Fig 1: Diminished RA signaling in PEX tissues. (A) Expression of CRBP1, CRABP2, RARA and RXRA mRNA in iris (n = 42) and ciliary body specimens (n = 22) from normal human donors (control) and donors with PEX syndrome (n = 24) using real-time PCR technology. Expression levels were significantly reduced in PEX specimens compared to control specimens. The relative expression levels were normalized relative to GAPDH and are represented as mean values ± SD. (B) Western blot analysis of CRBP1 protein expression in iris and ciliary body tissue from normal donors (control, n = 4) and patients with PEX syndrome (n = 4). Protein expression is normalized to the house-keeping gene ß-actin and is expressed as percent of the expression in controls. (C) Western blot analysis of RBP4 protein in serum and aqueous humor samples from cataract patients (control, n = 4) and patients with PEX syndrome (n = 4). RBP4 protein is normalized to total protein content and is expressed as percent of controls (data represent mean values ± SD of eight samples; *P < 0.05; **P < 0.01 ***P < 0.001). (D) Real-time PCR analysis of RXRA, LOXL1, ELN, FBN1 and TGFB1 mRNA in hTCFs (n = 4) and TM cells (hTMC, n = 4) transfected with RXRA-specific siRNA or scrambled control siRNA; expression levels were normalized relative to GAPDH and expressed relative to mock-transfected controls (dashed line; *P < 0.05; **P < 0.01; ***P < 0.001). (E) Real-time PCR analysis of RXRA, LOXL1, ELN, FBN1 and TGFB1 mRNA in hTCF and hTMC without and with stimulation by 2.5 μm RA for 48 h; expression levels were normalized relative to GAPDH and expressed relative to unstimulated controls (dashed line; *P < 0.05; **P < 0.001; ***P < 0.0001).
Fig 2: Diagnostic performance of the biomarker panel in validation cohort I and results compared to those for hemoglobin. Validation cohort I (n = 343) was randomly divided into a training set (n = 241) and a testing set (n = 102). The biomarker panel included CAT, LTF, MMP9, RBP4, and SERPINA3. (A) Receiver operating characteristic (ROC) curve analysis of the biomarker panel and hemoglobin in the training set. The area under the curve (AUC) was compared using the DeLong test. (B) Calibration curve analysis of the biomarker panel and hemoglobin in the training set. The closer the curve is to the reference line (black), the more accurate the prediction. (C) Decision curve analysis (DCA) for the biomarker panel and hemoglobin in the training set. The larger the area under the curve is, the greater the clinical benefit that can be expected. (D) ROC curves, (E) calibration curve, (F) DCA curve for the biomarker panel and hemoglobin in the testing set.
Fig 3: Diagnostic performance of the biomarker panel in Validation Cohort II and results compared to hemoglobin. Validation cohort II (n = 310) included 141 CRC patients, 82 CRA patients, and 87 HCs. The biomarker panel included CAT, LTF, MMP9, RBP4, and SERPINA3. (A) Receiver operating characteristic (ROC) curve analysis of biomarkers between the CRC and HC groups. The area under the curve (AUC) was compared using the DeLong test. (B) Calibration curve analysis of the biomarker panel between the CRC and HC groups. The closer the curve is to the reference line (black), the more accurate the prediction. (C) Decision curve analysis (DCA) for the biomarker panel between the CRC and HC groups. The larger the area under the curve is, the greater the clinical benefit that can be expected. (D) ROC curves, (E) calibration curve, (F) DCA curve for the biomarker panel and hemoglobin between the CRC+CRA and HC groups. (G) ROC curves, (H) calibration curve, (I) DCA curve for the biomarker panel and hemoglobin between the CRA and HC groups.
Fig 4: Protein abundance assays and differential analysis of the biomarker panel and hemoglobin were performed in Validation cohort II. The biomarker panel included CAT, LTF, MMP9, RBP4, and SERPINA3. Validation cohort II (n = 310) included 141 CRC patients (red), 82 CRA patients (green), and 87 HCs (blue), with each dot representing one sample. The unpaired Wilcoxon rank test was used to analyze differences in protein abundance. * P< 0.05, ** P< 0.01, *** P< 0.001, **** P< 0.0001, ns, not significant.
Fig 5: Validation of biomarker panel expression in tumor tissue. Tumor tissue samples from CRC patients in discovery cohort (n=14). The biomarker panel included CAT, LTF, MMP9, RBP4, and SERPINA3. (A) Immunohistochemistry (IHC)-based detection of biomarker panel expression in CRC tumor tissue and adjacent normal tissue. Black arrows indicate where the biomarker is significantly overexpressed. Scale bar represents 100 μm. (B) Differential analysis of IHC integrated optical density (IOD) values. Analyses were performed using paired t-test. * P< 0.05, ** P< 0.01.
Supplier Page from Abcam for Anti-RBP4 antibody [EPR5879]