Figure 5. Warmth and Warm–Microbiota Transplantation Ameliorate Ovariectomy–Induced Transcriptional Deregulation.

(A) Mean-difference plot (MD-plot) of the log fold change gene expression between tibias of 24 weeks old female mice that were ovariectomized or sham-operated at 16 weeks of age, and then kept at RT for two months (OvaRT vs ShamRT, respectively) shown as average count per million (CPM). Red dots show the increased and blue show the decreased genes selected for FDR<0.05.

(B) MD-plot of the log fold change of gene expression between tibias of 24 weeks old female mice that were ovariectomized at 16 weeks of age and then kept at 34°C for two months (Ova34), versus ShamRT, shown as average count per million (CPM). Red dots show increased and blue show decreased genes selected for FDR<0.05.

(C) Comparison between the log fold change of genes deregulated by ovariectomy at RT shown in red (|log₂FC| > 1; OVART vs ShamRT) and the same genes when exposing the ovariectomized-mice at 34°C shown in blue (|log₂FC| > 1; Ova34°C vs ShamRT).

(D) Top 10 most deregulated Reactome pathways between tibias of Ova RT and Ova34°C mice.

(E) Volcano-plot comparing the p-value and the log fold change of gene expression between tibias of 21 weeks old ovariectomized, microbiota recipient female mice (OvaTransp34°C, or OvaTranspRT) as in Figure 4A-E. Green dots: (|log₂FC| > 1; blue dots: P < 0.01; red dots: P <0.01 and (|log₂FC| > 1.

(F and G) Expression analysis of the ovariectomy-altered genes ((|log₂FC| > 1) at RT (OvaRT vs shamRT)) in tibia from OvaTransp34°C mice compared to the OvaTranspRT controls (OvaTransp34°C vs. OvaTranspRT). In (F) blue and red show genes (up- or downregulated, respectively) unaltered by microbiota transplantation. Green show genes with reduced or reverted expression when mice are transplanted with warm-microbiota. (G) Comparison between the log fold changes of genes (|log₂FC| > 1) as in (C) using the groups of mice from

(F): OvaRT vs ShamRT (red) and OvaTransp34°C vs. OvaTranspRT (blue).

(H) Top 10 most deregulated reactome pathways between tibias from OvaTransp34°C and OvaTranspRT mice.

(I) Top 10 most deregulated reactome pathways between tibias from 34°C and RT mice.