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Procell Inc panc 1 cell lines
Panc 1 Cell Lines, supplied by Procell Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 86 stars, based on 1 article reviews

panc 1 cell lines - by Bioz Stars, 2026-06

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( A ) Mitochondrial oxygen consumption rate (OCR) and ( B ) extracellular acidification rate (ECAR) in KRAS* cells 4 h after 10p IRE or 20p IRE (400 V). ( C ) Effect of IRE on mitochondrial ATP production. Data are normalized to cell number at the end of the assay (pmol/min/1000 cells). Data are expressed as mean ± SD (n=6/group). *p<0.05, **p<0.01, ***p<0.001 (one-way ANOVA). ( D ) Representative confocal fluorescence microscopic images of cytoplasmic dsDNA and corresponding quantification of fluorescence (FL) signal intensity in cytosol and nuclei. PANC-1 cells were stained with PicoGreen for dsDNA 24 h after IRE (400 V, 20 pulses). Untreated cells were used as a control. The negative control was cells not stained with PicoGreen. Data are expressed as mean ± SD (n=4). *p<0.05, *p<0.01, (Student’s t test). ( E ) Analysis of dsDNA from IRE-treated KRAS* tumors using the Qubit dsDNA HS assay kits. KRAS* tumor tissues were collected on day 1 and day 4 after IRE (1200 V, 99 pulses) and processed for dsDNA assays. *p<0.05, ****p<0.0001 (Student’s t test). ( F ) Real-time PCR analysis of the interferon-stimulated genes Irf1 , Cxcl10 , and Isg15 downstream of the cGAS-STING pathway. KRAS* cells were collected 4 h after IRE (400 V, 20 pulses). *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001 (one-way ANOVA). Data are expressed as mean ± SD (n=3/group).

Journal: bioRxiv

Article Title: The abscopal effect of IRE combined with anti–PD-1 achieves local ablation and systemic control of PDAC

doi: 10.64898/2026.05.04.722535

Figure Lengend Snippet: ( A ) Mitochondrial oxygen consumption rate (OCR) and ( B ) extracellular acidification rate (ECAR) in KRAS* cells 4 h after 10p IRE or 20p IRE (400 V). ( C ) Effect of IRE on mitochondrial ATP production. Data are normalized to cell number at the end of the assay (pmol/min/1000 cells). Data are expressed as mean ± SD (n=6/group). *p<0.05, **p<0.01, ***p<0.001 (one-way ANOVA). ( D ) Representative confocal fluorescence microscopic images of cytoplasmic dsDNA and corresponding quantification of fluorescence (FL) signal intensity in cytosol and nuclei. PANC-1 cells were stained with PicoGreen for dsDNA 24 h after IRE (400 V, 20 pulses). Untreated cells were used as a control. The negative control was cells not stained with PicoGreen. Data are expressed as mean ± SD (n=4). *p<0.05, *p<0.01, (Student’s t test). ( E ) Analysis of dsDNA from IRE-treated KRAS* tumors using the Qubit dsDNA HS assay kits. KRAS* tumor tissues were collected on day 1 and day 4 after IRE (1200 V, 99 pulses) and processed for dsDNA assays. *p<0.05, ****p<0.0001 (Student’s t test). ( F ) Real-time PCR analysis of the interferon-stimulated genes Irf1 , Cxcl10 , and Isg15 downstream of the cGAS-STING pathway. KRAS* cells were collected 4 h after IRE (400 V, 20 pulses). *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001 (one-way ANOVA). Data are expressed as mean ± SD (n=3/group).

Article Snippet: The human pancreatic cancer cell line PANC-1 was purchased from American Type Culture Collection.

Techniques: Fluorescence, Staining, Control, Negative Control, Real-time Polymerase Chain Reaction

Journal: bioRxiv

Article Title: Pancreatic cancer extracellular vesicles carry a time-of-day-regulated miRNA cargo that disrupts the skeletal muscle clock and bioenergetics

doi: 10.64898/2026.05.03.722338

Figure Lengend Snippet: (A) Clinical and molecular characterization of six pancreatic cancer patients (n=6) stratified by unsupervised hierarchical clustering of serum sEV miRNA expression profiles. Three clusters were resolved: Cluster 1 (P1, P3, P5; red), Cluster 2 (P4; blue), and Cluster 3 (P2, P6; green). Diagnoses are indicated above each patient identifier: PDAC, pancreatic ductal adenocarcinoma; NET, neuroendocrine tumor; Mesoth., malignant mesothelioma of the pancreas. Clinical parameters shown include age at diagnosis (dot plot, years), number of prior treatment lines (bar chart, top right), and comorbidity burden (bar chart, bottom right). The Jaccard similarity index between within-cluster patient pairs (CPM>0) is plotted for Clusters 1 and 3, demonstrating high intra-cluster miRNA-profile similarity (range 0.588–0.644). A Venn diagram illustrates the overlap between miRNAs detected in patient serum sEVs (T=428) and in PANC-1 sEVs across 9 time-points (T=1,426); 359 miRNAs are shared between the two datasets. Bar charts indicate the percentage of cluster-specific patient miRNAs detectable in the PANC-1 sEV secretome: Cluster 1, 84.6%; Cluster 2, 84.2%; Cluster 3, 83.9%. ( B ) UpSet plot (top) displaying the combinatorial overlap of miRNAs detected in each patient’s serum sEVs. Vertical bars indicate the number of miRNAs shared by each intersecting patient combination defined in the connected dot matrix below; horizontal bars on the right show the total number of miRNAs detected per patient. Below the UpSet plot, a heatmap shows expression levels (log2(CPM+1)) of the 11 candidate miRNAs functionally validated in this study across all six patients. Rows are ordered by hierarchical clustering of miRNA expression profiles; columns are arranged by patient cluster assignment (Cluster 1, red; Cluster 2, blue; Cluster 3, green). Color scale ranges from blue (low expression) to dark red (high expression). ( C ) Frequency of the 11 candidate miRNAs among the top 50 most highly expressed miRNAs in pancreatic cancer tumor specimens. For each of 495 tumor samples from the GDC data portal (TCGA-PAAD, CPTAC-3, and HCMI-CMDC), miRNAs were ranked by RPM, and a given miRNA was classified as highly expressed if any of its corresponding precursors ranked within the top 50. Bars show the percentage of samples meeting this criterion; absolute sample counts are given in parentheses. ( D ) Proposed mechanistic model. Within the PDAC tumor cell, a dysregulated clock (BMAL1↓, altered rhythmic output) drives miRNA sorting into EVs during biogenesis; miR-27b-3p is uniquely (among those tested) and rhythmically packaged, while other sEV miRNAs are loaded constitutively. Secreted sEVs exhibit variable particle number with stable size, and these miRNAs are detectable in patient serum (84– 85% overlap with PANC-1 sEV miRNAs). In the recipient skeletal muscle cell, the full sEV miRNA cargo collectively disrupts the circadian clock, altering period, phase, and amplitude. In parallel, individual miRNAs reprogram bioenergetics along four non-redundant trajectories: Energetic (miR-27b-3p + others; ↑OCR, aerobic/oxidative, ↑spare respiratory capacity); High metabolic (miR-191-5p + others; ↑OCR, ↑ECAR, high respiratory capacity); Quiescent (others; ↓OCR, ↓ECAR, low metabolic capacity); and Glycolytic (miR-183-5p + others; ↓OCR, ↑ECAR, glycolytic drift). The integration of circadian disruption, bioenergetic reprogramming, and proteostatic dysregulation collectively drives muscle-cell atrophy in vitro ; whether and how these molecularly distinct insults produce a systemic cachexia phenotype in vivo remains to be determined (gray box).

Article Snippet: The human pancreatic cancer cell line PANC-1, the murine fibroblast cell line NIH3T3, and the murine myoblast cell line C2C12 were purchased from American Type Culture Collection (ATCC, Manassas, VA).

Techniques: Expressing, Biomarker Discovery, Disruption, In Vitro, In Vivo

(A) Bioluminescence recording, ( B ) period analysis, and ( C ) phase and amplitude analysis of U2OS BMAL1 :Luc reporter cells treated with PANC-1 CM at 12.5%, 25%, 50%, and 100% of the recording media. ( D ) Bioluminescence recording, ( E ) period analysis, and ( F ) phase and amplitude analysis of NIH3T3 Bmal1 :Luc reporter cells treated with PANC-1 CM at the same concentrations. For all bioluminescence experiments, at least three complete oscillations were included in the period estimation, excluding the first 24 h. Mean ± SD of relative mRNA expression of NIH3T3 core clock genes Bmal1 ( G ), Per2 ( H ), and Cry2 ( I ) measured over 36 h in response to PANC-1 CM. Relative mRNA levels of core clock genes in synchronized C2C12 myotubes over 32 h following treatment with PANC-1 CM: ( J ) Bmal1 , ( K ) Per2 , and ( L ) Cry2 . Cosine curves were fit for visualization purposes only; solid lines represent rhythmic oscillations (p<0.05) detected by MetaCycle, while dashed lines indicate loss of statistical rhythmicity (Suppl. Table 1). ( G–L ) Black: Control; ( G–I ) Red: PANC-1 CM; ( J–L ) Blue: PANC-1 CM. ( M ) Schematic representation and representative images of mature C2C12 myotube atrophy in response to NIH3T3 or PANC-1 released factors using a Transwell co-culture system; three measurements per myotube (yellow arrows) were used to quantify shortening. ( N ) Quantification of normalized myotube diameter under NIH3T3 vs PANC-1 co-culture, normalized to NIH3T3 co-culture control. One-way ANOVA: ( B ) p=0.0009, ( E ) p=0.0087. ( B, E ) Dunnett’s post-hoc test: *p<0.05; **p<0.01; ***p<0.001. (N) Student’s t-test: ***p<0.001.

Journal: bioRxiv

Article Title: Pancreatic cancer extracellular vesicles carry a time-of-day-regulated miRNA cargo that disrupts the skeletal muscle clock and bioenergetics

doi: 10.64898/2026.05.03.722338

Figure Lengend Snippet: (A) Bioluminescence recording, ( B ) period analysis, and ( C ) phase and amplitude analysis of U2OS BMAL1 :Luc reporter cells treated with PANC-1 CM at 12.5%, 25%, 50%, and 100% of the recording media. ( D ) Bioluminescence recording, ( E ) period analysis, and ( F ) phase and amplitude analysis of NIH3T3 Bmal1 :Luc reporter cells treated with PANC-1 CM at the same concentrations. For all bioluminescence experiments, at least three complete oscillations were included in the period estimation, excluding the first 24 h. Mean ± SD of relative mRNA expression of NIH3T3 core clock genes Bmal1 ( G ), Per2 ( H ), and Cry2 ( I ) measured over 36 h in response to PANC-1 CM. Relative mRNA levels of core clock genes in synchronized C2C12 myotubes over 32 h following treatment with PANC-1 CM: ( J ) Bmal1 , ( K ) Per2 , and ( L ) Cry2 . Cosine curves were fit for visualization purposes only; solid lines represent rhythmic oscillations (p<0.05) detected by MetaCycle, while dashed lines indicate loss of statistical rhythmicity (Suppl. Table 1). ( G–L ) Black: Control; ( G–I ) Red: PANC-1 CM; ( J–L ) Blue: PANC-1 CM. ( M ) Schematic representation and representative images of mature C2C12 myotube atrophy in response to NIH3T3 or PANC-1 released factors using a Transwell co-culture system; three measurements per myotube (yellow arrows) were used to quantify shortening. ( N ) Quantification of normalized myotube diameter under NIH3T3 vs PANC-1 co-culture, normalized to NIH3T3 co-culture control. One-way ANOVA: ( B ) p=0.0009, ( E ) p=0.0087. ( B, E ) Dunnett’s post-hoc test: *p<0.05; **p<0.01; ***p<0.001. (N) Student’s t-test: ***p<0.001.

Techniques: Expressing, Control, Co-Culture Assay

(A) Concentration of sEVs and ( B ) mean particle size of sEVs isolated from CM of circadian-synchronized PANC-1 cells collected every 4 h for 32 h (n=3 biological replicates per time-point). ( C ) Heatmap displaying z-score-normalized log2(CPM+1) expression of 1,426 miRNAs detected in PANC-1 sEVs collected at 9 consecutive time-points. Hierarchical clustering used Ward.D2 linkage with Euclidean distance. Rows represent individual miRNAs; columns represent time-points. Color scale: red = above-mean expression; blue = below-mean expression. The dendrogram on the left shows hierarchical relationships among miRNAs. Five clusters were identified with distinct temporal expression patterns: C1 (Red, n=352), C2 (Blue, n=200), C3 (Green, n=426), C4 (Orange, n=181), and C5 (Purple, n=267). The optimal number of clusters (k=5) was determined by combining the elbow method and silhouette analysis (mean silhouette score = 0.187). ( D ) Dot plots showing KEGG enrichment for miRNA clusters C1–C5. Target genes were predicted using miRDB (score ≥80) for each cluster’s miRNA set. The x-axis represents Gene Ratio; dot size represents the number of genes enriched in each pathway (Gene count). Dot color (C1: Red, C2: Blue, C3: Green, C4: Orange, C5: Purple) indicates statistical significance as −log10(padj); only the top 20 significant terms are shown. ( E ) Representative bioluminescence curves of synchronized NIH3T3 Bmal1 :Luc cells transfected with 25 nM of miRNA mimics: hsa-miR-27b-3p, hsa-miR-99b-5p, hsa-miR-127-3p, hsa-miR-191-5p, hsa-miR-615-3p, or negative-transfection control (cel-miR-67). ( F ) Period, ( G ) phase, and ( H ) amplitude analyses of NIH3T3 Bmal1 :Luc cells transfected with the miRNAs in panel E. ( I ) Representative bioluminescence curves of synchronized NIH3T3 Bmal1 :Luc cells transfected with 25 nM of: hsa-let-7f-5p, hsa-miR-10a-5p, hsa-miR-26a-5p, hsa-miR-30c-5p, hsa-miR-92a-3p, hsa-miR-183-5p, or negative-transfection control (cel-miR-67). ( J ) Period, ( K ) phase, and ( L ) amplitude analyses of NIH3T3 Bmal1 :Luc cells transfected with the miRNAs in panel I. All data are presented as mean ± SD. Circadian period, phase, and amplitude were calculated using the FFT-NLLS algorithm in BioDare2 ( https://biodare2.ed.ac.uk/ ). Statistical analyses of period, phase, and amplitude were performed by one-way ANOVA with Dunnett’s post-hoc test in GraphPad Prism. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001; n.s., not significant.

Journal: bioRxiv

Article Title: Pancreatic cancer extracellular vesicles carry a time-of-day-regulated miRNA cargo that disrupts the skeletal muscle clock and bioenergetics

doi: 10.64898/2026.05.03.722338

Figure Lengend Snippet: (A) Concentration of sEVs and ( B ) mean particle size of sEVs isolated from CM of circadian-synchronized PANC-1 cells collected every 4 h for 32 h (n=3 biological replicates per time-point). ( C ) Heatmap displaying z-score-normalized log2(CPM+1) expression of 1,426 miRNAs detected in PANC-1 sEVs collected at 9 consecutive time-points. Hierarchical clustering used Ward.D2 linkage with Euclidean distance. Rows represent individual miRNAs; columns represent time-points. Color scale: red = above-mean expression; blue = below-mean expression. The dendrogram on the left shows hierarchical relationships among miRNAs. Five clusters were identified with distinct temporal expression patterns: C1 (Red, n=352), C2 (Blue, n=200), C3 (Green, n=426), C4 (Orange, n=181), and C5 (Purple, n=267). The optimal number of clusters (k=5) was determined by combining the elbow method and silhouette analysis (mean silhouette score = 0.187). ( D ) Dot plots showing KEGG enrichment for miRNA clusters C1–C5. Target genes were predicted using miRDB (score ≥80) for each cluster’s miRNA set. The x-axis represents Gene Ratio; dot size represents the number of genes enriched in each pathway (Gene count). Dot color (C1: Red, C2: Blue, C3: Green, C4: Orange, C5: Purple) indicates statistical significance as −log10(padj); only the top 20 significant terms are shown. ( E ) Representative bioluminescence curves of synchronized NIH3T3 Bmal1 :Luc cells transfected with 25 nM of miRNA mimics: hsa-miR-27b-3p, hsa-miR-99b-5p, hsa-miR-127-3p, hsa-miR-191-5p, hsa-miR-615-3p, or negative-transfection control (cel-miR-67). ( F ) Period, ( G ) phase, and ( H ) amplitude analyses of NIH3T3 Bmal1 :Luc cells transfected with the miRNAs in panel E. ( I ) Representative bioluminescence curves of synchronized NIH3T3 Bmal1 :Luc cells transfected with 25 nM of: hsa-let-7f-5p, hsa-miR-10a-5p, hsa-miR-26a-5p, hsa-miR-30c-5p, hsa-miR-92a-3p, hsa-miR-183-5p, or negative-transfection control (cel-miR-67). ( J ) Period, ( K ) phase, and ( L ) amplitude analyses of NIH3T3 Bmal1 :Luc cells transfected with the miRNAs in panel I. All data are presented as mean ± SD. Circadian period, phase, and amplitude were calculated using the FFT-NLLS algorithm in BioDare2 ( https://biodare2.ed.ac.uk/ ). Statistical analyses of period, phase, and amplitude were performed by one-way ANOVA with Dunnett’s post-hoc test in GraphPad Prism. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001; n.s., not significant.

Techniques: Concentration Assay, Isolation, Expressing, Transfection, Control

(A) Top 35 miRNAs by mean expression in PANC-1-derived sEVs. Bar plot of mean log2 CPM across 9 time-points (4–36 h). Red bars: miRNAs selected from the top 35 to be tested in the BMAL1 :Luc reporter and atrophy assays; grey bars: remaining top-35 miRNAs. miRNAs are ranked in descending order of EV expression. ( B ) GO Biological Process enrichment of the experimentally validated targets (miRTarBase) of the 11 selected miRNAs. Terms are grouped into functional categories. Dot size represents the number of validated target genes associated with each term; dot color indicates Gene Ratio (proportion of input genes annotated to the term), from light pink (low) to dark red (high). Analysis performed with clusterProfiler. ( C , top panel) Normalized C2C12 myotube diameter at 0, 24, and 48 h post-transfection with miR-27b-3p, miR-615-3p, miR-191-5p, miR-127-3p, miR-99b-5p, or negative-transfection control (NTC); dexamethasone (Dexa) included as positive control. ( C , lower panel) Normalized C2C12 myotube diameter at the same time-points after transfection with hsa-let-7f-5p, miR-183-5p, miR-92a-3p, miR-30c-5p, miR-26a-5p, miR-10a-5p, NTC, or Dexa. ( D ) Oxygen consumption rate (OCR; top), resting-phenotype plot of basal OCR vs ECAR (middle), and metabolic-capacity plot of maximal OCR vs ECAR following FCCP (lower) for mature C2C12 myotubes 48 h post-transfection with miR-27b-3p, miR-615-3p, miR-191-5p, or NTC (Control). ( E ) Same panels for myotubes transfected with miR-127-3p, miR-99b-5p, miR-183-5p, or NTC. Sequential injections of oligomycin, FCCP, and rotenone/antimycin A were used to dissect mitochondrial respiration. Data are presented as mean ± SEM. ( C ) Measurements were taken from at least 5 random fields per well in N=3 wells; statistical analysis used 2-way ANOVA with Dunnett’s post-hoc correction: *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001.

Journal: bioRxiv

Article Title: Pancreatic cancer extracellular vesicles carry a time-of-day-regulated miRNA cargo that disrupts the skeletal muscle clock and bioenergetics

doi: 10.64898/2026.05.03.722338

Figure Lengend Snippet: (A) Top 35 miRNAs by mean expression in PANC-1-derived sEVs. Bar plot of mean log2 CPM across 9 time-points (4–36 h). Red bars: miRNAs selected from the top 35 to be tested in the BMAL1 :Luc reporter and atrophy assays; grey bars: remaining top-35 miRNAs. miRNAs are ranked in descending order of EV expression. ( B ) GO Biological Process enrichment of the experimentally validated targets (miRTarBase) of the 11 selected miRNAs. Terms are grouped into functional categories. Dot size represents the number of validated target genes associated with each term; dot color indicates Gene Ratio (proportion of input genes annotated to the term), from light pink (low) to dark red (high). Analysis performed with clusterProfiler. ( C , top panel) Normalized C2C12 myotube diameter at 0, 24, and 48 h post-transfection with miR-27b-3p, miR-615-3p, miR-191-5p, miR-127-3p, miR-99b-5p, or negative-transfection control (NTC); dexamethasone (Dexa) included as positive control. ( C , lower panel) Normalized C2C12 myotube diameter at the same time-points after transfection with hsa-let-7f-5p, miR-183-5p, miR-92a-3p, miR-30c-5p, miR-26a-5p, miR-10a-5p, NTC, or Dexa. ( D ) Oxygen consumption rate (OCR; top), resting-phenotype plot of basal OCR vs ECAR (middle), and metabolic-capacity plot of maximal OCR vs ECAR following FCCP (lower) for mature C2C12 myotubes 48 h post-transfection with miR-27b-3p, miR-615-3p, miR-191-5p, or NTC (Control). ( E ) Same panels for myotubes transfected with miR-127-3p, miR-99b-5p, miR-183-5p, or NTC. Sequential injections of oligomycin, FCCP, and rotenone/antimycin A were used to dissect mitochondrial respiration. Data are presented as mean ± SEM. ( C ) Measurements were taken from at least 5 random fields per well in N=3 wells; statistical analysis used 2-way ANOVA with Dunnett’s post-hoc correction: *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001.

Techniques: Expressing, Derivative Assay, Functional Assay, Transfection, Control, Positive Control

( A ) Representative western blot for Calnexin, TSG101, CD9, and ApoA2 in PANC-1 whole-cell lysates (WCL) and isolated sEVs. ( B ) Negative-stain electron microscopy of PANC-1 sEVs resuspended in PBS, showing the characteristic cup-shaped morphology typical of EVs. Black arrows: representative sEVs displaying cup-shaped morphology. ( C ) Calcein acetoxymethyl-ester (Calcein-AM) staining of PANC-1 sEVs showing distinct fluorescent puncta indicative of intact vesicular membranes. (D) Representative fluorescent NTA (fNTA) of PANC-1 sEVs incubated with a fluorescently conjugated antibody against CD9. CMDR: CellMask Deep Red; Scatter: total particles; CD9-AF488: CD9-conjugated fluorescent-antibody-labeled particles. ( E ) Period, phase, and amplitude analyses of BMAL1:Luc oscillations in U2OS reporter cells treated with sEVs isolated from PANC-1 CM. ( F ) Period, phase, and amplitude analyses of Bmal1:Luc oscillations in NIH3T3 reporter cells treated with sEVs isolated from PANC-1 CM. For all bioluminescence experiments, at least three complete oscillations were included in the period estimation, excluding the first 24 h. Student’s t-test: ( E ) period p<0.01, amplitude p=0.0074; (F) period p<0.05, amplitude p<0.0001. ( G ) Myotube atrophy in response to PANC-1 sEVs over 72 h. Measurements were taken from at least 10 random fields in n=3 wells and normalized to control (without sEVs) for each time-point. Dexamethasone (Dexa) is included as a positive control for atrophy. ****p<0.0001; ***p<0.001.

Journal: bioRxiv

Article Title: Pancreatic cancer extracellular vesicles carry a time-of-day-regulated miRNA cargo that disrupts the skeletal muscle clock and bioenergetics

doi: 10.64898/2026.05.03.722338

Figure Lengend Snippet: ( A ) Representative western blot for Calnexin, TSG101, CD9, and ApoA2 in PANC-1 whole-cell lysates (WCL) and isolated sEVs. ( B ) Negative-stain electron microscopy of PANC-1 sEVs resuspended in PBS, showing the characteristic cup-shaped morphology typical of EVs. Black arrows: representative sEVs displaying cup-shaped morphology. ( C ) Calcein acetoxymethyl-ester (Calcein-AM) staining of PANC-1 sEVs showing distinct fluorescent puncta indicative of intact vesicular membranes. (D) Representative fluorescent NTA (fNTA) of PANC-1 sEVs incubated with a fluorescently conjugated antibody against CD9. CMDR: CellMask Deep Red; Scatter: total particles; CD9-AF488: CD9-conjugated fluorescent-antibody-labeled particles. ( E ) Period, phase, and amplitude analyses of BMAL1:Luc oscillations in U2OS reporter cells treated with sEVs isolated from PANC-1 CM. ( F ) Period, phase, and amplitude analyses of Bmal1:Luc oscillations in NIH3T3 reporter cells treated with sEVs isolated from PANC-1 CM. For all bioluminescence experiments, at least three complete oscillations were included in the period estimation, excluding the first 24 h. Student’s t-test: ( E ) period p<0.01, amplitude p=0.0074; (F) period p<0.05, amplitude p<0.0001. ( G ) Myotube atrophy in response to PANC-1 sEVs over 72 h. Measurements were taken from at least 10 random fields in n=3 wells and normalized to control (without sEVs) for each time-point. Dexamethasone (Dexa) is included as a positive control for atrophy. ****p<0.0001; ***p<0.001.

Techniques: Western Blot, Isolation, Staining, Electron Microscopy, Incubation, Labeling, Control, Positive Control

Verification of protein tyrosine phosphatase kappa (PTPRK) knockdown in pancreatic cancer cell lines. (A) QPCR results show the PTPRK expression in control cell line PANC-1 pEF and PTPRK knockdown cell line PANC-1 PTPRK kd . (B) PTPRK expression in CFPAC-1 pEF and CFPAC-1 PTPRK kd cell lines. (C) Western blot results show the PTPRK protein expression in both PANC-1 and CFPAC-1 cell lines with PTPRK nockdown. * p <0.05, ** p <0.01, *** p <0.001.

Journal: Cancer Diagnosis & Prognosis

Article Title: Elevated Protein Tyrosine Phosphatase Kappa Expression Is Associated With Disease Progression and Poor Prognosis of Pancreatic Cancer

doi: 10.21873/cdp.10560

Figure Lengend Snippet: Verification of protein tyrosine phosphatase kappa (PTPRK) knockdown in pancreatic cancer cell lines. (A) QPCR results show the PTPRK expression in control cell line PANC-1 pEF and PTPRK knockdown cell line PANC-1 PTPRK kd . (B) PTPRK expression in CFPAC-1 pEF and CFPAC-1 PTPRK kd cell lines. (C) Western blot results show the PTPRK protein expression in both PANC-1 and CFPAC-1 cell lines with PTPRK nockdown. * p <0.05, ** p <0.01, *** p <0.001.

Article Snippet: Human pancreatic cancer cell lines PANC-1 and CFPAC-1 were purchased from ATCC (American Type Culture Collection, Manassas, VA, USA).

Techniques: Knockdown, Expressing, Control, Western Blot

Protein tyrosine phosphatase kappa (PTPRK) and cell proliferation. (A, B) A proliferation was performed to examine whether PTPRK is associated with pancreatic cell proliferation. (C, D) QPCR results show the expression of CDK6 and CCND1 in control cell lines PANC-1 pEF /CFPAC-1 pEF and PTPRK knockdown cell lines PANC-1 PTPRK kd /CFPAC-1 PTPRK kd . (E) TCGA dataset is used to draw a scatter plot showing the association between CDK6 and PTPRK at transcripts level. (F) In the TCGA dataset, the association between CCND1 and PTPRK transcript levels is shown. * p <0.05, ** p <0.01, *** p <0.001.

Journal: Cancer Diagnosis & Prognosis

Article Title: Elevated Protein Tyrosine Phosphatase Kappa Expression Is Associated With Disease Progression and Poor Prognosis of Pancreatic Cancer

doi: 10.21873/cdp.10560

Figure Lengend Snippet: Protein tyrosine phosphatase kappa (PTPRK) and cell proliferation. (A, B) A proliferation was performed to examine whether PTPRK is associated with pancreatic cell proliferation. (C, D) QPCR results show the expression of CDK6 and CCND1 in control cell lines PANC-1 pEF /CFPAC-1 pEF and PTPRK knockdown cell lines PANC-1 PTPRK kd /CFPAC-1 PTPRK kd . (E) TCGA dataset is used to draw a scatter plot showing the association between CDK6 and PTPRK at transcripts level. (F) In the TCGA dataset, the association between CCND1 and PTPRK transcript levels is shown. * p <0.05, ** p <0.01, *** p <0.001.

Article Snippet: Human pancreatic cancer cell lines PANC-1 and CFPAC-1 were purchased from ATCC (American Type Culture Collection, Manassas, VA, USA).

Techniques: Expressing, Control, Knockdown

Response to cyclin-dependent kinase 6 (CDK6) inhibitors in the protein tyrosine phosphatase kappa (PTPRK) knockdown pancreatic cancer cell line models. Both CFPAC-1 and PANC-1 cell lines were treated with different concentration of the CDK6 inhibitor BSJ-03-123 (A and B), CDK4/6 inhibitor Palbociclib (C and D) and CDK4 inhibitor 3-ATA (E and F). Corresponding IC 50 test results are shown. Cell viability was determined following a 3-day treatment with the inhibitors. * p <0.05, ** p <0.01, *** p <0.001.

Journal: Cancer Diagnosis & Prognosis

Article Title: Elevated Protein Tyrosine Phosphatase Kappa Expression Is Associated With Disease Progression and Poor Prognosis of Pancreatic Cancer

doi: 10.21873/cdp.10560

Figure Lengend Snippet: Response to cyclin-dependent kinase 6 (CDK6) inhibitors in the protein tyrosine phosphatase kappa (PTPRK) knockdown pancreatic cancer cell line models. Both CFPAC-1 and PANC-1 cell lines were treated with different concentration of the CDK6 inhibitor BSJ-03-123 (A and B), CDK4/6 inhibitor Palbociclib (C and D) and CDK4 inhibitor 3-ATA (E and F). Corresponding IC 50 test results are shown. Cell viability was determined following a 3-day treatment with the inhibitors. * p <0.05, ** p <0.01, *** p <0.001.

Article Snippet: Human pancreatic cancer cell lines PANC-1 and CFPAC-1 were purchased from ATCC (American Type Culture Collection, Manassas, VA, USA).

Techniques: Knockdown, Concentration Assay

Protein tyrosine phosphatase kappa (PTPRK) and lymph node metastasis. (A) The scatter plot shows that the lymph angiogenesis marker VEGFC is inversely correlated with PTPRK in the TCGA cohort. QPCR shows the expression of VEGFC in pancreatic cancer cell lines PANC-1 (B) and CFPAC-1 (C) with PTPRK knockdown. * p <0.05, ** p <0.01, *** p <0.001.

Journal: Cancer Diagnosis & Prognosis

Article Title: Elevated Protein Tyrosine Phosphatase Kappa Expression Is Associated With Disease Progression and Poor Prognosis of Pancreatic Cancer

doi: 10.21873/cdp.10560

Figure Lengend Snippet: Protein tyrosine phosphatase kappa (PTPRK) and lymph node metastasis. (A) The scatter plot shows that the lymph angiogenesis marker VEGFC is inversely correlated with PTPRK in the TCGA cohort. QPCR shows the expression of VEGFC in pancreatic cancer cell lines PANC-1 (B) and CFPAC-1 (C) with PTPRK knockdown. * p <0.05, ** p <0.01, *** p <0.001.

Article Snippet: Human pancreatic cancer cell lines PANC-1 and CFPAC-1 were purchased from ATCC (American Type Culture Collection, Manassas, VA, USA).

Techniques: Marker, Expressing, Knockdown

Journal: Scientific Reports

Article Title: The C16orf87 protein is a subunit of the MIER corepressor complex controlling embryonic development and cell migration

doi: 10.1038/s41598-026-50740-7

Figure Lengend Snippet: Knockdown of C16orf87 causes minor changes in the host cell protein profile. ( A ) Alignment of human ( Homo sapiens , UniProtKB accession number Q6PH81 ), mouse ( Mus musculus , UniProtKB accession number Q9CR55 ), and zebrafish ( Danio rerio , UniProtKB accession number Q6DGQ4 ) C16orf87 amino acid sequences. Alignment mismatches are indicated in gray boxes. The underlined sequence represents a possible minimal Akt/PKB kinase consensus recognition motif. A Ser91(S91) phosphorylation site is marked with an asterisk. ( B ) Per-residue confidence (pLDDT) coloring of the top-ranked predicted model of C16orf87. In the inset, the predicted zinc-ribbon domain is shown with the zinc-interacting cysteines (Cys16, Cys19, Cys30, and Cys32) indicated around the zinc ion (Zn 2+ ). The position of the phosphorylated serine (Ser91), a putative alpha-helix between amino acid residues Ser-107 and Ala-126, and the confidently predicted C-terminal alpha-helix between amino acid residues Asp-130 and Ile-153 are also highlighted. The ipTM and pTM values are annotated. N, N-terminus; C, C-terminus. Figure was rendered using ChimeraX (version 1.8, https://www.rbvi.ucsf.edu/chimerax ) ( C ) Western blot (WB) analysis of C16orf87 siRNA (siC16) knockdown in Panc-01, MiaPaCa-2, and C2C12 cell lines. A non-specific, scrambled siRNA (siScr) was used as a control; the WB membrane was probed with the antibodies against C16orf87 and actin. MS-based proteomics analysis of siRNA-treated C2C12 ( D ), MiaPaCa-2 ( E ), and Panc-01 ( F ) cells. Data points corresponding to histones are colored in pink, and statistically significant ( P < 0.05, fold-change > 1) proteins are colored in yellow (mouse cell line C2C12) and green (human cell lines, Panc-01 and MiaPaCa-2).

Article Snippet: Human pancreatic cancer cell lines Panc-01 (ATCC, CRL-1469) and MiaPaCa-2 (ATCC, CRL-1420), mouse skeletal muscle cell line C2C12 (ATCC, CRL-1772), and human cervical cancer cell line HeLa S3 (ATCC, CCL-2.2) were used in this study.

Techniques: Knockdown, Sequencing, Phospho-proteomics, Residue, Western Blot, Control, Membrane

CRISPR-Cas9 knockout of C16orf87 does not affect Panc-01 cell viability. ( A ) Whole-cell lysates of Panc-01 KO and Panc-01 WT cells were analyzed by western blot (WB), and proteins were detected with the antibodies against C16orf87 and actin. An asterisk indicates the migration of the C16orf87 protein. Mw; molecular weight marker. ( B ) MS-based proteomics analysis of Panc-01 KO and Panc-01 WT cell lysates (FDR ≤ 0.05, n = 3). Data points representing histones and proteins of interest are highlighted in red. ( C ) Panc-01 KO and Panc-01 WT cell viability was analyzed by FACS after AnnexinV-FITC (AnnV) and DRAQ7 (Dq7) staining. ( D ) Panc-01 KO and Panc-01 WT cell proliferation was analyzed by FACS after EdU-A647 incorporation into cells. ( E ) Quantification of the FACS analysis ( P < 0.01 (**)).

Journal: Scientific Reports

Article Title: The C16orf87 protein is a subunit of the MIER corepressor complex controlling embryonic development and cell migration

doi: 10.1038/s41598-026-50740-7

Figure Lengend Snippet: CRISPR-Cas9 knockout of C16orf87 does not affect Panc-01 cell viability. ( A ) Whole-cell lysates of Panc-01 KO and Panc-01 WT cells were analyzed by western blot (WB), and proteins were detected with the antibodies against C16orf87 and actin. An asterisk indicates the migration of the C16orf87 protein. Mw; molecular weight marker. ( B ) MS-based proteomics analysis of Panc-01 KO and Panc-01 WT cell lysates (FDR ≤ 0.05, n = 3). Data points representing histones and proteins of interest are highlighted in red. ( C ) Panc-01 KO and Panc-01 WT cell viability was analyzed by FACS after AnnexinV-FITC (AnnV) and DRAQ7 (Dq7) staining. ( D ) Panc-01 KO and Panc-01 WT cell proliferation was analyzed by FACS after EdU-A647 incorporation into cells. ( E ) Quantification of the FACS analysis ( P < 0.01 (**)).

Techniques: CRISPR, Knock-Out, Western Blot, Migration, Molecular Weight, Marker, Staining

CRISPR-Cas9 knockout of C16orf87 reduces Panc-01 cell migration. ( A ) Microscopy images of the in vitro scratch assay in Panc-01 KO and Panc-01 WT cells. Images were taken at 0, 6, 12, and 24 h after scratches were applied. ( B ) The cell migration rate was calculated based on the extent of cell coverage within the scratched area ( P < 0.05 (*) and P < 0.01 (**)).

Journal: Scientific Reports

Article Title: The C16orf87 protein is a subunit of the MIER corepressor complex controlling embryonic development and cell migration

doi: 10.1038/s41598-026-50740-7

Figure Lengend Snippet: CRISPR-Cas9 knockout of C16orf87 reduces Panc-01 cell migration. ( A ) Microscopy images of the in vitro scratch assay in Panc-01 KO and Panc-01 WT cells. Images were taken at 0, 6, 12, and 24 h after scratches were applied. ( B ) The cell migration rate was calculated based on the extent of cell coverage within the scratched area ( P < 0.05 (*) and P < 0.01 (**)).

Techniques: CRISPR, Knock-Out, Migration, Microscopy, In Vitro, Wound Healing Assay

C16orf87 partially mediates HDAC1 and MIER1 protein interactions. ( A ) C16orf87 interacts with the HDAC and MIER proteins. Volcano plot of the IP-MS experiment showing identified proteins interacting with the Flag-C16orf87 protein in HeLa cells. An adjusted P -value cut-off of 0.05 and a log2 fold change cut-off of 2 were used. Data are shown from a biological triplicate experiment. ( B ) Lack of C16orf87 does not change HDAC and MIER protein accumulation. Soluble Panc-01 WT (WT) and Panc-01 KO (KO) whole-cell lysates were analyzed by WB with the indicated antibodies. ( C ) C16orf87 partially mediates HDAC1 and MIER1 interaction. Co-immunoprecipitation of Flag-HDAC1 from siRNA (siC16 and siScr) and pcDNA3-Flag-HDAC1-transfected HeLa cells. Isolated proteins were analyzed by WB with the indicated antibodies. An arrowhead indicates the migration of the MIER1 protein isoforms, whereas an asterisk indicates the migration of the C16orf87 isoforms. ( D ) HDAC1 interacts weakly with C16orf87 in vitro. GST (as a control) and GST-HDAC1 pull-down with bacterially purified 8 × His-tagged C16orf87(Wt, 5 × C > A, 1–130, and 5 × C > A/1–130) proteins. An asterisk indicates a degradation product/partially translated GST-HDAC1. Proteins were detected with the anti-His and anti-GST antibodies.

Journal: Scientific Reports

Article Title: The C16orf87 protein is a subunit of the MIER corepressor complex controlling embryonic development and cell migration

doi: 10.1038/s41598-026-50740-7

Figure Lengend Snippet: C16orf87 partially mediates HDAC1 and MIER1 protein interactions. ( A ) C16orf87 interacts with the HDAC and MIER proteins. Volcano plot of the IP-MS experiment showing identified proteins interacting with the Flag-C16orf87 protein in HeLa cells. An adjusted P -value cut-off of 0.05 and a log2 fold change cut-off of 2 were used. Data are shown from a biological triplicate experiment. ( B ) Lack of C16orf87 does not change HDAC and MIER protein accumulation. Soluble Panc-01 WT (WT) and Panc-01 KO (KO) whole-cell lysates were analyzed by WB with the indicated antibodies. ( C ) C16orf87 partially mediates HDAC1 and MIER1 interaction. Co-immunoprecipitation of Flag-HDAC1 from siRNA (siC16 and siScr) and pcDNA3-Flag-HDAC1-transfected HeLa cells. Isolated proteins were analyzed by WB with the indicated antibodies. An arrowhead indicates the migration of the MIER1 protein isoforms, whereas an asterisk indicates the migration of the C16orf87 isoforms. ( D ) HDAC1 interacts weakly with C16orf87 in vitro. GST (as a control) and GST-HDAC1 pull-down with bacterially purified 8 × His-tagged C16orf87(Wt, 5 × C > A, 1–130, and 5 × C > A/1–130) proteins. An asterisk indicates a degradation product/partially translated GST-HDAC1. Proteins were detected with the anti-His and anti-GST antibodies.

Techniques: Protein-Protein interactions, Immunoprecipitation, Transfection, Isolation, Migration, In Vitro, Control, Purification

Lack of C16orf87 alters chromatin accessibility. ( A ) Distribution of the more accessible chromatin genomic features identified by ATAC-seq in Panc-01 WT and Panc-01 KO cells. The X-axis shows values in percentage. Gene locus diagrams showing genomic regions near the WWOX ( B ) and NCOA7-HINT3 ( C ) genes, with peaks representing ATAC-seq reads indicating chromatin accessibility. Data were aligned to available tracks (ChIP-Atlas) of HDAC1 , HDAC2 , MIER1 , MIER2 , MIER3 , and H3K27ac markers. Panc-01 WT and Panc-01 KO peaks are shown in blue and green, respectively, and significant differences (FDR threshold: 0.05) in read quantities (peaks), observed in genomic intervals, are shown in red bars on the third track (top to bottom). WWOX represents one of the genes with a higher peak on the Panc-01 WT compared to Panc-01 KO . NCOA7-HINT3 represents one of the genes with higher peaks on the Panc-01 KO . ( D ) qRT-PCR analysis of the NCOA7 , HINT3 , WWOX , and C16orf87 mRNA expression. Relative mRNA expression in Panc-01 WT and Panc-01 KO cells after normalization to 18S rRNA and considering mRNA levels in Panc-01 WT cells as 1.

Journal: Scientific Reports

Article Title: The C16orf87 protein is a subunit of the MIER corepressor complex controlling embryonic development and cell migration

doi: 10.1038/s41598-026-50740-7

Figure Lengend Snippet: Lack of C16orf87 alters chromatin accessibility. ( A ) Distribution of the more accessible chromatin genomic features identified by ATAC-seq in Panc-01 WT and Panc-01 KO cells. The X-axis shows values in percentage. Gene locus diagrams showing genomic regions near the WWOX ( B ) and NCOA7-HINT3 ( C ) genes, with peaks representing ATAC-seq reads indicating chromatin accessibility. Data were aligned to available tracks (ChIP-Atlas) of HDAC1 , HDAC2 , MIER1 , MIER2 , MIER3 , and H3K27ac markers. Panc-01 WT and Panc-01 KO peaks are shown in blue and green, respectively, and significant differences (FDR threshold: 0.05) in read quantities (peaks), observed in genomic intervals, are shown in red bars on the third track (top to bottom). WWOX represents one of the genes with a higher peak on the Panc-01 WT compared to Panc-01 KO . NCOA7-HINT3 represents one of the genes with higher peaks on the Panc-01 KO . ( D ) qRT-PCR analysis of the NCOA7 , HINT3 , WWOX , and C16orf87 mRNA expression. Relative mRNA expression in Panc-01 WT and Panc-01 KO cells after normalization to 18S rRNA and considering mRNA levels in Panc-01 WT cells as 1.

Techniques: Quantitative RT-PCR, Expressing

Expression of EN2 in human pancreatic cancer tissues. (A, B), Pancreatic Tissue Arrays containing normal and cancerous tissues were purchased from Biomax. EN2 expression was measured by IHC. Representative photographs of 60 pancreatic tissues from various stages of pancreatic cancer. Blue = nuclei, Brown/pink colour = EN2. * = significantly different from normal, p = < 0.01. (C) TCGA data on the expression of EN2 mRNA. * = significantly different from normal.

Journal: Journal of Cellular and Molecular Medicine

Article Title: EN2 Regulates Pancreatic Cancer Initiation, Progression, and Epithelial‐Mesenchymal Transition Through the Notch Signalling Pathway

doi: 10.1111/jcmm.71158

Figure Lengend Snippet: Expression of EN2 in human pancreatic cancer tissues. (A, B), Pancreatic Tissue Arrays containing normal and cancerous tissues were purchased from Biomax. EN2 expression was measured by IHC. Representative photographs of 60 pancreatic tissues from various stages of pancreatic cancer. Blue = nuclei, Brown/pink colour = EN2. * = significantly different from normal, p = < 0.01. (C) TCGA data on the expression of EN2 mRNA. * = significantly different from normal.

Article Snippet: Human pancreatic cancer cell lines (PANC‐1 and AsPC‐1) and human normal pancreatic ductal epithelial cells (HPNE) were purchased from the American Type Culture Collection (ATCC, Manassas, VA).

Techniques: Expressing

The expression of EN2 in HPNE, pancreatic cancer cell lines, and pancreatic CSCs. (A), Protein expression of EN2 in HPNE, pancreatic cancer cell lines, and pancreatic CSCs. Crude proteins were isolated, and EN2 expression was measured by Western blot analysis. β‐Actin was used as a loading control. (B), Expression of EN2 mRNA in HPNE, pancreatic cancer cell lines, and pancreatic CSCs. RNA was isolated, and EN2 expression was measured by q‐RT‐PCR. GAPDH was used as an internal control. Data represent mean ( n = 4) ± SD. *, # and % = significantly different from HPNE ( p < 0.05). (C), Expression of EN2. Immunocytochemistry was performed to examine EN2 expression in HPNE, PANC‐1, and AsPC‐1 cells.

Journal: Journal of Cellular and Molecular Medicine

Article Title: EN2 Regulates Pancreatic Cancer Initiation, Progression, and Epithelial‐Mesenchymal Transition Through the Notch Signalling Pathway

doi: 10.1111/jcmm.71158

Figure Lengend Snippet: The expression of EN2 in HPNE, pancreatic cancer cell lines, and pancreatic CSCs. (A), Protein expression of EN2 in HPNE, pancreatic cancer cell lines, and pancreatic CSCs. Crude proteins were isolated, and EN2 expression was measured by Western blot analysis. β‐Actin was used as a loading control. (B), Expression of EN2 mRNA in HPNE, pancreatic cancer cell lines, and pancreatic CSCs. RNA was isolated, and EN2 expression was measured by q‐RT‐PCR. GAPDH was used as an internal control. Data represent mean ( n = 4) ± SD. *, # and % = significantly different from HPNE ( p < 0.05). (C), Expression of EN2. Immunocytochemistry was performed to examine EN2 expression in HPNE, PANC‐1, and AsPC‐1 cells.

Techniques: Expressing, Isolation, Western Blot, Control, Reverse Transcription Polymerase Chain Reaction, Immunocytochemistry

EN2 knockdown reduces motility, migration, invasion, and EMT marker expression in pancreatic cancer cells. (A) Cell Motility Assay. Pancreatic cancer cells expressing scrambled or EN2 shRNA were cultured in petri dishes. After 18 h, a linear scratch was generated using a fine pipette tip, and phase‐contrast images were captured at 0 and 48 h to assess wound closure. (B) Cell Migration Assay. Cells expressing scrambled or EN2 shRNA were seeded in six‐well plates, and migration was quantified as described in the Materials and Methods. Data represent mean ( n = 4) ± SD. * p < 0.05 compared with the scrambled control. (C) Cell Invasion Assay. Cells expressing scrambled or EN2 shRNA were seeded in six‐well plates, and invasion was measured as described in the Materials and Methods. Data represent mean ( n = 4) ± SD. * p < 0.05 compared with the scrambled control. (D, E) Total RNA was isolated, and the expression of E‐cadherin, N‐cadherin, Snail, Slug, and Zeb1 was quantified by qRT‐PCR. GAPDH served as the internal control. Data represent mean ( n = 4) ± SD. * p < 0.05 between groups.

Journal: Journal of Cellular and Molecular Medicine

Article Title: EN2 Regulates Pancreatic Cancer Initiation, Progression, and Epithelial‐Mesenchymal Transition Through the Notch Signalling Pathway

doi: 10.1111/jcmm.71158

Figure Lengend Snippet: EN2 knockdown reduces motility, migration, invasion, and EMT marker expression in pancreatic cancer cells. (A) Cell Motility Assay. Pancreatic cancer cells expressing scrambled or EN2 shRNA were cultured in petri dishes. After 18 h, a linear scratch was generated using a fine pipette tip, and phase‐contrast images were captured at 0 and 48 h to assess wound closure. (B) Cell Migration Assay. Cells expressing scrambled or EN2 shRNA were seeded in six‐well plates, and migration was quantified as described in the Materials and Methods. Data represent mean ( n = 4) ± SD. * p < 0.05 compared with the scrambled control. (C) Cell Invasion Assay. Cells expressing scrambled or EN2 shRNA were seeded in six‐well plates, and invasion was measured as described in the Materials and Methods. Data represent mean ( n = 4) ± SD. * p < 0.05 compared with the scrambled control. (D, E) Total RNA was isolated, and the expression of E‐cadherin, N‐cadherin, Snail, Slug, and Zeb1 was quantified by qRT‐PCR. GAPDH served as the internal control. Data represent mean ( n = 4) ± SD. * p < 0.05 between groups.

Techniques: Knockdown, Migration, Marker, Expressing, Motility Assay, shRNA, Cell Culture, Generated, Transferring, Cell Migration Assay, Control, Invasion Assay, Isolation, Quantitative RT-PCR

EN2 shRNA inhibits Notch‐target genes and Nanog expression and RBP JK transcription in pancreatic cancer cells. (A, B), Expression of Notch target genes. RNA was isolated, and the expression of cMyc, Cyclin D1, Bcl‐2, Hes 1 and Nanog was measured in cells by q‐RT‐PCR. GAPDH was used as an internal control. Data represent mean ( n = 4) ± SD. * = significantly different between groups ( p < 0.05). (C), RBP JK transcription. PANC‐1 and AsPC‐1 cells were transduced with RBP JK ‐responsive GFP/firefly luciferase viral particles (pGreen Fire1‐ RBP JK with EF1, System Biosciences) along with EN2/scrambled or EN2 shRNA viral particles. RBP JK reporter activity was measured as we described . Data represent mean ( n = 4) ± SD. * = significantly different from scrambled control group ( p < 0.05).

Journal: Journal of Cellular and Molecular Medicine

Article Title: EN2 Regulates Pancreatic Cancer Initiation, Progression, and Epithelial‐Mesenchymal Transition Through the Notch Signalling Pathway

doi: 10.1111/jcmm.71158

Figure Lengend Snippet: EN2 shRNA inhibits Notch‐target genes and Nanog expression and RBP JK transcription in pancreatic cancer cells. (A, B), Expression of Notch target genes. RNA was isolated, and the expression of cMyc, Cyclin D1, Bcl‐2, Hes 1 and Nanog was measured in cells by q‐RT‐PCR. GAPDH was used as an internal control. Data represent mean ( n = 4) ± SD. * = significantly different between groups ( p < 0.05). (C), RBP JK transcription. PANC‐1 and AsPC‐1 cells were transduced with RBP JK ‐responsive GFP/firefly luciferase viral particles (pGreen Fire1‐ RBP JK with EF1, System Biosciences) along with EN2/scrambled or EN2 shRNA viral particles. RBP JK reporter activity was measured as we described . Data represent mean ( n = 4) ± SD. * = significantly different from scrambled control group ( p < 0.05).

Techniques: shRNA, Expressing, Isolation, Reverse Transcription Polymerase Chain Reaction, Control, Transduction, Luciferase, Activity Assay

Effects of Cal on aPSC activation. (A) (Left) VDR mRNA expression in PDAC cell lines (AsPC‐1, MIA PaCa‐2, and PANC‐1) and aPSCs was determined by qRT‐PCR ( n = 3). (Right) CYP24A1 mRNA expression in PDAC or aPSCs treated with DMSO or Cal (100 nM and 48 h) was examined by qRT‐PCR ( n = 3). (B) VDR protein expression in PDAC cell lines (AsPC‐1, MIA PaCa‐2, and PANC‐1) and aPSCs was determined by western blot ( n = 3). (C) Correlation analysis between α‐SMA and VDR mRNA expression in aPSCs, with GAPDH normalization ( n = 9). (D) VDR and α‐SMA gene expression in aPSCs treated with DMSO or Cal (100 nM and 48 h) was evaluated by qRT‐PCR ( n = 3). (E) VDR and α‐SMA protein expression in aPSCs treated with DMSO or Cal (100 nM and 48 h) ( n = 4). (F) Immunocytochemistry showing α‐SMA expression in aPSCs treated with DMSO or Cal (100 nM and 48 hr) ( n = 3). (G) EZ4U assay indicating the impacts of Cal on the proliferation of aPSCs ( n = 3). (H) Transwell migration assay and (I) wound healing showing the effects of Cal on aPSCs’ migration ability ( n = 3). caPSCs, PSCs derived from pancreatic cancer; cpPSCs, PSCs derived from chronic pancreatitis; cuPSCs, culture‐activated PSCs derived from normal tissue; aPSCs, activated PSCs; HPF, high‐power field; Ctr, control group treated with DMSO. All experiments were conducted in triplicate. ns, not significant. ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001.

Journal: Mediators of Inflammation

Article Title: The Vitamin D3 Analog Calcipotriol Attenuates Pancreatic Cancer Malignancy via Downregulating Thrombospondin 1 in Pancreatic Stellate Cells

doi: 10.1155/mi/2632235

Figure Lengend Snippet: Effects of Cal on aPSC activation. (A) (Left) VDR mRNA expression in PDAC cell lines (AsPC‐1, MIA PaCa‐2, and PANC‐1) and aPSCs was determined by qRT‐PCR ( n = 3). (Right) CYP24A1 mRNA expression in PDAC or aPSCs treated with DMSO or Cal (100 nM and 48 h) was examined by qRT‐PCR ( n = 3). (B) VDR protein expression in PDAC cell lines (AsPC‐1, MIA PaCa‐2, and PANC‐1) and aPSCs was determined by western blot ( n = 3). (C) Correlation analysis between α‐SMA and VDR mRNA expression in aPSCs, with GAPDH normalization ( n = 9). (D) VDR and α‐SMA gene expression in aPSCs treated with DMSO or Cal (100 nM and 48 h) was evaluated by qRT‐PCR ( n = 3). (E) VDR and α‐SMA protein expression in aPSCs treated with DMSO or Cal (100 nM and 48 h) ( n = 4). (F) Immunocytochemistry showing α‐SMA expression in aPSCs treated with DMSO or Cal (100 nM and 48 hr) ( n = 3). (G) EZ4U assay indicating the impacts of Cal on the proliferation of aPSCs ( n = 3). (H) Transwell migration assay and (I) wound healing showing the effects of Cal on aPSCs’ migration ability ( n = 3). caPSCs, PSCs derived from pancreatic cancer; cpPSCs, PSCs derived from chronic pancreatitis; cuPSCs, culture‐activated PSCs derived from normal tissue; aPSCs, activated PSCs; HPF, high‐power field; Ctr, control group treated with DMSO. All experiments were conducted in triplicate. ns, not significant. ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001.

Article Snippet: Human PDAC cell lines PANC‐1 (male, American Type Culture Collection [ATCC] CRL‐1469, RRID: CVCL_0480), MIA PaCa‐2 (male, ATCC CRL‐1420, RRID: CVCL_0428), and AsPC‐1 (female, ATCC CRL‐1682, RRID: CVCL_0152) were purchased directly from the ATCC (Manassas, VA, USA) in 2015.

Techniques: Activation Assay, Expressing, Quantitative RT-PCR, Western Blot, Gene Expression, Immunocytochemistry, Transwell Migration Assay, Migration, Derivative Assay, Control

Dose‐dependent effect of rTHBS1 on PDAC malignancy. (A) Transwell migration assays showing the response of PDAC cell lines PANC‐1 and MIA PaCa‐2 to varying concentrations of rTHBS1 (0, 0.5, and 5 μg/mL). (B) Transwell invasion assays were used to quantify the invasive potential of the same PDAC cell lines under the same rTHBS1 treatments. (C) Proliferation of PDAC cells was assessed by EZ4U assay after treatment with rTHBS1 at 0, 0.5, 5, and 20 μg/mL. (D) Wound healing assays complement the migration analysis, with images and quantification of the migration area closure. (E) Representative micrographs depicting morphological alterations in PANC‐1 and MIA PaCa‐2 cells when cultured in standard medium, aPSCs‐CM, and standard medium supplemented with 5 μg/mL of rTHBS1. All experiments were conducted in triplicate. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001.

Journal: Mediators of Inflammation

Article Title: The Vitamin D3 Analog Calcipotriol Attenuates Pancreatic Cancer Malignancy via Downregulating Thrombospondin 1 in Pancreatic Stellate Cells

doi: 10.1155/mi/2632235

Figure Lengend Snippet: Dose‐dependent effect of rTHBS1 on PDAC malignancy. (A) Transwell migration assays showing the response of PDAC cell lines PANC‐1 and MIA PaCa‐2 to varying concentrations of rTHBS1 (0, 0.5, and 5 μg/mL). (B) Transwell invasion assays were used to quantify the invasive potential of the same PDAC cell lines under the same rTHBS1 treatments. (C) Proliferation of PDAC cells was assessed by EZ4U assay after treatment with rTHBS1 at 0, 0.5, 5, and 20 μg/mL. (D) Wound healing assays complement the migration analysis, with images and quantification of the migration area closure. (E) Representative micrographs depicting morphological alterations in PANC‐1 and MIA PaCa‐2 cells when cultured in standard medium, aPSCs‐CM, and standard medium supplemented with 5 μg/mL of rTHBS1. All experiments were conducted in triplicate. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001.

Techniques: Migration, Cell Culture

Inhibition of aPSCs‐CM–driven malignancy in PDAC by THBS1 neutralizing antibody. THBS1 neutralizing Ab diminished aPSCs‐CM–induced migration (A, C, D), invasion (B), proliferation (E–F), and EMT (G–H) of PDAC but had no effects on Cal‐aPSCs‐CM–induced malignancy of PDAC. aPSCs‐CM, CM from aPSCs pretreated with DMSO; Cal‐aPSCs‐CM, CM harvested from aPSCs pretreated with 100 nM Cal for 48 h. CM was then pretreated with 1 μg/mL of THBS1 Ab or control IgG and added to the PDAC. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001.

Journal: Mediators of Inflammation

Article Title: The Vitamin D3 Analog Calcipotriol Attenuates Pancreatic Cancer Malignancy via Downregulating Thrombospondin 1 in Pancreatic Stellate Cells

doi: 10.1155/mi/2632235

Figure Lengend Snippet: Inhibition of aPSCs‐CM–driven malignancy in PDAC by THBS1 neutralizing antibody. THBS1 neutralizing Ab diminished aPSCs‐CM–induced migration (A, C, D), invasion (B), proliferation (E–F), and EMT (G–H) of PDAC but had no effects on Cal‐aPSCs‐CM–induced malignancy of PDAC. aPSCs‐CM, CM from aPSCs pretreated with DMSO; Cal‐aPSCs‐CM, CM harvested from aPSCs pretreated with 100 nM Cal for 48 h. CM was then pretreated with 1 μg/mL of THBS1 Ab or control IgG and added to the PDAC. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001.

Techniques: Inhibition, Migration, Control

Attenuation of aPSCs‐CM–induced PDAC aggressiveness by CD47 blockade. CD47 blocking Ab diminished aPSCs‐CM–induced migration (A, C, D), invasion (B), proliferation (E–F), and EMT (G–H) of PDAC but had no effects on Cal‐aPSCs‐CM–induced aggressiveness of PDAC. PDAC were pretreated with 2 μg/mL CD47 blocking Ab or control IgG. All experiments were conducted in triplicate. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001.

Journal: Mediators of Inflammation

Article Title: The Vitamin D3 Analog Calcipotriol Attenuates Pancreatic Cancer Malignancy via Downregulating Thrombospondin 1 in Pancreatic Stellate Cells

doi: 10.1155/mi/2632235

Figure Lengend Snippet: Attenuation of aPSCs‐CM–induced PDAC aggressiveness by CD47 blockade. CD47 blocking Ab diminished aPSCs‐CM–induced migration (A, C, D), invasion (B), proliferation (E–F), and EMT (G–H) of PDAC but had no effects on Cal‐aPSCs‐CM–induced aggressiveness of PDAC. PDAC were pretreated with 2 μg/mL CD47 blocking Ab or control IgG. All experiments were conducted in triplicate. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001.

Techniques: Blocking Assay, Migration, Control

Differential impact on PDAC organoid morphology and EMT marker expression by aPSCs‐CM and antibody interventions. (A) Representative bright‐field images displaying PDAC organoids over 5 days in control (aPSCs‐CM), treated with Cal‐aPSCs‐CM, with THBS1 antibody‐depleted aPSCs‐CM, and with organoids where CD47 has been blocked, followed by treatment with aPSCs‐CM. (B) Western blot analysis of E‐cadherin and vimentin in organoids subjected to these varied treatments. (C) Protein expression quantification normalized to GAPDH, demonstrating the effect of THBS1 depletion and CD47 inhibition on EMT markers in PDAC organoids. All experiments were conducted in triplicate. ∗ p < 0.05 and ∗∗ p < 0.01. Scale bar: 100 μm.

Journal: Mediators of Inflammation

Article Title: The Vitamin D3 Analog Calcipotriol Attenuates Pancreatic Cancer Malignancy via Downregulating Thrombospondin 1 in Pancreatic Stellate Cells

doi: 10.1155/mi/2632235

Figure Lengend Snippet: Differential impact on PDAC organoid morphology and EMT marker expression by aPSCs‐CM and antibody interventions. (A) Representative bright‐field images displaying PDAC organoids over 5 days in control (aPSCs‐CM), treated with Cal‐aPSCs‐CM, with THBS1 antibody‐depleted aPSCs‐CM, and with organoids where CD47 has been blocked, followed by treatment with aPSCs‐CM. (B) Western blot analysis of E‐cadherin and vimentin in organoids subjected to these varied treatments. (C) Protein expression quantification normalized to GAPDH, demonstrating the effect of THBS1 depletion and CD47 inhibition on EMT markers in PDAC organoids. All experiments were conducted in triplicate. ∗ p < 0.05 and ∗∗ p < 0.01. Scale bar: 100 μm.

Techniques: Marker, Expressing, Control, Western Blot, Inhibition

Schematic overview of the study workflow. The MPS consisted of one‐chamber microfluidic devices used to generate two cancer‐on‐a‐chip models: PDAC and LAC. Cells were embedded in natural hydrogels composed of egg white (EW)‐gelatin and collagen type I, respectively. At the end of the culture period, devices were fixed, stained, dehydrated, disassembled, critically point‐dried, mounted, and coated for SEM and FIB‐SEM imaging. For LAC models, the FIB was additionally used to prepare thin lamellae for TEM. This workflow enabled high‐resolution observation of both external and internal organization of 3D multicellular structures, including cell–cell and cell–matrix interactions, matrix deposition, and intercellular communication.

Journal: Small Science

Article Title: Ultrastructural Study of Microphysiological Systems of the Tumor Microenvironment

doi: 10.1002/smsc.202500567

Figure Lengend Snippet: Schematic overview of the study workflow. The MPS consisted of one‐chamber microfluidic devices used to generate two cancer‐on‐a‐chip models: PDAC and LAC. Cells were embedded in natural hydrogels composed of egg white (EW)‐gelatin and collagen type I, respectively. At the end of the culture period, devices were fixed, stained, dehydrated, disassembled, critically point‐dried, mounted, and coated for SEM and FIB‐SEM imaging. For LAC models, the FIB was additionally used to prepare thin lamellae for TEM. This workflow enabled high‐resolution observation of both external and internal organization of 3D multicellular structures, including cell–cell and cell–matrix interactions, matrix deposition, and intercellular communication.

Article Snippet: Human PDAC cell line PANC‐1, and human LAC cell line A549 from the American Type Culture Collection (ATCC, USA) were used for this work.

Techniques: Staining, Imaging

Schematic of a generic MPS used in this study. The inner geometry consists of a single microfluidic channel with ports for hydrogel loading and perfusion of culture medium. The central chamber contains the 3D multicellular tumor culture embedded in a hydrogel matrix. Two distinct cancer‐on‐a‐chip models were implemented, each in a separate device: pancreatic ductal adenocarcinoma (PDAC, PANC‐1 cells) and lung adenocarcinoma (LAC, A549 cells). This schematic highlights the overall architecture of the chip and the spatial arrangement of the tumor constructs, hydrogel, and medium channels.

Journal: Small Science

Article Title: Ultrastructural Study of Microphysiological Systems of the Tumor Microenvironment

doi: 10.1002/smsc.202500567

Figure Lengend Snippet: Schematic of a generic MPS used in this study. The inner geometry consists of a single microfluidic channel with ports for hydrogel loading and perfusion of culture medium. The central chamber contains the 3D multicellular tumor culture embedded in a hydrogel matrix. Two distinct cancer‐on‐a‐chip models were implemented, each in a separate device: pancreatic ductal adenocarcinoma (PDAC, PANC‐1 cells) and lung adenocarcinoma (LAC, A549 cells). This schematic highlights the overall architecture of the chip and the spatial arrangement of the tumor constructs, hydrogel, and medium channels.

Article Snippet: Human PDAC cell line PANC‐1, and human LAC cell line A549 from the American Type Culture Collection (ATCC, USA) were used for this work.

Techniques: Construct

(A) Scheme of the MPS of the TME of PDAC, consisting of a hydrogel‐based cancer‐on‐a‐chip model where the PANC‐1 cell line was used. (B) Brightfield microscopy representative images of the PANC‐1 aggregates grown over 14 days of culture in EW/gelatin hydrogels. Zoomed area shows single aggregate. Dual beam FIB‐SEM representative image of EW/gelatin hydrogel after 14 days of culture, displaying the typical nanoglobular morphology of these hydrogels. (C) Dual beam FIB‐SEM representative images of the surface of PANC‐1 aggregates in EW/gelatin hydrogel. ECM: extracellular matrix produced by the cells. Orange asterisk: EW/gelatin hydrogel. Green arrows: unions between cells. Orange arrows: unions between the cells and the hydrogel. Yellow arrows: spherical particles. Blue arrows: large spherical particles.

Journal: Small Science

Article Title: Ultrastructural Study of Microphysiological Systems of the Tumor Microenvironment

doi: 10.1002/smsc.202500567

Figure Lengend Snippet: (A) Scheme of the MPS of the TME of PDAC, consisting of a hydrogel‐based cancer‐on‐a‐chip model where the PANC‐1 cell line was used. (B) Brightfield microscopy representative images of the PANC‐1 aggregates grown over 14 days of culture in EW/gelatin hydrogels. Zoomed area shows single aggregate. Dual beam FIB‐SEM representative image of EW/gelatin hydrogel after 14 days of culture, displaying the typical nanoglobular morphology of these hydrogels. (C) Dual beam FIB‐SEM representative images of the surface of PANC‐1 aggregates in EW/gelatin hydrogel. ECM: extracellular matrix produced by the cells. Orange asterisk: EW/gelatin hydrogel. Green arrows: unions between cells. Orange arrows: unions between the cells and the hydrogel. Yellow arrows: spherical particles. Blue arrows: large spherical particles.

Article Snippet: Human PDAC cell line PANC‐1, and human LAC cell line A549 from the American Type Culture Collection (ATCC, USA) were used for this work.

Techniques: Microscopy, Produced

(A) Dual beam FIB‐SEM representative images of the ECM secreted by the PANC‐1 cells. Image iii shows some measurements of the diameter of the fibers that comprise the ECM. (B) Measurements of the diameter and length of the ECM fibers secreted by the PANC‐1 cells. Data shown in violin plot as its distribution with median and the interquartile range (IQR), each dot represent a measurement ( n = 98 for fiber diameter and n = 46 for fiber length).

Journal: Small Science

Article Title: Ultrastructural Study of Microphysiological Systems of the Tumor Microenvironment

doi: 10.1002/smsc.202500567

Figure Lengend Snippet: (A) Dual beam FIB‐SEM representative images of the ECM secreted by the PANC‐1 cells. Image iii shows some measurements of the diameter of the fibers that comprise the ECM. (B) Measurements of the diameter and length of the ECM fibers secreted by the PANC‐1 cells. Data shown in violin plot as its distribution with median and the interquartile range (IQR), each dot represent a measurement ( n = 98 for fiber diameter and n = 46 for fiber length).

Article Snippet: Human PDAC cell line PANC‐1, and human LAC cell line A549 from the American Type Culture Collection (ATCC, USA) were used for this work.

Techniques:

(A) Dual beam FIB‐SEM representative images of the internal cell organization of the aggregates formed by PANC‐1 cells in EW/gelatin hydrogels. Image i shows the surface of the aggregate milled with the ion beam along the yellow line. Images ii–v show internal sections of the aggregate shown in image i. Image vi belongs to a different aggregate. Individual cells are indicated by red numbers. For additional guidance to identify cell boundaries, Figure S1 provides the same images with a colored overlay mask highlighting the cellular regions. Light blue arrows: darker and denser intracellular areas. Green arrows: membrane projections connecting adjacent cells. Yellow arrows: EVs secreted by the cells. (B) Measurements of the size (i.e., diameter) of the EVs secreted by the PANC‐1 cells. Data shown in violin plot as its distribution with median and the IQR, each dot represents a measurement ( n = 48).

Journal: Small Science

Article Title: Ultrastructural Study of Microphysiological Systems of the Tumor Microenvironment

doi: 10.1002/smsc.202500567

Figure Lengend Snippet: (A) Dual beam FIB‐SEM representative images of the internal cell organization of the aggregates formed by PANC‐1 cells in EW/gelatin hydrogels. Image i shows the surface of the aggregate milled with the ion beam along the yellow line. Images ii–v show internal sections of the aggregate shown in image i. Image vi belongs to a different aggregate. Individual cells are indicated by red numbers. For additional guidance to identify cell boundaries, Figure S1 provides the same images with a colored overlay mask highlighting the cellular regions. Light blue arrows: darker and denser intracellular areas. Green arrows: membrane projections connecting adjacent cells. Yellow arrows: EVs secreted by the cells. (B) Measurements of the size (i.e., diameter) of the EVs secreted by the PANC‐1 cells. Data shown in violin plot as its distribution with median and the IQR, each dot represents a measurement ( n = 48).

Article Snippet: Human PDAC cell line PANC‐1, and human LAC cell line A549 from the American Type Culture Collection (ATCC, USA) were used for this work.

Techniques: Membrane

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