Journal: Scientific Reports

Article Title: Increase of intracellular Zn 2+ concentration directly inhibits phospholipase Cε and suppresses inflammation and tumour formation in mice

doi: 10.1038/s41598-025-25886-5

Figure Lengend Snippet: Zinc as an Active Principle of ZPT. (A) Chemical structures of dipyrithione and sodium pyrithione are shown. The activities of varying concentrations of ZPT, ZnSO 4 , dipyrithione and sodium pyrithione to inhibit the hydrolysis of Compound 1 by purified PLCε were measured as described in Methods. ( B ) The activities of varying concentrations of ZnSO 4 to inhibit the hydrolysis of Compound 1 by purified PLCε, PLCβ4, PLCδ1 or PLCγ1 were measured as described in Methods. (C) The activities of 5 µM ZnSO 4 to inhibit the hydrolysis of Compound 1 by purified PLCε were measured in the presence of varying concentrations of sodium pyrithione. ( D ) Caco2 cells were treated with 5 µM ZnSO 4 in the presence or absence of 5 µM sodium pyrithione or with 5 µM ZPT for 30 min. Intracellular Zn 2+ concentrations of the treated cells were measured by using Zinquin ethyl ester as described in Methods. The values obtained are expressed as fold changes over the average value of untreated cells. All the experiments described above were performed three times in duplicates.

Article Snippet: A human colon cancer epithelial cell line Caco2 was purchased from ATCC (HTB-37) and maintained in 5% CO 2 at 37 °C in modified Eagle’s minimum essential medium (MEM) (Nacalai tesque) supplemented with 20% fetal bovine serum (FBS) (Sigma), non-essential amino acids (Gibco) and 100 μg/ml penicillin-streptomycin (Nacalai tesque).

Techniques: Purification

Journal: Scientific Reports

Article Title: Increase of intracellular Zn 2+ concentration directly inhibits phospholipase Cε and suppresses inflammation and tumour formation in mice

doi: 10.1038/s41598-025-25886-5

Figure Lengend Snippet: Effect of ZPT on PKD–NF-κB Signaling and Proinflammatory Gene Expression. (A) Caco2 and SW480 cells were serum-starved for 3 h in the presence of the indicated concentrations of ZPT or a vehicle (DMSO) and subsequently stimulated by 20 µM LPA for 30 min. PKD phosphorylated at Ser916 and total PKD in the cell lysates were quantified by immunoblotting with anti-phospho-PKD (Ser916) and anti-PKD Abs, respectively, as described in Methods. Numbers below the immunoblots indicate fold increase of the phospho-PKD signals divided by the total PKD signals over that at 0 min after LPA stimulation of the control cells. Each of the experiments was performed three times yielding equivalent results. A representative result is shown. The original blots are presented in Supplementary Figures S4 and S5. (B) Caco2 cells treated with 5 µM ZPT or the vehicle as described in (A) were subjected to subcellular fractionation and the resulting nuclear and cytoplasmic fractions were subjected to immunoblotting with the anti-NF-κB p65 Ab. TATA-binding protein (TBP) and α-tubulin were used as markers for the nuclear and the cytoplasmic fractions, respectively. A fold change of the nuclear NF-κB caused by 5 µM ZPT treatment is shown in numbers. Each of the experiments was performed three times yielding equivalent results. A representative result is shown. The original blots are presented in Supplementary Figure S6. (C) Total cellular RNAs isolated from Caco2 cells treated as described in (B) were subjected to qRT-PCR for quantification of the mRNA levels of the proinflammatory molecules: CXCL1, CXCL8, CCL2, CCL20, TNF-α and COX-2, by using the b- actin mRNA as an internal control. The obtained values are expressed as relative fold changes of nuclear NF-κB normalized to TATA-binding protein (TATA), with the value from the − LPA/−ZPT condition set to 1. The experiments were performed three times in duplicates.

Article Snippet: A human colon cancer epithelial cell line Caco2 was purchased from ATCC (HTB-37) and maintained in 5% CO 2 at 37 °C in modified Eagle’s minimum essential medium (MEM) (Nacalai tesque) supplemented with 20% fetal bovine serum (FBS) (Sigma), non-essential amino acids (Gibco) and 100 μg/ml penicillin-streptomycin (Nacalai tesque).

Techniques: Gene Expression, Western Blot, Control, Fractionation, Binding Assay, Isolation, Quantitative RT-PCR

Journal: Scientific Reports

Article Title: Increase of intracellular Zn 2+ concentration directly inhibits phospholipase Cε and suppresses inflammation and tumour formation in mice

doi: 10.1038/s41598-025-25886-5

Figure Lengend Snippet: Effect of ZPT Administration on Proliferation of Xenografted Caco2 cells. Caco2 cells (5 × 10 6 cells) were implanted subcutaneously into the right flanks of female nude mice. One week later, the mice were administered intraperitoneally with ZPT, dipyrithione or the vehicle 3 days per week for 8 weeks, during which the body weight (A) and the tumour volumes (B) were measured every 7 days. The number of mice ( n ) for each group is shown in parenthesis. The number of mice ( n ) for each group is shown in parenthesis. Thereafter, the tumours were excised and weighed (C) .

Article Snippet: A human colon cancer epithelial cell line Caco2 was purchased from ATCC (HTB-37) and maintained in 5% CO 2 at 37 °C in modified Eagle’s minimum essential medium (MEM) (Nacalai tesque) supplemented with 20% fetal bovine serum (FBS) (Sigma), non-essential amino acids (Gibco) and 100 μg/ml penicillin-streptomycin (Nacalai tesque).

Techniques: