Journal: Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology

Article Title: Inhibition of Lithium Sensitive Orai1/ STIM1 Expression and Store Operated Ca2+ Entry in Chorea-Acanthocytosis Neurons by NF-κB Inhibitor Wogonin.

doi: 10.1159/000495229

Figure Lengend Snippet: Fig. 2. Effect of lithium treatment without and with NFκB inhibitor wogonin on Orai1, STIM1 and STIM2 protein abundance in iPSC-derived neurons from ChAc patients. A,B,C. Original Western blot of (A) Orai1, (B) STIM1 and (C) STIM2 protein abundance in neurons differentiated from iPSCs derived from ChAc patients (ChAc) without or with lithium (24 h, 2 mM) treatment without and with presence of NFκB inhibitor wogonin (50 µM). D,E,F. Arithmetic means (± SEM, n = 5) of (D) Orai1, (E) STIM1 and (F) STIM2 protein abundance in neurons generated from iPSCs derived from ChAc patients without (blue bars) or with (red and grey bars) lithium (24 h, 2 mM) treatment without (red bars) and with (grey bars) presence of NFκB inhibitor wogonin (50 µM). #(p<0.05), ##(p<0.01) indicates statistically significant difference to respective value in the absence of treatment, §(p<0.05), §§§(p<0.001) indicates statistically significant difference to respective value in the absence of wogonin.

Article Snippet: The proteins were separated by 10% SDS-PAGE in Glycine-Tris buffer and electro-transferred onto nitrocellulose membranes for 90 min. After blocking with 5% milk in TBST at room temperature for 1 h, the membranes were incubated with primary anti-ORAI1 antibody (1:1000, Proteintech), anti-STIM1 antibody (1:1000, Cell Signaling), anti-STIM2 antibody (1:1000, Cell Signaling) and anti-GAPDH antibody (1:1000, Cell Signaling) at 4°C overnight.

Techniques: Quantitative Proteomics, Derivative Assay, Western Blot, Generated

Journal: Cell Communication and Signaling : CCS

Article Title: TRPP2 and STIM1 form a microdomain to regulate store-operated Ca 2+ entry and blood vessel tone

doi: 10.1186/s12964-020-00560-7

Figure Lengend Snippet: Co-localization and co-immunoprecipitation of TRPP2 and STIM1 in transfected HEK293 cells. a Upper penal: GFP-tagged TRPP2 (TRPP2-GFP) co-localized with endoplasmic reticulum maker (ER-DsRed); Lower penal: TRPP2-GFP co-localized with mCherry-tagged STIM1 (mCherry-STIM1). b Representative images showing co-immunoprecipitation followed by immunoblots [left, immunoblot with anti-GFP; right, immunoblot with anti-mCherry]. GFP or mCherry antibody pulled down the proteins from TRPP2-GFP and mCherry-STIM1 co-expressing HEK293 cells. The experiment was repeated 4 times

Article Snippet: The primary rabbit antibody against STIM1 was obtained from ProSci.

Techniques: Immunoprecipitation, Transfection, Western Blot, Expressing

Journal: Cell Communication and Signaling : CCS

Article Title: TRPP2 and STIM1 form a microdomain to regulate store-operated Ca 2+ entry and blood vessel tone

doi: 10.1186/s12964-020-00560-7

Figure Lengend Snippet: FRET efficiency of TRPP2-STIM1 interaction in TRPP2 and STIM1 co-expressing HEK293 cells. a Representative images showing GFP, mCherry and FRET efficiency channels. The cells expressed TRPP2-mCherry and STIM1-GFP (upper), mCherry-TRPP2 and STIM1-GFP (middle) or GFP-mCherry (lower) proteins respectively. b Summarized data showing FRET efficiency in transfected HEK293 cells (four groups: mCherry and GFP, mCherry-GFP, TRPP2-mCherry and STIM1-GFP, mCherry-TRPP2 and STIM1-GFP). Values are shown as the mean ± SEM ( n = 9–20 cells). * P < 0.05 for GFP and mCherry co-expression vs. GFP-mCherry expression; # P < 0.05 for TRPP2-mCherry and STIM1-GFP co-expression vs. mCherry-TRPP2 and STIM1-GFP co-expression. c Schematic diagram of full-length TRPP2 and its truncated derivatives: TRPP2 without N1 (2-111aa, ∆N1) and TRPP2 without N2 (112-221aa, ∆N2). d Representative images of co-immunoprecipitation experiments in HEK293 cells co-expressed with STIM1 plus GFP-tagged full-length TRPP2 (WT) or ∆N1 or ∆N2. IP, GFP antibody; IB, anti-EGFP antibody. The GFP-only vector was used as a negative control. e Summarized data showing the relative binding strength of STIM1 with TRPP2 (WT), or ∆N1 or ∆N2. The optical density of ∆N1 or ∆N2 blot was normalized to that of WT blot (= 100%) and expressed as the relative binding strength. Values are shown as the mean ± SEM ( n = 3). * P < 0.05 for STIM1 and ∆N1 or ∆N2 co-expression vs. STIM1 and TRPP2 co-expression

Article Snippet: The primary rabbit antibody against STIM1 was obtained from ProSci.

Techniques: Expressing, Transfection, Immunoprecipitation, Plasmid Preparation, Negative Control, Binding Assay

Journal: Cell Communication and Signaling : CCS

Article Title: TRPP2 and STIM1 form a microdomain to regulate store-operated Ca 2+ entry and blood vessel tone

doi: 10.1186/s12964-020-00560-7

Figure Lengend Snippet: Effect of TRPP2-STIM1 interaction on store-operated Ca 2+ entry (SOCE) in TRPP2 and STIM1 co-expressing HEK293 cells. a - d Summary of data showing changes in Ca 2+ release ( a , c ) and SOCE ( b , d ) in HEK293 cells transfected with TRPP2 siRNA, STIM1 siRNA, STIM1, TRPP2 and/or dominant negative TRPP2 (D511V), and treated by ATP (10 μmol/L), 2APB (100 μmol/L) + ATP (10 μmol/L) and thapsigargin (TG, 2.5 μmol/L) for 8 min in Ca 2+ -free solution. SOCE was evoked by extracellular Ca 2+ (1 mmol/L) application. Values are shown as the mean ± SEM ( n = 3–5). * P < 0.05 compared with scrambled siRNA in each treatment. e Summarized data showing ATP (10 μmol/L)-induced SOCE in HEK293 cells co-expressed with STIM1 and GFP-tagged full-length TRPP2 (GFP-TRPP2) or TRPP2 without N1 (GFP-TRPP2-∆N1, deletion of 2-111aa in TRPP2) or TRPP2 without N2 (GFP-TRPP2-∆N2, deletion of 112-221aa in TRPP2). Values are shown as mean ± SEM ( n = 5). * P < 0.05 for STIM1 and GFP-TRPP2 or GFP-TRPP2-∆N2 co-expression vs. STIM1 and GFP-TRPP2-∆N1 co-expression

Article Snippet: The primary rabbit antibody against STIM1 was obtained from ProSci.

Techniques: Expressing, Transfection, Dominant Negative Mutation

Journal: Cell Communication and Signaling : CCS

Article Title: TRPP2 and STIM1 form a microdomain to regulate store-operated Ca 2+ entry and blood vessel tone

doi: 10.1186/s12964-020-00560-7

Figure Lengend Snippet: Co-immunoprecipitation and in situ proximity ligation assay (PLA) of TRPP2 and STIM1 in mouse aortic smooth muscle cells. ( a , b ) Immunoblots showing that anti-TRPP2 and anti-STIM1 recognized TRPP2 and STIM1 proteins, and co-immunoprecipitation followed by immunoblots ( a , immunoblot with anti-TRPP2; b , immunoblot with anti-STIM1). Proteins from the mouse aortic smooth muscle cells were immunoprecipitated with indicated antibody (+) or no antibody (−). ( c ) PLA analysis was used to detect the interaction between TRPP2 and STIM1. Representative images were displayed in the presence of anti-STIM1 antibody alone ((a)-(c)), or in the presence of anti-TRPP2 and anti-STIM1 antibodies ((d)-(f)). ((c), (f)) were merged with bright view. Red doted fluorescence showing positive signal. Nuclei were marked by DAPI staining (blue color). Scale bar represents 5 μm. The experiment was repeated 4 times

Article Snippet: The primary rabbit antibody against STIM1 was obtained from ProSci.

Techniques: Immunoprecipitation, In Situ, Proximity Ligation Assay, Western Blot, Fluorescence, Staining

Journal: Cell Communication and Signaling : CCS

Article Title: TRPP2 and STIM1 form a microdomain to regulate store-operated Ca 2+ entry and blood vessel tone

doi: 10.1186/s12964-020-00560-7

Figure Lengend Snippet: Role of TRPP2 in store-operated Ca 2+ entry (SOCE) and STIM1 puncta formation in mouse aortic smooth muscle cells. a Representative traces showing ATP (10 μmol/L)-induced Ca 2+ release in Ca 2+ -free solution and SOCE in the mouse aortic smooth muscle cells transfected with TRPP2, STIM1, both TRPP2 and STIM1, or scrambled siRNAs. b Summary of data showing changes in intracellular Ca 2+ concentration increase in response to extracellular ATP (10 μmol/L) or thapsigargin (TG, 2.5 μmol/L) application in the mouse aortic smooth muscle cells treated with or without 2APB (100 μmol/L) for 10 min in Ca 2+ -free solution. c Summary of data showing changes in intracellular Ca 2+ concentration increase in response to extracellular Ca 2+ (1 mmol/L) application in the mouse aortic smooth muscle cells treated by ATP (10 μmol/L), 2APB (100 μmol/L) + ATP (10 μmol/L) and thapsigargin (TG, 2.5 μmol/L) for 10 min in Ca 2+ -free solution. Values are shown as mean ± SEM ( n = 3–6 experiments). * P < 0.05 for scrambled siRNA vs. TRPP2 or STIM1 or TRPP2 + STIM1 siRNA transfection in each treatment. d Representative images showing STIM1 puncta formation in the mouse aortic smooth muscle cells transfected with TRPP2 siRNA or scrambled siRNA and treated by ATP (10 μmol/L), 2APB (100 μmol/L) + ATP (10 μmol/L) and thapsigargin (TG, 2.5 μmol/L) for 10 min in Ca 2+ -free solution. Scale bar represents 10 μm. The experiment was repeated 4 times

Article Snippet: The primary rabbit antibody against STIM1 was obtained from ProSci.

Techniques: Transfection, Concentration Assay

Journal: Cell Communication and Signaling : CCS

Article Title: TRPP2 and STIM1 form a microdomain to regulate store-operated Ca 2+ entry and blood vessel tone

doi: 10.1186/s12964-020-00560-7

Figure Lengend Snippet: Role of TRPP2 and STIM1 in Ca 2+ release and store-operated Ca 2+ entry (SOCE)-induced mouse aorta contraction. a Representative traces showing phenylephrine (Phe, 10 μmol/L)-induced contraction in Ca 2+ -free solution and extracellular Ca 2+ (2.5 mmol/L) re-addition-induced contraction in mice aortae. b - e Summarized data showing Phe-induced contraction in Ca 2+ -free solution ( b , d ) and extracellular Ca 2+ re-addition ( c , e )-induced contractions in mice aortae, which were transfected with TRPP2 siRNA ( b , c ), SITM1 siRNA ( d , e ) or scrambled siRNA. f - i Summarized data showing endothelin 1 (ET-1, 100 nmom/L)-induced contraction in Ca 2+ -free solution ( f , h ) and extracellular Ca 2+ re-addition ( g , i )-induced contractions in mice aortae transfected with TRPP2 siRNA ( f , g ), SITM1 siRNA ( h , i ) or scrambled siRNA. Values are shown as mean ± SEM ( n = 3–4 mice). * P < 0.05 for scrambled siRNA vs. TRPP2 siRNA or STIM1 siRNA transfection. j Representative traces showing thapsigargin (TG, 2.5 μmol/L)-induced contraction in Ca 2+ -free solution and extracellular Ca 2+ (2.5 mmol/L) re-addition-induced contraction in mice aortae. k - n Summarized data showing TG-induced contraction in Ca 2+ -free solution ( k , m ) and extracellular Ca 2+ re-addition ( l , n )-induced contraction in mice aortae, which were transfected with TRPP2 siRNA ( j - l ), SITM1 siRNA ( m - n ) or scrambled siRNAs. Values are shown as the mean ± SEM ( n = 3–4 mice). * P < 0.05 for TRPP2 siRNA or STIM1 siRNA vs. scrambled siRNA

Article Snippet: The primary rabbit antibody against STIM1 was obtained from ProSci.

Techniques: Transfection

Journal: Cell Communication and Signaling : CCS

Article Title: TRPP2 and STIM1 form a microdomain to regulate store-operated Ca 2+ entry and blood vessel tone

doi: 10.1186/s12964-020-00560-7

Figure Lengend Snippet: Role of TRPP2 and STIM1 in agonist-induced mouse aorta contraction. Phenylephrine (10 μmol/L, a , b , e , f ) and endothelin 1 (100 nmol/L, c , d , g , h ) concentration-dependently induced the contraction of the mice aortae transfected with TRPP2 ( a - d ), STIM1 ( e - h ) or scrambled siRNA. b , d , f , h The mice aortae were pretreated by heparin (1 mg/ml) using a reversible permeabilization loading procedure. Values are shown as mean ± SEM ( n = 4–6 mice). * P < 0.05 for scrambled siRNA vs. TRPP2 or STIM1 siRNA transfection

Article Snippet: The primary rabbit antibody against STIM1 was obtained from ProSci.

Techniques: Concentration Assay, Transfection

Journal: Frontiers in Pharmacology

Article Title: Immune Checkpoint Inhibitors Regulate K + Channel Activity in Cytotoxic T Lymphocytes of Head and Neck Cancer Patients

doi: 10.3389/fphar.2021.742862

Figure Lengend Snippet: αPD-1 treatment increases K + channel activity in HNSCC T cells. (A) Representative current traces of KCa3.1 and Kv1.3 channels recorded in whole-cell mode of voltage-clamp configuration in activated CD8 + PBTs cells from a HNSCC patient in absence or presence of αPD-1 (10 μg/ml, for 6 h). Data are normalized to maximum current at +50 mV recorded using a ramp pulse protocol from −120 mV to +50 mV for 200 ms every 15 s. The holding potential used was −70 mV. (B,C) KCa3.1 (B) and Kv1.3 (C) conductance (G) measured in the absence or presence of αPD-1 (10 μg/ml, 6 h incubation) in CD8 + PBTs of HNSCC patients ( n = 68 cells without pembrolizumab and n = 55 cells with pembrolizumab from 14 patients). (D) Representative current traces of divalent free current (DVF) through CRAC channels recorded in whole-cell mode of voltage-clamp configuration in activated CD8 + PBTs from a HNSCC patient. Data were recorded using a ramp pulse protocol from −100 to +100 mV with at holding potential of +30 mV every 1.5 s. Cells were perfused with 0 mM Ca 2+ solution (1 min) followed by 20 mM Ca 2+ (1 min) and DVF solutions (2 min, see methods) to amplify currents during recordings. (E) Peak DVF current values measured in absence and presence of αPD-1 (10 μg/ml, 6 h incubation) in CD8 + PBTs of HNSCC patients ( n = 34 cells without αPD-1 and n = 31 cells with αPD-1 from 8 patients). The values in panels (B,C) and (E) are represented as box plots: the horizontal line indicates the median; the lower box is the 25 th percentile; the upper box is the 75 th percentile; and the whiskers represent the 10 th and 90 th percentiles. (F) Ion channel expression (KCa3.1, Kv1.3, Orai1 and STIM1) in HNSCC patient T cells after treatment with αPD-1 (10 μg/ml for 6 h). Effect of αPD-1 treatment is shown as ratio of mean fluorescence intensity (MFI, fold change) values of treatment versus control group. Data are represented as scatter plot where each symbol represents an individual patient ( n = 4–5). Horizontal line represents mean values for each group. Data in panels (B,C,E) were analyzed by Mann-Whitney rank sum test.

Article Snippet: Cells were then stained for rabbit anti-human STIM1 (Proteintech) primary antibodies followed by secondary antibodies (Alexa Fluor 594 goat anti-rabbit IgG/Thermo Fisher).

Techniques: Activity Assay, Incubation, Expressing, Fluorescence, Control, MANN-WHITNEY

Journal: Frontiers in Pharmacology

Article Title: Immune Checkpoint Inhibitors Regulate K + Channel Activity in Cytotoxic T Lymphocytes of Head and Neck Cancer Patients

doi: 10.3389/fphar.2021.742862

Figure Lengend Snippet: Differential time-dependent involvement of PI3K and calmodulin on PD-L1 mediated inhibition of KCa3.1 channels. (A,B) Representative recordings of KCa3.1 channels in activated CD8 + PBTs from HDs showing the effect of the PI3K inhibitor LY294002 (10 µM) +/− phosphatidylinositol-3 phosphatase (PI3P) (100 nM) (A) and PD-L1 (PD-L1-Fc, 10 μg/ml) +/− PI3P (100 nM) (B). (C) Summary of the pharmacological modulation of KCa3.1 channels byLY294002 and PI3P in the absence and presence of plate bound PD-L1 in activated CD8 + PBTs of HDs. Cells were activated using anti-CD3/CD28 antibodies for 72 h. Cells were perfused with LY294002 for 15 min followed by patch clamp recordings with and without PI3P, delivered intracellularly via patch pipette ( n = eight to nine cells per group from 3 HDs). All KCa3.1 conductance (G) values are normalized to the average G of the control group (drug-free). (D) KCa3.1 G measured in absence or presence of PD-L1 in activated CD8 + PBTs of HDs. Cells were treated with plate-bound PD-L1 (PD-L1-Fc, 10 μg/ml) and activated using anti-CD3/CD28 antibodies for 120 h PI3P was delivered intracellularly via the patch pipette during the electrophysiological experiments (drug-free control). Cells were held at −70 mV, n = four to five cells per group from one HD. The values in panel (C,D) are represented as box and whiskers plot. The lower and upper bound of the box represent 25 th and 75 th percentiles respectively. Median values are shown as horizontal line. The lower and upper error bars represents 10 th and 90 th percentile respectively. (E) Percentage change in mean fluorescence intensity (MFI) of ion channels (Kv1.3, KCa3.1, Orai1, Stim1) and Calmodulin (CaM) measured using flow cytometry. Each dot represents an individual HD and the horizontal black line represents the mean value. Data in panel (C) were analyzed by One Way ANOVA ( p < 0.001) followed by Holm-Sidak’s post hoc analysis. Data in (D) were analyzed by One way ANOVA followed by Holm-Sidak’s post hoc analysis.

Article Snippet: Cells were then stained for rabbit anti-human STIM1 (Proteintech) primary antibodies followed by secondary antibodies (Alexa Fluor 594 goat anti-rabbit IgG/Thermo Fisher).

Techniques: Inhibition, Patch Clamp, Transferring, Control, Fluorescence, Flow Cytometry

Journal: American Journal of Respiratory Cell and Molecular Biology

Article Title: Arachidonate-Regulated Ca2+Influx in Human Airway Smooth Muscle

doi: 10.1165/rcmb.2013-0144oc

Figure Lengend Snippet: Figure 3. Mechanisms of AA-induced Ca21 influx. (A) Role of receptor-operated Ca21 entry in AA-induced Ca21 influx. In the presence of the receptor- operated channel inhibitor, SKF96365 (10 mM), AA-induced [Ca21]i responses were not significantly altered, suggesting that the observed Ca21 influx was not via this mechanism. (B) Role of store-operated Ca21 entry (SOCE) in AA-induced Ca21 influx. La31 (1 mM; which inhibits SOCE) slightly decreased the amplitude of AA-induced [Ca21]i oscillations, but did not affect oscillation frequency. In contrast, 1 mM La31 completely eliminated [Ca21]i responses, suggesting that the observed influx is unlikely to be SOCE, especially because sarcoplasmic reticulum (SR) stores were not depleted during these experiments. (C) Role of stromal interaction molecule (STIM) 1 in AA-induced [Ca21]i responses. In the presence of STIM1 small interfering RNA (siRNA), AA-induced elevations in [Ca21]i levels were reduced and [Ca21]i oscillations decreased compared with lipofectamine (vehicle) control, whereas negative siRNA was without effect. These data suggest that STIM1 modulates AA-induced [Ca21]i responses. (D) Role of caveolae/caveolin-1 in AA-induced [Ca21]i responses. Suppression of caveolin-1 using siRNA eliminated AA-induced [Ca21]i oscillations, whereas lipofectamine or negative siRNA were without effect. These data suggest that caveolin-1 modulates AA-induced Ca21 influx in human ASM. (E) Role of Orai3 on AA-induced [Ca21]i responses. Orai3 siRNA reduced the frequency of AA-induced [Ca21]i oscillations and slightly altered the amplitudes of [Ca21]I compared with lipofectamine or negative siRNA. These data suggest a role for Orai 3 in modulating AA-induced [Ca21]i responses. (F) Western analysis confirmed siRNA knockdown of STIM1, caveolin-1, and Orai3, and lack of effect of negative siRNA. See Table 2 for summary and statistics of Ca21. *Significant siRNA effect (P , 0.05). Cav1, caveolin-1; Con, control; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; Neg, negative.

Article Snippet: Tissue culture reagents (DMEM F/12, FBS) were from Invitrogen, zileuton from R&D Systems Inc. (Minneapolis, MN), xestospongin C (XeC) and ryanodine from Millipore Inc. (Billerica, MA), caveolin-1 and secondary antibodies from Santa Cruz Biotechnology, Inc. (Dallas, TX), STIM1 antibody from Novus (Littleton, CO), and Orai3 antibody from Abcam (Cambridge, MA).

Techniques: Small Interfering RNA, Control, Western Blot, Knockdown

Journal: American Journal of Respiratory Cell and Molecular Biology

Article Title: Arachidonate-Regulated Ca2+Influx in Human Airway Smooth Muscle

doi: 10.1165/rcmb.2013-0144oc

Figure Lengend Snippet: Figure 7. Proposed model of ARC channels in human ASM. Regardless of the source (agonist- or cytokine-induced production), AA activates plasma membrane ARC channels that are within caveolae. Such activation is facilitated by plasma membrane STIM1, and involves repeated SR Ca21 release. IP3R, IP3 receptor; RyR, ryanodine receptor; SERCA, sarco/endoplasmic reticulum calcium ATPase.

Article Snippet: Tissue culture reagents (DMEM F/12, FBS) were from Invitrogen, zileuton from R&D Systems Inc. (Minneapolis, MN), xestospongin C (XeC) and ryanodine from Millipore Inc. (Billerica, MA), caveolin-1 and secondary antibodies from Santa Cruz Biotechnology, Inc. (Dallas, TX), STIM1 antibody from Novus (Littleton, CO), and Orai3 antibody from Abcam (Cambridge, MA).

Techniques: Clinical Proteomics, Membrane, Activation Assay

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: TRPV6-mediated Store-operated Ca 2+ Entry Participates in Pancreatic Acinar Cell Injury During Acute Pancreatitis

doi: 10.1016/j.jcmgh.2025.101620

Figure Lengend Snippet: TRPV6 and STIM1 interact in PACs. ( A ) Representative IHC staining images and quantification of STIM1 staining performed on mouse pancreas tissue (n = 4 sections from 3 mice per group). ( B ) Representative IHC staining images and quantification of STIM1 performed on human pancreas tissue (n = 3 sections from 3 patients per group). ( C and D ) Representative images ( C ) and quantification ( D ) of TRPV6 and STIM1 colocalization in PACs obtained by structured illumination microscopy (n = 7–8 regions of interest [ROIs] from 3 independent experiments per group). ( E and F ) Blots ( E ) and quantification ( F ) of Co-IP experiments performed by immunoprecipitating TRPV6 or STIM1 in freshly isolated PAC lysates (n = 3 blots from 3 independent experiments per group). ( G and H ) Representative whole-cell TRPV6 current traces ( G ) and quantification ( H ) of current density at −74 mV under different holding potentials in TRPV6- and STIM1-transfected HEK293T cells (n = 8–10 cells from 4 independent experiments per group). Data are the mean ± SEM; n.s., non-significance, ∗ P < .05, ∗∗ P < .01.

Article Snippet: Then, slides were incubated for 1 hour at room temperature using a primary antibody of STIM1 dilution of 1:100 (NB110-55300, Novus), and then for another hour with biotinylated secondary antibodies (1:1000).

Techniques: Immunohistochemistry, Staining, Microscopy, Co-Immunoprecipitation Assay, Isolation, Transfection

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: TRPV6-mediated Store-operated Ca 2+ Entry Participates in Pancreatic Acinar Cell Injury During Acute Pancreatitis

doi: 10.1016/j.jcmgh.2025.101620

Figure Lengend Snippet: Pharmacological TRPV6 inhibition ameliorates CER-induced AP. ( A ) Schematic of the in vivo experiments in AP and control mice. ( B ) Line chart displaying the effect of SORC27 on von Frey filament (VFF) test withdrawal thresholds at 0.5, 1.5, 135 2.5, and 3.5 hours following the last PBS injection (n = 6 mice per group). ( C ) Serum amylase, pancreatic MPO, and trypsin activity (n = 6 mice per group). ( D and E ) Representative images ( D ) and histological scores ( E ) of pancreas tissue sections (n = 6 mice per group). ( F ) Line chart displaying the effect of SORC27 on VFF test withdrawal thresholds at 0.5, 1.5, 2.5, and 3.5 hours following the last CER injection (n = 6 mice per group). ( G ) Serum amylase, pancreatic MPO, and trypsin activity (n = 6 samples per group). ( H ) Representative H&E-stained images and histological scores of pancreas tissue sections (n = 6 sections from 6 mice per group). ( I and J ) Representative images of pancreas ( I ) and lung ( J ) tissue section staining for F4/80 + macrophages ( left ) and quantitative results ( right ) (n = 6 sections from 6 mice per group). ( K ) Representative images of lung injury from CER-AP (CER), low (SOR-L) and high (SOR-H) -dose-SORC27 treated mice AP modeling. ( L ) Western blot (WB) images and quantification, and gene expression ( M ) of pancreatic STIM1 after SORC27 treatment (n = 4–6 mice per group). Data are the mean ± SEM; n.s., non-significance, ∗ P < .05, ∗∗ P < .01, ∗∗∗ P < .001.

Article Snippet: Then, slides were incubated for 1 hour at room temperature using a primary antibody of STIM1 dilution of 1:100 (NB110-55300, Novus), and then for another hour with biotinylated secondary antibodies (1:1000).

Techniques: Inhibition, In Vivo, Control, Injection, Activity Assay, Staining, Western Blot, Gene Expression

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: TRPV6-mediated Store-operated Ca 2+ Entry Participates in Pancreatic Acinar Cell Injury During Acute Pancreatitis

doi: 10.1016/j.jcmgh.2025.101620

Figure Lengend Snippet: Pancreatic Trpv6 knockdown alleviates CER-induced AP. ( A ) Schematic of the in vivo experiments in si Trpv6 and siNC mice. ( B ) Pancreatic Trpv6 gene expression (n = 5–6 mice per group). ( C ) Line chart displaying the effect of SORC27 on VFF test withdrawal thresholds at 0.5, 1.5, 2.5, and 3.5 hours following the last PBS injection (n = 6–7 mice per group). ( D ) Serum amylase, pancreatic MPO, and trypsin activity (n = 4–6 mice per group). ( E and F ) Representative images ( E ) and histological scores ( F ) of pancreas tissue sections (n = 5–6 mice per group). ( G ) Line chart indicating VFF withdrawal thresholds observed at 0.5, 1.5, 2.5, and 3.5 hours following the last CER injection (n = 8, 12 mice per group). ( H ) Serum amylase, pancreatic MPO, and trypsin activity (n = 8–12 samples per group). ( I and J ) Representative H&E-stained images ( I ) and histological scores ( J ) of pancreas tissue sections (n = 77–88 sections from at least 8 mice per group). ( K and L ) Representative images and quantitative results of pancreas ( K ) and lung tissue ( L ) F4/80 + staining for macrophages (n = 3 sections from 3 mice per group). ( M ) Representative images of lung injury. ( N ) Immunoblotting images and quantification of STIM1 levels in pancreas tissue (n = 4 samples per group). ( O ) Representative IHC staining images and quantitative results of STIM1 levels in pancreas tissue (n = 3 sections from 3 mice per group). Data are the mean ± SEM; n.s., non-significance, ∗ P < .05, ∗∗ P < .01, ∗∗∗ P < .001.

Article Snippet: Then, slides were incubated for 1 hour at room temperature using a primary antibody of STIM1 dilution of 1:100 (NB110-55300, Novus), and then for another hour with biotinylated secondary antibodies (1:1000).

Techniques: Knockdown, In Vivo, Gene Expression, Injection, Activity Assay, Staining, Western Blot, Immunohistochemistry