Fig 1: Anti-ENO1 antibodies are a member of ATAB. (a) The signals of ENO1 in JEG-3 cells were reduced by incubation with a mixture of antibodies (10 µg/ml rabbit anti-ENO1 antibodies and 10 µg/ml human ATAB-positive IgGs) compared with only 10 µg/ml incubating human purified IgGs or 10 µg/ml rabbit anti-ENO1 antibodies. Different secondary antibodies were used in western blots: goat-anti-human antibody, goat-anti-human antibody and goat-anti-rabbit antibody (from left to right rows). (b.c) The diagram shows that in the flow cytometry mixed antibodies (0·42 mg/ml anti-ENO1 antibodies with 2 mg/ml isolated IgGs) bind less JEG-3 cells as ATAB-positive IgGs (P < 0·05, Wilcoxon test, n = 3). (d.e) Pre-incubating 20 µg recombinant human ENO1 protein with 2 mg/ml ATAB-positive IgGs decreases the binding of ATAB-positive IgGs to JEG-3 cells (P < 0·05, Wilcoxon test, n = 3). The MCS values are showed with mean ± SD and P-values are shown individual treatment groups compared to incubation with ATAB-positive IgGs.
Fig 2: ENO1 coexpresses with ß-arrestin in the extravillous trophoblasts (EVT) of uRM patients and healthy controls. Confocal microscopy images showed a co-localization of ENO1 (red) and ß-arrestin (green) in EVT of the first trimester placenta in uRM women (a–d) and healthy controls (e–h), which are shown with white arrows. HLA-G is used as the marker for EVT (magenta), magnification x20, scale bar =20 µM. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)
Fig 3: Detection of nuclear Enolase-1/ MBP-1 in low/high-grade serous carcinoma. Notes: Higher nuclear ENO-1/ MBP-1 staining was detected in (A) low-grade serous carcinoma compared to (B) high-grade serous carcinoma (median IRS 3 (range 0–8) vs. median IRS 2 (range 0–4), p<0.001).
Fig 4: Anti-ENO1 titers are higher in the sera of ATAB-positive patients. (a) Anti-ENO1 antibodies were increased in the 16 uRM patients with positive ATAB compared with 17 cases with negative ATAB (P = 0·007, student's t-test). (b.c) The levels of anti-?-Enolase antibodies and anti-citrullinated fibrinogen (CFG) antibodies were not changed in 36 ATAB-positive patients and 30 ATAB-negative cases (both P > 0·05, student's t-test). (d) Anti-MCV antibodies were higher in the sera of 34 ATAB-positive patients than 28 ATAB-negative cases (P = 0·018, student's t-test). Data were shown as mean ± SEM and statistically significant differences between two uRM groups were marked with P values.
Fig 5: Anti-ENO1 antibodies inhibit the expression of ß-hCG and progesterone in human villous trophoblast (HVT) cells. (a) The production of ß-hCG in HVT cells was inhibited by 1, 10 and 100 ng/ml of anti-ENO1 antibodies in a dose dependent manner. (each P value <0·05, Wilcoxon test, n = 6). (b) The secretion of progesterone in HVT cells was suppressed by 1, 10 and 100 ng/ml of anti-ENO1 antibodies in a dose dependant manner. (each P value <0·05, Wilcoxon test, n = 6). Values are expressed as shown as mean ± SEM and P-values are shown individual treatment groups compared to isotype control.
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