Journal: Genetics and molecular research : GMR

Article Title: Increased survivin expression and its association with clinical parameters of congenital choledochal cysts.

doi: 10.4238/gmr.15028032

Figure Lengend Snippet: Figure 1. Immunohistochemical staining results for the protein expression of survivin. The cytoplasm showed high survivin expression, which presented as granular form, with yellow or dark brown staining, while the nuclei were hardly stained (A. cytoplasm, 200X; B. nuclei, 400X).

Article Snippet: The primary mouse anti-human survivin monoclonal antibody was purchased from Wuhan Boster Biological Engineering Co., Ltd. (China).

Techniques: Immunohistochemical staining, Staining, Expressing

Journal: Cellular and Molecular Life Sciences

Article Title: S100A16 promotes acute kidney injury by activating HRD1-induced ubiquitination and degradation of GSK3β and CK1α

doi: 10.1007/s00018-022-04213-5

Figure Lengend Snippet: HRD1 involves in the activation of Wnt/β-catenin signaling pathway and is regulated by S100A16 in injured renal fibroblasts. a Representative micrographs in the corticomedullary junction of mice kidneys showed the expression of HRD1 in WT mice and S100A16 +/− mice at 1 day after IRI, as determined by immunohistochemical staining. Scale bar, 50 μm, 20 μm (Enlarged). b HRD1, GSK3β and CK1α expressions were tested using kidney tissues from WT mice and S100A16 +/− mice at 1 day after IRI compared with sham mice by western blot assays. c Quantitation of immunoblot data for HRD1, GSK3β and CK1α proteins as in b . **** P < 0.0001, *** P < 0.001, ** P < 0.01, * P < 0.05. n = 3; each point represents the expression in a sample pooled from two mice. d Western blot analyses showed that ICG-001 blocked the increased HRD1 expression induced by H/R in NRK-49F cells, and ICG-001 also recovered the expressions of GSK3β and CK1α. Cell lysates after various treatments, as indicated, were immunoblotted with antibodies against HRD1, GSK3β, CK1α and β-actin. e Quantitation of western blot data for HRD1, GSK3β and CK1α proteins as in d . *** P < 0.001, ** P < 0.01, versus control; ## P < 0.01, # P < 0.05, n.s. not significant, versus H/R alone. n = 3. f Western blot analyses showed that knockdown of S100A16 inhibited the HRD1 expression and enhanced the expressions of GSK3β and CK1α in normal or hypoxia condition. g Quantitation of western blot data for HRD1, GSK3β and CK1α proteins as in f . *** P < 0.001, ** P < 0.01, versus scrambled shRNA; ## P < 0.01, versus H/R + scrambled shRNA. n = 3. h Western blots showed that overexpression of S100A16 increased the HRD1 expression, but it was impeded by ICG-001 in NRK-49F cells. The GSK3β and CK1α expressions were opposite to HRD1. i Quantified HRD1, GSK3β and CK1α protein levels in h . ** P < 0.01, versus pcDNA3.1; # P < 0.05, versus S100A16 OE. n = 3

Article Snippet: The primary antibodies against S100A16 (HPA045841; Sigma-Aldrich, St Louis, MO, USA), β-catenin (610153; BD Biosciences, San Jose, CA, USA), HRD1 (13473-1-AP; Proteintech, Chicago, IL, USA) were used with 1:100 dilution.

Techniques: Activation Assay, Expressing, Immunohistochemical staining, Staining, Western Blot, Quantitation Assay, Control, Knockdown, shRNA, Over Expression

Journal: Cellular and Molecular Life Sciences

Article Title: S100A16 promotes acute kidney injury by activating HRD1-induced ubiquitination and degradation of GSK3β and CK1α

doi: 10.1007/s00018-022-04213-5

Figure Lengend Snippet: HRD1 degrades both GSK3β and CK1α via the ubiquitin–proteasome pathway in NRK-49F cells. a The interaction between HRD1 and either GSK3β or CK1α was detected in the co-IP analysis in NRK-49F cells. b A cycloheximide chase was performed to establish the time course of GSK3β or CK1α biogenesis. NRK-49F cells were infected with or without Ad-HRD1 for 48 h, and then the cells were treated with CHX (100 μg/ml) for 0, 2,4 or 6 h. The expressions of GSK3β and CK1α in whole-cell lysates were measured by western blotting. c Quantitation of western blot data for GSK3β and CK1α proteins as in b . * P < 0.05. n = 3. d The presence of MG132 (20 µM) increased the expressions of GSK3β and CK1α with or without Ad-HRD1 infection compared with controls in NRK-49F cells. e Quantification of GSK3β and CK1α protein expressions from experiments as shown in d . *** P < 0.001, ** P < 0.01, * P < 0.05. n = 3. f More ubiquitin conjugated to GSK3β was detected in the cells overexpressing HRD1 compared with no HRD1 transfection. g More ubiquitin conjugated to CK1α was detected in the cells overexpressing HRD1 compared with no HRD1 transfection. h Quantification of ubiquitin conjugated to GSK3β as in f , normalized to GAPDH expression. *** P < 0.001. n = 3. i Quantification of ubiquitin conjugated to CK1α as in g , normalized to GAPDH expression. ** P < 0.01. n = 3

Article Snippet: The primary antibodies against S100A16 (HPA045841; Sigma-Aldrich, St Louis, MO, USA), β-catenin (610153; BD Biosciences, San Jose, CA, USA), HRD1 (13473-1-AP; Proteintech, Chicago, IL, USA) were used with 1:100 dilution.

Techniques: Ubiquitin Proteomics, Co-Immunoprecipitation Assay, Infection, Western Blot, Quantitation Assay, Transfection, Expressing

Journal: Cellular and Molecular Life Sciences

Article Title: S100A16 promotes acute kidney injury by activating HRD1-induced ubiquitination and degradation of GSK3β and CK1α

doi: 10.1007/s00018-022-04213-5

Figure Lengend Snippet: S100A16 down-regulates the expressions of both GSK3β and CK1α by HRD1 to affect downstream HGF in NRK-49F cells. a Western blot analyses showed that overexpression of S100A16 aggravated the downregulated expressions of GSK3β, CK1α, and HGF induced by Ad-HRD1 in NRK-49F cells. b Quantitation of western blot data for GSK3β, CK1α, and HGF proteins, as in a . * P < 0.05, versus pcDNA3.1; ## P < 0.01, versus pcDNA3.1 + Ad-HRD1. n = 3. c Real-time qPCR demonstrated that overexpression of S100A16 aggravated the reduction of relative HGF/β-actin mRNA level induced by Ad-HRD1 compared with the Ad-HRD1 treatment alone in NRK-49F cells. ** P < 0.01, versus pcDNA3.1; # P < 0.05, versus pcDNA3.1 + Ad-HRD1. n = 3. d Western blot analyses showed that knockdown S100A16 recovered Ad-HRD1-induced the down-regulated expressions of GSK3β, CK1α, and HGF. e Quantitation of western blot data for GSK3β, CK1α and HGF proteins, as in d . ** P < 0.01, * P < 0.05, versus scrambled shRNA; ## P < 0.01, versus scrambled shRNA + Ad-HRD1. n = 3. f Representative images showed the accumulation of β-catenin fluorescence in the nucleus of HRD1-overexpressing NRK-49F cells. The β-catenin fluorescence was amplified after overexpressing S100A16 in the HRD1-overexpressing NRK-49F cells. Scale bar, 20 μm

Article Snippet: The primary antibodies against S100A16 (HPA045841; Sigma-Aldrich, St Louis, MO, USA), β-catenin (610153; BD Biosciences, San Jose, CA, USA), HRD1 (13473-1-AP; Proteintech, Chicago, IL, USA) were used with 1:100 dilution.

Techniques: Western Blot, Over Expression, Quantitation Assay, Knockdown, shRNA, Fluorescence, Amplification

Journal: Cellular and Molecular Life Sciences

Article Title: S100A16 promotes acute kidney injury by activating HRD1-induced ubiquitination and degradation of GSK3β and CK1α

doi: 10.1007/s00018-022-04213-5

Figure Lengend Snippet: The Wnt/β-catenin signaling activation induced by S100A16–HRD1–GSK3β/CK1α pathway in renal fibroblasts under IRI condition promotes the occurrence of AKI. Schematic diagram shows the increased expression of S100A16 in fibroblasts during renal ischemia and hypoxia. The expression of the E3 ubiquitin ligase HRD1 is elevated, and the members of β-catenin degradation complex, GSK3β and CK1α, are destroyed via the ubiquitin–proteasome pathway, thereby promoting the accumulation of β-catenin and its transfer to the cell nucleus. Activation of the Wnt/β-catenin signaling pathway then inhibits the transcription of the downstream HGF that secreted by fibroblasts, and this procedure eventually leads to unrepairable kidney injury and renal dysfunction

Article Snippet: The primary antibodies against S100A16 (HPA045841; Sigma-Aldrich, St Louis, MO, USA), β-catenin (610153; BD Biosciences, San Jose, CA, USA), HRD1 (13473-1-AP; Proteintech, Chicago, IL, USA) were used with 1:100 dilution.

Techniques: Activation Assay, Expressing, Ubiquitin Proteomics

Journal: OncoTargets and therapy

Article Title: Piwi-Interacting RNA1037 Enhances Chemoresistance and Motility in Human Oral Squamous Cell Carcinoma Cells

doi: 10.2147/OTT.S233322

Figure Lengend Snippet: Targeting the upregulated expression of piR-1037 induced by CDDP enhanced the sensitivity of OSCC cells to CDDP. ( A ) Confirmation of the suppressive effect of piRNA-1037 antisense oligonucleotides (piR-1037 complementary DNA oligonucleotides) (500 nM) on the expression of piRNA-1037 in SCC4 and SCC9 cells receiving the indicated treatments. ( B ) Western blot analysis and quantification of the effect of piR-1037 antisense oligonucleotides on the expression of the chemoresistance biomarkers MDR1 and α-enolase in SCC4 and SCC9 cells treated with CDDP (10 μM). ( C ) Decreased cell viability and increased apoptosis (TUNEL assay) ( D ) in OSCC cells transfected with piR-1037 antisense oligonucleotides and treated with CDDP compared with CDDP-treated cells transfected with a scrambled control. Cell apoptosis was detected using the TUNEL-based TiterTACS™ Colorimetric Apoptosis Detection Kit (Trevigen, USA). ( E ) Activity of caspase-3, caspase-8 and caspase-1 ( F ). The activity of caspases measured using Caspase-3 and Caspase-1 assay kits (Abcam, USA) and a Caspase-8 colorimetric kit (Sigma Aldrich, USA). The data represent the mean ±SD from at least three independent replicates. Statistical analysis was performed using one-way ANOVA analysis or unpaired student’s t -test: * p <0.05; ** p < 0.01.

Article Snippet: The membrane was then incubated with 5% nonfat milk in TBST (10 mM Tris, pH 8.0, 150 mM NaCl, 0.5% Tween 20) for 1 h. The membrane was washed once with TBST, followed by incubation with primary antibodies against MDR1 (Novus Biologicals, USA) (1:1000), α-enolase (Boster, China) (1:1000), XIAP (Abcam, USA) (1 μg/mL), cleaved caspase-3 (R&D systems, USA) (0,5 μg/mL), E-Cadherin (1:500), N-Cadherin (1:1000) (Abcam, USA), and β-actin (1:10,000) (Sigma Aldrich, USA).

Techniques: Expressing, Western Blot, TUNEL Assay, Transfection, Control, Activity Assay

Journal: Nature Communications

Article Title: Macrophage lineage cells-derived migrasomes activate complement-dependent blood-brain barrier damage in cerebral amyloid angiopathy mouse model

doi: 10.1038/s41467-023-39693-x

Figure Lengend Snippet: a Plasma CD5L concentration of CAA patients ( N = 12) and HC ( N = 12) as well as CAA models ( N = 5 in each group) and WT C57/BL6 mice ( N = 5). Data are presented as mean values ± SEM with the indicated significance (by two-tailed Student’s t test). b Left: CD5L deposition in mice brain was evaluated with immunostaining. Right: Immunostaining of CD31 (red), CD5L (green), C5b-9 (blue) and TSPAN4 (purple). Data are representative of 4 biologically independent experiments. c Plasma C5-9 concentration of mice. N = 6 in PBS-M, Aβ40-M and CD5L KO Aβ40-M transferred mice and N = 5 in CD5L KO PBS-M treated group. Data are presented as mean values ± SEM with the indicated significance (by one-way ANOVA followed by Tukey’s post-test). d C5b-9 deposition in brain as assessed with immunostaining. Data are representative of 3 biologically independent experiments. e Fluorescent intensity of 3kDa-Dextran in the plasma and eyeballs of the recipients. N = 6 in PBS-M, Aβ40-M and CD5L KO Aβ40-M transferred mice and N = 5 in CD5L KO PBS-M treated group. Data are presented as mean values ± SEM with the indicated significance (by one-way ANOVA followed by Tukey’s post-test). f Leakage of 3kDa-Dextran in brain parenchyma was assessed with fluorescent microscopy. Data are representative of 3 biologically independent experiments. g Coronal brain sections of the recipients were subjected to immunostaining of CD31 (green) and ZO-1 (red). Data are representative of 3 biologically independent experiments. h Skin autopsy sample of a CAA patient (Left) and brain tissue of WT and Tg-SwDI/B +/+ mice (24w of age) (Right) were subjected to IEM with CD5L staining. Blue dash lines outline micro-vessel wall. Red stars emphasize blood vessel lumen. Red arrow heads emphasize peri-vessel macrophages. Red arrows emphasize CD5L deposition in blood vessels within migrasome-like structures. Data are representative of 3 biologically independent experiments. Source data are provided as a Source Data file.

Article Snippet: Concentration of mouse NEFL (FineTest, EM1688), mouse C5b-9 (FineTest, EM1392), mouse CD5L (Boster, EK1414), human CD5L (MEIMIAN, MM-50587H1), human Aβ40 (CUSABIO, CSB-E08299h), human Aβ42 (MAI Bio, LM-2685H), and human C5b-9 (FineTest, EH3858) in plasma was assessed with commercial kits according to instructions of manufacturers.

Techniques: Clinical Proteomics, Concentration Assay, Two Tailed Test, Immunostaining, Microscopy, Staining

Journal: Translational Andrology and Urology

Article Title: Identification of BIRC5 and HMMR as prognostic biomarkers for immune infiltration in prostate cancer

doi: 10.21037/tau-24-359

Figure Lengend Snippet: Association between the expression of BIRC5 and HMMR genes and cancer prognosis. (A) Venn diagram revealing the identification of 159 DEGs at the intersection among three subgroups of PCa. (B) KEGG pathway enrichment analysis results for the 159 DEGs. (C) Cytoscape MCODE module used to screen 10 hub-key genes. (D,E) Kaplan-Meier curves of BCRFS for BIRC5 and HMMR in TCGA-PRAD. (F,G) Kaplan-Meier curves of BCRFS for BIRC5 and HMMR in GSE70769. (H,I) Univariate Cox regression analysis for differentially expressed hub-key genes in TCGA-PRAD and GSE70769. PCa, prostate cancer; mPCa, metastatic prostate cancer; CSPC, castration-sensitive prostate cancer; CRPC, castration-resistant prostate cancer; NEPC, neuroendocrine prostate cancer; ECM, extracellular matrix; BCRFS, biochemical recurrence-free survival; HR, hazard ratio; TCGA, The Cancer Genome Atlas; PRAD, prostate adenocarcinoma; DEGs, differentially expressed genes; KEGG, Kyoto Encyclopedia of Genes and Genomes.

Article Snippet: Primary antibodies for BIRC5 (Survivin, dilution 1:100, A00379, Boster, Wuhan, China) and HMMR (dilution 1:100, PA12592, Boster, Wuhan, China) were applied and incubated overnight at 4 °C.

Techniques: Expressing

Journal: Translational Andrology and Urology

Article Title: Identification of BIRC5 and HMMR as prognostic biomarkers for immune infiltration in prostate cancer

doi: 10.21037/tau-24-359

Figure Lengend Snippet: Analysis of the expression levels of BIRC5 and HMMR. (A) BIRC5 expression in tumor and normal tissues from TCGA pan-cancer and GTEx data. (B) HMMR expression in tumor and normal tissues from TCGA pan-cancer and GTEx data. (C,D) BIRC5 and HMMR expression in mPCa, localized PCa, and benign tissues from GSE3325. (E,F) BIRC5 and HMMR expression in NEPC and CSPC/CRPC tissues from GSE59984. (G) BIRC5 expression in mCRPC and localized PCa tissues from GSE35988. (H) HMMR expression in mCRPC and localized PCa tissues from GSE32269. *, P<0.05; **, P<0.01; ***, P<0.001; ns, not significant. TPM, transcripts per million; PCa, prostate cancer; mPCa, metastatic prostate cancer; CSPC, castration-sensitive prostate cancer; CRPC, castration-resistant prostate cancer; NEPC, neuroendocrine prostate cancer; PC, principal component; mCRPC, metastatic castration-resistant prostate cancer; TCGA, The Cancer Genome Atlas; GTEx, Genotype-Tissue Expression.

Article Snippet: Primary antibodies for BIRC5 (Survivin, dilution 1:100, A00379, Boster, Wuhan, China) and HMMR (dilution 1:100, PA12592, Boster, Wuhan, China) were applied and incubated overnight at 4 °C.

Techniques: Expressing

Journal: Translational Andrology and Urology

Article Title: Identification of BIRC5 and HMMR as prognostic biomarkers for immune infiltration in prostate cancer

doi: 10.21037/tau-24-359

Figure Lengend Snippet: Correlation and enrichment analyses of BIRC5 and HMMR. (A,B) Correlation between BIRC5, HMMR, and various immune cells; (C-H) association between the expression levels of BIRC5, HMMR, and the enrichment scores of various immune cells. *, P<0.05; **, P<0.01; ***, P<0.001.

Article Snippet: Primary antibodies for BIRC5 (Survivin, dilution 1:100, A00379, Boster, Wuhan, China) and HMMR (dilution 1:100, PA12592, Boster, Wuhan, China) were applied and incubated overnight at 4 °C.

Techniques: Expressing

Journal: Translational Andrology and Urology

Article Title: Identification of BIRC5 and HMMR as prognostic biomarkers for immune infiltration in prostate cancer

doi: 10.21037/tau-24-359

Figure Lengend Snippet: Correlation analysis between BIRC5 and HMMR expression and T helper cell and CD4 + /CD8 + T cell infiltration. (A-C) Correlation between BIRC5 expression in distinct PCa subgroups and Th cell infiltration levels using the xCELL algorithm. (D-F) Correlation between HMMR expression in distinct PCa subgroups and Th cell infiltration levels using the xCELL algorithm. (G-I) Correlation between BIRC5 expression in distinct PCa subgroups and CD4 + /CD8 + T cell infiltration levels using the quanTIseq algorithm. (J-L) Correlation between HMMR expression in distinct PCa subgroups and CD4 + /CD8 + T cell infiltration levels using the quanTIseq algorithm. PCa, prostate cancer; mCRPC, metastatic castration-resistant PCa; NEPC, neuroendocrine prostate cancer.

Article Snippet: Primary antibodies for BIRC5 (Survivin, dilution 1:100, A00379, Boster, Wuhan, China) and HMMR (dilution 1:100, PA12592, Boster, Wuhan, China) were applied and incubated overnight at 4 °C.

Techniques: Expressing

Journal: Translational Andrology and Urology

Article Title: Identification of BIRC5 and HMMR as prognostic biomarkers for immune infiltration in prostate cancer

doi: 10.21037/tau-24-359

Figure Lengend Snippet: Protein expression of BIRC5 and HMMR in PCa. (A-F) IHC images demonstrating the staining intensity of BIRC5 and HMMR in normal prostate tissues, Gleason ≤7 group, and Gleason >7 group (400×). (G,H) The AOD of IHC images of BIRC5 and HMMR used for comparisons among normal prostate tissues, the Gleason ≤7 group, and the Gleason >7 group. *, P<0.05; **, P<0.01; ***, P<0.001; ****, P<0.0001; AOD, average optical density; IHC, immunohistochemistry.

Article Snippet: Primary antibodies for BIRC5 (Survivin, dilution 1:100, A00379, Boster, Wuhan, China) and HMMR (dilution 1:100, PA12592, Boster, Wuhan, China) were applied and incubated overnight at 4 °C.

Techniques: Expressing, Staining, Immunohistochemistry