31-Phosphorus Magnetic Resonance Spectroscopy in Evaluation of Glioma and Metastases in 3T MRI

Background: 31-Phosphorus magnetic resonance spectroscopy (31-P MRS) has excellent potential for clinical neurological practice because of its noninvasive in-vivo assessment of cellular energy metabolism and the indirect evaluation of the phospholipid composition of the cell membrane, intracellular pH, and intracellular Mg2+ concentration. Purpose: The aim of this study was to evaluate the metabolic characteristics of glioma and metastases using 31-P MRS and assess utility to differentiate both. Study Type: Prospective study. Population: Fifteen consecutive patients with brain tumor. Field Strength/Sequence: Three-tesla magnetic resonance imaging/three-dimensional MRS imaging sequence. Statistical Tests: Unpaired sample t-test, and one-way analysis of variance with Tukey's post-hoc test. Results: Significantly decreased values of phosphomonoesters/inorganic phosphate (PME/Pi) in the tumor group (1.22 ± 0.72) compared with controls (2.28 ± 1.44) with a p-value of 0.041 were observed. There is a significant decrease in phosphocreatine (PCr)/Pi values (energy demand) in the tumor group (2.76 ± 0.73) compared with controls (4.13 ± 1.75) with a p-value of 0.050. Significant increase in Pi/adenosine triphosphate (ATP) was noted in tumor group (0.28 ± 0.09) compared with controls (0.22 ± 0.08) with p-value 0.049. Among tumor group, PME/PCr values were significantly decreased in gliomas (0.35 ± 0.17) than metastasis (0.58 ± 0.23) compared with controls with a p-value of 0.047. A significant decrease in PME/ATP was noted in gliomas (0.25 ± 0.12) than metastasis (0.41 ± 0.14) compared with controls with a p-value of 0.034. The tumor group exhibits alkaline pH (7.12 ± 0.10) compared with the normal parenchyma (7.04 ± 0.06) with a significant p-value of 0.025. Glioma and metastasis could not be differentiated with pH. However, the perilesional edema of glioma shows alkaline pH (7.09 ± 0.06) and metastasis shows acidic pH (7.02 ± 0.05) with a significant p-value of 0.030. Conclusion: Our study provides new insight into the cellular constituents and pH of gliomas and metastases and results were significant in differentiation between these two. However, due to the additional high expense, it is available as a research tool in very few institutions in India.

Olive Oil Extracts and Oleic Acid Attenuate the LPS-Induced Inflammatory Response in Murine RAW264.7 Macrophages but Induce the Release of Prostaglandin E2

Olive oil contains high amounts of oleic acid (OA). Although OA has been described to inhibit inflammatory processes, the effects of olive oil on cellular mechanisms remain poorly understood. Therefore, we compared the effects of major fatty acids (FA) from olive oil with those of olive oil extracts (OOE) on inflammatory mediators and alterations in the cellular phospholipid composition in murine macrophages. Upon treatment with different OOE, FA compositions of lipopolysaccharide (LPS)-stimulated murine RAW264.7 macrophages were analyzed using gas chromatography. Olive oil extracts and OA significantly reduced the LPS-induced expression of inducible nitric oxide synthase (iNos), cyclooxygenase (Cox2), and interleukin-6 mRNA. In addition, a significant decrease in Cox2 and iNos protein expression was observed. The formation of nitric oxide was significantly reduced, while the formation of prostaglandin (PG) E2 from arachidonic acid significantly increased after treatment with OOE or OA. The latter was associated with a shift in the phospholipid FA composition from arachidonic acid to OA, resulting in an elevated availability of arachidonic acid. Together, OOE and OA mediate anti-inflammatory effects in vitro but increase the release of arachidonic acid and hereinafter PGE2, likely due to elongation of OA and competitive incorporation of fatty acids into membrane phospholipids.

How to Combat Gram-Negative Bacteria Using Antimicrobial Peptides: A Challenge or an Unattainable Goal?

Antimicrobial peptides (AMPs) represent a promising and effective alternative for combating pathogens, having some advantages compared to conventional antibiotics. However, AMPs must also contend with complex and specialised Gram-negative bacteria envelops. The variety of lipopolysaccharide and phospholipid composition in Gram-negative bacteria strains and species are decisive characteristics regarding their susceptibility or resistance to AMPs. Such biological and structural barriers have created delays in tuning AMPs to deal with Gram-negative bacteria. This becomes even more acute because little is known about the interaction AMP–Gram-negative bacteria and/or AMPs’ physicochemical characteristics, which could lead to obtaining selective molecules against Gram-negative bacteria. As a consequence, available AMPs usually have highly associated haemolytic and/or cytotoxic activity. Only one AMP has so far been FDA approved and another two are currently in clinical trials against Gram-negative bacteria. Such a pessimistic panorama suggests that efforts should be concentrated on the search for new molecules, designs and strategies for combating infection caused by this type of microorganism. This review has therefore been aimed at describing the currently available AMPs for combating Gram-negative bacteria, exploring the characteristics of these bacteria’s cell envelop hampering the development of new AMPs, and offers a perspective regarding the challenges for designing new AMPs against Gram-negative bacteria.

Bicarbonate-Stimulated Membrane Reorganization in Stallion Spermatozoa

Classical in vitro fertilization (IVF) is still poorly successful in horses. This lack of success is thought to be due primarily to inadequate capacitation of stallion spermatozoa under in vitro conditions. In species in which IVF is successful, bicarbonate, calcium, and albumin are considered the key components that enable a gradual reorganization of the sperm plasma membrane that allows the spermatozoa to undergo an acrosome reaction and fertilize the oocyte. The aim of this work was to comprehensively examine contributors to stallion sperm capacitation by investigating bicarbonate-induced membrane remodelling steps, and elucidating the contribution of cAMP signalling to these events. In the presence of capacitating media containing bicarbonate, a significant increase in plasma membrane fluidity was readily detected using merocyanine 540 staining in the majority of viable spermatozoa within 15 min of bicarbonate exposure. Specific inhibition of soluble adenylyl cyclase (sAC) in the presence of bicarbonate by LRE1 significantly reduced the number of viable sperm with high membrane fluidity. This suggests a vital role for sAC-mediated cAMP production in the regulation of membrane fluidity. Cryo-electron tomography of viable cells with high membrane fluidity revealed a range of membrane remodelling intermediates, including destabilized membranes and zones with close apposition of the plasma membrane and the outer acrosomal membrane. However, lipidomic analysis of equivalent viable spermatozoa with high membrane fluidity demonstrated that this phenomenon was neither accompanied by a gross change in the phospholipid composition of stallion sperm membranes nor detectable sterol efflux (p > 0.05). After an early increase in membrane fluidity, a significant and cAMP-dependent increase in viable sperm with phosphatidylserine (PS), but not phosphatidylethanolamine (PE) exposure was noted. While the events observed partly resemble findings from the in vitro capacitation of sperm from other mammalian species, the lack of cholesterol removal appears to be an equine-specific phenomenon. This research will assist in the development of a defined medium for the capacitation of stallion sperm and will facilitate progress toward a functional IVF protocol for horse gametes.

FADS1 rs174550 genotype and high linoleic acid diet modify plasma PUFA phospholipids in a dietary intervention study

Introduction Fatty acid desaturase 1 (FADS1) gene encodes for delta-5 desaturase enzyme which is needed in conversion of linoleic acid (LA) to arachidonic acid (AA). Recent studies have shown that response to dietary PUFAs differs between the genotypes in circulating fatty acids. However, interactions between the FADS1 genotype and dietary LA on overall metabolism have not been studied. Objectives We aimed to examine the interactions of FADS1 rs174550 genotypes (TT and CC) and high-LA diet to identify plasma metabolites that respond differentially to dietary LA according to the FADS1 genotype. Methods A total of 59 men (TT n = 26, CC n = 33) consumed a sunflower oil supplemented diet for 4 weeks. Daily dose of 30, 40, or 50 ml was calculated based on body mass index. It resulted in 17–28 g of LA on top of the usual daily intake. Fasting plasma samples at the beginning and at the end of the intervention were analyzed with LC–MS/MS non-targeted metabolomics method. Results At the baseline, the carriers of FADS1 rs174550-TT genotype had higher abundance of long-chain PUFA phospholipids compared to the FADS1 rs174550-CC one. In response to the high-LA diet, LA phospholipids and long-chain acylcarnitines increased and lysophospholipids decreased in fasting plasma similarly in both genotypes. LysoPE (20:4), LysoPC (20:4), and PC (16:0_20:4) decreased and cortisol increased in the carriers of rs174550-CC genotype; however, these genotype–diet interactions were not significant after correction for multiple testing. Conclusion Our findings show that both FADS1 rs174550 genotype and high-LA diet modify plasma phospholipid composition. Trial registration The study was registered to ClinicalTrials: NCT02543216, September 7, 2015 (retrospectively registered).

Defunctionalizing Intracellular Organelles with Genetically-Encoded Molecular Tools Based on Engineered Phospholipase A/Acyltransferases (PLAATs)

Organelles vitally achieve multifaceted functions to maintain cellular homeostasis. Genetic and pharmacological approaches to manipulate individual organelles are powerful in probing their physiological roles. However, many of them are either slow in action, limited to certain organelles, or rely on toxic agents. Here, we designed a generalizable molecular tool utilizing phospholipase A/acyltransferases (PLAATs) for rapid induction of organelle defunctionalization via remodeling of the membrane phospholipid composition. In particular, we identified a minimal, fully catalytic PLAAT with no unfavorable side effects. Chemically-induced translocation of the engineered PLAAT to the mitochondria surface resulted in their rapid deformation in a phospholipase activity dependent manner, followed by loss of luminal proteins as well as dissipated membrane potential, thus invalidating the functionality. To demonstrate wide applicability, we then adapted the molecular tool in peroxisomes, and observed leakage of matrix-resident functional proteins. The technique was compatible with optogenetic control, viral delivery and operation in primary neuronal cultures. Due to such versatility, the PLAAT strategy should present a novel utility in organelle biology of diverse contexts.

LPEATs Tailor Plant Phospholipid Composition through Adjusting Substrate Preferences to Temperature

Acyl-CoA:lysophosphatidylethanolamine acyltransferases (LPEATs) are known as enzymes utilizing acyl-CoAs and lysophospholipids to produce phosphatidylethanolamine. Recently, it has been discovered that they are also involved in the growth regulation of Arabidopsis thaliana. In our study we investigated expression of each Camelina sativa LPEAT isoform and their behavior in response to temperature changes. In order to conduct a more extensive biochemical evaluation we focused both on LPEAT enzymes present in microsomal fractions from C. sativa plant tissues, and on cloned CsLPEAT isoforms expressed in yeast system. Phylogenetic analyses revealed that CsLPEAT1c and CsLPEAT2c originated from Camelina hispida, whereas other isoforms originated from Camelina neglecta. The expression ratio of all CsLPEAT1 isoforms to all CsLPEAT2 isoforms was higher in seeds than in other tissues. The isoforms also displayed divergent substrate specificities in utilization of LPE; CsLPEAT1 preferred 18:1-LPE, whereas CsLPEAT2 preferred 18:2-LPE. Unlike CsLPEAT1, CsLPEAT2 isoforms were specific towards very-long-chain fatty acids. Above all, we discovered that temperature strongly regulates LPEATs activity and substrate specificity towards different acyl donors, making LPEATs sort of a sensor of external thermal changes. We observed the presented findings not only for LPEAT activity in plant-derived microsomal fractions, but also for yeast-expressed individual CsLPEAT isoforms.

Hepatic resistance to cold ferroptosis in a mammalian hibernator Syrian hamster depends on effective storage of diet-derived α-tocopherol

Mammalian hibernators endure severe and prolonged hypothermia that is lethal to non-hibernators, including humans and mice. The mechanisms responsible for the cold resistance remain poorly understood. Here, we found that hepatocytes from a mammalian hibernator, the Syrian hamster, exhibited remarkable resistance to prolonged cold culture, whereas murine hepatocytes underwent cold-induced cell death that fulfills the hallmarks of ferroptosis such as necrotic morphology, lipid peroxidation and prevention by an iron chelator. Unexpectedly, hepatocytes from Syrian hamsters exerted resistance to cold- and drug-induced ferroptosis in a diet-dependent manner, with the aid of their superior ability to retain dietary α-tocopherol (αT), a vitamin E analog, in the liver and blood compared with those of mice. The liver phospholipid composition is less susceptible to peroxidation in Syrian hamsters than in mice. Altogether, the cold resistance of the hibernator’s liver is established by the ability to utilize αT effectively to prevent lipid peroxidation and ferroptosis.