Succinate receptor 1 inhibits mitochondrial respiration in cancer cells addicted to glutamine

Cancer cells display metabolic alterations to meet the bioenergetic demands for their high proliferation rates. Succinate is a central metabolite of the tricarboxylic acid (TCA) cycle, but was also shown to act as an oncometabolite and to specifically activate the succinate receptor 1 (SUCNR1), which is expressed in several types of cancer. However, functional studies focusing on the connection between SUCNR1 and cancer cell metabolism are still lacking. In the present study, we analyzed the role of SUCNR1 for cancer cell metabolism and survival applying different signal transduction, metabolic and imaging analyses. We chose a gastric, a lung and a pancreatic cancer cell line for which our data revealed functional expression of SUCNR1. Further, presence of glutamine (Gln) caused high respiratory rates and elevated expression of SUCNR1. Knockdown of SUCNR1 resulted in a significant increase of mitochondrial respiration and superoxide production accompanied by an increase in TCA cycle throughput and a reduction of cancer cell survival in the analyzed cancer cell lines. Combination of SUCNR1 knockdown and treatment with the chemotherapeutics cisplatin and gemcitabine further increased cancer cell death. In summary, our data implicates that SUCNR1 is crucial for Gln-addicted cancer cells by limiting TCA cycle throughput, mitochondrial respiration and the production of reactive oxygen species, highlighting its potential as a pharmacological target for cancer treatment

Combining metabolic phenotype determination with metabolomics and transcriptional analyses to reveal pathways regulated by hydroxycarboxylic acid receptor 2

Background The adaptation of cellular metabolism is considered a hallmark of cancer. Oncogenic signaling pathways support tumorigenesis and cancer progression through the induction of certain metabolic phenotypes associated with altered regulation of key metabolic enzymes. Hydroxycarboxylic acid receptor 2 (HCA(2)) is a G protein-coupled receptor previously shown to act as a tumor suppressor. Here, we aimed to unveil the connection between cellular metabolism and HCA(2) in BT-474 cells. Moreover, we intend to clarify how well this metabolic phenotype is reflected in transcriptional changes and metabolite levels as determined by global metabolomics analyses. Methods We performed both, siRNA mediated knockdown of HCA(2) and stimulation with the HCA(2)-specific agonist monomethyl fumarate. Seahorse technology was used to determine the role of HCA(2) in BT-474 breast cancer cell metabolism and its potential to induce a switch in the metabolic phenotype in the presence of different energy substrates. Changes in the mRNA expression of metabolic enzymes were detected with real-time quantitative PCR (RT-qPCR). Untargeted liquid chromatography-mass spectrometry (LC-MS) metabolic profiling was used to determine changes in metabolite levels. Results Knockdown or stimulation of HCA(2) induced changes in the metabolic phenotype of BT474 cells dependent on the availability of energy substrates. The presence of HCA(2) was associated with increased glycolytic flux with no fatty acids available. This was reflected in the increased mRNA expression of the glycolytic enzymes PFKFB4 and PKM2, which are known to promote the Warburg effect and have been described as prognostic markers in different types of cancer. With exogenous palmitate present, HCA(2) caused elevated fatty acid oxidation and likely lipolysis. The increase in lipolysis was also detectable at the transcriptional level of ATGL and the metabolite levels of palmitic and stearic acid. Conclusions We combined metabolic phenotype determination with metabolomics and transcriptional analyses and identified HCA(2) as a regulator of glycolytic flux and fatty acid metabolism in BT-474 breast cancer cells. Thus, HCA(2), for which agonists are already widely used to treat diseases such as psoriasis or hyperlipidemia, may prove useful as a target in combination cancer therapy

Hydroxycarboxylic acid receptor 3 and GPR84: Two metabolite-sensing G protein-coupled receptors with opposing functions in innate immune cells

G protein-coupled receptors (GPCRs) are key regulatory proteins of immune cell function inducing signaling in response to extracellular (pathogenic) stimuli. Although unrelated, hydroxycarboxylic acid receptor 3 (HCA3) and GPR84 share signaling via Gαi/o proteins and the agonist 3-hydroxydecanoic acid (3HDec). Both receptors are abundantly expressed in monocytes, macrophages and neutrophils but have opposing functions in these innate immune cells. Detailed insights into the molecular mechanisms and signaling components involved in immune cell regulation by GPR84 and HCA3 are still lacking. Here, we report that GPR84-mediated pro-inflammatory signaling depends on coupling to the hematopoietic cell-specific Gα15 protein in human macrophages, while HCA3 exclusively couples to Gαi protein. We show that activated GPR84 induces Gα15-dependent ERK activation, increases intracellular Ca2+ and IP3 levels as well as ROS production. In contrast, HCA3 activation shifts macrophage metabolism to a less glycolytic phenotype, which is associated with anti-inflammatory responses. This is supported by an increased release of anti-inflammatory IL-10 and a decreased secretion of pro-inflammatory IL-1β. In primary human neutrophils, stimulation with HCA3 agonists counteracts the GPR84-induced neutrophil activation. Our analyses reveal that 3HDec acts solely through GPR84 but not HCA3 activation in macrophages. In summary, this study shows that HCA3 mediates hyporesponsiveness in response to metabolites derived from dietary lactic acid bacteria and uncovers that GPR84, which is already targeted in clinical trials, promotes pro-inflammatory signaling via Gα15 protein in macrophages

Ice-tethered platforms & ROV, Progress report 2018

Progress report on the work of FRAM Task 3.1 for the bi-annual FRAM workshop

Associated factors and comorbidities in patients with pyoderma gangrenosum in Germany: a retrospective multicentric analysis in 259 patients

Background: Pyoderma gangrenosum (PG) is a rarely diagnosed ulcerative neutrophilic dermatosis with unknown origin that has been poorly characterized in clinical studies so far. Consequently there have been significant discussions about its associated factors and comorbidities. The aim of our multicenter study was to analyze current data from patients in dermatologic wound care centers in Germany in order to describe associated factors and comorbidities in patients with PG. Methods: Retrospective clinical investigation of patients with PG from dermatologic wound care centers in Germany. Results: We received data from 259 patients with PG from 20 different dermatologic wound care centers in Germany. Of these 142 (54.8\%) patients were female, 117 (45.2\%) were male; with an age range of 21 to 95 years, and a mean of 58 years. In our patient population we found 45.6\% with anemia, 44.8\% with endocrine diseases, 12.4\% with internal malignancies, 9.3\% with chronic inflammatory bowel diseases and 4.3\% with elevated creatinine levels. Moreover 25.5\% of all patients had a diabetes mellitus with some aspects of potential association with the metabolic syndrome. Conclusions: Our study describes one of the world's largest populations with PG. Beside the well-known association with chronic bowel diseases and neoplasms, a potentially relevant new aspect is an association with endocrine diseases, in particular the metabolic syndrome, thyroid dysfunctions and renal disorders. Our findings represent clinically relevant new aspects. This may help to describe the patients' characteristics and help to understand the underlying pathophysiology in these often misdiagnosed patients

Standard for Synthesis of Customized Peptides by Non-Ribosomal Peptide Synthetases

The purpose of this RFC is to introduce a standardized framework for the engineering of customizable non-ribosomal peptide synthetases (NRPS) and their application for in vivo and in vitro synthesis of short non-ribosomal peptides (NRPs) of user-defined sequence and structure

HiCT: High Throughput Protocols For CPE Cloning And Transformation

The purpose of this RFC is to provide instructions for a rapid and cost efficient cloning and transformation method which allows for the manufacturing of multi-fragment plasmid constructs in a parallelized manner: High Throughput Circular Extension Cloning and Transformation (HiCT). Description of construct libraries generated by the HiCT method can be found at http://2013.igem.org/Team:Heidelberg/Indigoidine. This RFC also points out further optimization strategies with regard to construct stability, reduction of transformation background and the generation of competent cells

Clinical Frailty Scale (CFS) reliably stratifies octogenarians in German ICUs: a multicentre prospective cohort study

Background: In intensive care units (ICU) octogenarians become a routine patients group with aggravated therapeutic and diagnostic decision-making. Due to increased mortality and a reduced quality of life in this high-risk population, medical decision-making a fortiori requires an optimum of risk stratification. Recently, the VIP-1 trial prospectively observed that the clinical frailty scale (CFS) performed well in ICU patients in overall-survival and short-term outcome prediction. However, it is known that healthcare systems differ in the 21 countries contributing to the VIP-1 trial. Hence, our main focus was to investigate whether the CFS is usable for risk stratification in octogenarians admitted to diversified and high tech German ICUs. Methods: This multicentre prospective cohort study analyses very old patients admitted to 20 German ICUs as a sub-analysis of the VIP-1 trial. Three hundred and eight patients of 80 years of age or older admitted consecutively to participating ICUs. CFS, cause of admission, APACHE II, SAPS II and SOFA scores, use of ICU resources and ICU- and 30-day mortality were recorded. Multivariate logistic regression analysis was used to identify factors associated with 30-day mortality. Results: Patients had a median age of 84 [IQR 82–87] years and a mean CFS of 4.75 (± 1.6 standard-deviation) points. More than half of the patients (53.6%) were classified as frail (CFS ≥ 5). ICU-mortality was 17.3% and 30-day mortality was 31.2%. The cause of admission (planned vs. unplanned), (OR 5.74) and the CFS (OR 1.44 per point increase) were independent predictors of 30-day survival. Conclusions: The CFS is an easy determinable valuable tool for prediction of 30-day ICU survival in octogenarians, thus, it may facilitate decision-making for intensive care givers in Germany. Trial registration: The VIP-1 study was retrospectively registered on ClinicalTrials.gov (ID: NCT03134807 ) on May 1, 2017

Platelet ice under Arctic pack ice in winter

The formation of platelet ice is well known to occur under Antarctic sea ice, where subice platelet layers form from supercooled ice shelf water. In the Arctic, however, platelet ice formation has not been extensively observed, and its formation and morphology currently remain enigmatic. Here, we present the first comprehensive, long‐term in situ observations of a decimeter thick subice platelet layer under free‐drifting pack ice of the Central Arctic in winter. Observations carried out with a remotely operated underwater vehicle (ROV) during the midwinter leg of the MOSAiC drift expedition provide clear evidence of the growth of platelet ice layers from supercooled water present in the ocean mixed layer. This platelet formation takes place under all ice types present during the surveys. Oceanographic data from autonomous observing platforms lead us to the conclusion that platelet ice formation is a widespread but yet overlooked feature of Arctic winter sea ice growth