OR26-04 MKRN3 overexpression in the hypothalamic arcuate nucleus modulates the body temperature of adult female mice

Abstract Disclosure: D. Franssen: None. Y. Li: None. E. Torres Jimenez: None. M. Sena Esteves: None. S. Roberts: None. R.S. Carroll: None. U.B. Kaiser: None. Background and Objective: Menopause in women is characterized by estradiol depletion, leading to adverse health outcomes, including vasomotor symptoms (VMS). Recent evidence implicates hypothalamic neurokinin B (NKB) in triggering VMS. MKRN3, an E3 ubiquitin ligase expressed in the hypothalamus, is known for its role in puberty onset, with high expression in the juvenile period that decreases prior to puberty onset in mice. Recent in vivo and in vitro studies suggest that MKRN3 reduces NKB levels, indicating its potential as a novel approach to regulate the increase in NKB associated with menopausal VMS. To test this hypothesis, we overexpressed Mkrn3 via adenoviral-associated vector (AAV) injections in an ovariectomized (OVX), estrogen-deficient mouse model. Methods: Adult female wild-type mice received bilateral stereotactic injections of AAV-CMV-Mkrn3-EGFP (Mkrn3 mice) or AAV-CMV-EGFP (control mice) into the arcuate nucleus (ARC). Three weeks post-injection, the mice underwent OVX to induce estrogen deficiency. Skin (Tsk) and core body (Tc) temperatures were recorded using temperature data loggers (DST-nano probes) on the ventral tail surface and implanted in the abdomen, respectively. Measurements were taken every 10 minutes for one week pre-OVX and for two weeks post-OVX. Data were analyzed with two-way ANOVA with Fisher's post-hoc test. Upon completion, mice were perfused and their brains were harvested for IHC analysis of Mkrn3 and Nkb levels. Mice were housed in 12-hour light/dark cycle at 22.5°C. Results: Pre-OVX, Mkrn3 mice exhibited significantly higher Tc during the light phase compared to control mice (36.67°C vs. 36.36°C, n=4; p<0.05). Conversely, during the dark phase, Mkrn3 mice had lower Tc than controls (37.01°C vs. 37.39°C, n=4; p<0.05). As anticipated, OVX was associated with a significant rise in Tc in controls during the light phase (36.67°C vs. 36.36°C, n=4; p<0.05). However, Mkrn3 mice had no significant change in Tc post-OVX (36.82°C vs. 36.67°C). Consequently, there were no significant differences in Tc between Mkrn3 and control mice post-OVX. Tsk analyses revealed no significant differences between Mkrn3 and control mice, either pre- or post-OVX. IHC analysis two weeks post-OVX confirmed appropriate viral vector delivery, with the detection of EGFP in the ARC of control and Mkrn3 mice but with Mkrn3 expression only in the ARC of Mkrn3 mice. No significant differences in Nkb levels in the ARC were found between groups two weeks post-OVX. Conclusion: Our findings demonstrate that Mkrn3 overexpression in the ARC of adult female mice modulates core body temperature in a phase-dependent manner, with opposite effects during light and dark phases. These effects on temperature appear to be estrogen-dependent, as differences were abolished following estrogen depletion. In addition, the hypothesized decrease of Nkb in the presence of Mkrn3 was not seen after OVX. Presentation: Monday, July 14, 2025