Peripheral administration of lipidized NPAF and NPFF analogs does not influence central food intake regulation but induces anxiety-like behavior.

RF-amide peptides influence multiple physiological processes, including the regulation of appetite, stress responses, behavior, and reproductive and endocrine functions. In this study, we examined the roles of neuropeptide FF receptors (NPFFR1 and NPFFR2) by generating several lipidized analogs of neuropeptide AF (NPAF) and 1DMe, a stable analog of neuropeptide FF (NPFF). These analogs were administered peripherally for the first time to investigate their effects on food intake and other potential physiological outcomes. Lipidized NPAF and 1DMe analogs exhibited enhanced stability and increased pharmacokinetics. These analogs demonstrated preserved high affinity for NPFFR2 in the nanomolar range, while the binding affinity for NPFFR1 was tens of nanomoles. They activated the ERK and Akt signaling pathways in cells overexpressing the NPFFR1 and NPFFR2 receptors. Acute food intake in fasted mice decreased after the peripheral administration of oct-NPAF or oct-1DMe. However, this effect was not as pronounced as that observed after the injection of palm11-PrRP31, a potent anorexigenic compound used as a comparator that binds to GPR10 and the NPFFR2 receptor with high affinity. Neither oct-1DMe nor oct-NPAF decreased food intake or body weight in mice with diet-induced obesity during long-term treatment. In mice treated with oct-1DMe, we observed decreased activity in the central zone during the open field test and decreased activity in the open arms of the elevated plus maze. Furthermore, we observed a decrease in plasma noradrenaline levels and an increase in plasma corticosterone levels, as well as an increase in Crh expression in the hypothalamus. Moreover, neuronal activity in the hypothalamus was increased after treatment with oct-1DMe. In this study, we report that oct-1DMe did not have any long-term effects on the central regulation of food intake; however, it caused anxiety-like behavior.

Lipidization as a tool toward peptide therapeutics.

Peptides, as potential therapeutics continue to gain importance in the search for active substances for the treatment of numerous human diseases, some of which are, to this day, incurable. As potential therapeutic drugs, peptides have many favorable chemical and pharmacological properties, starting with their great diversity, through their high affinity for binding to all sort of natural receptors, and ending with the various pathways of their breakdown, which produces nothing but amino acids that are nontoxic to the body. Despite these and other advantages, however, they also have their pitfalls. One of these disadvantages is the very low stability of natural peptides. They have a short half-life and tend to be cleared from the organism very quickly. Their instability in the gastrointestinal tract, makes it impossible to administer peptidic drugs orally. To achieve the best pharmacologic effect, it is desirable to look for ways of modifying peptides that enable the use of these substances as pharmaceuticals. There are many ways to modify peptides. Herein we summarize the approaches that are currently in use, including lipidization, PEGylation, glycosylation and others, focusing on lipidization. We describe how individual types of lipidization are achieved and describe their advantages and drawbacks. Peptide modifications are performed with the goal of reaching a longer half-life, reducing immunogenicity and improving bioavailability. In the case of neuropeptides, lipidization aids their activity in the central nervous system after the peripheral administration. At the end of our review, we summarize all lipidized peptide-based drugs that are currently on the market.

Combination of quinoxaline with pentamethinium system: Mitochondrial staining and targeting.

Pentamethinium indolium salts are promising fluorescence probes and anticancer agents with high mitochondrial selectivity. We synthesized two indolium pentamethinium salts: a cyclic form with quinoxaline directly incorporated in the pentamethinium chain (cPMS) and an open form with quinoxaline substitution in the γ-position (oPMS). To better understand their properties, we studied their interaction with mitochondrial phospholipids (cardiolipin and phosphatidylcholine) by spectroscopic methods (UV-Vis, fluorescence, and NMR spectroscopy). Both compounds displayed significant affinity for cardiolipin and phosphatidylcholine, which was associated with a strong change in their UV-Vis spectra. Nevertheless, we surprisingly observed that fluorescence properties of cPMS changed in complex with both cardiolipin and phosphatidylcholine, whereas those of oPMS only changed in complex with cardiolipin. Both salts, especially cPMS, display high usability in mitochondrial imaging and are cytotoxic for cancer cells. The above clearly indicates that conjugates of pentamethinium and quinoxaline group, especially cPMS, represent promising structural motifs for designing mitochondrial-specific agents.

Search for lipidized PrRP analogs with strong anorexigenic effect: In vitro and in vivo studies.

Prolactin-releasing peptide (PrRP) is an anorexigenic neuropeptide that attenuates food intake and increases energy expenditure. We designed three series of new lipidized PrRP31 analogs of different lengths of fatty acids attached at amino acids 1 or 11 directly or via linkers, part of them acetylated at the N-terminus and/or modified with dichlorophenylalanine (PheCl2) at the C-terminus. We tested their affinity for and activation of signaling pathways relevant to receptors GPR10, NPFF-R2, and NPFF-R1, effect on food intake in fasted or freely fed mice and rats, and stability in rat plasma. We aimed to select a strong dual GPR10/NPFF-R2 agonist whose affinity for NPFF-1 was not enhanced. The selected potent analog was then tested for body weight-lowering potency after chronic administration in mice with diet-induced obesity. PrRP31 analogs lipidized by monocarboxylic fatty acids showed strong dual affinity for both GPR10 and NPFF-R2 and activated MAPK/ERK1/2, Akt and CREB in cells overexpressing GPR10 and NPFF-R2. The selected analog stabilized at N- and C-termini and palmitoylated through the TTDS linker to Lys11 is a powerful dual agonist GPR10/NPFF-R2 at not enhanced affinity for NPFF-R1. It showed strong anti-obesity properties in mice with diet-induced obesity and became a potential compound for further studies.

A Novel Truncated Liver Enriched Antimicrobial Peptide-2 Palmitoylated at its N-Terminal Antagonizes Effects of Ghrelin.

Ghrelin is secreted in the stomach during fasting and targets the growth hormone secretagogue receptor (GHSR1a) in the hypothalamus and brainstem to exert its orexigenic effect. Recently, liver enriched antimicrobial peptide-2 (LEAP2) was identified as an endogenous high-affinity GHSR1a antagonist. LEAP2 is a 40-amino acid peptide with two disulfide bridges and GHRS1a affinity in the N-terminal hydrophobic part. In this study, we tested modified truncated N-terminal peptide LEAP2 (1-14), along with its myristoylated, palmitoylated, and stearoylated analogs, to determine their affinity to and activation of GHSR1a and their anorexigenic effects after acute peripheral administration. The lipidized analogs bound GHSR1a with affinity similar to that of natural LEAP2, and lipidization significantly enhanced the affinity of LEAP2(1-14) to GHSR1a. According to the beta-lactamase reporter gene response, the natural GHSR1a agonist ghrelin activated the receptor with nanomolar EC50 LEAP2(1-14) analogs behaved as inverse agonists of GHSR1a and suppressed internal activity of the receptor with EC50 values in the 10-8 M range. LEAP2(1-14) analogs significantly lowered acute food intake in overnight fasted mice, and palmitoylated LEAP2(1-14) was the most potent. In free-fed mice, all LEAP2(1-14) analogs significantly decreased the orexigenic effect of the stable ghrelin analog [Dpr3]Ghrelin. Moreover, palmitoylated LEAP2(1-14) inhibited the growth hormone (GH) release induced by [Dpr3] Ghrelin and exhibited an increased stability in rat plasma compared with LEAP2(1-14). In conclusion, palmitoylated LEAP2(1-14) had the most pronounced affinity for GHSR1a, had an anorexigenic effect, exhibited stability in rat plasma, and attenuated [Dpr3]Ghrelin-induced GH release. Such properties render palmitoylated LEAP2(1-14) a promising substance for antiobesity treatment. SIGNIFICANCE STATEMENT: The agonist and antagonist of one receptor are rarely found in one organism. For ghrelin receptor (growth hormone secretagogue receptor, GHSR), endogenous agonist ghrelin and endogenous antagonist/inverse agonist liver enriched antimicrobial peptide-2 (LEAP2) co-exist and differently control GHSR signaling. As ghrelin has a unique role in food intake regulation, energy homeostasis, and cytoprotection, lipidized truncated LEAP2 analogs presented in this study could serve not only to reveal the relationship between ghrelin and LEAP2 but also for development of potential anti-obesity agents.

Advanced microextraction techniques for the analysis of amphetamines in human breast milk and their comparison with conventional methods.

Breast milk analysis provides useful information about acute newborn exposure to harmful substances, such as psychoactive drugs abused by a nursing mother. Since breast milk represents a complex matrix with large amounts of interfering compounds, a comprehensive sample pre-treatment is necessary. This work focuses on determination of amphetamines and synthetic cathinones in human breast milk by microextraction techniques (liquid-phase microextraction and electromembrane extraction), and their comparison to more conventional treatment methods (protein precipitation, liquid-liquid extraction, and salting-out assisted liquid-liquid extraction). The aim of this work was to optimize and validate all the extraction procedures and thoroughly assess their advantages and disadvantages with special regard to their routine clinical use. The applicability of the extractions was further verified by the analysis of six real samples collected from breastfeeding mothers suspected of amphetamine abuse. The membrane microextraction techniques turned out to be the most advantageous as they required low amounts of organic solvents but still provided efficient sample clean-up, excellent quantification limit (0.5 ng mL-1), and good recovery (81-91% and 40-89% for electromembrane extraction and liquid-phase microextraction, respectively). The traditional liquid-liquid extraction as well as the salting-out assisted liquid-liquid extraction showed comparable recoveries (41-85% and 63-88%, respectively), but higher quantification limits (2.5 ng mL-1 and 5 ng mL-1, respectively). Moreover, these methods required multiple operating steps and were time consuming. Protein precipitation was fast and simple, but it demonstrated poor sample clean-up, low recovery (56-58%) and high quantification limit (5 ng mL-1). Based on the overall results, microextraction methods can be considered promising candidates, even for routine laboratory use.

MALDI Mass Spectrometry Imaging of Lipids on Free-Floating Brain Sections and Immunohistochemically Colocalized Markers of Neurodegeneration.

In mass spectrometry imaging (MSI), the essential steps in sample preparation include collection and storage. The most widely used preservation procedure for MSI consists in freezing samples and storing them at temperatures below -80 °C. On the other hand, the most common method for preserving biological samples in clinical practice is their fixation in paraformaldehyde. The storage of free-floating sections is a particular type of the preservation of paraformaldehyde-fixed tissues that is used in immunohistochemistry. This chapter describes the approach of the multimodal imaging of free-floating brain sections using the MSI of lipids and the immunohistochemistry of neurodegeneration markers.

Stability Study of Cannabidiol in the Form of Solid Powder and Sunflower Oil Solution.

Stability studies represent an essential component of pharmaceutical development, enabling critical evaluation of the therapeutic potential of an active pharmaceutical ingredient (API) or a final pharmaceutical product under the influence of various environmental factors. The aim of the present study was to investigate the chemical stability of cannabidiol (CBD) in the form of a solid powder (hereinafter referred to as CBD powder) and also dissolved in sunflower oil. We performed stress studies in accordance with the International Conference on Harmonization (ICH) guidelines, where 5 mg of marketed CBD in the form of a solid powder and in form of oil solution were exposed for 7 and 14, 30, 60, 90, 180, 270, and 365 days to precisely defined temperature and humidity conditions, 25 °C ± 2 °C/60% RH ± 5% and 40 °C ± 2 °C/75% RH ± 5% in both open and closed vials in the dark. CBD powder was significantly more stable than CBD in oil solution. Such finding is important because CBD is often administered dissolved in oil matrix in practice due to very good bioavailability. Thus, the knowledge on admissible shelf time is of paramount importance.

Iron Complexes of Flavonoids-Antioxidant Capacity and Beyond.

Flavonoids are common plant natural products able to suppress ROS-related damage and alleviate oxidative stress. One of key mechanisms, involved in this phenomenon is chelation of transition metal ions. From a physiological perspective, iron is the most significant transition metal, because of its abundance in living organisms and ubiquitous involvement in redox processes. The chemical, pharmaceutical, and biological properties of flavonoids can be significantly affected by their interaction with transition metal ions, mainly iron. In this review, we explain the interaction of various flavonoid structures with Fe(II) and Fe(III) ions and critically discuss the influence of chelated ions on the flavonoid biochemical properties. In addition, specific biological effects of their iron metallocomplexes, such as the inhibition of iron-containing enzymes, have been included in this review.

Spectroscopic study of in situ-formed metallocomplexes of proton pump inhibitors in water.

Proton pump inhibitors, such as omeprazole, pantoprazole and lansoprazole, are an important group of clinically used drugs. Generally, they are considered safe without direct toxicity. Nevertheless, their long-term use can be associated with a higher risk of some serious pathological states (e.g. amnesia and oncological and neurodegenerative states). It is well known that dysregulation of the metabolism of transition metals (especially iron ions) plays a significant role in these pathological states and that the above drugs can form complexes with metal ions. However, to the best of our knowledge, this phenomenon has not yet been described in water systems. Therefore, we studied the interaction between these drugs and transition metal ions in the surrounding water environment (water/DMSO, 99:1, v/v) by absorption spectroscopy. In the presence of Fe(III), a strong redshift was observed, and more importantly, the affinities of the drugs (represented as binding constants) were strong enough, especially in the case of omeprazole, so that the formation of a metallocomplex cannot be excluded during the explanation of their side effects.

Comparison of Chemical Composition and Biological Activities of Eight Selaginella Species.

Selaginella P. Beauv. is a group of vascular plants in the family Selaginellaceae Willk., found worldwide and numbering more than 700 species, with some used as foods and medicines. The aim of this paper was to compare methanolic (MeOH) and dichloromethane (DCM) extracts of eight Selaginella species on the basis of their composition and biological activities. Six of these Selaginella species are underinvestigated. Using ultra-high performance liquid chromatography-high-resolution mass spectrometry (UHPLC-HRMS) analysis, we identified a total of 193 compounds among the tested Selaginella species, with flavonoids predominating. MeOH extracts recovered more constituents that were detected, including selaginellins, the occurrence of which is only typical for this plant genus. Of all the tested species, Selaginella apoda contained the highest number of identified selaginellins. The majority of the compounds were identified in S. apoda, the fewest compounds in Selaginella cupressina. All the tested species demonstrated antioxidant activity using oxygen radical absorption capacity (ORAC) assay, which showed that MeOH extracts had higher antioxidant capacity, with the half maximal effective concentration (EC50) ranging from 12 ± 1 (Selaginella myosuroides) to 124 ± 2 (Selaginella cupressina) mg/L. The antioxidant capacity was presumed to be correlated with the content of flavonoids, (neo)lignans, and selaginellins. Inhibition of acetylcholinesterase (AChE) was mostly discerned in DCM extracts and was only exhibited in S. myosuroides, S. cupressina, Selaginella biformis, and S. apoda extracts with the half maximal inhibitory concentration (IC50) in the range of 19 ± 3 to 62 ± 1 mg/L. Substantial cytotoxicity against cancer cell lines was demonstrated by the MeOH extract of S. apoda, where the ratio of the IC50 HEK (human embryonic kidney) to IC50 HepG2 (hepatocellular carcinoma) was 7.9 ± 0.2. MeOH extracts inhibited the production of nitrate oxide and cytokines in a dose-dependent manner. Notably, S. biformis halved the production of NO, tumor necrosis factor (TNF)-α, and interleukin (IL)-6 at the following concentrations: 105 ± 9, 11 ± 1, and 10 ± 1 mg/L, respectively. Our data confirmed that extracts from Selaginella species exhibited cytotoxicity against cancer cell lines and AChE inhibition. The activity observed in S. apoda was the most promising and is worth further exploration.

New multimodal stationary phases prepared by Ugi multicomponent approach.

Eight different stationary phases based on two aminopropyl silicas of different brands suitable for multimodal chromatography applications have been prepared by a four-component Ugi reaction. The intention was to synthesize stationary phases significantly differing in their properties hereby demonstrating flexibility of the Ugi synthetic protocol. Diverse functional groups including a nonpolar long aliphatic chain, phenyl moiety, cholic acid scaffold, phenylboronic and monosaccharide units, charged betaine, and arginine moieties were immobilized on a silica surface. The novel sorbents were extensively characterized by elemental analysis, Raman spectroscopy, and chromatography. Considering the anchored chemical structures covalently bonded to the silica surface, reversed-phase, hydrophilic, and ion-exchange separation modes were expected. The chromatographic evaluation was performed directed to map the potential of the individual columns specifically in the mentioned chromatographic modes. The Ugi synthetic protocol has proven to be a simple, feasible, and versatile tool for the synthesis of sorbents of variable properties. The newly prepared stationary phases differed considerably in hydrophobicity and ion-exchange ability. A significant influence of the supporting aminopropyl silica on the final chromatographic behavior was observed. Finally, one practical example confirming applicability of the newly prepared sorbents was demonstrated in separation of cytarabine.

Combination of UV and MS/MS detection for the LC analysis of cannabidiol-rich products.

Cannabidiol (CBD), a major non-psychoactive cannabinoid, has received a lot of attention due to its potential anti-inflammatory, pain-relieving, and anti-anxiety properties. This has led to a recent boom in CBD-rich commercial products, which are sold without prescription in the form of oils, candies, and cosmetics. Since these products are derived from cannabis, the presence of the psychoactive tetrahydrocannabinol (THC) has to be tested before they enter the market. Here, we present a high-throughput approach based on liquid chromatography coupled to UV and tandem mass spectrometric detection for the determination of CBD, THC, and six other minor cannabinoids (cannabigerolic acid, cannabidivarin, cannabinol, cannabigerol, cannabidiolic acid, and tetrahydrocannabinolic acid) in a wide range of concentrations and in a variety of matrices, including oils, hydrophobic ointments, water-soluble liquids, plant material and gelatinous gummies. Each product was dissolved in a suitable solvent and further diluted to avoid matrix interference. The diluted samples were analyzed by reversed-phase chromatography, coupled to a UV detector followed by a triple quadrupole mass spectrometer, used in the multiple reaction monitoring mode. The UV signal was utilized for the quantification of samples containing high levels of CBD, while the mass spectrometer was used for low levels of THC and other minor cannabinoids. This allowed us to meet the required sensitivity for THC while significantly expanding the range of analyzed CBD, all within an 8-min long chromatographic run. The samples were further quantified using calibration in solvent, the approach was validated, and the validation criteria were met for all matrices except for two (i.e., emulsions and gels). The lower limit of quantification for THC was 0.5 µg/g in gummies, 1.0 µg/g in oils, ointments and liquids, and 5.0 µg/g in plant material. CBD was analyzed in the range of 0.5-60,000 µg/g in gummies, 1-120,000 µg/g in oils, ointments and liquids, and 5-300,000 µg/g in plant material. The developed method was used for the analysis of thirteen real products with a wide range of CBD content, with positive THC findings in twelve of them.

Role of mtDNA disturbances in the pathogenesis of Alzheimer's and Parkinson's disease.

Neurodegenerative diseases (e.g. Alzheimer's and Parkinson's disease) are becoming increasingly problematic to healthcare systems. Therefore, their underlying mechanisms are trending topics of study in medicinal research. Numerous studies have evidenced a strong association between mitochondrial DNA disturbances (e.g. oxidative damage, mutations, and methylation shifts) and the initiation and progression of neurodegenerative diseases. Therefore, this review discusses the risk and development of neurodegenerative diseases in terms of disturbances in mitochondrial DNA and as a part of a complex ecosystem that includes other important mechanisms (e.g. neuroinflammation and the misfolding and aggregation of amyloid-ß peptides, α-synuclein, and tau proteins). In addition, the influence of individual mitochondrial DNA haplogroups on the risk and development of neurodegenerative diseases is also described and discussed.

Mass spectrometry imaging of free-floating brain sections detects pathological lipid distribution in a mouse model of Alzheimer's-like pathology.

Mass spectrometry imaging (MSI) is a modern analytical technique capable of monitoring the spatial distribution of compounds within target tissues. Collection and storage are important steps in sample preparation. The recommended and most widely used preservation procedure for MSI is freezing samples in isopentane and storing them at temperatures below -80 °C. On the other hand, the most common and general method for preserving biological samples in clinical practice is fixation in paraformaldehyde. Special types of samples prepared from these fixed tissues that are used for histology and immunohistochemistry are free-floating sections. It would be very beneficial if the latter procedure could also be applicable for the samples intended for subsequent MSI analysis. In the present work, we optimized and evaluated paraformaldehyde-fixed free-floating sections for the analysis of lipids in mouse brains and used the sections for the study of lipid changes in double transgenic APP/PS1 mice, a model of Alzheimer's-like pathology. Moreover, we examined the neuroprotective properties of palm11-PrRP31, an anorexigenic and glucose-lowering analog of prolactin-releasing peptide, and liraglutide, a type 2 diabetes drug. From the free-floating sections, we obtained lipid images without interference or delocalization, and we demonstrated that free-floating sections can be used for the MSI of lipids. In the APP/PS1 mice, we observed a changed distribution of various lipids compared to the controls. The most significant changes in lipids in the brains of APP/PS1 mice compared to wild-type controls were related to gangliosides (GM2 36:1, GM3 36:1) and phosphatidylinositols (PI 38:4, 36:4) in regions where the accumulation of senile plaques occurred. In APP/PS1 mice peripherally treated with palm11-PrRP31 or liraglutide for 2 months, we found that both peptides reduced the amount and space occupied by lipids, which were linked to the senile plaques. These results indicate that palm11-PrRP31 as well as liraglutide might be potentially useful in the treatment of neurodegenerative diseases.

Formaldehyde Reacts with Amino Acids and Peptides with a Potential Role in Acute Methanol Intoxication.

Methanol, an aliphatic alcohol widely used in the industry, causes acute and chronic intoxications associated with severe long-term health damage, including permanent visual impairment, brain damage, mainly necrosis of the basal ganglia and high mortality due to cancer. However, the role of formaldehyde, an intermediate metabolite of methanol oxidation, in methanol toxicity remains unclear. Thus, we studied the reactivity of several amino acids and peptides in the presence of formaldehyde by identifying products by direct infusion electrospray high-resolution mass spectrometry (MS) and matrix-assisted laser desorption-ionization MS. Cysteine, homocysteine and two peptides, CG and CGAG, provided cyclic products with a +12 amu mass shift with respect to the original compounds. The proposed structures of the products were confirmed by high-resolution tandem MS. Moreover, the formation of the products with +12 amu mass shift was also shown for two biologically relevant peptides, fragments of ipilimumab, which is a human IgG1 monoclonal antibody against cytotoxic T-lymphocyte-associated protein 4. Overall, our experimental results indicate that formaldehyde reacts with some amino acids and peptides, yielding covalently modified structures. Such chemical modifications may induce undesirable changes in the properties and function of vital biomolecules (e.g., hormones, enzymes) and consequently pathogenesis.

Synthesis and identification of deschloroketamine metabolites in rats' urine and a quantification method for deschloroketamine and metabolites in rats' serum and brain tissue using liquid chromatography tandem mass spectrometry.

Deschloroketamine (2-(methylamino)-2-phenyl-cyclohexanone) is a ketamine analog belonging to a group of dissociative anesthetics, which have been distributed within the illicit market since 2015. However, it was also being sold as 'ketamine' misleading people to believe that they were getting genuine ketamine. Dissociative anesthetics have also come to the attention of the psychiatric field due to their potential properties in the treatment of depression. At present, there is a dearth of information on deschloroketamine related to its metabolism, biodistribution, and its mechanism of action. We have therefore carried out a metabolomics study for deschloroketamine via non-targeted screening of urine samples employing liquid chromatography combined with high-resolution mass spectrometry. We developed and validated a multiple reaction monitoring method using a triple quadrupole instrument to track metabolites of deschloroketamine. Furthermore, significant metabolites of deschloroketamine, (trans-dihydrodeschloroketamine, cis- and trans-dihydronordeschloroketamine, and nordeschloroketamine), were synthesized in-house. The prepared standards were utilized in the developed multiple reaction monitoring method. The quantification method for serum samples provided intra-day accuracy ranging from 86% to 112% with precision of 3% on average. The concentrations of cis/trans-dihydronordeschloroketamines and trans-dihydrodeschloroketamine were lower than 10 ng/mL, nordeschloroketamine and deschloroketamine ranged from 0.5 to 860 ng/mL in real samples. The quantification method for brain tissue provided intra-day accuracy ranging from 80% to 125% with precision of 7% on average. The concentrations of cis/trans-dihydronordeschloroketamines and trans-dihydrodeschloroketamine ranged from 0.5 to 70 ng/g, nordeschloroketamine and deschloroketamine varied from 0.5 to 4700 ng/g in real samples.

Strategy for improved therapeutic efficiency of curcumin in the treatment of gastric cancer.

Gastric cancer is a common oncological disease. Although enormous efforts have been expended, possible therapeutic modalities are still limited. For this reason, new therapeutic approaches and agents are highly requested and intensively developed. One strategy is the application of natural agents, such as curcumin, with proven anticancer effects and low toxicity for patients. Therefore, this review discusses the potential application of curcumin in the therapy of gastric cancer and its potential incorporation in therapeutic regimens. Because one of the largest impediments for widespread curcumin application is its limited bioavailability (caused mainly by its very low water solubility), studied strategies (drug delivery systems and curcumin derivatization) aimed to solve this obstacle are discussed in more detail.

The use of 1,5-diaminonaphthalene for matrix-assisted laser desorption/ionization mass spectrometry imaging of brain in neurodegenerative disorders.

The selection of a suitable matrix and deposition technique constitutes a critical step in successful matrix-assisted laser desorption/ionization mass spectrometry imaging measurement. In the present work, we compared three techniques of matrix deposition, specifically, sublimation and spraying of 1,5-diaminonaphthalene with two automatic sprayers, ImagePrep and iMatrixSpray. The studied methods were evaluated in experiments for the analysis of lipid composition in the brains of two mouse models of neurodegeneration: APP/PS1 mice with plaques of amyloid ß (Aß) peptides and THY-Tau22 mice with pathologically hyperphosphorylated Tau protein, two hallmarks of Alzheimer's disease-like pathology. The sublimation method provided irreproducible results because of significant matrix loss due to the high vacuum in the ion source and laser irradiation. In contrast, the ImagePrep and iMatrixSpray provided stable film of the matrix. The deposited matrix was stable during the measurement, and highly reproducible datasets were obtained. Both spraying methods yielded similar results with approximately the same number of detected lipids and comparable signal intensity. However, iMatrixSpray has two main advantages: a faster matrix deposition and the formation of smaller matrix crystals leading to better spatial resolution. In the APP/PS1 mouse model at an age of 6 months, we found colocalization of Aß plaques with different phospholipids, sphingolipids and lysophospholipids. We did not find a difference in lipid composition between the THY-Tau22 mice and the wild-type controls. The results indicate that hyperphosphorylation of tau protein in the THY-Tau22 mouse model at the age of 6 months is not accompanied with a significant change in lipid content in the brain. However, considering limitations of the used method, a definitive conclusion in this respect will need further research.

Nuclear transport of nicotinamide phosphoribosyltransferase is cell cycle-dependent in mammalian cells, and its inhibition slows cell growth.

Nicotinamide phosphoribosyltransferase (NAMPT) is located in both the nucleus and cytoplasm and has multiple biological functions including catalyzing the rate-limiting step in NAD synthesis. Moreover, up-regulated NAMPT expression has been observed in many cancers. However, the determinants and regulation of NAMPT's nuclear transport are not known. Here, we constructed a GFP-NAMPT fusion protein to study NAMPT's subcellular trafficking. We observed that in unsynchronized 3T3-L1 preadipocytes, 25% of cells had higher GFP-NAMPT fluorescence in the cytoplasm, and 62% had higher GFP-NAMPT fluorescence in the nucleus. In HepG2 hepatocytes, 6% of cells had higher GFP-NAMPT fluorescence in the cytoplasm, and 84% had higher GFP-NAMPT fluorescence in the nucleus. In both 3T3-L1 and HepG2 cells, GFP-NAMPT was excluded from the nucleus immediately after mitosis and migrated back into it as the cell cycle progressed. In HepG2 cells, endogenous, untagged NAMPT displayed similar changes with the cell cycle, and in nonmitotic cells, GFP-NAMPT accumulated in the nucleus. Similarly, genotoxic, oxidative, or dicarbonyl stress also caused nuclear NAMPT localization. These interventions also increased poly(ADP-ribosyl) polymerase and sirtuin activity, suggesting an increased cellular demand for NAD. We identified a nuclear localization signal in NAMPT and amino acid substitution in this sequence (424RSKK to ASGA), which did not affect its enzymatic activity, blocked nuclear NAMPT transport, slowed cell growth, and increased histone H3 acetylation. These results suggest that NAMPT is transported into the nucleus where it presumably increases NAD synthesis required for cell proliferation. We conclude that specific inhibition of NAMPT transport into the nucleus might be a potential avenue for managing cancer.