Journal: Molecular Medicine Reports

Article Title: Estradiol inhibits autophagy of Mycobacterium tuberculosis -infected 16HBE cells and controls the proliferation of intracellular Mycobacterium tuberculosis

doi: 10.3892/mmr.2022.12712

Figure Lengend Snippet: Mtb-infected epithelial cells and the expression of ERα and ERβ in granulomatous tissues of patients with TBTB. (A) Mtb-positive epithelial cells were analyzed using double-labeled immunofluorescence. Mtb was observable in goblet cells and ciliated cells. (B) Only ERα was strongly expressed in tissue samples from patients with TBTB. MUC5AC is commonly used to identify goblet cells. FOXJ1 is a marker of ciliated cells. Magnification, ×100 and ×400 for the low and high-magnification panels, respectively. Mtb, Mycobacterium tuberculosis ; TBTB, tracheobronchial tuberculosis; ER, estrogen receptor; MUC5AC, mucin 5AC; FOXJ1, forkhead box J1.

Article Snippet: Mouse anti-human ERα (1:500; cat. no. NBP2-61764; Novus Biologicals, Ltd.) and mouse anti-human ERβ (1:500; cat. no. NBP2-44366; Novus Biologicals, Ltd.) monoclonal antibodies were added and incubated overnight at 4°C.

Techniques: Infection, Expressing, Labeling, Immunofluorescence, Marker

Journal: Molecular Medicine Reports

Article Title: Estradiol inhibits autophagy of Mycobacterium tuberculosis -infected 16HBE cells and controls the proliferation of intracellular Mycobacterium tuberculosis

doi: 10.3892/mmr.2022.12712

Figure Lengend Snippet: ERα and ERβ are expressed in 16HBE cells. 16HBE cells were treated with E1 or E2 for 24 h and then ER expression was examined by immunofluorescence. 16HBE cells strongly expressed ERα and ERβ. (A) Compared with the PBS group, the fluorescence intensity of ERα in the E1 group was markedly decreased, while it did not significantly change in the E2 group. (B) Compared with PBS group, the fluorescence intensity of ERβ exhibited no significant change in the E1 group, while the fluorescence intensity of ERβ was markedly decreased in the E2 group. Magnification, ×100. E1, 10 −8 M estradiol; E2, 10 −6 M estradiol.

Article Snippet: Mouse anti-human ERα (1:500; cat. no. NBP2-61764; Novus Biologicals, Ltd.) and mouse anti-human ERβ (1:500; cat. no. NBP2-44366; Novus Biologicals, Ltd.) monoclonal antibodies were added and incubated overnight at 4°C.

Techniques: Expressing, Immunofluorescence, Fluorescence

Journal: Molecular Medicine Reports

Article Title: Estradiol inhibits autophagy of Mycobacterium tuberculosis -infected 16HBE cells and controls the proliferation of intracellular Mycobacterium tuberculosis

doi: 10.3892/mmr.2022.12712

Figure Lengend Snippet: Inhibition of autophagy by estradiol promotes ROS production and phosphorylation of AKT/mTOR by binding to ERα. (A) E2 inhibited autophagy by binding to ERα. Infected 16HBE cells were incubated with PPT or DPN for 24 h in the absence of estradiol. Moreover, infected 16HBE cells were pre-treated with AZD9496 or BAZ for 1 h and co-incubated with estradiol for 24 h. (B) ROS levels in infected 16HBE cells treated with ER agonist or antagonist for 24 h were detected using the DCFH-diacetate probe (10 µM). Mean fluorescence intensity of ROS was analyzed by flow cytometry. (C) Protein levels of p-AKT, total AKT, p-AMPK, total AMPK, p-mTOR and total mTOR in Mtb-infected 16HBE cells were analyzed using western blotting. *P<0.05 and **P<0.01 vs. PBS. E2, 10 −6 M estradiol; PPT, 4, 4, 4, 4(4-propyl-[1H]-pyrazole-1, 3, 5-triyl) trisphenol or ERα-specific agonist (10 −6 M); DPN, diarylpropionitrile or ERβ-specific agonist (10 −6 M); AZD9496, ERα-specific antagonist (10 −6 M); BZA, Bazedoxifene or nonspecific ER modulator (0.5 µM); Mtb, Mycobacterium tuberculosis ; p, phosphorylated; DCFH, 2,7-dichlorodihydrofluorescein.

Article Snippet: Mouse anti-human ERα (1:500; cat. no. NBP2-61764; Novus Biologicals, Ltd.) and mouse anti-human ERβ (1:500; cat. no. NBP2-44366; Novus Biologicals, Ltd.) monoclonal antibodies were added and incubated overnight at 4°C.

Techniques: Inhibition, Binding Assay, Infection, Incubation, Fluorescence, Flow Cytometry, Western Blot