Journal: International Journal of Molecular Sciences

Article Title: Structural Identification of the Pacemaker Cells and Expression of Hyperpolarization-Activated Cyclic Nucleotide-Gated (HCN) Channels in the Heart of the Wild Atlantic Cod, Gadus morhua (Linnaeus, 1758)

doi: 10.3390/ijms22147539

Figure Lengend Snippet: Primary and secondary antibodies used in the present study.

Article Snippet: Anti-HCN1 , Polyclonal , Alomone Labs , APC-056 , 1:100 , AB_2039900.

Techniques:

Journal: Nature Communications

Article Title: Sfrp5 identifies murine cardiac progenitors for all myocardial structures except for the right ventricle

doi: 10.1038/ncomms14664

Figure Lengend Snippet: ( a ) Representative photographs of LacZ-stained sections from Sfrp5-ERT2Cre / Rosa-lacZ embryos at E10.5, labelled by tamoxifen at E7.5. ( b ) Schematic drawing for the expression of Sfrp5 (orange) and the cardiac contribution of its lineages (red) when cells were labelled at E7.5 and analysed at E10.5. ( c ) Double fluorescent whole-mount ISH of mouse embryos at E7.5 for detection of Sfrp5 (green) and Hcn4 , Tbx5 or Isl1 (red), as indicated in each panel. ( d ) Schematic drawings of gene expression of Sfrp5 , Isl1 , Hcn4 and Tbx5 . ( e ) Double immunohistochemistry of Sfrp5-venusYFP at E8.5 using anti-GFP (green) and anti-HCN4 (red) antibodies. Co-expression with HCN4 was found in the VP. Weak expression of YFP also co-distributed with HCN4-positive cells in the IFT and part of the primitive ventricle (PV). ( f , g ) Double immunohistochemistry of Sfrp5-venusYFP ( e ), Sfrp5-venusYFP/Isl1 - Cre / CAG-(floxed)-CAT-mRFP ( f ) or Sfrp5 - Cre / CAG-(floxed)-CAT-eGFP ( g ) embryos at E9.5 using anti-GFP (green) antibodies and anti-HCN4, anti-RFP, anti - Isl1 or anti-Tbx1 (red) antibodies. Rectangles in upper panels are magnified in lower panels (v: ventral; d: dorsal). Co-expression with Isl1 was detected (arrows in panel ( f ) and parenthesis in panel ( g )). Boundaries for the VP from the atrium are indicated by white dotted lines ( f , g ). Signal for Tbx1, but not GFP, was observed in the anterior splanchnic mesoderm and anterior OFT (arrowheads in panel ( g )). A, atrium.

Article Snippet: After washing with PBS and 1 h of preincubation with 1% BSA in PBS for blocking, sections were incubated with 500 × diluted primary antibodies (anti-GFP: Nakalai GF090R or Abcam ab290; anti-Tbx1: Santa Cruz sc-17874; anti-Tbx18: Santa Cruz SC17869; anti-Wt1: Abcam ab10670; anti-Nkx2-5: Abcam ab35842; anti-TnT: Thermo MS-295-P0; anti-Hcn4: Alomone Labs APC052; anti-RFP: MBL PM005; anti-β-catenin: Millipore 05-665; anti-Ki67: Abcam ab15580) at 4 °C overnight.

Techniques: Staining, Expressing, Gene Expression, Immunohistochemistry, Immunofluorescence

Journal: CNS Neuroscience & Therapeutics

Article Title: HCN channels in the lateral habenula regulate pain and comorbid depressive‐like behaviors in mice

doi: 10.1111/cns.14831

Figure Lengend Snippet: Four isoforms of HCN channels are enriched in the LHb. (A–D) Representative images of HCN channels (green), NeuN (red) and merged images of the HCN channel, and NeuN in LHb ( n = 9 slices from 3 mice; Up, scale bar, 200 μm, Bottom, scale bar, 20 μm). White arrows indicate the HCN channel and soma co‐labeled neurons and yellow arrows indicate the HCN channel distributing on the non‐neuronal sites. (A) HCN1; (B) HCN2; (C) HCN3; (D) HCN4. (E–H) Percentage of co‐localized NeuN + and HCN + neurons ( n = 9 slices from 3 mice). (E) HCN1 (48.82 ± 3.550%); (F) HCN2 (86.95 ± 2.160%); (G) HCN3 (87.71 ± 1.576%); (H) HCN4 (34.70 ± 4.315%). Data are shown as mean ± SEM.

Article Snippet: Thereafter, the membranes were incubated with rabbit polyclonal anti‐HCN1 (1:1000; Proteintech, USA), HCN2 (1:1000; proteintech, USA), HCN3 (1:500; Alomone Labs, Israel), HCN4 (1:500; Alomone Labs, Israel), and rabbit polyclonal anti‐β‐actin (1:1000; Proteintech, USA) antibodies overnight at 4°C.

Techniques: Labeling, Immunofluorescence

Journal: CNS Neuroscience & Therapeutics

Article Title: HCN channels in the lateral habenula regulate pain and comorbid depressive‐like behaviors in mice

doi: 10.1111/cns.14831

Figure Lengend Snippet: Increased Ih and up‐regulation of HCN2 isoforms of the LHb in SNI 6w mice. (A) Pie charts illustrating the percentage of LHb neurons without Ih and with Ih in Sham and SNI 6w mice (Sham, n = 59 neurons without Ih and 24 neurons with Ih from 36 mice; SNI, n = 47 neurons without Ih and 32 neurons from 17 mice, p = 0.1210, Chi‐square test). (B) Proportion of three firing types of neurons without Ih and neurons with Ih in the LHb of Sham 6w and SNI 6w mice, respectively (without Ih neurons in Sham 6w mice: n = 26 silent neurons, n = 26 tonic neurons, n = 7 burst neurons; without Ih neurons in SNI 6w mice: n = 14 silent neurons, n = 19 tonic neurons, n = 14 burst neurons from 29 mice, p = 0.0051, Chi‐square test; with Ih neurons in Sham 6w mice: n = 14 silent neurons, n = 8 tonic neurons, n = 2 burst neurons; with Ih neurons in SNI 6w mice: n = 14 silent neurons, n = 13 tonic neurons, n = 5 burst neurons from 21 mice, p = 0.1027, Chi‐square test.). (C) Representative traces of Ih currents of LHb neurons in Sham and SNI 6w mice. (D) The I‐V curves showing the amplitude of Ih responding to different voltages of LHb neurons in Sham and SNI 6w mice (Sham, n = 24 neurons from 12 mice; SNI, n = 31 neurons from 15 mice, F (1.046,21.21) = 13.25, p (−70 mV) = 0.0188, p (−80 mV) = 0.0034, p (−90 mV) = 0.0009, p (−100 mV) = 0.0013, p (−110 mV) = 0.0011, p (−120 mV) = 0.0009, p (−130 mV) = 0.0009, two‐way ANOVA). (E) Amplitude of Ih at −130 mV of LHb neurons in Sham and SNI 6w mice (Sham, n = 24 neurons from 12 mice; SNI, n = 31 neurons from 15 mice, p = 0.0049, Mann–Whitney test). (F) Time constant of Ih of LHb neurons in Sham and SNI 6w mice (Sham, n = 24 neurons from 12 mice; SNI, n = 31 neurons from 15 mice, p < 0.0001, Mann–Whitney test). (G) Cm of LHb neurons with Ih in Sham and SNI 6w mice (Sham, n = 24 neurons from 12 mice; SNI, n = 31 neurons from 15 mice, p = 0.0576, Mann–Whitney test). (H) Ih current density versus voltage in Sham and SNI 6w mice (Sham, n = 24 neurons from 12 mice; SNI, n = 31 neurons from 15 mice, F (0.9957,20.20) = 7.095, p (−60 mV) = 0.0282, p (−70 mV) = 0.0252, p (−80 mV) = 0.0140, p (−90 mV) = 0.0106, p (−100 mV) = 0.0142, p (−110 mV) = 0.0147, p (−120 mV) = 0.0133, p (−130 mV) = 0.0141, two‐way ANOVA). (I) Ih current density at −130 mV in Sham and SNI 6w mice (Sham, n = 24 neurons from 12 mice; SNI, n = 31 neurons from 15 mice, p = 0.0019, Mann–Whitney test). (J) Activation curves in Sham and SNI 6w mice. (K) Half‐activation voltage of Ih in Sham and SNI 6w mice (Sham, n = 11 neurons from 8 mice; SNI, n = 16 neurons from 8 mice, p = 0.0079, Mann–Whitney test). (L–O), Representative images (UP) and quantitative analysis (Bottom) of western blot showing the protein level of HCN1‐4 from Sham and SNI mice. (L) HCN1 ( n = 5 for each group, p = 0.3807, unpaired t ‐test). (M) HCN2 ( n = 7 for each group, p = 0.0305, unpaired t ‐test). (N) HCN3 ( n = 6 for each group, p = 0.3416, unpaired t ‐test). (O) HCN4 ( n = 6 for each group, p = 0.9736, unpaired t ‐test). Data are shown as mean ± SEM or Min to Max. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: Thereafter, the membranes were incubated with rabbit polyclonal anti‐HCN1 (1:1000; Proteintech, USA), HCN2 (1:1000; proteintech, USA), HCN3 (1:500; Alomone Labs, Israel), HCN4 (1:500; Alomone Labs, Israel), and rabbit polyclonal anti‐β‐actin (1:1000; Proteintech, USA) antibodies overnight at 4°C.

Techniques: Chi-Square, MANN-WHITNEY, Activation Assay, Analysis, Western Blot, Patch Clamp

Journal: Journal of neurophysiology

Article Title: The HCN4 channel modulates external tufted cell burst firing

doi: 10.1152/jn.00194.2026

Figure Lengend Snippet: Effect of intracellular delivery of HCN blocking antisera on external tufted cell firing. A : External tufted cell firing 1 minute after whole cell configuration baseline ( top ). After 6 min intracellular diffusion with HCN4 blocking antisera external tufted cell spiking was terminated in this cell ( bottom ); representative biocytin filled external tufted cell shown. B : External tufted cell firing 1 minute after whole cell configuration baseline ( top ). After 6 min (shown here, to longer time 30 min recorded in this cell) intracellular diffusion with HCN2 blocking antisera external tufted cell spiking was unaffected ( bottom ). C : Group data showing firing rates with blocking antisera represented as a percent of the pre-equilibration baseline.

Article Snippet: Membrane blots were incubated in TBST buffer with 5% powder milk for 1 h, and then probed in 4 °C overnight with one of the following primary antibodies in same buffer: rabbit anti-HCN1 (#APC-056, 1:200), -HCN2 (#APC-030, 1:200), -HCN3 (#APC-057, 1:200) and -HCN4 (#APC-052, 1:200) antibody (Alomone Labs, Jerusalem, Israel) and rabbit anti-β-actin (1:2500; Cell Signaling Technology, Danvers, MA, USA).

Techniques: Blocking Assay, Diffusion-based Assay

Journal: The Journal of Neuroscience

Article Title: Persistent Discharges in Dentate Gyrus Perisoma-Inhibiting Interneurons Require Hyperpolarization-Activated Cyclic Nucleotide-Gated Channel Activation

doi: 10.1523/JNEUROSCI.3671-14.2015

Figure Lengend Snippet: Persistent firing (PF) requires HCNC activation. A, Left, Confocal image from a biocytin-filled PII. Right, Antibodies against HCN1 label PII soma and axon collaterals. B, Left, Traces show persistent firing activity in the control condition (gray) and after bath application of 30 μm ZD7288 (blue). Coapplication of ZD7288 and 5 mm K+ does not recover persistent firing (red). Top right, Hyperpolarizing current injection in a PII (−200 pA, 1 s) in the control condition and in the presence of ZD7288. Bottom right, Probability of evoking antidromic action potentials (AP) as a function of extracellular stimulus intensity in control (ctrl; gray), in the presence of ZD7288 (blue), or in the presence of ZD7288 and 5 mm K+ (red). Note that elevated K+ compensates the effects of ZD7288 on axonal excitability. C–E, Bar graphs summarize the influence of ZD7288 (blue) or ZD7288 with K+ (C, red), 10–30 μm ivabradine (Ivab), 200 μm Cd2+, and intracellularly applied 10 mm BAPTA on PF duration or number of action potentials generated during persistent firing. Circles connected by lines represent individual experiments. Bars with lines represent mean ± SEM. *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001.

Article Snippet: Selected slices were incubated with primary antibodies against PV (polyclonal rabbit, 1:1000; Swant), HCN1 (rabbit polyclonal, 1:100; Alomone Labs), or Ankyrin-G (mouse, 1:100; Life Technologies) in PBS containing 10% goat serum and 0.2% Triton X-100 for 36 h (4°C).

Techniques: Activation Assay, Confocal, Activity Assay, Control, Injection, Generated, Immunofluorescence, Patch Clamp

Journal: Communications Biology

Article Title: Primary dendrites of mitral cells synapse unto neighboring glomeruli independent of their odorant receptor identity

doi: 10.1038/s42003-018-0252-y

Figure Lengend Snippet: Matching of mitral cells (MCs) with the CNG − glomeruli. a MOR29A glomeruli in the hemizygous female CNG-A2. Image: mitral-cell dendrites are compared between the CNG-positive (+) and -negative (–) glomeruli with the same odorant receptor identity, MOR29A. OSN axons of MOR29A glomeruli (magenta for CFP) and connecting MCs (green for YFP) were analyzed by two-photon laser microscopy. Three-dimensional videos are available in the supplementary information. GL, glomerular layer; EPL, external plexiform layer: MCL, mitral cell layer; FP, fluorescent protein. Scale bar, 100 μm. Graph: The glomerular sizes and numbers of connecting MCs are compared between the CNG + and CNG − glomeruli at P1, 7, and 14. n = 3 animals, n = 5–8 glomeruli for each time point. Error bar, S.E.; n.s., p > 0.05; *0.05 > p > 0.01; ** p < 0.01; Welch’s t -test. b MOR29B glomeruli in the hemizygous female CNG-A2 KO. Dendrite connection and synapse formation of MCs were compared between the CNG + and CNG − glomeruli of MOR29B as shown for the MOR29A glomeruli in a . MOR29B glomeruli are shown in green, because they were labeled with YFP whose fluorescence cannot be distinguished from that of YFP for MCs. c Synapse formation in the CNG + and CNG − glomeruli. Olfactory bulb (OB) sections containing the MOR29A glomeruli at P14 were immunostained with antibodies against synaptic markers, Synaptophysin (Syn, pre-synaptic marker) and post-synaptic density 95 (PSD95, post-synaptic marker). Serial OB sections were stained with antibodies against CNG-A2 (CNG-channel marker) and Pcdh21 (M/T-cell marker). No differences were found in staining between the CNG + and CNG − glomeruli. Scale bar, 100 μm. Error bar, S.E.; n.s., p > 0.05; Welch’s t -test

Article Snippet: Antibodies used in this study are as follows: rabbit anti-CNG-A2 antibodies (1:200, Alomone Labs); rabbit anti-PSD95 antibodies (1:1000, Cell Signaling); mouse anti-Synaptophysin (Syn) antibodies (1:1000, Millipore); goat anti-OMP antibodies (1:1000, Wako Pure Chemical); rabbit anti-NQO1 antibodies (1:300, Abcam); guinea pig anti-vGlut2 antibodies (1:300, Millipore); rabbit anti-GluR1 antibodies (1:300, Abcam); rabbit anti-GFP antibodies (1:300, Invitrogen); chicken anti-GFP antibodies (1:300, Abcam).

Techniques: Microscopy, Labeling, Fluorescence, Olfactory, Marker, Staining, Immunofluorescence

Journal: Communications Biology

Article Title: Primary dendrites of mitral cells synapse unto neighboring glomeruli independent of their odorant receptor identity

doi: 10.1038/s42003-018-0252-y

Figure Lengend Snippet: Activity-dependent degeneration and regeneration of synapses. a Generation of empty glomeruli. Coronal olfactory bulb (OB) sections (8 weeks) from the WT and CNG-A2 +/− female mice were stained with anti-OMP antibodies to detect OSN axons. Due to the competition in the hemizygous KO, CNG − OSNs were eliminated and empty glomeruli (encircled) devoid of OSN axons were generated. Sections were also stained with DAPI. n = 5 animals. GL, glomerular layer; EPL, external plexiform layer; MCL, mitral cell layer. Scale bar, 50 μm. b Refilling of empty glomeruli. Coronal sections (15 weeks) of the WT and CNG-A2 +/− female mice were immunostained with antibodies against OMP and CNG-A2. Refilled glomeruli (CNG − ) are encircled. CNG + glomeruli are encircled by dotted lines. n = 5 animals. GL, glomerular layer; EPL, external plexiform layer; MCL, mitral cell layer. Scale bar, 50 μm. c Septum structures in the refilled glomeruli. Coronal sections of the Tg MOR29A mice (15 weeks) with the background of CNG-A2 +/− KO were immunostained with antibodies against GFP, CNG-A2 and OMP. Sections were also stained with DAPI. Glomerular structures are encircled. In the refilled MOR29A glomeruli, septum structures (dotted line) were often formed separating the CNG + and CNG − OSN axons of regenerated MOR29A + OSNs (i). In some cases, the vacated CNG − glomeruli were refilled with CNG + axons, mostly MOR29A + , without forming a septum (ii), or with MOR29A and other odorant receptor species forming a septum (iii). d Rations of the CNG + and CNG − glomeruli. MOR29A glomeruli were analyzed at 2, 8, and 15 weeks. At 8 weeks, most CNG − MOR29A glomeruli were devoid of OSN axons. e Schematic diagrams of degeneration and regeneration of CNG + and CNG − axons. CNG − axons are eliminated due to the competition with CNG + axons and the vacated CNG − glomeruli are partly occupied by CNG + OSN axons forming the septum structure within glomeruli. GL, glomerular layer; EPL, external plexiform layer; MCL, mitral cell layer. Scale bars, 50 μm

Article Snippet: Antibodies used in this study are as follows: rabbit anti-CNG-A2 antibodies (1:200, Alomone Labs); rabbit anti-PSD95 antibodies (1:1000, Cell Signaling); mouse anti-Synaptophysin (Syn) antibodies (1:1000, Millipore); goat anti-OMP antibodies (1:1000, Wako Pure Chemical); rabbit anti-NQO1 antibodies (1:300, Abcam); guinea pig anti-vGlut2 antibodies (1:300, Millipore); rabbit anti-GluR1 antibodies (1:300, Abcam); rabbit anti-GFP antibodies (1:300, Invitrogen); chicken anti-GFP antibodies (1:300, Abcam).

Techniques: Activity Assay, Olfactory, Staining, Generated, Immunofluorescence

Journal: iScience

Article Title: SLC17A9-PTHLH-EMT axis promotes proliferation and invasion of clear renal cell carcinoma

doi: 10.1016/j.isci.2022.105764

Figure Lengend Snippet: SLC17A9 expression level was upregulated in ccRCC tissues (A) SLC17A9 mRNA expression levels in the ccRCC tissues were higher than those in the normal tissues in the TCGA cohort. (B) SLC17A9 expression in normal tissues was lower than that in paired ccRCC tissues. (C) High expression of SLC17A9 was associated with the T stage and showed an increasing trend. (D) SLC17A9 expression levels in N0 and N1 stages. (E) mRNA expression of SLC17A9 in M stage. (F) SLC17A9 expression levels in different TNM stages in TCGA. (G) SLC17A9 expression levels in G grade. (H) SLC17A9 was upregulated in tumor samples compared with normal samples in the ICGC-RECA-EU cohort. (I) SLC17A9 expression levels in tumor samples were higher than those in paired normal kidney tissues. ∗∗∗∗p < 0.0001; ∗∗∗p < 0.001; ∗∗p < 0.01; and ∗p < 0.05. SLC17A9, Solute Carrier Family 17 Member 9; ccRCC, clear cell renal cell carcinoma; KIRC, kidney renal clear cell carcinoma; TCGA, The Cancer Genome Atlas; and ICGC, International Cancer Genome Consortium. Data are represented as mean ± SD.

Article Snippet: Next, the sections were incubated with rabbit SLC17A9 primary antibody (1:100; 26731-1-AP; Proteintech, USA) at 4 °C overnight.

Techniques: Expressing

Journal: iScience

Article Title: SLC17A9-PTHLH-EMT axis promotes proliferation and invasion of clear renal cell carcinoma

doi: 10.1016/j.isci.2022.105764

Figure Lengend Snippet: Diagnostic and prognostic values of SLC17A9 (A) SLC17A9 effectively distinguished para-cancer tissues from ccRCC tissues in TCGA-KIRC (AUC = 0.9459, p < 0.0001). (B) SLC17A9 effectively distinguished para-cancer tissues from ccRCC tissues in ICGC (AUC = 0.9228, p < 0.0001). (C) Diagnostic value of SLC17A9 for ccRCC patients in TCGA-KIRC increased with time. AUC at 2 years was 0.6691, AUC at 4 years was 0.6728, and AUC at 6 years was 0.6864. (D) High SLC17A9 expression group had poorer OS than the low expression group in the TCGA cohort, p < 0.0001. (E) High SLC17A9 expression indicated poorer DFS in the TCGA cohort, p = 0.0059. (F) Survival analysis of E-MTAB-1980 validation cohort with low SLC17A9 expression group presenting better survival status, p = 0.0247. (G) Construction of a nomogram for predicting 1-, 3-, and 5-year OS possibilities of individual ccRCC patients. (H–J) Calibration curves of (H) 1-year, (I) 3-year, and (J) 5-year OS of ccRCC patients. Light-colored diagonal represented predicted results, whereas the red-colored broken line represented actual results. Data are represented as mean ± SD.

Article Snippet: Next, the sections were incubated with rabbit SLC17A9 primary antibody (1:100; 26731-1-AP; Proteintech, USA) at 4 °C overnight.

Techniques: Diagnostic Assay, Expressing, Biomarker Discovery

Journal: iScience

Article Title: SLC17A9-PTHLH-EMT axis promotes proliferation and invasion of clear renal cell carcinoma

doi: 10.1016/j.isci.2022.105764

Figure Lengend Snippet: Univariate and multivariate analyses of SLC17A9 expression and patient overall survival using TCGA-KIRC data (n = 522)

Article Snippet: Next, the sections were incubated with rabbit SLC17A9 primary antibody (1:100; 26731-1-AP; Proteintech, USA) at 4 °C overnight.

Techniques: Expressing

Journal: iScience

Article Title: SLC17A9-PTHLH-EMT axis promotes proliferation and invasion of clear renal cell carcinoma

doi: 10.1016/j.isci.2022.105764

Figure Lengend Snippet: Univariate and multivariate analyses of SLC17A9 expression and patient disease free survival. using TCGA-KIRC data (n = 522)

Article Snippet: Next, the sections were incubated with rabbit SLC17A9 primary antibody (1:100; 26731-1-AP; Proteintech, USA) at 4 °C overnight.

Techniques: Expressing

Journal: iScience

Article Title: SLC17A9-PTHLH-EMT axis promotes proliferation and invasion of clear renal cell carcinoma

doi: 10.1016/j.isci.2022.105764

Figure Lengend Snippet: SLC17A9 was upregulated in ccRCC cell lines and tissues (A and B) Relative mRNA and protein levels of SLC17A9 in ccRCC cell lines. (C and D) Relative mRNA and protein levels of SLC17A9 in ccRCC tissues and adjacent normal kidney tissues. (E) Immunohistochemistry (IHC) results (200x,400x) of SLC17A9 in ccRCC tissues and para-cancer tissues. ∗∗∗∗p < 0.0001; ∗∗∗p < 0.001; ∗∗p < 0.01; and ∗p < 0.05. Data are represented as mean ± SD. The length of scale bars are 50 and 20 μm.

Article Snippet: Next, the sections were incubated with rabbit SLC17A9 primary antibody (1:100; 26731-1-AP; Proteintech, USA) at 4 °C overnight.

Techniques: Immunohistochemistry

Journal: iScience

Article Title: SLC17A9-PTHLH-EMT axis promotes proliferation and invasion of clear renal cell carcinoma

doi: 10.1016/j.isci.2022.105764

Figure Lengend Snippet: Effects of SLC17A9 on the proliferation of renal cancer cells (A and B) Successful knockdown of SLC17A9 mRNA in 786-O and A498 cells. (C and D) SLC17A9 mRNA levels were significantly overexpressed in 786-O and A498 cells. (E and F) Protein levels after SLC17A9 knockdown in 786-O and A498 cells. (G and H) Protein levels after SLC17A9 overexpression in 786-O and A498 cells. (I–L) Cell counting kit-8 assays detected the effects of SLC17A9 overexpression and knockdown on the proliferation of 786-O and A498 cells. ∗∗∗∗p < 0.0001; ∗∗∗p < 0.001; ∗∗p < 0.01; and ∗p < 0.05. Data were presented as the mean ± standard deviation (SD) from three independent experiments.

Article Snippet: Next, the sections were incubated with rabbit SLC17A9 primary antibody (1:100; 26731-1-AP; Proteintech, USA) at 4 °C overnight.

Techniques: Knockdown, Over Expression, Cell Counting, Standard Deviation

Journal: iScience

Article Title: SLC17A9-PTHLH-EMT axis promotes proliferation and invasion of clear renal cell carcinoma

doi: 10.1016/j.isci.2022.105764

Figure Lengend Snippet: SLC17A9 affected renal cancer cells via EMT signaling pathways (A) GSEA results of SLC17A9 using TCGA data. (B) Correlation analyses of Snail, N-cadherin, E-cadherin, and vimentin with SLC17A9 were performed using the TCGA-KIRC data. (C–F) Effects of SLC17A9 overexpression and knockdown on E-cadherin, N-cadherin, vimentin, and snail-1 in renal cancer cells. (G–J) Effects of SLC17A9 overexpression and knockdown on cell migration and invasion. ∗∗∗∗p < 0.0001. Data are represented as mean ± SD. The length of scale bar is 100 μm.

Article Snippet: Next, the sections were incubated with rabbit SLC17A9 primary antibody (1:100; 26731-1-AP; Proteintech, USA) at 4 °C overnight.

Techniques: Protein-Protein interactions, Over Expression, Knockdown, Migration

Journal: iScience

Article Title: SLC17A9-PTHLH-EMT axis promotes proliferation and invasion of clear renal cell carcinoma

doi: 10.1016/j.isci.2022.105764

Figure Lengend Snippet: SLC17A9-activated EMT pathway was dependent on PTHLH (A) Four potential downstream genes of SLC17A9. (B and C) Regulation of PTHLH by SLC17A9 was verified at the mRNA level. (D) Regulation of PTHLH by SLC17A9 was verified at the protein level. (E and F) Knockdown of PTHLH could reverse the effect of SLC17A9 overexpression on EMT markers. (G and H) Migration and invasion assays (200x) for the indicated ccRCC cells. ∗∗∗∗p < 0.0001; ∗∗∗p < 0.001; ∗∗p < 0.01; and ∗p < 0.05. Data are represented as mean ± SD. The length of scale bar is 100 μm.

Article Snippet: Next, the sections were incubated with rabbit SLC17A9 primary antibody (1:100; 26731-1-AP; Proteintech, USA) at 4 °C overnight.

Techniques: Knockdown, Over Expression, Migration

Journal: iScience

Article Title: SLC17A9-PTHLH-EMT axis promotes proliferation and invasion of clear renal cell carcinoma

doi: 10.1016/j.isci.2022.105764

Figure Lengend Snippet: Upregulation of SLC17A9 promoted tumor progression in vivo (A) Prediction of SLC17A9 associated drugs. (B and C) IC 50 of vorinostat in A498 and 786-O. (D) Protein expression levels of SLC17A9 in cells cultivated at different vorinostat concentrations. (E–I) Subcutaneous tumor in mice, its growth curve, and final volume and weight (n = 5). ∗∗∗∗p < 0.0001; ∗∗∗p < 0.001; ∗∗p < 0.01; and ∗p < 0.05. Data are represented as mean ± SD.

Article Snippet: Next, the sections were incubated with rabbit SLC17A9 primary antibody (1:100; 26731-1-AP; Proteintech, USA) at 4 °C overnight.

Techniques: In Vivo, Expressing

Journal: iScience

Article Title: SLC17A9-PTHLH-EMT axis promotes proliferation and invasion of clear renal cell carcinoma

doi: 10.1016/j.isci.2022.105764

Figure Lengend Snippet:

Article Snippet: Next, the sections were incubated with rabbit SLC17A9 primary antibody (1:100; 26731-1-AP; Proteintech, USA) at 4 °C overnight.

Techniques: Recombinant, Software

Journal: Discover. Oncology

Article Title: Pyroptosis-related genes regulate proliferation and invasion of pancreatic cancer and serve as the prognostic signature for modeling patient survival.

doi: 10.1007/s12672-022-00495-0

Figure Lengend Snippet: Fig. 5 Expression and survival analysis of PRGs in PDAC. A The protein expression score of staining and intensity of NLRP1 and CASP4 pro- teins in all PDAC samples (HPA database). Protein expression score is based on immunohistochemical data manually scored with regard to staining intensity (negative, weak, moderate or strong) and fraction of stained cells (< 25%, 25–75% or > 75%). B Immunohistochemi- cal staining of NLRP1 and CASP4 proteins in PDAC. C Methylation levels of CASP4 at chr11:104838825–104841325 and chr11:104826922– 104829422 in PDAC and para-cancer tissues. D The OS survival curves of NLRP1 and CASP4 mRNA expression level in PDAC. E CASP4 and NLRP1 immunohistochemistry for patients from Zhongnan Hospital (×200)

Article Snippet: To test whether the expression of CASP4 or NLRP1 affects the sensitivity of pancreatic cancer cells to MEK inhibitors, Trametinib (MedChemExpress, USA) was added to siRNA-transfected PANC-1 and Aspc-1 cells.

Techniques: Expressing, Staining, Immunohistochemical staining, Methylation, Immunohistochemistry

Journal: Discover. Oncology

Article Title: Pyroptosis-related genes regulate proliferation and invasion of pancreatic cancer and serve as the prognostic signature for modeling patient survival.

doi: 10.1007/s12672-022-00495-0

Figure Lengend Snippet: Fig. 6 PRGs regulated the proliferation and invasion of pancreatic cancer cells in vitro. A, B After siRNA transfection, qPCR and western blot was used to detect RNA and protein expression levels of CASP4 and NLRP1 respectively in PANC-1 and Aspc-1 cells. C CCK8 assay was used to detect the proliferation of PANC-1 and Aspc-1 cells after transfection with siRNA. D Transwell was used to detect the change of cell inva- sion ability after transfection with siRNA (*P < 0.05, **P < 0.01, ***P < 0.001)

Article Snippet: To test whether the expression of CASP4 or NLRP1 affects the sensitivity of pancreatic cancer cells to MEK inhibitors, Trametinib (MedChemExpress, USA) was added to siRNA-transfected PANC-1 and Aspc-1 cells.

Techniques: In Vitro, Transfection, Western Blot, Expressing, CCK-8 Assay

Journal: Discover. Oncology

Article Title: Pyroptosis-related genes regulate proliferation and invasion of pancreatic cancer and serve as the prognostic signature for modeling patient survival.

doi: 10.1007/s12672-022-00495-0

Figure Lengend Snippet: Fig. 9 NLRP1 was associated with KRAS and P53 mutations potentially. A The expression levels of NLRP1 were different in KRAS mutation, P53 mutation and wild-type tumor tissues respectively. B Correlation curve between NLRP1, KRAS and P53 expression levels based on TCGA database. C GSEA analysis of DEGs in high NLRP1 expression group. D PANC-1 and Aspc-1 cells were tested for LDH release 24 h after siNLRP1 transfection. E Cleavage of GSDMD and p-ERK were monitored by immunoblot analysis 48 h after siNLRP1 transfection (*P < 0.05, ***P < 0.001)

Article Snippet: To test whether the expression of CASP4 or NLRP1 affects the sensitivity of pancreatic cancer cells to MEK inhibitors, Trametinib (MedChemExpress, USA) was added to siRNA-transfected PANC-1 and Aspc-1 cells.

Techniques: Expressing, Mutagenesis, Transfection, Western Blot

Journal: Discover. Oncology

Article Title: Pyroptosis-related genes regulate proliferation and invasion of pancreatic cancer and serve as the prognostic signature for modeling patient survival.

doi: 10.1007/s12672-022-00495-0

Figure Lengend Snippet: Fig. 10 Analysis of drug sensitivity associated with PEGs. A Correlation of CASP4 and NLRP1 expression with multidrug sensitivity in GDSC. Red is positive correlation, which means the higher the gene expression, the more sensitive to the drug, while the blue is the opposite. The Spearman correlation represent the gene expression correlates with the drug. The positive correlation means that the gene high expression is resistant to the drug, vise verse. B Survival rate of cells exposed to trametinib at the concentrations of 10 µM/L for 48 or 72 h (*P < 0.05, **P < 0.01, ***P < 0.001)

Article Snippet: To test whether the expression of CASP4 or NLRP1 affects the sensitivity of pancreatic cancer cells to MEK inhibitors, Trametinib (MedChemExpress, USA) was added to siRNA-transfected PANC-1 and Aspc-1 cells.

Techniques: Expressing, Gene Expression