Objective:To investigate the effects of trazodone on the growth, motility and ferroptosis of endometrial carcinoma cells, and to study its effect on phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/ mechanistic target of rapamycin (mTOR) signaling pathway.Methods:Human endometrial carcinoma HEC-1A cells were divided into a control group and an experimental group. HEC-1A cells in the control group and in the experimental group were treated with 0 and 2 μmol/L trazodone dimethyl sulfoxide solution, respectively, for 24 h. Cell growth was evaluated by cell counting kit-8 and colony formation assay, cell motility was evaluated by scratch assay, Transwell assay and Western blotting, ferroptosis was evaluated by Western blotting and iron detection kit, and the effect of trazodone on PI3K/Akt/mTOR signaling pathway was evaluated by Western blotting. Analysis and comparisons were made using one-way analysis of variance and Tukey′s multiple comparisons.Results:The cell survival rate [(32.2±3.2%)] and the number of cell clones (18.0±4.0) in the experimental group were lower than those in the control group [(99.2±4.3)% and 35.0±5.0], and the differences were statistically significant (both P<0.01). The relative scratch width of the experimental group (0.57±0.07) was higher than that of the control group (0.24±0.05), and the difference was statistically significant ( P<0.01). The number of invasive cells in the experimental group (7.0±1.0) was lower than that in the control group (15.0±2.0), the difference was statistically significant ( P<0.01). The relative expression levels of matrix metalloproteinase-9 (0.50±0.05) and matrix metalloproteinase-2 in the experimental group (0.75±0.08) were lower than those in the control group (0.82±0.07 and 1.25±0.15), and the differences were statistically significant (both P<0.01). The iron level [(190.5±18.5)%] and the relative expression level of acyl-CoA synthetase long-chain family member 4 (0.63±0.06) in the experimental group were higher than those in the control group [(99.2±8.9)% and 0.38±0.05)], and the differences were statistically significant (both P<0.05). The relative expression level of glutathione peroxidase 4 in the experimental group (0.22±0.05) was lower than that in the control group (1.22±0.13) ( P<0.01). The levels of phosphorylated PI3K/PI3K, phosphorylated Akt/Akt and phosphorylated mTOR/mTOR in the experimental group (0.62±0.08, 0.35±0.05, and 1.46±0.18) were lower than those in the control group (1.47±0.16, 1.32±0.11, and 2.34±0.11), and the differences were statistically significant (all P<0.01). Conclusions:Trazodone may have an anti-tumor effect on endometrial carcinoma cells by inhibiting the growth and motility of tumor cells, promoting ferroptosis, and inhibiting the PI3K/Akt/mTOR signaling pathway.

Objective:To investigate the effect and possible mechanism of the X-ray radiation-induced abscopal effects (X-RIAEs) on the ovarian reserve of mice.Methods:Totally sixteen female C57BL/6J mice aged 6-8 weeks with regular estrous cycle were randomly divided into the sham group ( n=8) and irradiation group ( n=8). After anesthesia, the mice in the irradiation group were irradiated with 8 Gy X-ray on the local area of the chest every day for 3 d, while the mice in the sham group were not irradiated. After irradiation 21 d, the estrous cycle, serum hormones, serum pro-inflammatory factors, and ovarian morphological changes were detected. Ribonucleic acid sequencing (RNA-seq) was used to detect the expression of transcriptional levels in mouse ovarian tissues. The differentially expressed genes (DEGs) were screened and analyzed by gene ontology-biological process (GO_BP). Real-time quantitative polymerase chain reaction (RT-qPCR) verified the sequencing results. The expression and localization of spermatogenesis- and oogenesis-specific basic helix-loop-helix-containing protein 1 (SOHLH1) and neutrophil elastase (NE) in ovarian tissues were detected by immunohistochemistry (IHC). Results:Compared with mice in the sham group, the irradiation group had a disordered estrous cycle, reduced primordial follicles[10.50 (1.25, 12.75) vs. 60.00 (30.00, 90.25), P<0.001] and growing follicles [(4.50 (2.50, 9.00) vs. 18.50 (18.00, 20.75), P<0.001], significantly increased atretic follicles [56.00 (45.25, 98.75) vs. 12.50 (5.25, 20.25), P<0.001]. The levels of serum estradiol [(70.28±5.27) pmol/L] and anti-Müllerian hormone [(104.00±6.98) μg/L] in the irradiation group were significantly lower than those in the sham group [(97.58±7.25) pmol/L, P=0.016; (129.70±8.39) μg/L, P=0.046], but the follicle-stimulating hormone (FSH) level in the irradiation group was not significantly different from that in the sham group ( P>0.05). Compared with the sham group, the serum levels of TNF-α [(488.30±36.20) μg/L vs. (31.61±12.89) μg/L, P<0.001] and IL-1β [(62.37±2.50) μg/L vs. (52.75±2.06) μg/L, P=0.018] in the irradiation group were significantly increased, and the serum level of interleukin (IL)-6 in the irradiation group was also increased compared with the sham group, but the difference was not statistically significant ( P>0.05). The results of GO_BP analysis showed that the down-regulated DGEs were mainly involved in the process of follicular development, and the up-regulated DGEs were involved in the inflammation process. The results of RT-qPCR were consistent with those of sequencing. The immunohistochemistry results showed that the positive expression area of SOHLH1 in the irradiation group [(23.18±4.00)%] was significantly lower than that of the sham group [(65.90±6.28)%, P=0.005], while the positive expression area of NE [(30.73±4.00)%] was significantly higher than that of the sham group [(14.47±2.22)%, P=0.024]. Conclusion:X-RIAEs can induce an inflammatory reaction in ovarian tissue and inhibit the growth and development of ovarian follicles in mice, which leads to a decrease in ovarian reserve.

Objective:To investigate the effect and possible mechanism of the X-ray radiation-induced abscopal effects (X-RIAEs) on the ovarian reserve of mice.Methods:Totally sixteen female C57BL/6J mice aged 6-8 weeks with regular estrous cycle were randomly divided into the sham group ( n=8) and irradiation group ( n=8). After anesthesia, the mice in the irradiation group were irradiated with 8 Gy X-ray on the local area of the chest every day for 3 d, while the mice in the sham group were not irradiated. After irradiation 21 d, the estrous cycle, serum hormones, serum pro-inflammatory factors, and ovarian morphological changes were detected. Ribonucleic acid sequencing (RNA-seq) was used to detect the expression of transcriptional levels in mouse ovarian tissues. The differentially expressed genes (DEGs) were screened and analyzed by gene ontology-biological process (GO_BP). Real-time quantitative polymerase chain reaction (RT-qPCR) verified the sequencing results. The expression and localization of spermatogenesis- and oogenesis-specific basic helix-loop-helix-containing protein 1 (SOHLH1) and neutrophil elastase (NE) in ovarian tissues were detected by immunohistochemistry (IHC). Results:Compared with mice in the sham group, the irradiation group had a disordered estrous cycle, reduced primordial follicles[10.50 (1.25, 12.75) vs. 60.00 (30.00, 90.25), P<0.001] and growing follicles [(4.50 (2.50, 9.00) vs. 18.50 (18.00, 20.75), P<0.001], significantly increased atretic follicles [56.00 (45.25, 98.75) vs. 12.50 (5.25, 20.25), P<0.001]. The levels of serum estradiol [(70.28±5.27) pmol/L] and anti-Müllerian hormone [(104.00±6.98) μg/L] in the irradiation group were significantly lower than those in the sham group [(97.58±7.25) pmol/L, P=0.016; (129.70±8.39) μg/L, P=0.046], but the follicle-stimulating hormone (FSH) level in the irradiation group was not significantly different from that in the sham group ( P>0.05). Compared with the sham group, the serum levels of TNF-α [(488.30±36.20) μg/L vs. (31.61±12.89) μg/L, P<0.001] and IL-1β [(62.37±2.50) μg/L vs. (52.75±2.06) μg/L, P=0.018] in the irradiation group were significantly increased, and the serum level of interleukin (IL)-6 in the irradiation group was also increased compared with the sham group, but the difference was not statistically significant ( P>0.05). The results of GO_BP analysis showed that the down-regulated DGEs were mainly involved in the process of follicular development, and the up-regulated DGEs were involved in the inflammation process. The results of RT-qPCR were consistent with those of sequencing. The immunohistochemistry results showed that the positive expression area of SOHLH1 in the irradiation group [(23.18±4.00)%] was significantly lower than that of the sham group [(65.90±6.28)%, P=0.005], while the positive expression area of NE [(30.73±4.00)%] was significantly higher than that of the sham group [(14.47±2.22)%, P=0.024]. Conclusion:X-RIAEs can induce an inflammatory reaction in ovarian tissue and inhibit the growth and development of ovarian follicles in mice, which leads to a decrease in ovarian reserve.

Objective To study the inhibitory effect of antimicrobial peptide LL-37 on Candida albicans through its ability to promote the secretion of immune factors by vaginal epithelial cells. Methods (1) LL-37 prokaryotic expression vector pET-Duet/LL-37 was constructed and its expression was induced in Escherichia coli M15. The expressed LL-37 fusion protein was purified and identified by western blot. Antifungal activity of the purified protein was initially identified by Kirby-Bauer (K-B) method. (2) Purified LL-37 protein was added to human vaginal epithelial cells co-cultured with Candida, and inhibitory effect on Candida growth was determined by the glucose consumption method. Interferon γ (IFN-γ), interleukin 10 (IL-10) concentration and IFN-γ/IL-10 ratio were measured by ELISA at different time points. Results (1) LL-37 fusion protein was purified to 96% purity at a concentration of 433.92 μg/ml, and was shown to possess anti-fungal activity confirmed by the K-B method. (2) A Candida-vaginal epithelial cells co-culture system was successfully constructed. LL-37 recombinant protein inhibited the growth of Candida with absorbance values significantly higher in the treatment group compared to the control group at all measured time points (12-hour:3.008±0.003 versus 2.967±0.003, 24-hour:2.941±0.003 versus 2.601±0.003, 48-hour:2.893 ± 0.004 versus 2.409 ± 0.003; all P<0.01). Furthermore, the rate of decrease was also much slower compared to the control group. In both control and experimental groups, IFN-γ and IL-10 secretion levels were observed to rise at first peaking at 24 hours and subsequently decrease. For each time period, IFN-γconcentration in the experimental group was significantly higher at 24 hours compared to the control group [(104.00 ± 1.07) versus (85.17 ± 0.28) pg/ml, P<0.01]. In contrast, IL-10 concentrations were significantly lower than the control group at all time points (P<0.01). IFN-γ/IL-10 ratio was also observed to be significantly higher than the control group at all measured time points (P<0.01). Conclusions (1) Recombinant protein LL-37 could significantly inhibit the growth of Candida. (2) By influencing the secretion of immune factors such as IFN-γ, IL-10, etc, recombinant protein LL-37 is able to adjust vaginal epithelial cells local immunity, and enhance resistance to Candida infection.