Trizol buffer

The qPCR experiments were performed as described previously [19 (link), 20 (link)]. The total RNA (Ribonucleic Acid) of cells were extracted using the Trizol buffer (Beyotime) and were reverse-transcribed by Multiscribe™ Reverse Transcriptase (Applied Biosystems, Thermo Scientific Corporation) according to the manufacturer’s instructions. The relative expression was calculated by the comparative Ct method. The primers used for qPCR analysis were shown in Table 1.Table 1

The primers used in this work

Targets	Primer sequence (5′–3′)
MMP-9	Forward: 5′-GGGACGCAGACATCGTCATC-3′
	Reverse: 5′-TCGTCATCGTCGAAATGGGC-3′
Bax	Forward: 5′-TGAAGCGACTGATGTCCCTG-3′
	Reverse: 5′-CAAAGATGGTCACGGTCTGC-3′
Caspase-3	Forward: 5′-CCTGGTTCATCCAGTCGCTT-3′
	Reverse: 5′-TCTGTTGCCACCTTTCGGTT-3′
Bcl-2	Forward: 5′-GTGAAGTCAACATGCCTGCC-3′
	Reverse: 5′-ACAGCCTGCAGCTTTGTTTC-3′

Furosine was purchased from PolyPeptide (Strasbourg, France); the purity was above 95%. Dulbecco’s Modified Eagle Medium (DMEM), fetal bovine serum (FBS), and non-essential amino acids (NEAA) were obtained from GIBCO (Waltham, MA, USA), 1% penicillin/streptomycin was purchased from Thermo Fisher (Waltham, MA, USA). Lactoferrin was purchased from Sigma (St. Louis, MO, USA), with the purity of above 95%.
Cell counting kit-8 (CCK-8 kit) was purchased from Solarbio (Beijing, China), Elisa detection kits of T, FSH and LH in mice serum were purchased from Jiancheng (Nanjing, China). Hematoxylin-Eosin (HE) staining kit was purchased from Solarbio (Beijing, China).
Trizol buffer, RIPA lysate buffer (including proteases and PMSF), and protein concentration detection kit were obtained from Beyotime (Nanjing, China). Primers of GAPDH, Cep55, NF-κB, PI3K, Akt, FOX01 and TNF-α were synthesized from Sangon (Shanghai, China), reagents related with quantitative real time polymerase chain reaction (q-PCR) were purchased from Sangon. Primary antibodies of β-actin, Cep55, NF-κB, PI3K, p-PI3K, Akt, p-Akt, FOX01, p-FOX01, and TNF-α, as well as the secondary antibodies, were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Reagents related with western blotting were purchased from Solarbio. Enhanced chemiluminescence (ECL) reagent was purchased from Tanon (Shanghai, China).

Total RNA of KCs (5 × 10⁶) was extracted using Trizol buffer (R0016, Beyotime, China). The total RNA samples were reverse transcribed into cDNA strictly following the protocol of the Primescript™ RT reagent kit with the gDNA Eraser (RR047A, Takara, Japan). PCR was conducted using the SYBR premix Ex Taq II kit (RR820A, Takara, Japan). The relative expression of the targeted gene was determined using the –(delta delta C(T)) method after normalizing against the β-actin gene. The primers for the targeted genes are shown in Supplementary Figure 2.