Journal: BMC Cancer

Article Title: A proteomic strategy to identify novel serum biomarkers for liver cirrhosis and hepatocellular cancer in individuals with fatty liver disease

doi: 10.1186/1471-2407-9-271

Figure Lengend Snippet: Serum proteins separated by two dimensional gel electrophoresis in early and advanced NAFLD, with and without HCC . These are representative blots comparing immune depleted and desalted serum from a patient with NAFLD and no fibrosis, a patient with NAFLD cirrhosis, and a patient with NAFLD cirrhosis and cancer. Serum profiles separated by 2D gel electrophoresis from 5 patients in each group, run in duplicate, were compared using Progenesis software. 5 spots were characterised by MS after excision. Spots 1,2 and 3 were isoforms of apolipoprotein A1, spot 4 was apolipoprotein A4, and spot 5 was identified as CD5 antigen like, or CD5L.

Article Snippet: A direct ELISA for novel candidate CD5L was optimised using different concentrations of serum (raw, 1;10, 1:100, 1:500 and 1:1000), primary anti-CD5L antibody (R&D systems) and recombinant CD5L protein (R&D systems).

Techniques: Two-Dimensional Gel Electrophoresis, Electrophoresis, Software

Journal: BMC Cancer

Article Title: A proteomic strategy to identify novel serum biomarkers for liver cirrhosis and hepatocellular cancer in individuals with fatty liver disease

doi: 10.1186/1471-2407-9-271

Figure Lengend Snippet: CD5L discriminates poorly between cirrhotic patients with and without HCC . Serum levels of CD5L were measured by ELISA assay in cirrhotic patients with (218 ± 221; n = 45) and without (194 ± 166; n = 49) HCC, as shown in 2A. Characteristics of the individuals are shown in Table 1. There was no significant difference between the two groups. ROC analyses is shown in 2B. The area under the curve is 0.495 (95% confidence intervals 0.376 and 0.614). A level > 400 ng/ml has a sensitivity of only 20%, but a specificity of 88%. Levels > 500 ng/ml have a specificity of 96%, > 600 ng/ml of 98% and > 700 ng/ml of 100%. For comparison, the mean CD5L serum level from patients without cirrhosis (detailed in figure 3) is also shown.

Article Snippet: A direct ELISA for novel candidate CD5L was optimised using different concentrations of serum (raw, 1;10, 1:100, 1:500 and 1:1000), primary anti-CD5L antibody (R&D systems) and recombinant CD5L protein (R&D systems).

Techniques: Enzyme-linked Immunosorbent Assay, Comparison

Journal: BMC Cancer

Article Title: A proteomic strategy to identify novel serum biomarkers for liver cirrhosis and hepatocellular cancer in individuals with fatty liver disease

doi: 10.1186/1471-2407-9-271

Figure Lengend Snippet: CD5L discriminates between steatohepatitis patients with and without cirrhosis . Serum levels of CD5L were measured in a total of 113 patients with either ALD or NAFLD. Fibrosis was scored histologically on liver biopsy as per the Brunt scoring system. The numbers of individuals in each group are shown within the boxes of chart 3A. The difference in CD5L levels between the different stages of fibrosis is statistically significant by univariate analysis (p = 0.004) controlled for both age and sex. This difference The ROC analyses for the identification of those individuals with stage 4 fibrosis (cirrhosis) is depicted in 3B. The area under the curve is 0.719 (95% confidence intervals 0.623 and 0.816), p < 0.0001. A level of CD5L 50 ng/ml has a sensitivity of 78% and a specificity of 46%, while a level of 100 ng/ml has a sensitivity of 63% and a specificity of 72%. A level greater than 200 ng/ml has a specificity of 95% (sensitivity 41%), and greater than 300 ng/ml of 97% (sensitivity 27%).

Article Snippet: A direct ELISA for novel candidate CD5L was optimised using different concentrations of serum (raw, 1;10, 1:100, 1:500 and 1:1000), primary anti-CD5L antibody (R&D systems) and recombinant CD5L protein (R&D systems).

Techniques:

Journal: BMC Cancer

Article Title: A proteomic strategy to identify novel serum biomarkers for liver cirrhosis and hepatocellular cancer in individuals with fatty liver disease

doi: 10.1186/1471-2407-9-271

Figure Lengend Snippet: CD5L mRNA expression is not altered in association with either fat, inflammation or fibrosis scores in pre-cirrhotic NAFLD liver tissues . mRNA CD5L expression was quantified by real-time PCR, relative to GAPDH and a normal liver sample, in 21 pre-cirrhotic NAFLD biopsy tissues, 13 normal liver samples taken at the time of liver resection, one cirrhotic liver and one HCC. As shown in 4A-C, there was no difference in any pre-cirrhotic NAFLD biopsy tissues in association with the degree of fat, inflammation or fibrosis. There was a significant increase in the NAFLD tissues as a group (n = 21), compared with normal liver tissues (n = 13) as represented in 4D (6.945 ± 0.722 versus1.68 ± 0.269; p = 0.000, ***). The elevated CD5L mRNA expression in one cirrhotic and HCC tissue pair, obtained at the time of laparoscopic radiofrequency ablation, is also presented in 4C and is in keeping with the elevated serum CD5L levels identified in the larger cohort of patients studied.

Article Snippet: A direct ELISA for novel candidate CD5L was optimised using different concentrations of serum (raw, 1;10, 1:100, 1:500 and 1:1000), primary anti-CD5L antibody (R&D systems) and recombinant CD5L protein (R&D systems).

Techniques: Expressing, Real-time Polymerase Chain Reaction

Journal: BMC Cancer

Article Title: A proteomic strategy to identify novel serum biomarkers for liver cirrhosis and hepatocellular cancer in individuals with fatty liver disease

doi: 10.1186/1471-2407-9-271

Figure Lengend Snippet: The identification of the differentiating protein spots on 2D gels.

Article Snippet: A direct ELISA for novel candidate CD5L was optimised using different concentrations of serum (raw, 1;10, 1:100, 1:500 and 1:1000), primary anti-CD5L antibody (R&D systems) and recombinant CD5L protein (R&D systems).

Techniques: Sequencing

Journal: bioRxiv

Article Title: Bimolecule detection for Extracellular Vesicle Screening

doi: 10.1101/2020.07.23.217018

Figure Lengend Snippet: ( A ) The EMARS products for serum EVs from EML4-ALK transgenic mouse. To average experimental results over each group, an aliquot of the serum (10 μL each) from 10 animals in each group (WT), large lung tumor-bearing (TL), and small lung tumor-bearing mice (TS) was mixed in equal proportions, and then applied to EV purification and EMARS. The EMARS products were concentrated and purified by immunoprecipitation with the anti-fluorescein antibody Sepharose. The resulting samples were subjected to SDS-PAGE analysis with fluorescence detection. “IP” indicates the immunoprecipitated samples, and “Lys” indicates the lysate samples before immunoprecipitation. The right panel indicates the same gel as the left panel, but exposed for a longer time. ( B ) Confirmation of candidate partner molecules (CD5L and PZP) with mouse CHL1 in EVs. The EMARS products of WT, TL, and TS were respectively applied to immunoprecipitation (anti-fluorescence antibody Sepharose) and western blot analysis with anti-CD5L (left panel) antibody. After the stripping as described in the “ Materials and methods ”, the membranes were re-stained with anti-PZP antibody (right panel). Arrows indicate the detected band of CD5L and PZP proteins (including predicted dimer). Asterisk indicates unknown bands (predicted as non-specific or partial fragments). ( C, D ) Confirmation of candidate partner molecules (SLC4A1 and THBS1) with mouse CHL1 in EVs. The western blot analysis was performed with anti-SLC4A1 antibody ( C ). After stripping, the membranes were re-stained with anti-THBS1 antibody ( D ). Arrows indicate the detected band of SLC4A1 and THBS1 proteins (including predicted dimers). Asterisks indicate unknown bands (predicted as non-specific or partial fragments). ( E ) Expression of SLC4A1 (left column) and CHL1 proteins (right column) in tumor tissues from two male and two female EML4-ALK transgenic mice. The fragments of lung cancer tissues were mashed and washed gently with PBS, and then lysed with SDS-PAGE sample buffer directly. The resulting samples were subjected to Western blot analysis with anti-SLC4A1 antibody and anti-CHL1 antibody. Arrows indicate the detected band of monomer SLC4A1 and CHL1 proteins.

Article Snippet: After blocking with 5% skim milk solution, transferred membrane was reacted with first antibodies as follows: anti-mouse and human CHL1 antibody (AF2147 and MAB2126; R&D systems, MN; 1 μg/ml), HRP-conjugated anti-mouse and human CHL1 antibody (described above), anti-α2 integrin antibody (ab133557; Abcam; 1 μg/ml), anti-β1 integrin antibody (610467; BD transduction laboratories, NJ; 0.25 μg/ml), anti-FGFR3 antibody (c-15; Santa cruz, TX; 0.2 μg/ml 5% BSA-PBS), anti-TSG101 antibody (EXOAB-TSG101-1; System Biosciences, CA; 1:500), anti-CD63 antibody (EXOAB-CD63A-1; System Biosciences, CA; 1:500), anti-CD5L antibody (AF2834; R&D systems, MN; 0.4 μg/ml), anti-Pregnancy Zone Protein (PZP) antibody (PAG324Ra01; CLOUD-CLONE, TX; 0.5 μg/ml), anti-SLC4A1 antibody (18566-1-AP; PROTEINTECH, IL; 0.6 μg/ml), anti-Thrombospondin 1 (THBS1) antibody (PAA611Hu01; CLOUD-CLONE, TX; 0.5 μg/ml), and anti-caspase 14 antibody (MAB8215; R&D systems, MN; 0.5 μg/ml).

Techniques: Transgenic Assay, Purification, Immunoprecipitation, SDS Page, Fluorescence, Western Blot, Stripping Membranes, Staining, Expressing

Journal: Biochemical and biophysical research communications

Article Title: CD5L is a target of transcription factor Nrf2.

doi: 10.1016/j.bbrc.2025.152225

Figure Lengend Snippet: Fig. 4. Recombinant CD5L (rCD5L) suppresses TGFβ-mediated fibrogenic responses in human lung fibroblast cells. Protein levels of COL1A1, FN1, and α-SMA in IMR-90 cells pre-treated with vehicle or rCD5L (0.4 μg/mL) for 3 h, followed by treatment with vehicle or TGFβ (2 ng/mL) for a further 24 h. Representative blots and quantification relative to α-tubulin are shown. Unpaired two-sample Student’s t-test (2-tailed) was used to analyze the difference between rCD5L-treated and vehicle-treated cells, either without or with TGFβ treatment.

Article Snippet: Human rCD5L (2797-CL, R&D Systems) was reconstituted in sterile PBS.

Techniques: Recombinant

Journal: Journal of Translational Medicine

Article Title: Hypoxic migrasomes drive colorectal cancer liver metastasis by mediating CD5L + macrophage efferocytosis via NRP2/PROX1 axis

doi: 10.1186/s12967-025-07485-0

Figure Lengend Snippet: Characterization of myeloid cell subsets and identification of CD5L⁺ macrophages as potential migrasome-responsive cells. ( A ) UMAP visualization showing the reclustering of myeloid cells into 10 distinct subpopulations. ( B ) Heatmap displaying the expression profiles of representative marker genes for each myeloid subset. ( C ) Bar plot showing the relative abundance of each myeloid subpopulation in normoxic versus hypoxic migrasome-treated groups. A notable 8-fold increase in the proportion of CD5L⁺ macrophages was observed in the hypoxic group. ( D ) Gene Set Variation Analysis (GSVA) indicating significant activation of the complement signaling pathway in CD5L⁺ macrophages. ( E ) Chord diagram depicting inferred cell–cell communication networks, highlighting potential interactions between CD5L⁺ macrophages and fibroblasts, T cells, and other immune cells via GDF signaling. ( F ) Activation IGF signaling pathway network mediated crosstalk between CD5L⁺ macrophages and epithelial tumor cells. ( G ) In the IGF signaling pathway network, CD5L⁺ macrophages exhibit specific overexpression of IGF1. ( H ) Among the six macrophage cell-types, CD5L⁺ macrophages exhibit specific overexpression of CD5L

Article Snippet: The pellet was resuspended in 200 μL antibody dilution buffer (2% FBS in 1× PBS) and incubated with 0.5 μg CD5L antibody (BAF2834, R&D Systems) at 4 °C for 60 min in the dark.

Techniques: Expressing, Marker, Activation Assay, Over Expression

Journal: Journal of Translational Medicine

Article Title: Hypoxic migrasomes drive colorectal cancer liver metastasis by mediating CD5L + macrophage efferocytosis via NRP2/PROX1 axis

doi: 10.1186/s12967-025-07485-0

Figure Lengend Snippet: Phagocytic activity and immunofluorescence validation of CD5L⁺ macrophages in CRC liver metastases following migrasome treatment. ( A ) UMAP blot showing the expression of migrasome marker TSPAN4 in myeloid subsets. ( B ) Boxplot showing efferocytosis scores across the 10 identified myeloid cell subtypes. ( C ) Violin plots depicting the expression of efferocytosis markers CD300B, MERTK, and CD300D across 10 distinct myeloid cell subtypes. ( D ) UMAP plots displaying the expression patterns of three efferocytosis-associated marker genes specifically enriched in CD5L⁺ macrophages. ( E ) Immunofluorescence staining of tumor tissues from MC38-tumor bearing mice showing colocalization of CD163, CD5L, and the migrasome marker MERTK in both treatment groups. Increased MERTK expression is observed in the hypoxic group, indicating enhanced migrasome targeting of CD5L⁺ macrophages

Article Snippet: The pellet was resuspended in 200 μL antibody dilution buffer (2% FBS in 1× PBS) and incubated with 0.5 μg CD5L antibody (BAF2834, R&D Systems) at 4 °C for 60 min in the dark.

Techniques: Activity Assay, Immunofluorescence, Biomarker Discovery, Expressing, Marker, Staining

Journal: Biomedicines

Article Title: Concomitant High Apoptosis Inhibitor of Macrophage (AIM) and Low Prostate-Specific Antigen (PSA) Indicates Activated T Cell-Mediated Anticancer Immunity, Enhance Sensitivity to Pembrolizumab, and Elicit Good Prognosis in Prostate Cancer

doi: 10.3390/biomedicines9091225

Figure Lengend Snippet: AIM and PSA expression levels are inversely correlated in patients with prostate cancer. Box and whisker plots of the differential expression of ( A ) AIM/CD5L and ( B ) PSA/KLK3 transcript expression in the prostate gland or prostate adenocarcinoma from the GSE6956 and GSE68907 cohort, respectively. ( C ) Comparative heatmap of the expression of KLK3 and CD5L in TCGA PRAD normal and tumor samples. ( D ) Box and whisker plots comparing the mRNA expression levels of AIM/CD5L and PSA/KLK3 in the TCGA PRAD cohort. ( E ) Representative IHC images showing the differential expression of AIM and AIM in normal prostate, low-grade, medium grade, and high-grade PCa samples from the TMU-SHH cohort. ( F ) Line and dots plot of the correlation between AIM and PSA transcript expression in prostate cancer cell lines from the DepMap Public 21Q2 dataset ( left ). Line and dot plot showing the effect of CRISPR-mediated aim gene knockout, on PSA transcript expression in prostate cancer cell lines from the DepMap Public 21Q2 dataset ( right ). TCGA, the cancer genome atlas; PRAD, prostate adenocarcinoma; IHC, immunohistochemistry; TPM, transcripts per million.

Article Snippet: Ectopic expression of AIM/CD5L was achieved using the AIM (CD5L) (NM_005894) Human (Myc-DDK)-Tagged ORF Clone (#RC206528L2, OriGene Technologies, Inc., Rockville, MD, USA) in pCMV6-Entry vector transfected into PC-3 cells using Lipofectamine 2000 reagent (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions.

Techniques: Expressing, Whisker Assay, Quantitative Proteomics, CRISPR, Gene Knockout, Immunohistochemistry

Journal: Biomedicines

Article Title: Concomitant High Apoptosis Inhibitor of Macrophage (AIM) and Low Prostate-Specific Antigen (PSA) Indicates Activated T Cell-Mediated Anticancer Immunity, Enhance Sensitivity to Pembrolizumab, and Elicit Good Prognosis in Prostate Cancer

doi: 10.3390/biomedicines9091225

Figure Lengend Snippet: Aberrantly expressed PSA and significantly suppressed AIM expression characterize advanced stage prostate cancer, regardless of mutation status. Box and whisker and dot plots of the expression profiles of ( A ) PSA/KLK3 and ( B ) AIM/CD5L in normal tissue, early stage, or advanced stage disease across 32 cancer types from the ICR-curated dataset. Graphical representation of cases affected by ( C ) psa/klk3 and ( D ) aim/cd5l mutation ( upper ) or CNV ( lower ) in ICR-curated pan-cancer TCGA cohort data. ICR, Institute of Cancer Research; CNV, copy number variation; TCGA, the cancer genome atlas; PRAD, prostate adenocarcinoma; red box, PCa/PRAD cohort.

Article Snippet: Ectopic expression of AIM/CD5L was achieved using the AIM (CD5L) (NM_005894) Human (Myc-DDK)-Tagged ORF Clone (#RC206528L2, OriGene Technologies, Inc., Rockville, MD, USA) in pCMV6-Entry vector transfected into PC-3 cells using Lipofectamine 2000 reagent (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions.

Techniques: Expressing, Mutagenesis, Whisker Assay

Journal: Biomedicines

Article Title: Concomitant High Apoptosis Inhibitor of Macrophage (AIM) and Low Prostate-Specific Antigen (PSA) Indicates Activated T Cell-Mediated Anticancer Immunity, Enhance Sensitivity to Pembrolizumab, and Elicit Good Prognosis in Prostate Cancer

doi: 10.3390/biomedicines9091225

Figure Lengend Snippet: The differential expression of PSA and AIM is associated with disease recurrence in patients with prostate cancer but is equivocal for overall survival. Kaplan–Meier curves showing the effect of high or low ( A ) PSA and ( B ) AIM expression on BCR-free survival in the TCGA PRAD cohort. Kaplan–Meier curves showing the effect of high or low ( C ) PSA and ( D ) AIM expression on overall survival in the TCGA PRAD cohort. BCR risk group-stratified heatmap ( left ) and box-and-whisker plots ( right ) of ( E ) PSA/KLK3 or ( F ) AIM/CD5L expression levels in the GSE40272 cohort. ( G ) XY plot showing the differential expression of AIM and PSA in the GSE within the Broad_2020_09_c7_immunologic gene set. BCR, biochemical recurrence; TCGA, the cancer genome atlas; PRAD, prostate adenocarcinoma.

Article Snippet: Ectopic expression of AIM/CD5L was achieved using the AIM (CD5L) (NM_005894) Human (Myc-DDK)-Tagged ORF Clone (#RC206528L2, OriGene Technologies, Inc., Rockville, MD, USA) in pCMV6-Entry vector transfected into PC-3 cells using Lipofectamine 2000 reagent (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions.

Techniques: Quantitative Proteomics, Expressing, Whisker Assay

Journal: Biomedicines

Article Title: Concomitant High Apoptosis Inhibitor of Macrophage (AIM) and Low Prostate-Specific Antigen (PSA) Indicates Activated T Cell-Mediated Anticancer Immunity, Enhance Sensitivity to Pembrolizumab, and Elicit Good Prognosis in Prostate Cancer

doi: 10.3390/biomedicines9091225

Figure Lengend Snippet: PSA-associated suppression of AIM is implicated in the enhanced metastability of prostate cancer and a high AIM/PSA ratio is associated with strong castration-induced regression. Histograms showing the differential expression of ( A ) cd5l/aim and ( B ) klk3/psa in poorly metastatic or highly metastatic sample from the GDS2865 dataset. ( C ) Histogram comparing the cd5l and klk3 expression values in the poorly or highly metastatic samples from the GDS2865 dataset. Histograms showing the differential expression of ( D ) cd5l/aim and ( E ) klk3/psa in androgen-dependent growth, castration-induced regression nadir, or castration-resistant regrowth samples from the GDS4107 dataset. ( F ) Histograms of the cd5l/klk3 ratio in androgen-dependent growth, castration-induced regression nadir, or castration-resistant regrowth samples from the GDS4107 dataset. ( G ) Sample percentage bin plot of cd5l and klk3 expression in the GSE46691 cohort stratified by propensity for metastasis. ( H ) Box and whisker plots of the differential expression levels of CD5L ( upper left ) and KLK3 ( upper right ) in patients with or without AR-amplification in the MET500 PRAD cohort. Box and whisker plots of the differential expression levels of CD5L ( lower left ) and KLK3 ( lower right ) in patients with or without ERG fusion in the MET500 PRAD cohort. ( I ) Representative photomicrographs ( upper ) and quantitative chart ( lower ) showing the effect of CD5L_OE on invasion of PC3 or DU145 cells. ( J ) Representative photomicrographs ( upper ) and histograms ( lower ) showing the effect of CD5L_OE on migration of PC3 cells at indicated time-points. CD5L_OE, overexpression of cd5l ; AR, androgen receptor; F.C., fold change; ** p < 0.01; *** p < 0.001.

Article Snippet: Ectopic expression of AIM/CD5L was achieved using the AIM (CD5L) (NM_005894) Human (Myc-DDK)-Tagged ORF Clone (#RC206528L2, OriGene Technologies, Inc., Rockville, MD, USA) in pCMV6-Entry vector transfected into PC-3 cells using Lipofectamine 2000 reagent (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions.

Techniques: Quantitative Proteomics, Expressing, Whisker Assay, Amplification, Migration, Over Expression

Journal: Biomedicines

Article Title: Concomitant High Apoptosis Inhibitor of Macrophage (AIM) and Low Prostate-Specific Antigen (PSA) Indicates Activated T Cell-Mediated Anticancer Immunity, Enhance Sensitivity to Pembrolizumab, and Elicit Good Prognosis in Prostate Cancer

doi: 10.3390/biomedicines9091225

Figure Lengend Snippet: The functional association between PSA and AIM modulates metastasis and mirrors the immunogenicity in patients with prostate cancer. Box and whisker plots showing the differential expression of ( A ) cd5l/aim and ( B ) psa/klk3 genes normal, primary tumor, or metastatic samples from the TNMplot gene chip data. ( C ) Expression heatmap showing the correlation between CD5L , KLK3 , with biomarkers of cancer stemness, epithelial-to-mesenchymal transition, metastasis, or T-cell activation in normal, localized PCa, and bone metastatic CRPC in the GSE32269. ( D ) Graphical representation of the differential expression of CD5L , KLK3 , biomarkers of M1 phenotype, M2 phenotype, and T-cell activation in non-tumor or tumor samples from the TCGA PRAD cohort.

Article Snippet: Ectopic expression of AIM/CD5L was achieved using the AIM (CD5L) (NM_005894) Human (Myc-DDK)-Tagged ORF Clone (#RC206528L2, OriGene Technologies, Inc., Rockville, MD, USA) in pCMV6-Entry vector transfected into PC-3 cells using Lipofectamine 2000 reagent (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions.

Techniques: Functional Assay, Immunopeptidomics, Whisker Assay, Quantitative Proteomics, Expressing, Activation Assay

Journal: Biomedicines

Article Title: Concomitant High Apoptosis Inhibitor of Macrophage (AIM) and Low Prostate-Specific Antigen (PSA) Indicates Activated T Cell-Mediated Anticancer Immunity, Enhance Sensitivity to Pembrolizumab, and Elicit Good Prognosis in Prostate Cancer

doi: 10.3390/biomedicines9091225

Figure Lengend Snippet: Heatmap showing the correlation between CD5L , KLK3 , with ( A ) biomarkers of positive regulation of T-cell responses, pro-inflammatory markers, Treg markers, ( B ) T-cell activation markers, and ( C ) macrophage markers, in normal, localized PCa, and bone metastatic CRPC in the GSE32269. Columns with similar annotations are collapsed by taking mean inside each group. Rows are centered; unit variance scaling is applied to rows. Both rows and columns are clustered using correlation distance and average linkage. Varied rows, three columns. ( D ) Graphical visualization of the five clusters formed by cd5l and klk3 gene-enrichment-based molecular or biological process functional annotation clustering using a medium classification stringency. ( E ) Gene ontology visualization of reported gene-term association. Treg, regulatory T cell; CRPC, castration-resistant prostate cancer.

Article Snippet: Ectopic expression of AIM/CD5L was achieved using the AIM (CD5L) (NM_005894) Human (Myc-DDK)-Tagged ORF Clone (#RC206528L2, OriGene Technologies, Inc., Rockville, MD, USA) in pCMV6-Entry vector transfected into PC-3 cells using Lipofectamine 2000 reagent (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions.

Techniques: Activation Assay, Functional Assay

Journal: Biomedicines

Article Title: Concomitant High Apoptosis Inhibitor of Macrophage (AIM) and Low Prostate-Specific Antigen (PSA) Indicates Activated T Cell-Mediated Anticancer Immunity, Enhance Sensitivity to Pembrolizumab, and Elicit Good Prognosis in Prostate Cancer

doi: 10.3390/biomedicines9091225

Figure Lengend Snippet: AIM binds directly to PSA, and the ectopic (re)expression of AIM enhances the anticancer therapeutic effect of Pembrolizumab against PCa cells. 2D visualization of the auto-generated ( A ) CD5L-KLK3 interactome showing complicity in the immune landscape, macrophages, EMT and CSC regulation, ( B ) CD5L interactome, or KLK3 interactome. ( C ) Three-dimensional visualization of the direct complex formation by AIM/CD5L (surface model) and PSA/KLK3 (ribbon model). ( D ) Line graph showing the effect of CD5L_OE with or without 5 μM Pembrolizumab on PC3 cell proliferation. ( E ) Representative photomicrographs ( upper ) and histograms ( lower ) showing the effect of CD5L_OE with or without 5 μM Pembrolizumab on PC3 cell invasion. ( F ) Representative photomicrographs ( upper ) and quantitative chart ( lower ) showing the effect of CD5L_OE with or without 5 μM Pembrolizumab on the tumorsphere formation capability of PC3 cells. ( G ) Representative IHC photomicrographs ( upper ) and histograms ( lower ) of the immunoreactivity of AIM, PSA, and PDL1 in pembro_responders or pembro_non-responders from the TMU-SHH cohort. ( H ) Histograms showing the differential CD8 + T-cell count/field in pembro_responders or pembro_non-responders from the TMU-SHH cohort. WT, wild type; OE, overexpression; * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: Ectopic expression of AIM/CD5L was achieved using the AIM (CD5L) (NM_005894) Human (Myc-DDK)-Tagged ORF Clone (#RC206528L2, OriGene Technologies, Inc., Rockville, MD, USA) in pCMV6-Entry vector transfected into PC-3 cells using Lipofectamine 2000 reagent (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instructions.

Techniques: Expressing, Generated, Cell Counting, Over Expression