Journal: Frontiers in Immunology

Article Title: Granzyme B from mast cells contributes to choroidal neovascularization in a model of wet age-related macular degeneration

doi: 10.3389/fimmu.2026.1710965

Figure Lengend Snippet: Mast cell stabilizer prevents mast cell degranulation and reduces sprouting. (A) Day 8 representative explant images showing choroidal sprouting under the indicated conditions. Yellow outline indicates sprouting area, center black outline indicates explant. (B) Time-course analysis of choroidal sprouting in explants treated with mast cell degranulator (48/80) in the presence or absence of the KF. 48/80 was applied on days 2, 4, and 6 (arrows marked “T”) alongside KF. Significant differences in sprouting were observed at 4, 6 and 8 days. (C, D) Quantification of sprouting in WT explants treated with 48/80 +/- KF or just KF vs control shows significantly more sprouting when the mast cell degranulator is not present. (N=3). (E) Tryptase ELISA on CSA supernatants from indicated conditions. 48/80 treated explant supernatants contain significantly more tryptase than KF treated supernatant samples. (N=3). (F) Bleached CSA choroids stained with Toluidine blue after the treatments indicated. Arrows point to mast cells (or degranulated mast cells). Scale bar represents 20µm. Data are presented as mean ± SEM. N = 3 per condition. Statistical analysis was performed using unpaired t-tests. *p<0.05, **p<0.01, ***p<0.001, ns = not significant.

Article Snippet: Using the Mouse Mast Cell Tryptase ELISA Kit (CUSABIO, Cat. CSB-E14326m-1), supernatants were added neat into wells and incubated for 1 hour at 37 °C.

Techniques: Control, Enzyme-linked Immunosorbent Assay, Staining

Journal: PLOS One

Article Title: Differential sensitivity of MCPH1- and BRCA2-deficient cancer cells to PARP-1 inhibition

doi: 10.1371/journal.pone.0345514

Figure Lengend Snippet: (A) . U2OS or Hela cells were transfected with Control (siCON), MCPH1 (siMCPH1) or BRCA2 (siBRCA2) siRNA for 48h or 120h and protein expression analysed via Western blotting with the indicated antibodies. MCPH1 expression in sgCON (control) and sgMCPH1 (MCPH1 knockout) cells was also investigated by Western blotting. (B) Quantification of MCPH1 or BRCA2 protein bands normalised to GAPDH and expressed as % change relative to siCON or sgCON. Data is the mean and standard error of three (48h) or two (120h and sgCON/sgMCPH1) independent experiments. (C) U2OS or Hela cells were transfected with MCPH1 siRNA for 72h before analysis of nuclear morphology (premature chromosome condensation, PCC) via DAPI staining and fluorescence microscopy. (D) The % of cells displaying PCC, with at least 100 cells analysed.

Article Snippet: Membranes were blocked with 5% dried skimmed milk/TBS-T (50mM Tris pH 7.6, 150mM NaCl and 0.2% v/v Tween-20) for a minimum of 1-hour prior to overnight incubation at 4°C with primary antibodies; MCPH1 (11962–1-AP, Proteintech), BRCA2 (29450–1-AP, Proteintech) and GAPDH (60004–1-Ig, Proteintech).

Techniques: Transfection, Control, Expressing, Western Blot, Knock-Out, Staining, Fluorescence, Microscopy

Journal: PLOS One

Article Title: Differential sensitivity of MCPH1- and BRCA2-deficient cancer cells to PARP-1 inhibition

doi: 10.1371/journal.pone.0345514

Figure Lengend Snippet: U2OS cells were transfected with control (siCON), BRCA2 (siBRCA2) or BRCA2 + MCPH1 (siBRCA2 + siMCPH1) siRNA for 48h before (A) Western blot analysis or (B) seeding into 96-well plates and treating with the indicated doses of AZD2461 or Talazoparib, with each treatment performed in triplicate. After 96-hours, cell viability was measured using an MTS assay. MTS data represents the mean and standard error from at least three independent experiments, with cell viability expressed as a % relative to the corresponding untreated sample. Statistical significance was measured using two-way ANOVA (p-values described in text, with significance set at p < 0.05).

Article Snippet: Membranes were blocked with 5% dried skimmed milk/TBS-T (50mM Tris pH 7.6, 150mM NaCl and 0.2% v/v Tween-20) for a minimum of 1-hour prior to overnight incubation at 4°C with primary antibodies; MCPH1 (11962–1-AP, Proteintech), BRCA2 (29450–1-AP, Proteintech) and GAPDH (60004–1-Ig, Proteintech).

Techniques: Transfection, Control, Western Blot, MTS Assay

Journal: Pharmaceutical research

Article Title: Participation of Monocarboxylate Transporter 8, But Not P-Glycoprotein, in Carrier-Mediated Cerebral Elimination of Phenytoin across the Blood-Brain Barrier

doi: 10.1007/s11095-021-03003-1

Figure Lengend Snippet: Conditions for RT-PCR

Article Snippet: The blocked membrane was incubated with rabbit-derived anti-MCT8 (20676–1-AP; Proteintech, Rosemont, IL, USA) for approximately 12 h at 4°C or mouse-derived anti-Na + , K + -ATPase α1 antibodies (05–369; Merck) for 2 h at 20°C.

Techniques:

Journal: Pharmaceutical research

Article Title: Participation of Monocarboxylate Transporter 8, But Not P-Glycoprotein, in Carrier-Mediated Cerebral Elimination of Phenytoin across the Blood-Brain Barrier

doi: 10.1007/s11095-021-03003-1

Figure Lengend Snippet: Expression of monocarboxylate transporter 8 (MCT8) mRNA in mouse brain capillaries. RT-PCRwas performed in the absence (−) or presence (+) of reverse transcriptase. Total RNA from the mouse brain was used as a positive control for MCT8 mRNA expression. This image was representative of the images from 3 replicates.

Article Snippet: The blocked membrane was incubated with rabbit-derived anti-MCT8 (20676–1-AP; Proteintech, Rosemont, IL, USA) for approximately 12 h at 4°C or mouse-derived anti-Na + , K + -ATPase α1 antibodies (05–369; Merck) for 2 h at 20°C.

Techniques: Expressing, Positive Control

Journal: Pharmaceutical research

Article Title: Participation of Monocarboxylate Transporter 8, But Not P-Glycoprotein, in Carrier-Mediated Cerebral Elimination of Phenytoin across the Blood-Brain Barrier

doi: 10.1007/s11095-021-03003-1

Figure Lengend Snippet: Protein expression of monocarboxylate transporter 8 (MCT8) in rat tran capillaries. Crude membrane fraction (25 μg) from rat brain capillaries and liver was used for the Western blot analysis of MCT8 proteins. Na+, K+-ATPase α1 was used for the loading control of the crude membrane fraction (21). An arrowhead indicates the expected size of the rat MCT8 proteins (14).

Article Snippet: The blocked membrane was incubated with rabbit-derived anti-MCT8 (20676–1-AP; Proteintech, Rosemont, IL, USA) for approximately 12 h at 4°C or mouse-derived anti-Na + , K + -ATPase α1 antibodies (05–369; Merck) for 2 h at 20°C.

Techniques: Expressing, Western Blot

Journal: Pharmaceutical research

Article Title: Participation of Monocarboxylate Transporter 8, But Not P-Glycoprotein, in Carrier-Mediated Cerebral Elimination of Phenytoin across the Blood-Brain Barrier

doi: 10.1007/s11095-021-03003-1

Figure Lengend Snippet: [l4C]Phenytoin uptake in X. oocytes expressing MCT5 (a), MCT8 (b), or MCT9 (c). [l4C]Phenytoin uptake (0.1 μCi/200μL; 9.09μM) in X. oocytes expressing rat MCT5 (a), MCT8 (b), or MCT9 (c) or in X. oocytes injected with nuclease-free water (a, b, and c) In the absence (−, b) or presence of 400μM unlabeled phenytoin, 1 mM bromosulfophthalein (BSP), or 1 mM desipramine (b). Each open circle represents an individual sample. Each column represents the mean ± S.D. (n = 5–20). **p < 0.01, a significant difference.

Article Snippet: The blocked membrane was incubated with rabbit-derived anti-MCT8 (20676–1-AP; Proteintech, Rosemont, IL, USA) for approximately 12 h at 4°C or mouse-derived anti-Na + , K + -ATPase α1 antibodies (05–369; Merck) for 2 h at 20°C.

Techniques: Expressing, Injection

Journal: Cell Death Discovery

Article Title: MCT4-driven CAF-mediated metabolic reprogramming in breast cancer microenvironment is a vulnerability targetable by miR-425-5p

doi: 10.1038/s41420-024-01910-x

Figure Lengend Snippet: Representative Western blot analysis of MCT4 performed on plasmamembrane fractions of Cancer-Associated Fibroblasts (CAFs) ( n = 4) and Normal Fibroblasts (NFs) ( n = 3). Na + /K+ ATPase was used for data normalization of the fractionation in the sample preparation. Densitometry data are expressed after normalization on Na + /K+ ATPase band intensity A . A representative confocal microscopy image of MCT4 expression in CAFs ( n = 3) (DAPI blue, MCT4 red) B . Three different subpopulations of two primary fibroblasts were collected at different growing distances from the tumor (intratumoral, IT; peritumoral, PT; normal, NF) and MCT4 gene expression was evaluated by quantitative real-time PCR. Values are expressed as a mean of two biological replicates of technical duplicates C . Culture media from NFs ( n = 2) and CAFs ( n = 2) were collected and assessed by a colorimetric L-lactate assay. Values are expressed as mean of two technical replicates D . Graph shows mean ± SD. * P values < 0.05; *** p < 0.001. Bar = 100 μm.

Article Snippet: Plasmid transfection in MS-5 (2500 ng of SLC16A3 tGFP-tagged supplied by Origene CAT#: RG211987) was carried out using Lipofectamine 3000 reagent (Invitrogen).

Techniques: Western Blot, Fractionation, Sample Prep, Confocal Microscopy, Expressing, Real-time Polymerase Chain Reaction, Lactate Assay

Journal: Cell Death Discovery

Article Title: MCT4-driven CAF-mediated metabolic reprogramming in breast cancer microenvironment is a vulnerability targetable by miR-425-5p

doi: 10.1038/s41420-024-01910-x

Figure Lengend Snippet: Bioinformatics predictions (TargetScan and miRANDA) indicated miR-425-5p as a putative miRNA targeting MCT4 mRNA A . Representative Real Time PCR for miR-425-5p upon miR-425-5p and anti miR-425-5p transfection in Cancer-Associated Fibroblasts (CAF n = 1) and Normal Fibroblasts (NF n = 1) respectively. Data are expressed as mean of two technical replicates B . Representative Western Blot analysis of MCT4 upon miR-425-5p and anti-miR-425 transfection in CAFs ( n = 4) and NFs ( n = 2) respectively C . Dual luciferase assay of the predicted binding site for miR-425-5p on the 3’UTR region of SLC16A3. Data are expressed as mean of two technical replicates D . Quantitative Real Time PCR for miR-425-5p performed on CAFs ( n = 3) and NFs ( n = 2), to evaluate miR-425-5p levels. Values are expressed as mean of two technical replicates E . Graph shows mean ± SD over control. * P values < 0.05.

Article Snippet: Plasmid transfection in MS-5 (2500 ng of SLC16A3 tGFP-tagged supplied by Origene CAT#: RG211987) was carried out using Lipofectamine 3000 reagent (Invitrogen).

Techniques: Real-time Polymerase Chain Reaction, Transfection, Western Blot, Luciferase, Binding Assay, Control

Journal: Cell Death Discovery

Article Title: MCT4-driven CAF-mediated metabolic reprogramming in breast cancer microenvironment is a vulnerability targetable by miR-425-5p

doi: 10.1038/s41420-024-01910-x

Figure Lengend Snippet: Metabolic changes in MCT4-transfected MS-5 cells with or without miR-425-5p overexpression, as assessed by glycolytic proton efflux rate (glycoPER) kinetics A , proton efflux rate (PER) B , extracellular acidification rate (ECAR) C , oxygen consumption rate OCR D , and ATP production E . Data are presented as a mean of biological duplicates of eight technical replicates ± SD over control.

Article Snippet: Plasmid transfection in MS-5 (2500 ng of SLC16A3 tGFP-tagged supplied by Origene CAT#: RG211987) was carried out using Lipofectamine 3000 reagent (Invitrogen).

Techniques: Transfection, Over Expression, Control

Journal: Cell Death Discovery

Article Title: MCT4-driven CAF-mediated metabolic reprogramming in breast cancer microenvironment is a vulnerability targetable by miR-425-5p

doi: 10.1038/s41420-024-01910-x

Figure Lengend Snippet: MiR-425-5p alters the metabolism of breast Cancer-associated Fibroblasts (CAFs) by downregulating MCT4 and indirectly reducing GLUT1, leading to decreased lactate extrusion. The interaction between miR-425-5p reprogrammed CAFs and breast cancer epithelial cells has a significant impact on breast cancer cell viability, proliferation, and migration. Angiogenesis is also influenced. This graphical abstract was drawn by using and adapting pictures from Servier Medical Art (Smart.Servier.com), provided by Servier, licensed under a Creative Commons Attribution 3.0 unported license ( http://creativecommons.org/licenses/by/3.0/ ).

Article Snippet: Plasmid transfection in MS-5 (2500 ng of SLC16A3 tGFP-tagged supplied by Origene CAT#: RG211987) was carried out using Lipofectamine 3000 reagent (Invitrogen).

Techniques: Migration

Journal: Journal of Inflammation Research

Article Title: Exploring Long Non-Coding RNAs Associated with IP3/DAG Signaling Pathway as Potential Biomarkers Involved in Mast Cell Degranulation in Chronic Spontaneous Urticaria with 2-Year Follow-Up

doi: 10.2147/JIR.S343826

Figure Lengend Snippet: Serum concentrations of hs-CRP, LTB4, PGD2, MCT, D-dimer and HIS in the CSU (n = 56) and healthy control (n = 13) groups. Differences between groups were assessed using Mann–Whitney U -tests; experiments were repeated three times.

Article Snippet: The levels of HIS (ab213975, Abcam, Cambridge, UK), MCT (CSB-E09012h, CUSABIO, Wuhan, China), LTB4 (CSB-E08033h), PGD2 (CSB-E13898h), hs-CRP (CSB-E08617h), and D-dimer (CSB-E05175h) were assessed in peripheral blood serum using human ELISA kits according to the manufacturer’s instructions.

Techniques: Control, MANN-WHITNEY