Fig. 7: Overexpression of FTO in the ARC promoted early puberty onset through the PLCβ3/Ca2+/CAMK signalling pathway. | Communications Biology

Fig. 7: Overexpression of FTO in the ARC promoted early puberty onset through the PLCβ3/Ca2+/CAMK signalling pathway.

From: FTO-mediated m6A demethylation regulates GnRH expression in the hypothalamus via the PLCβ3/Ca2+/CAMK signalling pathway

Fig. 7

a Left panel: schematic illustrations of scAAV microinjection. The picture material of mouse was created with BioRender.com. Right panel: transduction of ARC cells with scAAV delivered to the ARC in female rats. Red triangle: microinjection site. Green: scAAV-infected cells; blue: DAPI-stained cell nuclei in the hypothalamus; dotted line: ARC. 3 V, third ventricle. Scale bars, 100 μm. b Cumulative percentages of vaginal opening (VO) after intra-ARC AAV-FTO and AAV-control transfection. Blue shading depicts the time when all AAV-FTO animals showed VO. c GnRH abundance in serum at 4 weeks (n = 8) and 5 weeks (n = 6) in female rats as determined by ELISA. d Uterine/ovarian coefficients in the rats transfected with AAV-control or AAV-FTO at different stages (n = 5–8). e, f Pathological assessment of follicular development by H&E staining in the ARC-transfected group (e) and the number of corpora lutea (f) (n = 5). Scale bars, 500 μm. g, h Serum LH (g) and FSH (h) levels in 4-week-old (n = 8) and 5-week-old (n = 6) female rats as determined by ELISA in the ARC-transfected group. i–k Pathological assessment of the uterus by H&E staining in the ARC-transfected group (i) and the myometrial thickness index (j) and endometrial thickness index (k) between the groups. Scale bars, 500 μm. l Liver coefficient in the rats transfected with AAV-control or AAV-FTO at different stages (n = 6–8). m, o FTO, PLCβ3, CAM, and CAMKII mRNA levels detected by qPCR in the ARC in 4-week (m) or 5-week (o) female rats receiving AAV-control (n = 3) or AAV-FTO (n = 3). n, p FTO, PLCβ3, CAM, CAMKII and phosphorylated CAMKII proteins detected by western blotting in the ARC in 4-week (n) or 5-week (p) female rats receiving AAV-control (n = 3) or AAV-FTO (n = 3). Uterine/ovarian organ coefficient = [uterine/ovarian weight (g)]/body weight (g). Liver organ coefficient = liver weight (g)]/body weight (g). AAV-FTO for the FTO-overexpression group, AAV-control for the AAV negative control group. The bars represent the means ± SEMs (n = 3). *P < 0.05, **P < 0.01, ***P < 0.001, and ns, P > 0.05 versus the AAV-control group by Student’s t test.

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