Immunoliposomes That Target Endothelium In Vitro Are Dependent on Lipid Raft Formation

2010 ◽  
Vol 7 (5) ◽  
pp. 1569-1575 ◽  
Author(s):  
Rico C. Gunawan ◽  
Debra T. Auguste
Keyword(s):  
Lipid Raft

scholarly journals Regulation of Gap Junctions in Porcine Cumulus-Oocyte Complexes: Contributions of Granulosa Cell Contact, Gonadotropins, and Lipid Rafts

2009 ◽  
Vol 23 (5) ◽  
pp. 700-710 ◽  
Author(s):  
Maxime Sasseville ◽  
Marie-Claude Gagnon ◽  
Christine Guillemette ◽  
Robert Sullivan ◽  
Robert B. Gilchrist ◽  
...  
Keyword(s):  
Gap Junctions ◽  
Lipid Rafts ◽  
Lipid Raft ◽  
Connexin 43 ◽  
In Vitro Maturation ◽  
Cell Contact ◽  
Germinal Vesicle Breakdown ◽  
Oocyte Growth ◽  
Junctional Communication

Abstract Gap-junctional communication (GJC) plays a central role in oocyte growth. However, little is known about the regulation of connexin 43 (Cx43)-based gap-junction channels in cumulus-oocyte complexes (COCs) during in vitro maturation. We show that rupture of COCs from mural granulosa cells up-regulates Cx43-mediated GJC and that gonadotropins signal GJC breakdown by recruiting Cx43 to lipid rafts when oocyte meiosis resumes. Oocyte calcein uptake through gap junctions increases during early in vitro oocyte maturation and remains high until 18 h, when it falls simultaneously with the oocyte germinal vesicle breakdown. Immunodetection of Cx43 and fluorescence recovery after photobleaching assays revealed that the increase of GJC is independent of gonadotropins but requires RNA transcription, RNA polyadenylation, and translation. GJC rupture, in contrast, is achieved by a gonadotropin-dependent mechanism involving recruitment of Cx43 to clustered lipid rafts. These results show that GJC up-regulation in COCs in in vitro culture is independent of gonadotropins and transcriptionally regulated. However, GJC breakdown is gonadotropin dependent and mediated by the clustering of Cx43 in lipid raft microdomains. In conclusion, this study supports a functional role of lipid raft clustering of Cx43 in GJC breakdown in the COCs during in vitro maturation.

scholarly journals Rnd1 and Rnd3 targeting to lipid raft is required for p190 RhoGAP activation

2012 ◽  
Vol 23 (8) ◽  
pp. 1593-1604 ◽  
Author(s):  
Izumi Oinuma ◽  
Kana Kawada ◽  
Kiyoka Tsukagoshi ◽  
Manabu Negishi
Keyword(s):  
Lipid Raft ◽  
Cell Types ◽  
Functional Difference ◽  
Distinct Subgroup ◽  
Functional Differences ◽  
P190 Rhogap ◽  
Rho Family ◽  
Rhoa Signaling

The Rnd proteins Rnd1, Rnd2, and Rnd3/RhoE are well known as key regulators of the actin cytoskeleton in various cell types, but they comprise a distinct subgroup of the Rho family in that they are GTP bound and constitutively active. Functional differences of the Rnd proteins in RhoA inhibition signaling have been reported in various cell types. Rnd1 and Rnd3 antagonize RhoA signaling by activating p190 RhoGAP, whereas Rnd2 does not. However, all the members of the Rnd family have been reported to bind directly to p190 RhoGAP and equally induce activation of p190 RhoGAP in vitro, and there is no evidence that accounts for the functional difference of the Rnd proteins in RhoA inhibition signaling. Here we report the role of the N-terminal region in signaling. Rnd1 and Rnd3, but not Rnd2, have a KERRA (Lys-Glu-Arg-Arg-Ala) sequence of amino acids in their N-terminus, which functions as the lipid raft-targeting determinant. The sequence mediates the lipid raft targeting of p190 RhoGAP correlated with its activation. Overall, our results demonstrate a novel regulatory mechanism by which differential membrane targeting governs activities of Rnd proteins to function as RhoA antagonists.

scholarly journals PKC-α Triggers EGFR Ubiquitination, Endocytosis and ERK Activation in Podocytes Stimulated with High Glucose

2017 ◽  
Vol 42 (1) ◽  
pp. 281-294 ◽  
Author(s):  
Chun-Tao Lei ◽  
Yan-Hong Wei ◽  
Hui Tang ◽  
Qian Wen ◽  
Chen Ye ◽  
...  
Keyword(s):  
Lipid Raft ◽  
High Glucose ◽  
Growth Factor Receptor ◽  
In Vitro Study ◽  
Diabetic Rats ◽  
Kidney Cortex ◽  
Podocyte Injury ◽  
Erk Activation ◽  
Signaling Activation

Background: Protein Kinase C-α (PKC-α) and epidermal growth factor receptor (EGFR) are both involved in diabetic kidney disease; however, the connection between these two proteins during high glucose-induced podocyte injury remains uncertain. Methods: Diabetes was induced in SD rats by streptozotocin (STZ). Fourteen days later, the kidney cortex was removed and subjected to plasma membrane isolation and lipid raft fractionation. In vitro study human podocyte cell line was differentiated and subjected to various treatments. The levels of membranous protein and endocytosis were assessed by biotinylation and sodium 2-mercaptoethane sulfonate (MesNa) treatment. Gö6976 and PYR-41 were used as inhibitors of PKC-α and ubiquitin activating E1 enzyme, respectively. Results: In diabetic rats, the abundance of PKC-α in the membranous fraction and the lipid raft domain is elevated, whereas the EGFR level is reduced. Consistently, in vitro high glucose treated podocytes, membranous EGFR is downregulated with increased PKC-α. Furthermore, the ubiquitination and endocytosis of EGFR are enhanced accompanied by extracellular signal–regulated kinase (ERK) signaling activation and podocyte damage during hyperglycemia. However, these processes can be ameliorated by inhibition of either PKC-α or ubiquitin activating E1 enzyme. Conclusion: During hyperglycemia, PKC-α mediates podocytic EGFR ubiquitination, endocytosis from cell surface and the subsequent ERK activation, which contributes to podocyte injury.

Iron-mediated interaction of alpha synuclein with lipid raft model membranes

Nanoscale ◽  
2020 ◽  
Vol 12 (14) ◽  
pp. 7631-7640 ◽  
Author(s):  
Fabio Perissinotto ◽  
Chiaramaria Stani ◽  
Elena De Cecco ◽  
Lisa Vaccari ◽  
Valeria Rondelli ◽  
...  
Keyword(s):  
Cell Membrane ◽  
Lipid Raft ◽  
Alpha Synuclein ◽  
Model Membranes ◽  
Membrane Systems ◽  
Pathological Conditions ◽  
Model Cell ◽  
Raft Domains ◽  
Model Cell Membrane

We demonstrated that pathological conditions as accumulation of iron cations promote fast formation of α-synuclein aggregation in vitro, which preferentially interact with lipid-raft domains in model cell membrane systems.

scholarly journals Identification of in vitro HSC fate regulators by differential lipid raft clustering

Cell Cycle ◽  
2012 ◽  
Vol 11 (8) ◽  
pp. 1535-1543 ◽  
Author(s):  
Nicola Vannini ◽  
Aline Roch ◽  
Olaia Naveiras ◽  
Alessandra Griffa ◽  
Stefan Kobel ◽  
...  
Keyword(s):  
Lipid Raft ◽  
Raft Clustering

scholarly journals Nystatin Modulates Genes in Immunity and Wingless Signaling Pathways in Cow Blood

2016 ◽  
Vol 7 (1) ◽  
pp. 1 ◽  
Author(s):  
Emmanuel K Asiamah ◽  
Sarah Adjei-Fremah ◽  
Kingsley Ekwemalor ◽  
Mulumebet Worku
Keyword(s):  
Signaling Pathway ◽  
Lipid Rafts ◽  
Adaptive Immunity ◽  
Lipid Raft ◽  
Wnt Signaling Pathway ◽  
Transcript Abundance ◽  
Innate And Adaptive Immunity ◽  
Total Plasma Protein ◽  
Wingless Signaling

Nystatin is an antifungal agent isolated from bacteria found in the dairy cow environment. It disrupts small platforms in the cell membrane, composed of sphingolipids and cholesterol known as lipid rafts. Pathogen recognition receptors (PRR) may be embedded in these lipid rafts. This study was conducted to evaluate the in vitro effects of the lipid raft inhibitor Nystatin, on the expression of genes in the innate and adaptive immunity and wnt signaling pathway in cow peripheral blood. Blood collected from four adult female Holstein-Friesian cows (n=4) was treated with 100ng/mL of Nystatin in vitro. Samples treated with Phosphate Buffer Saline served as control. Total protein concentration and prostaglandin E2in plasma were determined. Total RNA was isolated from cells and was used for cDNA synthesis. The effect of Nystatin on the expression of 84 genes on the cow Wingless signaling pathway and human innate and adaptive immunity arrays were assessed in cow blood using real-time PCR. Fold change in transcript abundance was calculated using Livak’s method. Nystatin was found to modulate transcription and translation of genes involved in homeostasis and immunity in cow blood. It also increased the concentration of total plasma protein and PGE2in cow blood and may thus have had a pro-inflammatory effect. This study provides evidence for the association between lipid raft inhibition and alterations in the wingless signaling pathway in ruminant blood. Furthermore, the results presented may inform antifungal drug design and use in cows.

scholarly journals The Antipsychotic Risperidone Alters Dihydroceramide and Ceramide Composition and Plasma Membrane Function in Leukocytes In Vitro and In Vivo

2021 ◽  
Vol 22 (8) ◽  
pp. 3919
Author(s):  
Alberto Canfrán-Duque ◽  
Óscar Pastor ◽  
David García-Seisdedos ◽  
Yessenia L. Molina ◽  
Bohdan Babiy ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Lipid Raft ◽  
Older Patients ◽  
In Vitro Assays ◽  
Fatty Acid Elongase ◽  
Membrane Function ◽  
Increased Risk ◽  
Raft Domains ◽  
Long Chain

Atypical or second-generation antipsychotics are used in the treatment of psychosis and behavioral problems in older persons with dementia. However, these pharmaceutical drugs are associated with an increased risk of stroke in such patients. In this study, we evaluated the effects of risperidone treatment on phospholipid and sphingolipid composition and lipid raft function in peripheral blood mononuclear cells (PBMCs) of older patients (mean age >88 years). The results showed that the levels of dihydroceramides, very-long-chain ceramides, and lysophosphatidylcholines decreased in PBMCs of the risperidone-treated group compared with untreated controls. These findings were confirmed by in vitro assays using human THP-1 monocytes. The reduction in the levels of very-long-chain ceramides and dihydroceramides could be due to the decrease in the expression of fatty acid elongase 3, as observed in THP-1 monocytes. Moreover, risperidone disrupted lipid raft domains in the plasma membrane of PBMCs. These results indicated that risperidone alters phospholipid and sphingolipid composition and lipid raft domains in PBMCs of older patients, potentially affecting multiple signaling pathways associated with these membrane domains.

scholarly journals The major soyabean allergen P34 resists proteolysis in vitro and is transported through intestinal epithelial cells by a caveolae-mediated mechanism

2012 ◽  
Vol 108 (9) ◽  
pp. 1603-1611 ◽  
Author(s):  
Eva Sewekow ◽  
Diane Bimczok ◽  
Thilo Kähne ◽  
Heidi Faber-Zuschratter ◽  
Lars Christian Kessler ◽  
...  
Keyword(s):  
Lipid Raft ◽  
Intestinal Barrier ◽  
Dietary Exposure ◽  
Soya Protein ◽  
The Body ◽  
Food Allergies ◽  
Epithelial Cell Line ◽  
Intestinal Epithelial ◽  
Lipid Raft Microdomains

Soya is considered to be one of the eight most significant food allergens. Among the allergenic soya proteins determined to date, P34 has been identified as one of the immunodominant soya antigens. Sensitisation to a specific food antigen like P34 generally follows the transit of intact antigens across the intestinal barrier and usually occurs in infants, who are most susceptible to food allergies. In the present study, we used the intestinal epithelial cell line IPEC-J2, which was originally derived from the jejunum of a neonatal piglet, to recapitulate the infant intestinal epithelium and study the binding and uptake of P34 protein. P34 was partially resistant to degradation in an in vitro proteolysis assay. IPEC-J2 cells were able to endocytose intact P34, as shown by immunofluorescence and immunoelectronmicroscopy methods. P34 associated with lipid raft microdomains of IPEC-J2 cells, and disruption of caveolae/lipid raft microdomains using methyl-β-cyclodextrin abolished P34 endocytosis, indicating that the observed endocytosis was mediated by caveolae. Using IPEC-J2 cells grown on Transwell filters, we further demonstrated that P34 is transported through the epithelial monolayer by transcytosis. Piglets frequently show hypersensitivity to soya antigens, and in this study, we show that healthy adult pigs with dietary exposure to soya protein mount an antibody response to soyabean protein P34, suggesting that this protein has entered the body, probably through gastrointestinal uptake. In summary, our data suggest that soya P34 resists proteolysis in the gastrointestinal tract and is transported through the intestinal epithelial barrier, thereby allowing sensitisation of immune cells in the sub-epithelial compartment.

The Effects of Alemtuzumab (A) Dose-Sequence upon Rituximab (R)-Induced Reorganization of Lipid Raft Domains and In-Vitro/In Vivo Anti-Tumor Activity.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2289-2289
Author(s):  
Francisco J. Hernandez-Ilizaliturri ◽  
Aruna C. Gowda ◽  
Dana Brombos ◽  
Dan Spear ◽  
Myron S. Czuczman
Keyword(s):  
Lipid Raft ◽  
In Vivo Studies ◽  
Rank Test ◽  
Raji Cells ◽  
Sequential Administration ◽  
In Vitro Exposure ◽  
Raft Domains ◽  
Anti Tumor Activity

Abstract Following [R] binding, CD20 antigen redistributes into lipid raft domains (LRD) and initiates signaling events leading to apoptosis in malignant B-cells. In addition, the clustering of CD20 receptors into LRD results in a relative increase in the antigen density within a given area of the cell membrane, thus facilitating [R]-mediated complement mediated cytotoxicity (CMC) and/or antibody dependent cellular cytotoxicity (ADCC). Combinations of monoclonal antibodies (mAbs) directed against unique tumor-associated targets can potentially alter the reorganization of LRD and modify their anti-tumor activity. Our main objective was to study the effects of [A] upon [R]-mediated anti-tumor effects. For ADCC/CMC studies, 51Cr-labeled NHL cells (Raji, DHL-4, DHL-10, Karpas422 or Ramos cells) were exposed to concurrent [A+R], sequential [R→A] or [A→R] prior to the addition of peripheral blood mononuclear cells (effector:target ratio of 40:1) or human serum, respectively. Trastuzumab [I] served as isotype control. Following a 6-hour period incubation, supernatant was harvested and % lysis calculated. In addition, LRD of 1x108 Raji cells exposed to similar mAb-combinations were extracted by sucrose gradient ultracentrifugation. Reorganization of CD20 into LRD was determined by Western blotting. For in vivo studies six to 8 week old SCID mice were inoculated by tail vein injection (iv) with 1x106 Raji cells (day 0). After tumor engraftment (Day +7), animals were divided in seven cohorts to receive 8 doses of vehicle control, [R], [A], [I], alternating doses of [R+A], or sequential dosing of [A→R] or [R→A]. MAb were administered IV at 5mg/kg/dose. The end point of the study was overall survival. Statistical analysis was performed with Kaplan-Meier survival curves and P values calculated by log rank test. In vitro exposure to [A] prior to [R] therapy resulted a 30 to 50% decrease in rituximab-mediated ADCC. In addition, exposure of Raji cells to [A] led to a decrease in the amount of CD20 reorganized into LRD following rituximab exposure. No decrease in [R]-mediated ADCC or reorganization of CD20 into LRD was observed when [A] was administered following [R] therapy. In vivo studies demonstrated a better anti-tumor activity in animals treated with alternating doses of [R + A], when compared to [R→A], [A→R], [R] or [A] therapy. The median survival for mice treated with combination mAb therapy [R+A] was 90 days, compared to 43 and 70 days for those treated with [R→A] and [A→R] respectively. Combination therapy with [R+A] resulted in a statistically significant longer survival when compared to [R], [A], or [I] (P<0.009). In conclusion, alternating administration of [R] and [A] was more effective in controlling lymphoma growth than either mAb monotherapy or sequential administration of [R→A]/[A→R]. In addition, in vitro exposure of lymphoma cells to [A] prior to [R] resulted in decreased anti-tumor activity that may be explained by impairment in the polarization of CD20 into LRD. The sequence of administration of mAbs being used in combination mAb clinical trials may strongly influence the degree of total anti-tumor activity achieved.

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