 128) coronary artery disease (CAD, p = 0.002). A high level of chemerin increases the risk for CAD in patients (p = 0.0013).Conclusions: Circulating chemerin is increased and correlates to inflammatory parameters in patients with advanced carotid stenosis. [ABSTRACT FROM AUTHOR]

Trauma triggers a rapid innate immune response to aid the clearance of damaged/necrotic cells and their released damage-associated molecular pattern (DAMP). Here, we monitored the expression of EMR2/ADGRE2, involved in the functional regulation of innate immune cells, on circulating neutrophils in very severely and moderately/severely injured patients up to 240 h after trauma. Notably, neutrophilic EMR2 showed a uniform, injury severity- and type of injury-independent posttraumatic course in all patients. The percentage of EMR2+ neutrophils and their EMR2 level increased and peaked 48 h after trauma. Afterwards, they declined and normalized in some, but not all, patients. Circulating EMR2+ compared to EMR2− neutrophils express less CD62L and more CD11c, a sign of activation. Neutrophilic EMR2 regulation was verified in vitro. Remarkably, it increased, depending on extracellular calcium, in controls as well. Cytokines, enhanced in patients immediately after trauma, and sera of patients did not further affect this neutrophilic EMR2 increase, whereas apoptosis induction disrupted it. Likely the damaged/necrotic cells/DAMPs, unavoidable during neutrophil culture, stimulate the neutrophilic EMR2 increase. In summary, the rapidly increased absolute number of neutrophils, especially present in very severely injured patients, together with upregulated neutrophilic EMR2, may expand our in vivo capacity to react to and finally clear damaged/necrotic cells/DAMPs after trauma. [ABSTRACT FROM AUTHOR]

(f) The percentages of GPR115+ and KRT1/10+ cells were determined in these constructs built with HaCaT WT cells (n = 10 optical fields, n = 20-27 cells/field, means ± SEM). (c) Western blot analysis of lysates of HA-GPR115 Cos-7 cells treated with PNGase F (left) or of HA-GPR115 Cos-7 cells cultured with tunicamycin (right); the Abs used are indicated. (b) HaCaT WT cells and Cos-7 cells, transfected with GPR115 pcDNA3.1, were cell-surface-stained with the GPR115ECD Ab and analyzed by flow cytometry. [Extracted from the article]

Circulating sterols result either from cholesterol (CH) synthesis or intestinal uptake. They are mainly esterified and can be oxygenated. Sterols accumulate in atherosclerotic plaques whereby their clinical impact is uncertain. Here, we determined associations between circulating and plaque sterol levels in patients with advanced carotid artery stenosis in respect to a prior ischemic event and statin treatment. Free and esterified CH, CH precursors and plant sterols as well as oxysterols were quantified by liquid chromatography-tandem mass spectrometry in 63 consecutive patients undergoing carotid endarterectomy. CH, CH precursors, plant sterols and oxysterols accumulated in carotid artery plaques. Absolute circulating sterol levels were not predictive for their corresponding plaque levels. After normalisation to CH, plant sterol but not oxysterol levels correlated between plasma and plaques. Among the circulating sterols, oxysterols occurred proportionally less in plaques. Furthermore, CH and plant sterols were less esterified in plaques than in plasma. Patients who experienced a prior ischemic event (n = 29) and asymptomatic patients had, except for lanosterol, comparable circulating sterol levels. In contrast, the absolute plaque levels of free CH, CH precursors and plant sterols as well as oxysterols were increased in symptomatic compared to asymptomatic patients. These differences remained significant for free CH, precursors and 3 out of 4 analyzed plant sterols after adjustment to the most influencing covariates - statin treatment, type 2 diabetes and age. Increased absolute plaque levels of free CH, precursors and plant sterols predict an ischemic event in patients with advanced carotid artery stenosis. Image 1 • Carotid artery plaques contain cholesterol (CH), CH precursors, plant and oxysterols. • CH and plant sterols are less esterified in plaques than in plasma. • Plasma and plaque CH-normalized plant sterol but not oxysterol levels correlate. • Plaque sterol levels are higher in symptomatic compared with asymptomatic patients. • Plaque free CH, CH precursor and plant sterol levels predict an ischemic event. [ABSTRACT FROM AUTHOR]

Background: Tumor escape mechanisms mediated in the tumor microenvironment can significantly reduce the capacity of the anti-tumor function of the immune system. TIE2-expressing monocytes (TEMs), related angiopoietins, and tumor necrosis are considered to have a key role in this process. We aimed to investigate the abundance and clinical significance of these biomarkers in hepatocellular carcinoma (HCC). Methods: In this retrospective study, 58 HCC patients received surgery with a curative intent. The abundance of TEMs, angiopoietin-1 and -2 were detected in tumor specimens of the HCC patients (n = 58), and together with the occurrence of histologic tumor necrosis, were associated with established clinicopathological characteristics and survival. Results: Patients with HCC characterized by necrosis and TEMs revealed reduced both overall survival and recurrence-free survival (all p < 0.05). Angiopoietins and TEMs were associated with metastatic and recurrent HCC. Furthermore, the formation of histologic tumor necrosis was associated with advanced tumor stage and density of TEMs (all p < 0.05). Conclusions: Histologic tumor necrosis, TEMs, and related angiopoietins were associated with multiple HCC parameters and patient survival. The tumor necrosis–TEM–angiopoietin axis may offer a novel diagnostic modality to predict patient outcome after surgery for HCC. [ABSTRACT FROM AUTHOR]

The adhesion class of G protein-coupled receptors (GPCRs) is the second largest family of GPCRs (33 members in humans). Adhesion GPCRs (aGPCRs) are defined by a large extracellular N-terminal region that is linked to a C-terminal seven transmembrane (7TM) domain via aGPCR-autoproteolysis inducing (GAIN) domain containing a GPCR proteolytic site (GPS).Most aGPCRs undergo autoproteolysis at the GPS motif, but the cleaved fragments stay closely associated, with the N-terminal fragment (NTF) bound to the 7TMof the C-terminal fragment (CTF). The NTFs ofmost aGPCRs contain domains known to be involved in cell-cell adhesion, while the CTFs are involved in classical G protein signaling, as well as other intracellular signaling. In this workshop report, we review themost recent findings on the biology, signaling mechanisms, and physiological functions of aGPCRs. [ABSTRACT FROM AUTHOR]

Intestinal stem cells (ISCs) require well-defined signals from their environment in order to carry out their specific functions. Most of these signals are provided by neighboring cells that form a stem cell niche, whose shape and cellular composition self-organize. Major features of this self-organization can be studied in ISC-derived organoid culture. In this system, manipulation of essential pathways of stem cell maintenance and differentiation results in well-described growth phenotypes. We here provide an individual cell-based model of intestinal organoids that enables a mechanistic explanation of the observed growth phenotypes. In simulation studies of the 3D structure of expanding organoids, we investigate interdependences between Wnt- and Notch-signaling which control the shape of the stem cell niche and, thus, the growth pattern of the organoids. Similar to in vitro experiments, changes of pathway activities alter the cellular composition of the organoids and, thereby, affect their shape. Exogenous Wnt enforces transitions from branched into a cyst-like growth pattern; known to occur spontaneously during long term organoid expansion. Based on our simulation results, we predict that the cyst-like pattern is associated with biomechanical changes of the cells which assign them a growth advantage. The results suggest ongoing stem cell adaptation to in vitro conditions during long term expansion by stabilizing Wnt-activity. Our study exemplifies the potential of individual cell-based modeling in unraveling links between molecular stem cell regulation and 3D growth of tissues. This kind of modeling combines experimental results in the fields of stem cell biology and cell biomechanics constituting a prerequisite for a better understanding of tissue regeneration as well as developmental processes. [ABSTRACT FROM AUTHOR]

The Adhesion G protein-coupled receptor (GPCR) CD97/ADGRE5 is induced, upregulated, and/or biochemically modified in various malignancies, compared to the corresponding normal tissues. As tumor cells are generally more resistant to apoptosis, we here studied the ability of CD97 to regulate tumor cell survival under apoptotic conditions. Stable overexpression of wild-type CD97 reduced serum starvation- and staurosporine-induced intrinsic and tumor necrosis factor (TNF)/cycloheximide-induced extrinsic apoptosis, indicated by an increase in cell viability, a lower percentage of cells within the subG0/G1 phase, expressing annexin V, or having condensed nuclei, and a reduction of DNA laddering. Protection from cell death by CD97 was accompanied by an inhibition of caspase activation and modulation of anti- and pro-apoptotic members of the BCL-2 superfamily. shRNA-mediated knockdown of CD97 and, in part, truncation of the seven-span transmembrane (TM7) region of CD97 increased caspase-mediated apoptosis. Protection from apoptosis required not only the TM7 region but also cleavage of the receptor at its GPCR proteolysis site (GPS), whereas alternative splicing of its extracellular domain had no effect. Together, our data indicate a role of CD97 in tumor cell survival. [ABSTRACT FROM AUTHOR]

Background: A rapid liquid chromatography–tandem mass spectrometric (LC–MS/MS) method was developed and validated for the quantification of reactive oxygen species (ROS) derived free oxysterols and cholesterol in human plasma and atherosclerotic plaque. Method: In vitro autoxidation of cholesterol during sample pretreatment was avoided by applying only one protein precipitation and re-concentration step using 80μl plasma. For preparation of 10mg atherosclerotic plaques an additional liquid–liquid extraction was included. Free 7-keto-, 7-α/ß-hydroxy-, 5,6-α-epoxy-, 5,6-β-epoxycholesterol, cholestane-3ß,5α,6ß-triol and cholesterol were separated within 7min on a monolithic column. An API 4000 tandem mass spectrometer was applied in positive ionization mode using atmospheric pressure chemical ionization. Results: The detection limit was 0.1ng/ml and the linearity ranged from 0.5 to 0.75 to 2000ng/ml for the oxysterols and from 50 to 1000μg/ml for cholesterol. Recovery was between 80.9 and 107.9%. Between-run imprecision ranged from 7.9 to 11.7%. Analysis of plasma samples from additional 50 middle-aged volunteers revealed a large inter-individual variability (e.g. 7-ketocholesterol 2.63-30.47ng/ml). Oxysterol concentrations normalized to cholesterol were about 43 times higher in carotid plaque compared to plasma (n=5). Conclusion: This rapid LC–MS/MS method enables reliable quantification focused on especially ROS-derived oxysterols in human plasma and atherosclerotic plaque samples under high-throughput conditions. [ABSTRACT FROM AUTHOR]

The implementation of stem-cell-based organoid culture more than ten years ago started a development that created new avenues for diagnostic analyses and regenerative medicine. In parallel, computational modelling groups realized the potential of this culture system to support their theoretical approaches to study tissues in silico. These groups developed computational organoid models (COMs) that enabled testing consistency between cell biological data and developing theories of tissue self-organization. The models supported a mechanistic understanding of organoid growth and maturation and helped linking cell mechanics and tissue shape in general. What comes next? Can we use COMs as tools to complement the equipment of our biological and medical research? While these models already support experimental design, can they also quantitatively predict tissue behavior? Here, we review the current state of the art of COMs and discuss perspectives for their application.

The heterogeneous response of acute myeloid leukemia (AML) to current anti-leukemic therapies is only partially explained by mutational heterogeneity. We previously identified GPR56 as a surface marker associated with poor outcome across genetic groups, which characterizes two leukemia stem cell (LSC)-enriched compartments with different self-renewal capacities. How these compartments self-renew remained unclear. Here, we show that GPR56 + LSC compartments are promoted in a complex network involving epithelial-to-mesenchymal transition (EMT) regulators besides Rho, Wnt, and Hedgehog (Hh) signaling. Unexpectedly, Wnt pathway inhibition increased the more immature, slowly cycling GPR56 + CD34 + fraction and Hh/EMT gene expression, while Wnt activation caused opposite effects. Our data suggest that the crucial role of GPR56 lies in its ability to co-activate these opposing signals, thus ensuring the constant supply of both LSC subsets. We show that CDK7 inhibitors suppress both LSC-enriched subsets in vivo and synergize with the Bcl-2 inhibitor venetoclax. Our data establish reciprocal transition between LSC compartments as a novel concept underlying the poor outcome in GPR56 high AML and propose combined CDK7 and Bcl-2 inhibition as LSC-directed therapy in this disease., (©2022 The Authors. Published under the terms of the CC BY 4.0 license.)