formalin-fixed and paraffin-embedded tissue microarrays of human pdac tissue microarrays  (U.S Biomax Inc)

Journal: Cell Death Discovery

Article Title: ALYREF-JunD-SLC7A5 axis promotes pancreatic ductal adenocarcinoma progression through epitranscriptome-metabolism reprogramming and immune evasion

doi: 10.1038/s41420-024-01862-2

Figure Lengend Snippet: A The mRNA expression profiles of m 5 C regulators in PDAC and normal tissues were based on TCGA+GTEx databases. B The mRNA expression of ALYREF in PDAC and adjacent tissues were obtained from GSE15471 and GSE16515 datasets. C ALYREF protein levels were analyzed in PDAC and paired normal tissues by western blotting (n = 9). D Representative IHC images and IHC scores of ALYREF staining in 20 pairs PDAC and normal tissues (scale bar, 250 μm (100×), 50 μm (400 ×)). E , F mIHC was performed on a tissue microarray to detect the relationship between ALYREF and CD8 + T cells ( n = 135, scale bar, 50 μm). G Kaplan–Meier curve of overall survival was performed based on the follow-up data of TMA assay. H Multivariate analysis of several factors was performed in TMA assay. Data are presented the mean ± SD. **** p < 0.0001.

Article Snippet: The human PDAC tissue microarray was created by Wuhan Servicebio technology (Wuhan, China) using 156 PDAC tissue specimens from the First Affiliated Hospital, School of Medicine, Zhejiang University, China.

Techniques: Expressing, Western Blot, Staining, Microarray

Journal: Cell Death Discovery

Article Title: ALYREF-JunD-SLC7A5 axis promotes pancreatic ductal adenocarcinoma progression through epitranscriptome-metabolism reprogramming and immune evasion

doi: 10.1038/s41420-024-01862-2

Figure Lengend Snippet: A Venn Diagram showing the intersection of RIP-BisSeq, RNA-seq and JASPAR database. JunD is identified as a potential target. B , C Expression of JunD following ALYREF knockdown was detected by RT-qPCR and western blotting. D RIP assay was used to validate the direct binding between ALYREF and JunD. E The mRNA decay rate of JunD in shNC and shALYREF PDAC cells treated with Actinomycin D. F The dot blot assay was used to assess the change of mRNA m 5 C level after NSUN2 knockdown in PDAC cells. G MeRIP-qPCR assay was conducted using m 5 C-specific antibody to measure the m 5 C levels. H RIP-qPCR assay was performed using anti-ALYREF antibody to determine the binging efficiency between ALYREF and JunD mRNA. I , J Expression of SLC7A5 following JunD knockdown was detected by RT-qPCR and western blotting. K Schematic illustration showing the position of ChIP- qPCR primers. L , M ChIP assay was performed to test whether JunD could bind to the promoter of SLC7A5. Dual-luciferase assays verified that whether JunD could promote the transcription of SLC7A5 mRNA. Data are presented the mean ± SD of 3 independent experiments, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: The human PDAC tissue microarray was created by Wuhan Servicebio technology (Wuhan, China) using 156 PDAC tissue specimens from the First Affiliated Hospital, School of Medicine, Zhejiang University, China.

Techniques: RNA Sequencing, Expressing, Knockdown, Quantitative RT-PCR, Western Blot, Binding Assay, Dot Blot, ChIP-qPCR, Luciferase

Journal: Cell Death Discovery

Article Title: ALYREF-JunD-SLC7A5 axis promotes pancreatic ductal adenocarcinoma progression through epitranscriptome-metabolism reprogramming and immune evasion

doi: 10.1038/s41420-024-01862-2

Figure Lengend Snippet: A The expression levels of ALYREF, JunD and SLC7A5 in TMA were measured by multiplex immunohistochemical and two groups of representative multiplexes immunohistochemical images are shown (scale bar, 50 μm). B The correlation of among the expression levels of ALYREF, JunD and SLC7A5 in TMA samples was analyzed by Pearson correlation coefficient ( n = 139). C Overall survival analysis based on TMA cohort showed that patients with high co-expression of ALYREF, JunD and SLC7A5 have poor prognosis. D The graphic illustration of ALYREF modulating tumor progression and immune escape in PDAC.

Article Snippet: The human PDAC tissue microarray was created by Wuhan Servicebio technology (Wuhan, China) using 156 PDAC tissue specimens from the First Affiliated Hospital, School of Medicine, Zhejiang University, China.

Techniques: Expressing, Multiplex Assay, Immunohistochemical staining