Fig 1: (A) Concentration of annexin A1 (ANXA1) released in the culture media surrounding U87-MG multicellular spheroid (grey) or over monolayer U87-MG cells (black) at different cell numbers. (B,C) Expression of ANXA1 by IHC in sections from a U87-MG xenotransplanted tumour tissue (with and without antibody). Basophilic structure of cell was stained in blue with haematoxylin solution.
Fig 2: ANXA1 is associated with poor survival of patients with breast cancer. (A) Kaplan-Meier survival curves show disease-free survival in patients with breast cancer with low and high ANXA1 (high ANXA1, n=22; low ANXA1, n=2663, p=0.023, log-rank test). (B) Plasma ANXA1 level was higher in malignant patients (n=66) than in benign patients (n=8) (plasma, tested by ELISA). (C) Patients with TNBC (n=12) have higher ANXA1 levels than luminal A (n=17), luminal B (n=28) and Her-2 like (n=9) patients (plasma, tested by ELISA). (D) The tumor specimens were prepared into cell suspension and tested by flow cytometry; figures show how CD4+ T cells and Treg cells (CD4+Foxp3+) are gated. (E) The number of Treg cells in different subtypes of tumor specimens. All data represent mean±SD. *p<0.05, **p<0.01, ***p<0.001 as determined by Student’s t-test. (F) Percentage of Treg cells in different subtypes of breast cancer. (?) Luminal A; (?) luminal B; (?) Her-2 like; (?) TNBC. TNBC, triple-negative breast cancer.
Fig 3: ANXA1 blocker inhibits breast tumor growth in vivo. (A) Images of resected tumors at the end point (n=8). (B) The excised tumors were weighed at the end point. The tumor weights of each group were compared using two-tailed unpaired t-tests. Data are presented as mean±SD (n=8). (C) The tumor growth curve; the arrow indicates the time starting drug administration. (D) Liver and lung lesions of both groups (×400); green arrows indicate metastasis lesions. (E) Number of metastasis lesions of both groups. *p<0.05, **p<0.01, ***p<0.001 as determined by Student’s t-test.
Fig 4: (A) Mean optical density of ANXA1 in glomeruli of AAV patients, LN, MCD and healthy controls respectively. (B) Mean optical density of ANXA1 in tubulointerstitium of AAV patients, LN, MCD and healthy controls respectively. (C) Comparison of ANXA1 expression of patients achieving complete renal recovery with those achieving partial renal recovery. (D–F) Correlation analyses of ANXA1 expression in glomeruli of AAV patients with serum creatinine (D), eGFR (E) and the proportion of crescents (F), respectively. AAV, anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis; ANXA1, annexin A1; eGFR, estimated glomerular filtration rate; HC, healthy control; LN, lupus nephritis; MCD, minimal change disease; ns, no significance (P > 0.05). **P < 0.01, ***P < 0.001.
Fig 5: ANXA1 enhances the suppression function of Treg cells. FPR2 expression in cells (A) compared with Teff cells, Treg cells have higher Fpr2 mRNA levels (tested by qPCR). (B) Compared with Teff cells, Treg cells have higher FPR2 levels (tested by flow cytometry). (C) Expression of ANXA1, FPR2 and FOXP3 in T effector cells and Treg cells. ?) ANXA1 and FOXP3 in T effector cells; ?) FPR2 and FOXP3 in T effector cells; ?) ANXA1 and FOXP3 in Treg cells; (iv) FPR2 and FOXP3 in T effector cells. (D) In vitro suppressive assays were performed. When Teff:Treg=2:1, 4:1, 8:1 and 16:1, the proliferation rates of Teff cells were 11.9%, 22.9%, 65.7% and 70.3%, respectively, compared with 17%, 33.1%, 76.3% and 79.4% in the control group. (E) Negative control: the proliferation rate of Teff cells without stimulation. (F) Comparison of proliferation rates between activated Teff cells (Teff:Treg=1:0) with and without Ac2-26. All data represent mean±SD. (G) Percentage of proliferated Teff cells was assessed. All data represent mean±SD. *p<0.05, **p<0.01, ***p<0.001 as determined by Student’s t-test.
Supplier Page from Abcam for Human Annexin A1 ELISA Kit