Rhri404

To induce the rat model of POF, 20 rats were subjected to the intraperitoneally (IP) administration of cyclophosphamide (CTX; Cat no: RHRI404, Supelco) at a dose of 200 mg/kg on day 1 and 8 mg/kg on days 2 to day 14. CTX is an active substance and can destroy follicles by the mechanism of apoptosis and tissue necrosis [38 ]. According to previous protocols, 21 days after the last injections rats can exhibit POF features [39 ]. To confirm POF status, 3 rats were randomly selected from both POF and the control groups and euthanized using an overdose of Ketamine and Xylazine. The left 18 rats were arbitrarily allocated into Control, POF, and POF plus music therapy. The rats in the experimental group were kept in the music box for 4 and 8 weeks (Fig. 1A).
Fig. 1

Timeline of the experimental procedure (A), and the design and development of acoustic music box (B)

Seven days after acclimation, a POI rat model was induced by the administration of cyclophosphamide (CTX; Cat no: RHRI404, Supelco). In short, CTX was injected intraperitoneally (i.p.) at a dose of 200 mg/kg BW on day 1 and 8 mg/kg bw from days 2 to 14. To confirm the POI status, three rats from the CTX-injected and control groups were randomly selected and subjected to histopathological examination and hormonal evaluations. The remaining POI rats were randomly allocated into three groups as follows: Control (Control-matched POI group, n = 7), Sham (POI + normal saline, n = 7), and AF-Exos (POI + AF-Exos, n = 7) (Fig. 1A).
Fig. 1

Schematic presentation experimental setup. Animal modeling procedure, exosome enrichment, intervention, sampling and mating (A), characterization of exosomes, size distribution (B) and zeta potential (C) assessment by dynamic light scattering method, scanning electron microscopy (D), transmission electron microscopy (E) and western blot analysis (F)