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The Korean Journal of Physiology &... Jul 2022Severe bacterial infections are frequently accompanied by depressed neutrophil functions. Thus, agents that increase the microbicidal activity of neutrophils could add...
Severe bacterial infections are frequently accompanied by depressed neutrophil functions. Thus, agents that increase the microbicidal activity of neutrophils could add to a direct antimicrobial therapy. Lysophosphatidylcholine augments neutrophil bactericidal activity the glycine (Gly)/glycine receptor (GlyR) α2/TRPM2/p38 mitogen-activated protein kinase (MAPK) pathway. However, the direct effect of glycine on neutrophil bactericidal activity was not reported. In this study, the effect of glycine on neutrophil bactericidal activity was examined. Glycine augmented bactericidal activity of human neutrophils (EC = 238 μM) in a strychnine (a GlyR antagonist)-sensitive manner. Glycine augmented bacterial clearance in mice, which was also blocked by strychnine (0.4 mg/kg, s.c.). Glycine enhanced NADPH oxidase-mediated reactive oxygen species (ROS) production and TRPM2-mediated [Ca] increase in neutrophils that had taken up E. coli . Glycine augmented Lucifer yellow uptake (fluid-phase pinocytosis) and azurophil granule-phagosome fusion in neutrophils that had taken up E. coli in an SB203580 (a p38 MAPK inhibitor)-sensitive manner. These findings indicate that glycine augments neutrophil microbicidal activity by enhancing azurophil granule-phagosome fusion the GlyRα2/ROS/calcium/p38 MAPK pathway. We suggest that glycine could be a useful agent for increasing neutrophil bacterial clearance.
PubMed: 35766001
DOI: 10.4196/kjpp.2022.26.4.229 -
Frontiers in Cell and Developmental... 2022MFSD8 is a transmembrane protein that has been reported to transport chloride ions across the lysosomal membrane. Mutations in are associated with a subtype of Batten...
MFSD8 is a transmembrane protein that has been reported to transport chloride ions across the lysosomal membrane. Mutations in are associated with a subtype of Batten disease called CLN7 disease. Batten disease encompasses a family of 13 inherited neurodegenerative lysosomal storage diseases collectively referred to as the neuronal ceroid lipofuscinoses (NCLs). Previous work identified an ortholog of human MFSD8 in the social amoeba (gene: , protein: Mfsd8) reported its localization to endocytic compartments, and demonstrated its involvement in protein secretion. In this study, we further characterized the effects of loss during growth and early stages of multicellular development. During growth, cells displayed increased rates of proliferation, pinocytosis, and expansion on bacterial lawns. Loss of also increased cell size, inhibited cytokinesis, affected the intracellular and extracellular levels of the quorum-sensing protein autocrine proliferation repressor A, and altered lysosomal enzyme activity. During the early stages of development, loss of delayed aggregation, which we determined was at least partly due to impaired cell-substrate adhesion, defects in protein secretion, and alterations in lysosomal enzyme activity. Overall, these results show that Mfsd8 plays an important role in modulating a variety of processes during the growth and early development of .
PubMed: 35756993
DOI: 10.3389/fcell.2022.930235 -
Frontiers in Nutrition 2022The Mediterranean diet is regarded as one of the most healthful dietary patterns in the world, owing to a combination of foods high in antioxidants and anticancer...
The Mediterranean diet is regarded as one of the most healthful dietary patterns in the world, owing to a combination of foods high in antioxidants and anticancer constituents. Barley bran is one of the components of the Mediterranean diet. It has nutritional and beneficial effects in different pathological conditions. Many studies were achieved to assess the nutritious values of barley bran, but there is no research indicating immunomodulatory and anticancer activities of barley bran grown in Jordan. The present study aims to examine and assess the potential immunomodulatory and anti-tumor activities of ethanol, -hexane, aqueous/methanol, and water extracts obtained from barley bran. The Maceration method was utilized to prepare ethanol, -hexane, aqueous/methanol, and water extracts. Various phytochemical groups were determined by using qualitative phytochemical tests. The antiproliferative activity of extracts was determined against MCF-7, HCT-116, A549, and EMT6/p by the MTT assay. The Folin-Ciocalteu reagent was used to detect the total phenolic content in extracts. Furthermore, immunomodulatory activity was assessed by determining the effect of extracts on splenocytes proliferation in the presence and absence of mitogens. The nitro blue tetrazolium assay and the neutral red method were used to assess the effect of each extract on the phagocytic activity of macrophages and pinocytosis, respectively. For the part, three different concentrations (10, 20, and 30% w/v) of barley bran were used to test the prophylactic effect in four Balb/C mice groups inoculated with EMT6/p cell-line subcutaneously. Also, serum samples were collected to assess the effect on cytokines (IFN-gamma, IL-2, IL-4, and IL-10). Barley bran extracts inhibited cancer cell proliferation. According to immunoassays, hexane and aqueous/methanol extracts could significantly rise lymphocyte proliferation and pinocytosis activity of macrophages. The activity of phagocytosis was increased by hexane and ethanol extracts. For the part, the average tumor size and weight of mice given the 30% barley bran group was significantly reduced ( < 0.05) compared with the control group. During our study, higher levels of TH1 cytokines (IFN- γ, IL-2) and lower levels of TH2 cytokine (IL-4) and T regulatory cytokine (IL-10) were obtained due to consumption of barley bran in food. Barley bran can be used as a prophylactic agent because it has anti-cancer and immunomodulatory activities.
PubMed: 35662936
DOI: 10.3389/fnut.2022.838373 -
PLoS Pathogens May 2022Entamoeba histolytica is a protozoan responsible for several pathologies in humans. Trophozoites breach the intestinal site to enter the bloodstream and thus traverse to...
Entamoeba histolytica is a protozoan responsible for several pathologies in humans. Trophozoites breach the intestinal site to enter the bloodstream and thus traverse to a secondary site. Macropinocytosis and phagocytosis, collectively accounting for heterophagy, are the two major processes responsible for sustenance of Entamoeba histolytica within the host. Both of these processes require significant rearrangements in the structure to entrap the target. Rho GTPases play an indispensable role in mustering proteins that regulate cytoskeletal remodelling. Unlike phagocytosis which has been studied in extensive detail, information on machinery of macropinocytosis in E. histolytica is still limited. In the current study, using site directed mutagenesis and RNAi based silencing, coupled with functional studies, we have demonstrated the involvement of EhRho5 in constitutive and LPA stimulated macropinocytosis. We also report that LPA, a bioactive phospholipid present in the bloodstream of the host, activates EhRho5 and translocates it from cytosol to plasma membrane and endomembrane compartments. Using biochemical and FRAP studies, we established that a PI Kinase acts upstream of EhRho5 in LPA mediated signalling. We further identified EhGEF2 as a guanine nucleotide exchange factor of EhRho5. In the amoebic trophozoites, EhGEF2 depletion leads to reduced macropinocytic efficiency of trophozoites, thus phenocopying its substrate. Upon LPA stimulation, EhGEF2 is found to sequester near the plasma membrane in a wortmannin sensitive fashion, explaining a possible mode for activation of EhRho5 in the amoebic trophozoites. Collectively, we propose that LPA stimulated macropinocytosis in E. histolytica is driven by the PI Kinase-EhGEF2-EhRho5 axis.
Topics: Animals; Entamoeba histolytica; Humans; Lipopolysaccharides; Phagocytosis; Pinocytosis; Protozoan Proteins; Trophozoites; rho GTP-Binding Proteins
PubMed: 35594320
DOI: 10.1371/journal.ppat.1010550 -
Proceedings of the National Academy of... May 2022Mammalian target of rapamycin complex 1 (mTORC1) senses amino acids to control cell growth, metabolism, and autophagy. Some amino acids signal to mTORC1 through the Rag...
Mammalian target of rapamycin complex 1 (mTORC1) senses amino acids to control cell growth, metabolism, and autophagy. Some amino acids signal to mTORC1 through the Rag GTPase, whereas glutamine and asparagine activate mTORC1 through a Rag GTPase-independent pathway. Here, we show that the lysosomal glutamine and asparagine transporter SNAT7 activates mTORC1 after extracellular protein, such as albumin, is macropinocytosed. The N terminus of SNAT7 forms nutrient-sensitive interaction with mTORC1 and regulates mTORC1 activation independently of the Rag GTPases. Depletion of SNAT7 inhibits albumin-induced mTORC1 lysosomal localization and subsequent activation. Moreover, SNAT7 is essential to sustain KRAS-driven pancreatic cancer cell growth through mTORC1. Thus, SNAT7 links glutamine and asparagine signaling from extracellular protein to mTORC1 independently of the Rag GTPases and is required for macropinocytosis-mediated mTORC1 activation and pancreatic cancer cell growth.
Topics: Amino Acid Transport Systems, Neutral; Asparagine; Glutamine; Humans; Lysosomes; Mechanistic Target of Rapamycin Complex 1; Pancreatic Neoplasms; Pinocytosis; Signal Transduction
PubMed: 35561222
DOI: 10.1073/pnas.2123261119 -
Bioconjugate Chemistry May 2022Macropinocytosis is a form of endocytosis that allows massive uptake of extracellular materials and is a promising route for intracellular delivery of biofunctional...
Macropinocytosis is a form of endocytosis that allows massive uptake of extracellular materials and is a promising route for intracellular delivery of biofunctional macromolecules and nanoparticles. Our laboratory developed a potent macropinocytosis-inducing peptide named P4A. However, the ability of this peptide is not apparent in the presence of serum. This study aims to endow P4A and related peptides with the ability to induce macropinocytosis in the presence of serum by N-terminal acylation with long-chain fatty acids (i.e., decanoic, myristic, and stearic acids). Stearylated P4A (stearyl-P4A) had the highest effect on stimulating macropinocytotic uptake. Moreover, the intramolecularly disulfide-bridged analogue, stearyl-oxP4A, showed an even higher ability. The effect of stearyl-oxP4A to facilitate the intracellular delivery of small extracellular vesicles (sEVs) was evaluated in terms of (i) cellular uptake using sEVs labeled with an enhanced green fluorescent protein (EGFP) and (ii) cytosolic liberation and expression of sEV-encapsulated luciferase mRNA in recipient cells. The two- to threefold uptake of both sEVs in the presence of stearyl-oxP4A suggests the potential of the peptide for sEV delivery in the presence of serum.
Topics: Biological Transport; Endocytosis; Extracellular Vesicles; Peptides; Pinocytosis
PubMed: 35506582
DOI: 10.1021/acs.bioconjchem.2c00113 -
Frontiers in Nutrition 2022The Mediterranean diet includes the consumption of various fruits and vegetables. Lemon and ginger are highly popular in Mediterranean cuisine. The current study aims to...
The Mediterranean diet includes the consumption of various fruits and vegetables. Lemon and ginger are highly popular in Mediterranean cuisine. The current study aims to evaluate both anticancer and immunomodulatory activities of lemon and ginger combination. The antiproliferative activities of the combination were tested against different cancer cell lines using 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay. The degree of apoptosis induction and vascular endothelial growth factor expression were detected using ELISA. Balb/C mice were inoculated with the EMT6/P breast cancer cells and received combination water extract orally for 14 days. The effect of the water extract on splenocytes proliferation was measured using the mitogen proliferation assay. Macrophage function was evaluated using the nitro blue tetrazolium assay and pinocytosis was assessed using the neutral red method. Gas chromatography coupled to the tandem mass spectrometry was used to determine the composition of the combination. The lemon and ginger combination showed significant apoptosis induction and angiogenesis suppression effects. Fifty percent of the mice taking this combination did not develop tumors with a percentage of tumor reduction of 32.8%. This combination showed a potent effect in stimulating pinocytosis. Alpha-pinene and α-terpineol were detected in high percentages in the combination water extract. The lemon and ginger combination represents promising options to develop anticancer infusions for augmenting conventional anticancer therapies. Further testing is required to understand the exact molecular mechanisms of this combination.
PubMed: 35495945
DOI: 10.3389/fnut.2022.829101 -
Experimental and Therapeutic Medicine Jun 2022Water-soluble polysaccharide isolated from soybean hull and fractionated using ion-exchange chromatography were investigated to determine their molecular characteristics...
Water-soluble polysaccharide isolated from soybean hull and fractionated using ion-exchange chromatography were investigated to determine their molecular characteristics and immunostimulating activity. In the present study, soybean hull polysaccharide (SHP) was separated and purified to obtain three main fractions (F1, F2 and F3), and their chemical and monosaccharide compositions were analyzed. SHP was mainly composed of carbohydrates (64.3%), proteins (16.2%) and sulfates (12.5%), with minor levels of uronic acid (3.2%), and predominantly contained glucose and mannose as monosaccharides. Moreover, when compared with cells treated with RPMI medium, SHP was revealed to promote the proliferation and pinocytosis of RAW264.7 cells, and to enhance the production of nitric oxide (NO), tumor necrosis factor (TNF)-α, interleukin (IL)-1β and IL-6. Furthermore, flow cytometry demonstrated that CD11b and CD40 were involved in the immune regulation of RAW264.7 cells by SHP. Moreover, western blotting and other experiments revealed that SHP, a type of pathogen-associated molecular pattern, was specifically recognized by the Toll-like receptor 2, which, in turn, upregulated the expression levels of proteins downstream of the mitogen-activated protein kinase and nuclear factor κB pathways. Notably, the immune activity of the F2 fraction was markedly higher than that of the crude polysaccharides. In summary, the purified F2 fraction of SHP may be an effective nutritional supplement for human disorders associated with low immunity.
PubMed: 35495602
DOI: 10.3892/etm.2022.11316 -
Bioengineered Apr 2022Ovulation-inducing drugs such as endogenous steroids could reduce endometrial receptivity during the implantation window, resulting in lower clinical pregnancy rates and...
Electroacupuncture improves endometrial receptivity through miRNA-223-3p-mediated regulation of leukemia inhibitory factor / signal transducer and activator of transcription 3 signaling pathway.
Ovulation-inducing drugs such as endogenous steroids could reduce endometrial receptivity during the implantation window, resulting in lower clinical pregnancy rates and higher miscarriage rates. The present study employed electroacupuncture therapy along with different frequencies on elevating impaired endometrial receptivity to elucidate the mechanism therein. The rats were split up into seven groups of normal, model, low-frequency electroacupuncture (LF-EA), high-frequency electroacupuncture (HF-EA), LF-EA+anti-miRNA, HF-EA+anti-miRNA, and anti-miRNA. PCR assays were used to detect miR-223-3p expressions. The effects of electroacupuncture and miR-223-3p on rats' endometrial membrane cell-drinking process in a manner of scanning electron microscopy were recorded. After that we observed on the electroacupuncture effects on the conditions of adhesion molecules and LIF/STAT3 signaling pathway with assays of immunofluorescence and Western Blot. This study was end up with dual luciferase assay, where combination of miR-233-3p onto the 3'-UTR sequence of LIF was determined. PCR assay demonstrated that HF-EA procured an inhibition in miR-223-3p expression, whereas scanning electron microscopy turned out that both electroacupuncture and miR-223-3p were capable of raising the amount of intrauterine pinocytosis and the number of blastocyst implantation in rats. Additionally, assays of Western Blot and immunofluorescence showed that therapy of electroacupuncture brought about decreasing expressions in adhesion molecules of E-cadherin, β-catenin and claudin-1 (CLDN1). We found that both electroacupuncture and miR-223-3p were able to fortify LIF/STAT3 signaling pathway, then the fact of miR-223-3p combination to LIF 3'-UTR sequence was validated via our dual-luciferase assay. Electroacupuncture therapy inhibited the miR-223-3p expression upon LIF/STAT3 signaling pathway to elevate endometrial receptivity.
Topics: Animals; Electroacupuncture; Female; Leukemia Inhibitory Factor; MicroRNAs; Pregnancy; Rats; STAT3 Transcription Factor; Signal Transduction
PubMed: 35435116
DOI: 10.1080/21655979.2022.2062524 -
Scientific Reports Apr 2022Macropinocytosis is a type of endocytosis accompanied by actin rearrangement-driven membrane deformation, such as lamellipodia formation and membrane ruffling, followed...
Macropinocytosis is a type of endocytosis accompanied by actin rearrangement-driven membrane deformation, such as lamellipodia formation and membrane ruffling, followed by the formation of large vesicles, macropinosomes. Ras-transformed cancer cells efficiently acquire exogenous amino acids for their survival through macropinocytosis. Thus, inhibition of macropinocytosis is a promising strategy for cancer therapy. To date, few specific agents that inhibit macropinocytosis have been developed. Here, focusing on the mechanosensitive ion channel Piezo1, we found that Yoda1, a Piezo1 agonist, potently inhibits macropinocytosis induced by epidermal growth factor (EGF). The inhibition of ruffle formation by Yoda1 was dependent on the extracellular Ca influx through Piezo1 and on the activation of the calcium-activated potassium channel KCa3.1. This suggests that Ca ions can regulate EGF-stimulated macropinocytosis. We propose the potential for macropinocytosis inhibition through the regulation of a mechanosensitive channel activity using chemical tools.
Topics: Biological Transport; Calcium; Carcinoma, Squamous Cell; Cell Line, Tumor; Epidermal Growth Factor; Humans; Ion Channels; Pinocytosis; Pyrazines; Thiadiazoles
PubMed: 35428847
DOI: 10.1038/s41598-022-10153-8