Adverse Effects of Avobenzone on Boar Sperm Function: Disruption of Protein Kinase A Activity and Tyrosine Phosphorylation.

Avobenzone (AVO), an ultraviolet (UV) filter, is frequently used as an ingredient in personal cosmetics. This UV filter has been found to be easily exposed in swimming pools and beaches, and it has been detected in human urine and blood. Moreover, numerous studies have demonstrated that AVO exhibits endocrine-disrupting properties. Nevertheless, the effects of AVO on male fertility have not yet fully understood. Therefore, this study aimed to assess the effects of AVO on various sperm functions during capacitation. First, boar spermatozoa were treated with various AVO concentrations. After treatment, sperm motility and kinetic characteristics, capacitation status, intracellular adenosine triphosphate (ATP) levels, and sperm viability were evaluated. Moreover, Western blot analysis w.as conducted to evaluate protein kinase A (PKA) activity and tyrosine phosphorylation. As a result, AVO treatment significantly decreased total motility, progressive motility, and several kinetic characteristics at high concentrations (50 and 100 µM). Furthermore, the capacitation status dose-dependently decreased. Conversely, no significant differences in acrosome reaction, cell viability, and intracellular ATP levels were observed. However, the intracellular ATP level tended to decrease. In addition, AVO dose-dependently induced abnormal changes in PKA activity and tyrosine phosphorylation. Although AVO did not directly exert a toxic effect on cell viability, it ultimately negatively affected sperm functions through abnormal alterations in PKA activity and tyrosine phosphorylation. Thus, the potential implications on male fertility must be considered when contemplating the safe utilization of AVO.

Ritonavir Has Reproductive Toxicity Depending on Disrupting PI3K/PDK1/AKT Signaling Pathway.

Ritonavir (RTV) is an antiviral and a component of COVID-19 treatments. Moreover, RTV demonstrates anti-cancer effects by suppressing AKT. However, RTV has cytotoxicity and suppresses sperm functions by altering AKT activity. Although abnormal AKT activity is known for causing detrimental effects on sperm functions, how RTV alters AKT signaling in spermatozoa remains unknown. Therefore, this study aimed to investigate reproductive toxicity of RTV in spermatozoa through phosphoinositide 3-kinase/phosphoinositide-dependent protein kinase-1/protein kinase B (PI3K/PDK1/AKT) signaling. Duroc spermatozoa were treated with various concentrations of RTV, and capacitation was induced. Sperm functions (sperm motility, motion kinematics, capacitation status, and cell viability) and expression levels of tyrosine-phosphorylated proteins and PI3K/PDK1/AKT pathway-related proteins were evaluated. In the results, RTV significantly suppressed sperm motility, motion kinematics, capacitation, acrosome reactions, and cell viability. Additionally, RTV significantly increased levels of phospho-tyrosine proteins and PI3K/PDK1/AKT pathway-related proteins except for AKT and PI3K. The expression level of AKT was not significantly altered and that of PI3K was significantly decreased. These results suggest RTV may suppress sperm functions by induced alterations of PI3K/PDK1/AKT pathway through abnormally increased tyrosine phosphorylation. Therefore, we suggest people who use or prescribe RTV need to consider its male reproductive toxicity.

Perfluorooctanoic acid suppresses sperm functions via abnormal Protein Kinase B activation during capacitation.

Perfluorooctanoic acid (PFOA) is a perfluorinated compound, a synthesized chemical, and has been used in several industrial products for more than 70 years. Although PFOA is known to exert toxic effects in normal cells, there is no detailed information on its reproductive toxicity and its effects on sperm functions related to protein kinase B (AKT). Therefore, this study was conducted to explore the effects of PFOA on sperm functions via AKT. Boar spermatozoa were incubated with different concentrations of PFOA (0, 0.1, 1, 10, and 100 µM) to induce capacitation. Sperm functions (sperm motility, motion kinematic parameters, capacitation status, cell viability, and intracellular ATP levels) were evaluated. In addition, the expression levels of AKT, phospho-AKT, phospho-PKA, and tyrosine phosphorylated proteins were evaluated by western blotting. Results showed significant decreases in sperm motility and motion kinematic parameters. PFOA treatment significant suppressed spermatozoa capacitation and intracellular ATP levels. Furthermore, it significantly decreased the levels of phospho-PKA and tyrosine phosphorylated proteins. The levels of AKT phosphorylation at Thr308 and Ser473 also significantly decreased. These findings suggest that PFOA diminishes sperm functions during capacitation and induces unnatural phosphorylation in AKT, leading to reproductive toxicity. Therefore, people should be aware of reproductive toxicity when using PFOA.

The natural flavonoid compound deguelin suppresses sperm (Sus Scrofa) functions through abnormal activation of the PI3K/AKT pathway.

Deguelin is a natural flavonoid extracted from plants belonging to the Lonchocarpus, Derris, or Tephrosia genera. It inhibits AKT activity in tumors and has the potential to be used as a treatment for malignant tumors. However, the risks associated with the use of deguelin on male fertility have not yet been explained in detail. Therefore, this study was conducted to investigate the effects of deguelin on sperm functions during capacitation. First, boar spermatozoa were exposed to different concentrations of deguelin (0.1, 1, 10, 50, and 100 µM). Next, sperm functional assessments, such as sperm motility, capacitation status, intracellular ATP level, and cell viability, were performed. The expression levels of PI3K/AKT-related proteins and the phosphorylation of their tyrosine residues were also evaluated by western blotting. No significant difference was observed in cell viability; however, deguelin considerably decreased sperm motility and motion kinematics in a dose-dependent manner. Although no significant difference was observed in the capacitation status, acrosome reaction decreased at high concentrations of deguelin (50 and 100 µM). Furthermore, intracellular ATP levels were significantly decreased in all deguelin treatment groups compared with those in the control group. Results of western blotting revealed that deguelin substantially diminished tyrosine phosphorylation. Interestingly, in contrast to previous studies showing that deguelin inhibits AKT activity, our results showed that it increased the expression of PI3K/AKT pathway-related proteins. Collectively, these findings indicate that deguelin exerts negative effects on sperm functions due to abnormal PI3K/AKT signaling activation. We believe that this is the first study to provide evidence that deguelin can regulate sperm functions independent of PI3K/AKT pathway inhibition. Furthermore, its detrimental effects on male fertility should be considered while developing or using deguelin as a therapeutic agent.

Comparison of Four Swab and Transport Media Combinations for the Detection of Respiratory Viruses.

BACKGROUND: We compared four combinations of nasopharyngeal swabs and transport media for their ability to transfer and recover viruses under different storage conditions. METHODS: Each swab was immersed in culture supernatants of influenza A virus (IAV), respiratory syncytial virus, and adenovirus, placed in transport medium, and stored at -20℃, +4℃, +20 to 25℃, and +37℃ for 5 days. On each day, virus culture and real-time PCR were performed for each virus. RESULTS: All samples under different storage conditions showed positive results up to 5 days using both virus culture and real-time PCR. Real-time PCR showed that samples stored at -20℃, 4℃, and 20 - 25℃ were within two cycle thresholds (Cts) up to 5 days, but IAV at 37℃ showed that viral titer decreased after 3 days. CONCLUSIONS: Our results indicate that these swab and transport media maintained the stability of the above viruses for 5 days at room temperature, refrigerated, and frozen storage conditions.

Reproductive toxicity of ritonavir in male: Insight into mouse sperm capacitation.

Since COVID-19 began in 2019, therapeutic agents are being developed for its treatment. Among the numerous potential therapeutic agents, ritonavir (RTV), an anti-viral agent, has recently been identified as an important element of the COVID-19 treatment. Moreover, RTV has also been applied in the drug repurposing of cancer cells. However, previous studies have shown that RTV has toxic effects on various cell types. In addition, RTV regulates AKT phosphorylation within cancer cells, and AKT is known to control sperm functions (motility, capacitation, and so on). Although deleterious effects of RTV have been reported, it is not known whether RTV has male reproduction toxicity. Therefore, in this study, we aimed to investigate the effects of RTV on sperm function and male fertility. In the present study, sperm collected from the cauda epididymis of mice were incubated with various concentrations of RTV (0, 0.1, 1, 10, and 100 µM). The expression levels of AKT, phospho-AKT (Thr308 and Ser473), and phospho-tyrosine proteins, sperm motility, motion kinematics, capacitation status, and cell viability were assessed after capacitation. The results revealed that AKT phosphorylation at Thr308 and Ser473 was significantly increased, and the levels of tyrosine-phosphorylated proteins (at approximately 25 and 100 kDa) were significantly increased in a dose-dependent manner. In addition, RTV adversely affected sperm motility, motion kinematics, and cell viability. Taken together, RTV may have negative effects on sperm function through an abnormal increase in tyrosine phosphorylation and phospho-AKT levels. Therefore, individuals taking or prescribing RTV should be aware of its reproductive toxicity.

GRP78 plays a key role in sperm function via the PI3K/PDK1/AKT pathway.

Glucose-regulated protein 78 (GRP78), which is commonly found in the endoplasmic reticulum (ER), is involved in stabilizing ER proteins and inducing the unfolded protein response. Furthermore, GRP78 is expressed on the surface of most common cancer cells, such as cells of breast, lung, liver, and prostate cancers, and plays a role in apoptosis and cell proliferation via the PI3K/PDK1/AKT signaling pathway. Therefore, various trials have been performed for evaluating cancer treatment by inhibiting GRP78. Moreover, GRP78 is expressed on the surface of spermatozoa; however, its role in spermatozoa physiology remains unclear. Therefore, this study was designed to investigate the effects of GRP78 on sperm function during capacitation and elucidate the underlying mechanisms. Boar spermatozoa were exposed to various concentrations of HA15, a GRP78 antagonist, and sperm kinematic parameters, capacitation status, cell viability, levels of PI3K/PDK1/AKT-pathway related proteins, and tyrosine phosphorylation were evaluated. GRP78 inhibition significantly decreased sperm motility, kinematic parameters, capacitated and acrosome-reacted spermatozoa counts, and cell viability. Moreover, GRP78 expression was significantly decreased in HA15-treated spermatozoa compared to that in the control group, and levels of PI3K/PDK1/AKT-pathway related proteins changed significantly. Furthermore, tyrosine phosphorylation was significantly altered in the HA15-treated group. The results of this study suggest that GRP78 inhibition in cancer therapy may negatively affect sperm function. These results lay a strong foundation for future studies aiming to identify the molecular mechanisms related to GRP78 in spermatozoa.

Bioassay-Guided Separation of Centipeda minima Using Comprehensive Linear Gradient Centrifugal Partition Chromatography.

A comprehensive linear gradient solvent system for centrifugal partition chromatography (CPC) was developed for the bioassay-guided isolation of natural compounds. The gradient solvent system consisted of three different ternary biphasic solvents types: n-hexane-acetonitrile-water (10:2:8, v/v), ethyl acetate-acetonitrile-water (10:2:8, v/v), and water-saturated n-butanol-acetonitrile-water (10:2:8, v/v). The lower phase of the n-hexane-acetonitrile-water (10:2:8, v/v) was used as the stationary phase, while its upper phase, as well as ethyl acetate-acetonitrile-water (10:2:8), and water-saturated n-butanol-acetonitrile-water (10:2:8, v/v) were pumped to generate a linear gradient elution, increasing the mobile phase polarity. We used the gradient CPC to identify antioxidant response elements (AREs), inducing compounds from Centipeda minima, using an ARE-luciferase assay in HepG2 cells, which led to the purification of the active molecules 3-methoxyquercetin and brevilin A. The developed CPC solvent systems allow the separation and isolation of compounds with a wide polarity range, allowing active molecule identification in the complex crude extract of natural products.

Anal Human Papillomavirus Infection among HIV-Infected Men in Korea.

BACKGROUND: Little is known about the epidemiology on human papillomavirus (HPV) infection among HIV-infected men in Korea. The objective of this study was to determine the prevalence, genotype distribution and risk factors associated with anal HPV infection among HIV-infected men in Korea. METHODS: A single-center cross-sectional study was conducted with HIV-infected men in Korea. Participants completed a detailed sexual behavior risk factor questionnaire. Anal samples were collected for cytology and HPV genotyping. Factors associated with anal HPV infection were assessed using multivariable logistic regression, stratifying by sexual behaviour. RESULTS: A total of 201 HIV-infected men were included in the study: 133 were from men who have sex with men (MSM) and 68 from men who have sex with women (MSW). Any anal HPV infection was detected in 82.7% of HIV-infected MSM and in 51.5% of HIV- infected MSW (P < 0.001). High-risk HPV (HR-HPV) prevalence was higher among MSM (47.4%) than MSW (25.0%; P = 0.002). The HR-HPV types identified most frequently were HPV 16 (11%), HPV 18 (9.9%), and HPV 58 (5%) in MSM, and HPV 58(11%) and HPV 16 (8.9%) in MSW. Prevalence of any HPV types in 9-valent vaccine types was higher among MSM than MSW (47.4% vs 22.1%. P = 0.001). Abnormal anal cytology was more commonly detected in MSM than MSW (42.9% vs.19.1%, P < 0.001). In HIV-infected MSM, higher number of lifetime male sex partners was significantly associated with any anal HPV infection, but age was a significant risk factor associated with anal HR-HPV infection. CONCLUSION: Anal HPV infection was highly prevalent in HIV-infected MSM in Korea, and also commonly found in HIV-infected MSW. In HIV-infected MSM, the significant risk factor for being infected with any HPV infection was lifetime number of male sexual partners, and with anal oncogenic HPV infection was age.

Chronic cavitary pulmonary histoplasmosis in a non-HIV and immunocompromised patient without overseas travel history.

Korea is not known as an endemic area for Histoplasma. However, we experienced a case of histoplasmosis in a person who had never been abroad. A 65-year-old female was admitted to the hospital for evaluation of multiple lung nodules. A computed tomography (CT) scan of the chest showed multiple ill-defined consolidations and cavitations in all lobes of both lungs. The patient underwent a CT-guided lung biopsy, and a histopathology study showed findings compatible with histoplasmosis. Based on biopsy results and clinical findings, the patient was diagnosed with chronic cavitary pulmonary histoplasmosis. The patient recovered completely following itraconazole treatment. This is the first case report of pulmonary histoplasmosis unconnected with either HIV infection or endemicity in Korea.

Identification of ultraviolet B-sensitive genes in human peripheral blood cells.

Ultraviolet B (UVB) is a serious irritant for the skin and increases a risk for skin cancer. To identify UVB-sensitive genes in peripheral blood, 11 healthy male volunteers were exposed to 0.3 J/cm(2) of narrow-band (NB)-UVB, about half of minimal erythema dose (MED) in Japanese, and gene expression in blood was analyzed at 4 h, 24 h, 4 d and 7 d after the irradiation using microarray carrying oligonucleotide probes for 2,000 stress-responsive genes. RNA prepared before the irradiation was used as a reference control. Microarray analysis identified 21 genes as UVB-responsive genes with a peak at 24 h in 6 subjects, and real-time PCR validated the significant down-regulation of 9 (ABCB10, ATF1, ABCD3, TANK, FAS, SLC30A9, CHUK, CASP1, and ABCE1) out of the 21 genes in 11 subjects. Considering sensitive and characteristic features of 9 marker genes, they may be useful indicators for monitoring systemic response to UVB irradiation.