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. 2022 May;13(5):11611-11623.
doi: 10.1080/21655979.2022.2070584.

MicroRNA-146 attenuates lipopolysaccharide induced ovarian dysfunction by inhibiting the TLR4/NF- κB signaling pathway

Fengping He  1 , Yanhui Liu  1 , Tang Li  1 , Qiulin Ma  1 , Zhang Yongmei  1 , Peiqing He  1 , Chuanyin Xiong  2
Affiliations

MicroRNA-146 attenuates lipopolysaccharide induced ovarian dysfunction by inhibiting the TLR4/NF- κB signaling pathway

Fengping He et al. Bioengineered. 2022 May.

Abstract

Premature ovarian insufficiency (POI) is a disease that seriously affects women's reproductive function and even leads to lifelong infertility. Little is known about the mechanism of lipopolysaccharide (LPS)-induced ovarian dysfunction. Thus, we aimed to identify the role of the up-regulation of microRNA (miRNA)-146 expression offered protection against ovarian dysfunction by inhibiting the toll-like receptor (TLR) 4, TLR4/phosphorylated (p)-nuclear factor (NF)-κB signaling pathway and inflammatory cytokine tumor necrosis factor (TNF)-a and Interleukin (IL)-6. In an in vivo study, we established an LPS-induced ovarian dysfunction mouse model. The mouse ovarian granulosa cells were transfected with miR-146 mimic or negative controls or inhibitor and then treated with LPS. Therefore, cell viability, cells apoptosis, IL-6 and TNF-a, TLR4, NF- κB were assessed, respectively. These results demonstrated that the up-regulation of miRNA-146 expression may protect against LPS-induced ovarian dysfunction and markedly increased the cell viability, and significantly reduced the ovarian granulosa cells apoptotic rate, and down-regulated IL-6 and TNF-a expression. In addition, miRNA-146 exerted protective ovarian functions might be via inhibiting TLR4/NF-κB signaling pathway. In summary, we reveal the up-regulation of miRNA-146 expression mitigated ovarian dysfunction by negatively regulating expression of the IL-6 and TNF-a, which may shed light on the potential molecular mechanisms of overexpression of miRNA-146 may reversed the ovarian dysfunction by inhibiting the TLR4/ NF-κB signaling pathway.

Keywords: apoptosis; lipopolysaccharide (LPS); miRNA-146; ovarian dysfunction; ovarian insufficiency (POI).

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Conflict of interest statement

The authors declared that they do not have any commercial or associative interest that represents a conflict of interest in connection with the work submitted.

Figures

None
Graphical abstract
Figure 1.
Figure 1.
LPS suppresses viability of granulosa cells.
Figure 2.
Figure 2.
Transfection efficiency of miRNA-146 in granulosa cells. a: miNA-146 mimics, inhibitors and transfection efficiency. The expression levels of miRNA-146 in granulosa cells and negative control group were detected by PCR. GAPDH is quality control, *** P < 0.001.
Figure 3.
Figure 3.
The levels of miRNA-146 expression and IL-6, and TNF-a in granulosa cells. C: Granulosa cells were transfected using miR-146 mimics, miRNA-146 inhibitors or negative control (NC). The effect of miRA-146 on siTLR4 expression in granulosa cells was measured by qRT PCR and Western blot. *P < 0.01.
Figure 4.
Figure 4.
The levels of Bcl-2 and Bax protein were analyzed by western blotting. A) Overexpression of miR-146 repress apoptosis of OGCs by increasing Bcl-2, ** P < 0.01, * P < 0.05, B) versus the control. C) Overexpression of miR-146 inhibited apoptosis of OGCs by reducing Bax, ***P < 0.001, ** P < 0.01, * P < 0.05, versus the control.
Figure 5.
Figure 5.
The levels of miR-146 expression and TLR4 and NF-ĸB expression. a: TLR4 expression. These granulosa cells were transfected with miR-146 mimics, miR-146 inhibitors, control, or TLR4 siRNA (si-TLR4), and were treated using concentration of 10 μg/mL LPS for 12 h by qPCR. * P < 0.05; ** P < 0.01; *** P < 0.001. B: NF-ĸB expression. These granulosa cells were transfected with miR-146 mimics, miR-146 inhibitors, control, or TLR4 siRNA (si-TLR4), and were treated using concentration of 10 μg/mL LPS for 12 h by qPCR. * P < 0.05; ** P < 0.01; *** P < 0.001. C: The levels of TLR4 and NF-ĸB expression in granulosa cells were detected by western blot analysis.
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