Small Peptides from Periplaneta americana Inhibits Oxidative Stress-Induced KGN Cell Apoptosis by Regulating Mitochondrial Function Through Bcl2L13.

This study examined the protective effects of small peptides from Periplaneta americana against H2O2-induced mitochondrial injury in human ovarian granulosa cells. The ATP level and mitochondrial membrane potential as well as the quantity and ultrastructure of mitochondria in cells were detected. Mitochondrial DNA copy number and expression levels of Bcl2L13, LC3B, and p62 were tested. Targeted silencing of Bcl2L13 expression in KGN cells. The expression levels of Bcl2L13 and LC3B as well as interaction were evaluated. The ATP level, mtDNA-CN, and MMP of the H2O2 group were significantly lower than those of the normal control group (P < 0.05), accompanied by a reduction in mitochondrial mass and mitochondrial fluorescence intensity (P < 0.05). However, the ATP level, mtDNA, and MMP in KGN cells were increased after SPPA treatment (P < 0.05). Scanning electron microscopy shows that SPPA ameliorates H2O2-induced structural damage to mitochondria. Moreover, the expression levels of Bcl2L13 and p62 in the H2O2 group were downregulated significantly compared with those of the normal control group (P < 0.05), while LC3B was upregulated (P < 0.05). After SPPA treatment, the expression levels of Bcl2L13 and p62 were upregulated (P < 0.05), while LC3B was downregulated (P < 0.05). The Co-IP results indicated that Bcl2L13 and LC3B interacted, and this interaction was weakened after cell treatment with H2O2, and dissociation between Bcl2L13 and LC3B declined after SPPA treatment. SPPA inhibits KGN cell apoptosis induced by oxidative stress via inhibition of mitochondrial injury Bcl2L13-mediated mitochondrial autophagy might participate in the regulation process.

Signaling pathways of Periplaneta americana peptide resist H2O2-induced apoptosis in pig-ovary granulosa cells through FoxO1.

Granulosa cell apoptosis induced by oxidative stress is an important cause of follicular atresia. Our previous studies found that Periplaneta americana peptide (PAP) decreased H2O2-induced apoptosis of pig-ovary granulosa cells (PGCs) through FoxO1. The aim of this study is to investigate the signaling pathways involved in PAP resistance against H2O2-induced apoptosis of PGCs. PGCs obtained from the follicles of non-estrous Duroc × Landrace × Yorkshire gilts (5 months old, 50-55 kg) were treated with H2O2 and PAP, or together with inhibitors against PI3K and JNK, and then collected for ROS levels and SOD activities detection, TUNEL staining, qRT-PCR, western blotting, immunofluorescence or coimmunoprecipitation. Results showed that the increased ROS levels and decreased activities of SOD caused by H2O2 stimulation were reversed by PAP. Additionally, PAP downregulated the differential abundance of mRNA of Bax and FasL, thus inhibiting H2O2-induced apoptosis of PGCs. PAP significantly reduces p-JNK expression and increases the p-FoxO1/FoxO1 expression ratio, thereby decreasing caspase-3 expression and cell apoptosis in H2O2-induced PGCs. PAP promotes the combination of FoxO1 with the 14-3-3 protein, increases FoxO1 translocation to the cytoplasm, and decreases FoxO1 acetylation. Therefore, PAP regulates FoxO1 expression through the JNK/FoxO1 signaling pathway and effects the translocation of FoxO1 to the cytoplasm by the FoxO1 interaction with 14-3-3, enabling reversal of the H2O2-induced apoptosis of PGCs. Acetylation of FoxO1 is also involved in the antiapoptotic effect of PAP.

Mechanism of action and the uses betaine in pig production.

Betaine, the trimethyl derivative of glycine, is a good methyl group donor, and an important component in pig production. However, betaine has not been extensively studied in this field. Therefore, in this study, we reviewed the effects of betaine in pig production performance, meat quality and reproductive performance, as well as its mechanisms, to provide a theoretical basis for the optimal use and development of this compound.

Protective effect of small peptides from Periplaneta americana on cyclophosphamide-induced premature ovarian failure.

AIM: To investigate the protective effect of small peptides from Periplaneta americana (SPPA) on cyclophosphamide (CP)-induced premature ovarian failure (POF) in mice. Silent mating type information regulation 2 homolog 1 (SIRT1) /tumor-associated protein 53 (p53) signaling pathway plays an important role in delaying POF. Hematopoietic progenitor cell antigen (CD34) reflects ovarian aging from the side. However, whether SPPA inhibits POF in mice by influencing the SIRT1/p53 pathway and CD34 expression remains to be studied. METHODS: Forty female Kun Ming (KM) mice were divided into four groups: a control group (normal saline, n = 10), POF model group (160 mg/kg CP, n = 10), SPPA low-dosage group (160 mg/kg CP + 100 mg/kg SPPA, n = 10), and SPPA high-dosage group (160 mg/kg CP + 200 mg/kg SPPA, n = 10). CP administration route is intraperitoneal injection, and SPPA administration route is intragastric. Eyeball enucleation blood samples and the ovaries of mice were collected by midline laparatomy and oopherectomy, and the malondialdehyde (MDA), nitric oxide (NO), superoxide dismutase (SOD), follicle-stimulating hormone (FSH), and anti-Müllerian hormone (AMH) concentrations were tested. Immunohistochemical tests for the expressions of SIRT1, p53, and CD34 were carried out. Finally, ovarian mRNA levels of SIRT1 and p53 were detected with real-time fluorescence quantification PCR (qRT-PCR). RESULTS: A mouse model of POF was generated using 160 mg/kg of CP. Compared with POF group, we found that plasma NO, MDA, and FSH decreased, while AMH and SOD increased in the SPPA low-dose group. Compared with the POF group, the SPPA low- and high-dosage groups achieved significant growth in the number of primordial, primary, and total number of healthy follicles at all levels, but sharp reductions in the number of atretic follicles. In addition, we found downregulated protein and mRNA expression of SIRT1, and upregulated that of p53 were observed in ovarian tissues of treated mice with POF, in immunohistochemistry experiments and qPCR experiments. In contrast, high protein and mRNA expression of SIRT1, and low that of p53 were observed in SPPA treatment groups. And the results of CD34 protein expression were consistent with that of SIRT1. CONCLUSION: In total, SPPA significantly inhibited POF caused by CP in mice via activation of the SIRT1/p53 signaling pathway in the mouse ovary.

Periplaneta americana peptide decreases apoptosis of pig-ovary granulosa cells induced by H2 O2 through FoxO1.

Oxidative stress can induce apoptosis of granulosa cells and lead to follicular atresia, thereby reducing the number of pigs giving birth. The aim of this study was to investigate the protective effect of Periplaneta americana peptide (PAP) on the apoptosis of the granulosa cells of pig ovaries (PGCs) induced by hydrogen peroxide (H2 O2 ) via FoxO1. PGCs were treated with H2 O2 to establish a cell apoptosis model. Cell viability was measured using the cell counting kit-8 (CCK-8) assay, and cell apoptosis was detected using flow cytometry. The malondialdehyde (MDA) level and nitric oxide (NO) content were detected to reflect the oxidative stress. Western blotting, qRT-PCR and overexpression were undertaken to determine the expression of FoxO1 and caspase-3, and immunofluorescence was used to detect FoxO1 in the nucleus and cytoplasm. PGCs were treated with 100 µM H2 O2 for 6 hr, which resulted in oxidative damage and apoptosis and an apoptosis rate for PGCs of 32.95%. Next, PGCs were treated with 400 µg/ml PAP for 24 hr to repair the apoptosis induced by H2 O2 . PAP improved cell viability in H2 O2 -stimulated PGCs, the increased MDA level and NO content caused by H2 O2 stimulation were reversed and the apoptotic rate of PGCs was reduced. The qRT-PCR and Western blotting results indicated that PAP decreased the H2 O2 -induced apoptosis and the expression of FoxO1 and caspase-3 in PGCs. The effect of PAP was the same following FoxO1 overexpression. FoxO1 was expressed in the nucleus when stimulated by H2 O2 or overexpression; however, it migrated to the cytoplasm following PAP treatment. PAP decreased the apoptosis of PGCs induced by H2 O2 by regulating FoxO1 expression and nuclear translocation.

The protective effect of small peptides from Periplaneta americana on hydrogen peroxide-induced apoptosis of granular cells.

This study investigates the protective effect of small peptides from Periplaneta americana (SPPA) on hydrogen peroxide (H2O2)-induced apoptosis of ovarian granular cells. H2O2 was applied to human ovarian granular cells (KGN cell strains). Cell viability was tested by cell counting Kit-8 (CCK-8). Cell apoptosis was tested by flow cytometry, and a cell apoptosis model was established. The model cells were treated with SPPA, and the cell survival rate was monitored using the CCK-8 method. The oxidative stress state of cells was examined using SOD, ROS, MDA, and NO kits. The protein expression levels of SIRT1, p53, and the apoptosis-related gene Caspase3 were measured using Western Blot methodology. Relative to the control group, cell viability declined significantly after the H2O2 treatment only (P < 0.01), while the apoptosis rate increased significantly (P < 0.01). The activity of SOD was weakened significantly (P < 0.01), while the cell levels of ROS, MDA, and NO increased dramatically (P < 0.01). Cell viability dramatically recovered (P < 0.01), and the SOD activity is hugely increased (P < 0.01) after SPPA treatment. In contrast, contents of ROS, MDA, and NO decreased sharply (P < 0.01), and significant dose-response relationships are characterized. Moreover, the H2O2 treatment group showed significantly downregulated expression of SIRT1 (P < 0.01) but significantly upregulated expressions of p53 and Caspase3 (P < 0.01) compared to the control group. Following the SPPA treatment of apoptosis cells, expression of SIRT1 increased significantly, while expressions of p53 and Caspase3 declined significantly (P < 0.01). This study suggests that SPPA inhibits H2O2-induced human KGN cell apoptosis through antioxidation, and the SIRT1/p53 signal pathway mediates the antioxidation.

Copper oxide nanoparticles alter cellular morphology via disturbing the actin cytoskeleton dynamics in Arabidopsis roots.

Copper oxide nanoparticles (CuO NPs) have severe nano-toxic effects on organisms. Limited data is available on influence of CuO NPs on plant cells. Here, the molecular mechanisms involved in the toxicity of CuO NPs are studied. Exposure to CuO NPs significantly increased copper content in roots (0.062-0.325 mg/g FW), but CuO NPs translocation rates from root to shoot were low (1.1-2.8%). Presented data were significant at p < 0.05 compared to control. CuO NPs inhibited longitudinal growth and promoted transverse growth in root tip cells. However, CuO NPs did not affect the leaf cells, implying that the transfer ability of CuO NPs was weak, and toxicity mainly affected roots. CuO NPs can conjugate with actin protein. The actin cytoskeleton experienced reorganization in the presence of CuO NPs. The longitudinal filamentous actin (F-actin) decreased, and the transverse F-actin increased. CuO NPs inhibited actin polymerization and promoted depolymerization. The behavior of individual F-actin was at steady state with time-lapse under CuO NPs treatment by time-lapse reflection fluorescence (TIRF) microscopy. The growth rate of actin filaments was weakened by CuO NPs. CuO NPs disturbed the subcellular localization of PINs and the gradient of auxin distribution in root tips in an actin-dependent manner. In conclusion, CuO NPs conjugated with actin and disturbed F-actin dynamics, triggering abnormal cell growth in the root tip, and findings provide theoretical basis for further study nano-toxicity in plants.

Reliability and validity of the Chinese pediatric voice handicap index.

OBJECTIVES: To evaluate the reliability and validity of the Chinese version of pediatric voice handicap index (pVHI). MATERIAL AND METHODS: The original English version-pVHI was translated into Chinese. Parents of 52 children with voice dysphonia and 43 children with no history or symptoms of voice problems were asked to fill the Chinese pVHI questionnaires twice with an interval of 2 weeks. GRB (Grade, Roughness, Breathiness) scale was used for perceptual assessment by two otolaryngologists and one speech pathologist for each child's voice. The internal consistency was assessed using Cronbach's alpha coefficient. Pearson's correlation coefficient was used to evaluate the test-retest reliability. The Kendall's coefficient of concordance W was used to assess the consistency of GRB scores of 3 voice specialists. The nonparametric Mann-Whitney test was used to assess the differences between the dysphonia group and controls. The correlation between pVHI and GRB scores were assessed using Pearson's correlation coefficient. RESULTS: The internal consistency of total score and three subscales scores of Chinese pVHI were 0.788-0.944. The test-retest reliability was 0.631-0.887(P < .001). The pVHI scores of control group significantly were lower than the pathological group (P = .000). The GRB scores of 3 voice specialists have an excellent consistency (W = 0.694-0.807, P = .000). The pVHI scores positively correlated with GRB assessment (P < .01). CONCLUSIONS: The Chinese version of pVHI had a good reliability and validity. It can be applicable and useful supplementary tool for evaluating parents' perception of their children's dysphonia.

Survival in Vivo Canine Phonation Model Without Stimulation.

OBJECTIVES: We describe a survival nonstimulated in vivo canine phonation model using distending laryngoscope, cramp frame, and constant humidified glottal airflow to elicit phonation. METHODS: Five beagle dogs were involved in this study. One cuffed endotracheal tube was placed below the glottis through the tracheotomy and delivered humidified airflow to the glottis. Arytenoids approximation was maintained using a clamp under the distending laryngoscope. Acoustic and aerodynamic parameters were measured using synchronous signal collection system and analysis software. Vocal oscillation also was examined using stroboscope laryngeal imaging. RESULTS: For the nonstimulated in vivo phonation animal, the sound intensity and fundamental frequency were 78.3 ± 6.8 dB and 127.6 ± 29.2 Hz in the first experiment and 82.9 ± 6.6 dB and 175.2 ± 4.4 Hz 4 weeks later. The aerodynamic analysis revealed the mean subglottal phonation threshold pressure (PTP) and phonation threshold flow (PTF) were 8.5 ± 4.0 cmH20 and 683.0 ± 356.4 mL/s in the first experiment and 16.1 ± 8.6 cmH20 and 384.8.0 ± 230.6 mL/s in the second experiment 4 weeks later. Stroboscope image revealed sustained vocal vibration during great airflow delivery to glottis in the phonation animal model. CONCLUSIONS: We developed a survival nonstimulated in vivo phonation canine model that allows the study of long-term animal phonation study as its own control.