Journal: Archives of dermatological research

Article Title: The effect of selinexor on prostaglandin synthesis in virus-positive Merkel cell carcinoma cell lines.

doi: 10.1007/s00403-024-03108-8

Figure Lengend Snippet: Fig. 3 The prostaglandin synthe sis pathway is responsible for converting arachidonic acid to prostaglandins and other lipids. Abbreviations cyclooxygenase (COX), cytosolic phospholipase A2 (cPLA2), prostaglandin (PG), 2 synthase (2 S), prostaglandin receptor (PTG R), Thromboxane- A (TXA), E prostanoid (EP)

Article Snippet: Lysates were loaded onto 4–12% bis-tris gels and 6% tris-glycine, and gel electrophoresis was performed using the Laemmli sodium dodecyl sulfate-polyacrylamide gel electrophoresis system and transferred to a polyvinylidene difluoride membrane. cPLA2 (cst-#5249; Cell Signaling Technology (CST), Danvers, MA, USA), COX1 (sc-70,878; SCB), COX2 (cst-#12,282; CST), PGD2S (sc-390,717; SCB), PGE2S (sc-514,224; SCB), PGF2S (#MAB7678; R&D Systems (RND), Minneapolis, MN, USA), PGI2S (#MAB7788, RND), TXA2S (PA545676, TFS), EP1 (#ab217925, Abcam, Waltham, MA, USA), EP2 (#ab167171, Abcam), EP3 (PA5-92124, TFS), EP4 (sc-55,596, SCB), TBXA2R (#27159-1-AP, Proteintech, Rosemont, IL, USA), PTGFR (PA570674, TFS), PTGIR (PA5-102240, TFS), and PTGDR (#orb315671, Biorbyt, Durham, NC, USA) were used.

Techniques:

Journal: Journal of Clinical Investigation

Article Title: Inhibition of 11β–hydroxysteroid dehydrogenase type II selectively blocks the tumor COX-2 pathway and suppresses colon carcinogenesis in mice and humans

doi: 10.1172/jci37398

Figure Lengend Snippet: Figure 5 Inhibition of 11βHSD2 activity with GA suppressed tumor growth through activation of glucocorticoid receptors. (A) GA augmented low-dose CS–induced inhibition of CT26 cell COX-2 expression. (B) Treatment with 10 mg/kg/d GA i.p. led to significant decreases in tumor size, COX-2 expression, and PGE2 levels, which were completely reversed by the glucocorticoid receptor inhibitor RU486 (7.5 mg/kg/d i.m.). n = 5–6 per group. (C) GA reduced CT26 tumor expression of phosphorylated cPLA2 (P-cPLA2) and mPGES-1, but had no effect on tumor expression of COX-1 (P = 0.23) and cPGES (P = 0.055). Original magnification, ×400. (D) GA treatment significantly increased corticosterone levels and decreased 11-keto-corticosterone levels in kidney, colon, and tumors. n = 5. *P < 0.0001, **P < 0.001, ***P < 0.02, #P < 0.05, ##P < 0.01, and ###P < 0.005 versus vehicle.

Article Snippet: Affinity-purified rabbit anti-mouse 11βHSD2 (catalog no. BHSD22-A) was purchased from Alpha Diagnostic International; rabbit anti-murine COX-2 (catalog no. 160106) and COX-1 (catalog no. 160109), rabbit anti-human mPGES-1 (catalog no. 160140) and cPGES (catalog no. 160150), and rabbit anti-rat 15-PGDH (catalog no. 160615) were from Cayman Chemicals; rabbit anti-human cPLA2 (catalog no. 2382) and phosphorylated cPLA2 (Ser505; catalog no. 2831) were from Cell Signaling; and goat anti-rat VEGF (catalog no. AF564) was from R&D Systems.

Techniques: Inhibition, Activity Assay, Activation Assay, Expressing

Journal: Journal of Clinical Investigation

Article Title: Inhibition of 11β–hydroxysteroid dehydrogenase type II selectively blocks the tumor COX-2 pathway and suppresses colon carcinogenesis in mice and humans

doi: 10.1172/jci37398

Figure Lengend Snippet: Figure 7 Proposed mechanism underlying 11βHSD2 activity and colorectal tumorigenesis. In tumor cells, glucocorticoids are converted to inac- tive keto-forms by 11βHSD2, reducing glucocorticoid receptor activa- tion, while inhibition of 11βHSD2 activity (by GA and its analogs or by gene knockdown) leads to increased levels of intracellular active glucocorticoids. The consequent inhibition of cPLA2 activity and COX-2 and mPGES-1 expression results in blockade of PGE2 production and inhibition of tumor growth.

Article Snippet: Affinity-purified rabbit anti-mouse 11βHSD2 (catalog no. BHSD22-A) was purchased from Alpha Diagnostic International; rabbit anti-murine COX-2 (catalog no. 160106) and COX-1 (catalog no. 160109), rabbit anti-human mPGES-1 (catalog no. 160140) and cPGES (catalog no. 160150), and rabbit anti-rat 15-PGDH (catalog no. 160615) were from Cayman Chemicals; rabbit anti-human cPLA2 (catalog no. 2382) and phosphorylated cPLA2 (Ser505; catalog no. 2831) were from Cell Signaling; and goat anti-rat VEGF (catalog no. AF564) was from R&D Systems.

Techniques: Activity Assay, Inhibition, Knockdown, Expressing

Journal: Cancer Research

Article Title: Lysophosphatidic Acid Is Constitutively Produced by Human Peritoneal Mesothelial Cells and Enhances Adhesion, Migration, and Invasion of Ovarian Cancer Cells

doi: 10.1158/0008-5472.can-05-1292

Figure Lengend Snippet: Figure 3. iPLA2 and cPLA2 activities are involved in generating the chemotactic activity of peritoneal mesothelial cell conditioned medium. A, total LPA production in the conditioned medium from LP9 was inhibited by HELSS (1 Amol/L) and AACOCF3 (25 Amol/L). ***, P < 0.001 versus the corresponding control (Student’s t test). The LPA in 24 hr-CM from LP9 is f80 nmol/L/106 cells/24 hours. B, PLA2 inhibitors reduced the chemotactic activity of 24 hr-CM from LP9 on SKOV3 cell migration. When LP9 cells were incubated for conditioned medium production, AACOCF3 (25 Amol/L), HELSS (1 Amol/L), or thioether amide-PC (TAPC; 10 Amol/L) was added to the medium during the incubation for 24 hours before collecting conditioned medium (Before). We also collected conditioned medium produced in the absence of inhibitors, but added inhibitors into the lower chamber after conditioned medium were collected to assess the direct effects of these inhibitors during the migration assays (After). The effects of these conditioned medium on migration of SKOV3 cells were analyzed. Relative fold changes in migration were determined by setting cell migration in response to conditioned medium without treatment at 1.0. ***, P < 0.001 versus the control (Student’s t test). C, a to d, cell-based enzymatic activities of PLA2 in peritoneal mesothelial cells with or without overnight serum (FBS) starvation. Cells (104) were incubated with DBPC for 30 minutes. See Materials and Methods for the assay conditions. e, summary of three independent cell-based enzymatic assays. Intensities were measured as described in Materials and Methods. D, cPLA2 is constitutively phosphorylated in peritoneal mesothelial cells. Phosphorylated cPLA2 and total cPLA2 expression in peritoneal mesothelial cells were detected by corresponding antibodies in Western blot analyses. h-actin was used as a loading control. Cell lysates from LP9 and LP3 grown with or without overnight starvation from FBS were used for Western blot analyses.

Article Snippet: Cytosolic phospholipase A2 (cPLA2) antibodies were from Upstate Cell Signaling Solutions (Lake Placid, NY) or Cell Signaling Technology (Beverly, MA).

Techniques: Activity Assay, Control, Migration, Incubation, Produced, Expressing, Western Blot

Journal: Cancer Research

Article Title: Lysophosphatidic Acid Is Constitutively Produced by Human Peritoneal Mesothelial Cells and Enhances Adhesion, Migration, and Invasion of Ovarian Cancer Cells

doi: 10.1158/0008-5472.can-05-1292

Figure Lengend Snippet: Figure 6. Invasion of SKOV3 cells into peritoneal mesothelial cells requires LPA2, LPA3, iPLA2, cPLA2, and MEK-ERK activity. A, invasion assays were conducted in the ECIS system (see Materials and Methods). LP9 (1 105 cells in 100 AL medium) were added to the gelatin-coated wells of the ECIS chambers and allowed to form a monolayer covering the gold electrode in each well. On the next day, SKOV3, NIH3T3, or MCF7 cells (1 105 cells in 300 AL medium) were added on top of the LP9 cell layer. In the control, 300 AL medium without cells was added on top of the LP9 monolayer. B, SKOV3 cells were transiently transfected with siRNAs against LPA1-3, GAPDH, or GFP, and tested for invasion into LP9 in the ECSI system. C, for inhibitor treatments, LP9 cell monolayers were pretreated with HELSS (1 Amol/L, 30 minutes) or AACOCF3 (100 Amol/L, 30 minutes) before addition of SKOV3 cells. D, SKOV3 cells were pretreated with the MEK inhibitor PD098059 (10 Amol/L, 30 minutes; top) or an Akt inhibitor (Akti, 10 Amol/L, 30 minutes; middle) before their application to the LP9 monolayers in the ECIS wells. Bottom, SKOV3 cells were transient transfected with dominant negative (dn) MEK or Akt and tested for invasion into LP9.

Article Snippet: Cytosolic phospholipase A2 (cPLA2) antibodies were from Upstate Cell Signaling Solutions (Lake Placid, NY) or Cell Signaling Technology (Beverly, MA).

Techniques: Activity Assay, Control, Transfection, Dominant Negative Mutation

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Glutamine suppresses airway neutrophilia by blocking cytosolic phospholipase A(2) via an induction of MAPK phosphatase-1.

doi: 10.4049/jimmunol.1201599

Figure Lengend Snippet: FIGURE 3. cPLA2 inhibitors suppress air- way neutrophilia. (A) Varying concentrations of AACOCF3 were administered 30 min before the secondary airway challenge. (B) Then 0.4 nmol of cPLA2 siRNA or control siRNA was administered i.v. 24 h before the secondary airway challenge, and cPLA2 protein and phospho-cPLA2 were detected 30 min after the secondary airway challenge. (C) Mice were pretreated with siRNA, and the number of air- way neutrophils was measured 8 h after the secondary airway challenge. Data represent (A, C) the mean 6 SD and (B) a representative of three separate experiments (n = 3–5 per group). *p , 0.05 versus control group, **p , 0.05 versus OVA group.

Article Snippet: Small interfering RNA (siRNA) strands for cPLA2 and controls were obtained from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Control

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Glutamine suppresses airway neutrophilia by blocking cytosolic phospholipase A(2) via an induction of MAPK phosphatase-1.

doi: 10.4049/jimmunol.1201599

Figure Lengend Snippet: FIGURE 4. LTB4 induces airway neutrophilia and Gln inhibits LTB4 production. (A) Varying concentrations of the 5-LO inhibitors AA-861 and MK-886 were administered 1 h before the secondary airway challenge. (B) BAL fluid levels of LTB4 were determined after a second OVA airway challenge at the time indicated. (C) Effects of Gln and cPLA2 siRNA on BAL fluid levels of LTB4 at 1 h. (D) Varying concentrations of LTB4 were instilled i.t. 1 h after the secondary airway challenge. (E) The Gln-treated group was instilled i.t. with 1 mg of LTB4 at 1 h. The number of airway neutrophils was measured at 8 h. Data represent the mean 6 SD of three to five separate experiments (n = 5 per group).

Article Snippet: Small interfering RNA (siRNA) strands for cPLA2 and controls were obtained from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques:

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Glutamine suppresses airway neutrophilia by blocking cytosolic phospholipase A(2) via an induction of MAPK phosphatase-1.

doi: 10.4049/jimmunol.1201599

Figure Lengend Snippet: FIGURE 5. Gln inhibits the phosphorylation of cPLA2 and p38, and p38 inhibitor inhibits cPLA2 phosphorylation and airway neutrophilia. (A and B) Gln and Ala were administered 20 min post challenge. SB202190 (5 mg/kg) was injected i.p., and (C) the levels of cPLA2 phosphorylation and (D) neutrophil numbers in BAL fluid were measured 30 min and 8 h after the secondary airway challenge, respectively. (E) SB202190 was injected i.p. and BAL levels of MIP-2, KC, CXCL5, and LTB4 were measured at 2 h. (A–C) A representative of three independent experiments with three to five mice per time point per experiment is shown. (D and E) Data represent the mean 6 SD of two separate experiments (n = 5 per group). *p , 0.05 versus OVA group.

Article Snippet: Small interfering RNA (siRNA) strands for cPLA2 and controls were obtained from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Phospho-proteomics, Injection

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Glutamine suppresses airway neutrophilia by blocking cytosolic phospholipase A(2) via an induction of MAPK phosphatase-1.

doi: 10.4049/jimmunol.1201599

Figure Lengend Snippet: FIGURE 6. Gln-induced inhibition of p38 is asso- ciated with the early induction of MKP-1. (A) Gln and Ala were administered 20 min post challenge. (B) MKP-1 siRNA and control siRNA were administered i.v. 24 h before the secondary airway challenge. MKP-1 was measured at 90 min. (C) Then 0.4 nmol of siRNAs was given, and phospho-p38, phospho-cPLA2, and MKP-1 induction were measured at 30 min. A repre- sentative of two or three independent experiments with three to five mice per time point per experiment is shown. **p , 0.05 versus OVA group.

Article Snippet: Small interfering RNA (siRNA) strands for cPLA2 and controls were obtained from Santa Cruz Biotechnology (Santa Cruz, CA).

Techniques: Inhibition, Control

Journal: The Journal of cell biology

Article Title: Tip60-mediated Rheb acetylation links palmitic acid with mTORC1 activation and insulin resistance.

doi: 10.1083/jcb.202309090

Figure Lengend Snippet: Figure 6. Tip60 interacts with and acetylates Rheb to regulate mTORC1 activity and insulin sensitivity under PA condition. (A) Relative mRNA levels of acetyltransferase genes (cbp, gcn5, pcaf, and tip60) were analyzed by quantitative PCR in fish myocytes under control or 500 μM PA treatment for 12 h (n = 3

Article Snippet: To inhibit Tip60, 150 μMMG149 was added to the culture medium of fish myocytes or C2C12 myotubes for 12 h in the presence or absence of PA. For TFEB activator 1 treatment, TFEB activator 1 (#HY135825; Med Chem Express) was dissolved in DMSO (Solarbio) to obtain 10 mM stock solution.

Techniques: Activity Assay, Real-time Polymerase Chain Reaction, Control

Journal: The Journal of cell biology

Article Title: Tip60-mediated Rheb acetylation links palmitic acid with mTORC1 activation and insulin resistance.

doi: 10.1083/jcb.202309090

Figure Lengend Snippet: Figure 9. A working model of how Tip60-mediated Rheb acetylation links PA with mTORC1 activation and insulin resistance. Created with https:// BioRender.com.

Article Snippet: To inhibit Tip60, 150 μMMG149 was added to the culture medium of fish myocytes or C2C12 myotubes for 12 h in the presence or absence of PA. For TFEB activator 1 treatment, TFEB activator 1 (#HY135825; Med Chem Express) was dissolved in DMSO (Solarbio) to obtain 10 mM stock solution.

Techniques: Activation Assay

Journal: Journal of Global Health

Article Title: Unveiling the genetic biomarkers for ageing: evidence from a large sample genome-wide association study and in vivo validation

doi: 10.7189/jogh.15.04279

Figure Lengend Snippet: Panel A. Representative Nissl-stained images of the cerebral cortex and hippocampus from 3-month-old and 20-month-old mice. Panel B. Quantitative analysis of Nissl-stained images comparing the mean density of Nissl-positive signals in the cerebral cortex and hippocampus of 3m and 20m mice (n = 3). Panel C. Western blot analysis of UBA7 and PLA2G4B expression across major organs and in the brains of three- and 20-month-old mice (n = 10). Panel D. Quantification of Western blot results for UBA7 and PLA2G4B in the brains of three- and 20-month-old mice (n = 10). * P ≤ 0.05. † P ≤ 0.01. au – arbitrary units, m – month.

Article Snippet: Subsequently, we blocked the membranes using 5% skim milk and incubated them with primary antibodies targeting UBA7 (Abmart, T59622 ), PLA2G4B (Affinity, DF9429), and GAPDH (Proteintech, 10494-1-AP) overnight at 4°C.

Techniques: Staining, Western Blot, Expressing