Journal: Cell Reports Medicine
Article Title: Profiling the metabolome of uterine fluid for early detection of ovarian cancer
doi: 10.1016/j.xcrm.2023.101061
Figure Lengend Snippet: Increased 4-OHE 2 in OC cells disturbs the tyrosine catabolism pathway (A) An illustration of the tyrosine catabolism pathway in which tyrosine is catabolized to diverse intermediates including L-DOPA, dopamine, NE, epinephrine, normetanephrine, 3,4-dihydroxymandelate, MHPG, etc., and to the end product of VMA. These processes are catalyzed by several enzymes, among which COMT participates in multiple steps. MHPG, 3-Methoxy-4-hydroxyphenyl glycolaldehyde. (B) Concentrations of NE and VMA in 12 pairs of para-tumor and tumor tissues were measured with an NE ELISA kit and a VMA ELISA kit, respectively. Data are presented as mean values ± SD. p values were calculated by unpaired two-tailed Student’s t tests. p <0.05 was considered statistically significant. (C) Expression levels of CYP1B1 between normal ovarian tissues (88 available) and serous OC tissues (426 available) were compared through an integrated analysis of TCGA and GTEx databases. The values of log 2 (normalized count) are used to measure the gene expression levels. p values were calculated by unpaired two-tailed Welch’s t test. p <0.05 was considered statistically significant. (D) The protein level of CYP1B1 in 12 pairs of para-tumor and tumor tissues examined by western blot. Beta-actin was used as a loading control. (E) 4-OHE 2 in para-tumor tissues and tumor tissues was extracted, and the corresponding levels were measured by targeted LC-MS analysis. Data are presented as mean values ± SD. p values were calculated by unpaired two-tailed Student’s t tests. p <0.05 was considered statistically significant. (F) Schematic diagram of dual functions of COMT in tyrosine catabolism and 4-OHE 2 degradation. (G) In vitro binding affinity of 4-OHE 2 or NE to COMT was measured by microscale thermophoresis (MST). F norm = F 1 / F 0 ( F norm, normalized fluorescence; F 1 , fluorescence after thermodiffusion; F 0 , initial fluorescence or fluorescence after T-jump). K D , dissociation constant. (H) Concentrations of NE and VMA in primary ovarian cancer cells and T1074, SKOV3, and A2780 cells treated with or without 5 μM 4-OHE 2 for 12 h. Three biologically independent replicates were performed. Data are presented as mean values ± SD. p values were calculated by unpaired two-tailed Student’s t tests. p <0.05 was considered statistically significant.
Article Snippet: The levels of NE and VMA were determined by Human ELISA kits following manufacturer’s instructions (Elabscience, E-EL-0047c; Elabscience, E-EL-0132c).
Techniques: Enzyme-linked Immunosorbent Assay, Two Tailed Test, Expressing, Gene Expression, Western Blot, Control, Liquid Chromatography with Mass Spectroscopy, In Vitro, Binding Assay, Microscale Thermophoresis, Fluorescence
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