Loss of Pip4k2c confers liver-metastatic organotropism through insulin-dependent PI3K-AKT pathway activation.

Liver metastasis (LM) confers poor survival and therapy resistance across cancer types, but the mechanisms of liver-metastatic organotropism remain unknown. Here, through in vivo CRISPR-Cas9 screens, we found that Pip4k2c loss conferred LM but had no impact on lung metastasis or primary tumor growth. Pip4k2c-deficient cells were hypersensitized to insulin-mediated PI3K/AKT signaling and exploited the insulin-rich liver milieu for organ-specific metastasis. We observed concordant changes in PIP4K2C expression and distinct metabolic changes in 3,511 patient melanomas, including primary tumors, LMs and lung metastases. We found that systemic PI3K inhibition exacerbated LM burden in mice injected with Pip4k2c-deficient cancer cells through host-mediated increase in hepatic insulin levels; however, this circuit could be broken by concurrent administration of an SGLT2 inhibitor or feeding of a ketogenic diet. Thus, this work demonstrates a rare example of metastatic organotropism through co-optation of physiological metabolic cues and proposes therapeutic avenues to counteract these mechanisms.

Fatty acids are crucial to fuel NK cells upon acute retrovirus infection.

Natural killer (NK) cells are cytotoxic innate immune cells, able to recognize and eliminate virus-infected as well as cancer cells. Metabolic reprogramming is crucial for their activity as they have enhanced energy and nutritional demands for their functions during an infection. Fatty acids (FAs) represent an important source of cellular energy and are essential for proliferation of immune cells. However, the precise role of FAs for NK cells activity in retrovirus infection was unknown. Here we show that activated NK cells increase the expression of the FA uptake receptor CD36 and subsequently the uptake of FAs upon acute virus infection. We found an enhanced flexibility of NK cells to utilize FAs as source of energy compare to naïve NK cells. NK cells that were able to generate energy from FAs showed an augmented target cell killing and increased expression of cytotoxic parameters. However, NK cells that were unable to generate energy from FAs exhibited a severely decreased migratory capacity. Our results demonstrate that NK cells require FAs in order to fight acute virus infection. Susceptibility to severe virus infections as it is shown for people with malnutrition may be augmented by defects in the FA processing machinery, which might be a target to therapeutically boost NK cell functions in the future.

Post-fast refeeding enhances intestinal stem cell-mediated regeneration and tumourigenesis through mTORC1-dependent polyamine synthesis.

For more than a century, fasting regimens have improved health, lifespan, and tissue regeneration in diverse organisms, including humans. However, how fasting and post-fast refeeding impact adult stem cells and tumour formation has yet to be explored in depth. Here, we demonstrate that post-fast refeeding increases intestinal stem cell (ISC) proliferation and tumour formation: Post-fast refeeding augments the regenerative capacity of Lgr5+ intestinal stem cells (ISCs), and loss of the tumour suppressor Apc in ISCs under post-fast refeeding leads to a higher tumour incidence in the small intestine and colon than in the fasted or ad libitum (AL) fed states. This demonstrates that post-fast refeeding is a distinct state. Mechanistically, we discovered that robust induction of mTORC1 in post-fast-refed ISCs increases protein synthesis via polyamine metabolism to drive these changes, as inhibition of mTORC1, polyamine metabolite production, or protein synthesis abrogates the regenerative or tumourigenic effects of post-fast refeeding. Thus, fast-refeeding cycles must be carefully considered when planning diet-based strategies for regeneration without increasing cancer risk, as post-fast refeeding leads to a burst not only in stem cell-driven regeneration but also in tumourigenicity.

Review on atmospheric pressure ionization sources for gas chromatography-mass spectrometry. Part I: Current ion source developments and improvements in ionization strategies.

The use of atmospheric pressure ionization (API) sources has become very popular for gas chromatography-mass spectrometry (GC-MS) determinations. GC-API-MS shows important advantages over traditional vacuum ionization sources such as a lower fragmentation preserving the molecular or quasi-molecular ion, the combination of GC separations with advanced mass spectrometers typically developed for liquid chromatography (LC) systems, a significantly higher sensitivity, or the reduction of costs due to the capability to use the same MS for both LC- and GC- couplings. For these reasons, the development of new API sources and GC-API-MS platforms has exponentially increased during the last years. Thus, this review is mainly focused on the last advances in GC-API-MS instrumentation. New setups and couplings on extensively reported API techniques as well as the development of new API sources for GC-MS coupling are thoroughly discussed. Moreover, novel ionization strategies have been reviewed to overcome some of the drawbacks of GC-API-MS methodologies.

Review on atmospheric pressure ionization sources for gas chromatography-mass spectrometry. Part II: Current applications.

The application of atmospheric pressure ionization (API) sources in gas chromatography-mass spectrometry (GC-MS) determinations is becoming more popular since they have shown great capabilities to sort out the main drawbacks of vacuum ionization techniques like electron ionization (EI) and chemical ionization (CI). The development of new API techniques and set-ups have grown in the last decades, opening the field of GC-MS to new applications and facing some of the major issues in current analytical methodologies such as the requirement of a compromise between sensitivity and selectivity. Thus, this review is mainly focused on the use of GC-API-MS in different application fields such as food analysis (food safety and food metabolomics), environmental analysis, clinical analysis, drug and pharmaceutical analysis, and petroleomics, among others. The methodologies have been critically reviewed to compare the performance of different API sources and approaches, highlighting the main contributions to overcoming some of the major limitations of the current methodologies as well as the new perspectives that GC-API-MS might open in the different fields.

Potential of atmospheric pressure ionization sources for the analysis of free fatty acids in clinical and biological samples by gas chromatography-mass spectrometry.

Because of the central role of fatty acids in biological systems, their accurate quantification is still important. However, the impact of the complex matrix of biologically and clinically relevant samples such as plasma, serum, or cells makes the analysis still challenging, especially, when free non-esterified fatty acids have to be quantified. Here we developed and characterized a novel GC-MS method using pentafluorobenzyl bromide as a derivatization agent and compared different ionization techniques such as atmospheric pressure chemical ionization (APCI), atmospheric pressure chemical photoionization (APPI), and negative ion chemical ionization (NICI). The GC-APCI-MS showed the lowest limits of detection from 30 to 300 nM for a broad range of fatty acids and a similar response for various fatty acids from a chain length of 10 to 20 carbon atoms. This allows the number of internal standards necessary for accurate quantification to be reduced. Moreover, the use of pentafluorobenzyl bromide allows the direct derivatization of free fatty acids making them accessible for GC-MS analysis without labor-intense sample pretreatment.

Characterization of the Extracellular Volatile Metabolome of Pseudomonas Aeruginosa Applying an in vitro Biofilm Model under Cystic Fibrosis-Like Conditions.

BACKGROUND: Cystic fibrosis (CF) is an autosomal recessive hereditary disease that leads to the production of thickened mucus in the lungs, favouring polymicrobial infections, such as chronic lung infections with the bacterial opportunistic pathogen Pseudomonas aeruginosa. METHOD: A biofilm model in combination with an adapted sampling and GC-MS analysis method were applied to in vitro studies on different variables influencing the composition of the extracellular volatile metabolome of P. aeruginosa. RESULTS: A significant influence on the metabolome could be demonstrated for the culture medium as well as the atmosphere during cultivation (aerobic or anaerobic). Furthermore, a significant influence of the mucoid (alginate-overproducing) phenotype of the bacterium on quantity and composition of volatile organic compounds could be observed. Based on the results a solid culture medium was developed to simulate the nutrient conditions in the lungs of a CF patient. The extracellular volatile metabolome of bacterial strains P. aeruginosa ATCC 10145, PAO1 and FRD1 was characterized under CF-like conditions. CONCLUSIONS: Bacterial strain-dependent metabolites were identified. When P. aeruginosa PAO1 and FRD1 clinical isolates were compared, 36 metabolites showed significant variations in intensities. When the clinical isolates were compared with the reference strain (P. aeruginosa ATCC 10145), 28 metabolites (P. aeruginosa PAO1) and 70 metabolites (P. aeruginosa FRD1) were determined whose peaks showed significant deviation (p > 95%) in intensity. Furthermore, the bacterial strains could be differentiated from each other by means of two principal components.

Oxylipin metabolism is controlled by mitochondrial ß-oxidation during bacterial inflammation.

Oxylipins are potent biological mediators requiring strict control, but how they are removed en masse during infection and inflammation is unknown. Here we show that lipopolysaccharide (LPS) dynamically enhances oxylipin removal via mitochondrial ß-oxidation. Specifically, genetic or pharmacological targeting of carnitine palmitoyl transferase 1 (CPT1), a mitochondrial importer of fatty acids, reveal that many oxylipins are removed by this protein during inflammation in vitro and in vivo. Using stable isotope-tracing lipidomics, we find secretion-reuptake recycling for 12-HETE and its intermediate metabolites. Meanwhile, oxylipin ß-oxidation is uncoupled from oxidative phosphorylation, thus not contributing to energy generation. Testing for genetic control checkpoints, transcriptional interrogation of human neonatal sepsis finds upregulation of many genes involved in mitochondrial removal of long-chain fatty acyls, such as ACSL1,3,4, ACADVL, CPT1B, CPT2 and HADHB. Also, ACSL1/Acsl1 upregulation is consistently observed following the treatment of human/murine macrophages with LPS and IFN-γ. Last, dampening oxylipin levels by ß-oxidation is suggested to impact on their regulation of leukocyte functions. In summary, we propose mitochondrial ß-oxidation as a regulatory metabolic checkpoint for oxylipins during inflammation.

Effect of Sampling Rate and Data Pretreatment for Targeted and Nontargeted Analysis by Means of Liquid Chromatography Coupled to Drift Time Ion Mobility Quadruple Time-of-Flight Mass Spectrometry.

Ion mobility as an additional separation dimension can help to resolve and annotate metabolite and lipid biomarkers and provides important information about the components in a sample. Identifying relevant information in the resulting data is challenging because of the complexity of the data and data evaluation strategies for both targeted or nontargeted workflows. Frequently, feature analysis is used as a first step to search for differences between samples in discovery workflows. However, follow-up experimentation often leads to more targeted data extraction methods. In both cases, optimizing data sets for data extraction can make an important contribution to the overall results. In this work, we evaluate the effect of experimental conditions including acquisition sampling rate and data pretreatment on lipid standards and lipid extracts as examples of complex biological samples analyzed by liquid chromatography coupled to drift time ion mobility quadrupole time-of-flight mass spectrometry. The results show that a reduction of both peak variation and background noise can be achieved by optimizing the sampling rate. The use of data pretreatment including data smoothing, intensity thresholding, and spike removal also play an important role in improving detection and annotation of analytes from complex biological samples, whereas nonoptimal data sampling rates and preprocessing can lead to adverse effects including the loss or alternation of small, or closely eluting, low-abundant peaks.

Chemical characterization of eight herbal liqueurs by means of liquid chromatography coupled with ion mobility quadrupole time-of-flight mass spectrometry.

Herbal liqueurs are a large group of diverse alcoholic beverages with an ancient tradition produced by maceration of various herbs and spices and are commonly drunken before or after a meal to aid in the digestion because of their potential functional properties. In the presented work, eight different commercial herbal liqueurs were investigated with regard to their composition of phenolic compounds by liquid chromatography ion mobility quadrupole time-of-flight mass spectrometry (LC-IM-QTOF-MS). This multidimensional analytical platform uses all-ion fragmentation for a deep coverage of the foodome. After an extensive data clean-up, 3225 features were found. 213 features were manually annotated due to the absence of databases and software tools able to consider the drift time or Collison Cross Section (CCS) together with high resolution MS/MS spectra for identification. The identified compounds reflected the large variance between the investigated samples and a wealth of potential bioactive compounds that these liqueurs harbor.

Compositional characterization of native Peruvian chili peppers (Capsicum spp.).

The national Capsicum germplasm bank of Peru at INIA holds a unique collection of more than 700 Capsicum accessions, including many landraces. These conserved accessions have never been thoroughly characterized or evaluated. Another smaller collection exists at UNALM, and CIDRA provided taxonomically characterized fruits from the Amazon region of Ucayali. Of these collections, 147 accessions have been selected to represent the biodiversity of Peruvian Capsicum annuum , Capsicum baccatum , Capsicum chinense , and Capsicum frutescens by morphological traits as well as by agronomic characteristics and regional origin. All fruits from the selected accessions have been oven-dried and ground in Peru and analyzed in Germany. Results are reported for each accession by total capsaicinoids and capsaicinoid pattern, total polyphenol content, antioxidant capacity, specific flavonoids (quercetin, kaempferol, luteolin, apigenin), fat content, vitamin C, surface color, and extractable color. A wide variability in phytochemical composition and concentration levels was found.