AMPK regulates immature boar Sertoli cell proliferation through affecting CDK4/Cyclin D3 pathway and mitochondrial function.

Sertoli cell (SC) proliferation plays an important role in sperm production and quality; however, the regulatory mechanism of SC proliferation is not well understood. This study investigated the role of adenosine monophosphate-activated protein kinase (AMPK) in the regulation of immature boar SC activity. Cell counting kit-8, Seahorse XFe96, mitochondrial respiratory enzyme-related assay kits, and transmission electron microscopy were used to detect SC proliferative viability, oxygen consumption rate (OCR), mitochondrial respiratory enzyme activity, and the ultrastructure of primary cultured SCs in vitro from the testes of 21-day-old boars. A dual luciferase reporter assay was performed to determine the miRNA-mRNA target interaction. Western blotting was used to analyze cell proliferation-related protein expression of p38, p21, proliferating cell nuclear antigen (PCNA), Cyclin-dependent kinase 4 (CDK4), Cyclin D3, and phosphorylated retinoblastoma protein (Rb). Each experiment had a completely randomized design, with three replicates in each experiment. The results showed that the AMPK inhibitor (Compound C, 20 µM-24 h) increased cell proliferation viability, ATP production, and maximal respiration of SCs by 0.64-, 0.12-, and 0.08-fold (p < 0.05), respectively; increased the SC protein expression of PCNA, CDK4, Cyclin D3, and p-Rb by 0.13-, 0.09-, 0.88-, and 0.12-fold (p < 0.05), respectively; and decreased the SC protein expression of p38 and p21 by 0.36- and 0.27-fold (p < 0.05), respectively. The AMPK agonist AICAR (2 mM-6 h) significantly inhibited SC ultrastructure, OCR, mitochondrial respiratory enzyme activity, and cell proliferation-related protein levels. AMPK was validated to be a target gene of miR-1285 based on the result in which the miR-1285 mimic inhibited the luciferase activity of wild-type AMPK by 0.54-fold (p < 0.001). MiR-1285 mimic promoted the OCR of SCs, with 0.45-, 0.15-, 0.21-, and 0.30-fold (p < 0.01) increases in ATP production, basal and maximal respiration, and spare capacity, respectively. MiR-1285 mimic increased the mitochondrial respiratory enzyme activity of SCs, with 0.63-, 0.70-, and 0.97-fold (p < 0.01) increases in NADH-Q oxidoreductase, cytochrome c oxidase, and ATP synthase, respectively. Moreover, the miR-1285 mimic increased the protein expression of PCNA, CDK4, Cyclin D3, and p-Rb by 0.24-, 0.30-, 0.22-, and 0.13-fold (p < 0.05), respectively, and reduced the protein expression of p38 and p21 by 0.58- and 0.66-fold (p < 0.001). MiR-1285 inhibitor showed opposite effects on the above indicators and induced numerous autophagosomes and large lipid droplets in SCs. A high dose of estradiol (10 µM-6 h, showed a promotion of AMPK activation in a previous study) significantly inhibited SC ultrastructure, mitochondrial function, and proliferation-related pathways, while these adverse effects were weakened by Compound C treatment or miR-1285 mimic transfection. Our findings suggest that the activation and inhibition of AMPK induced by specific drugs or synthesized targeted miRNA fragments could regulate immature boar SC proliferative activity by influencing the CDK4/Cyclin D3 pathway and mitochondrial function; this helps to provide a basis for the prevention and treatment of male sterility in clinical practice.

Glutamate rescues heat stress-induced apoptosis of Sertoli cells by enhancing the activity of antioxidant enzymes and activating the Trx1-Akt pathway in vitro.

Heat stress reduces the number of Sertoli cells, which is closely related to an imbalanced redox status. Glutamate functions to maintain the equilibrium of redox homeostasis. However, the role of glutamate in heat treated Sertoli cells remains unclear. Herein, Sertoli cells from 3-week-old piglets were treated at 44 °C for 30 min (heat stress). Glutamate levels increased significantly following heat stress treatment, followed by a gradual decrease during recovery, while glutathione (GSH) showed a gradual increase. The addition of exogenous glutamate (700 µM) to Sertoli cells before heat stress significantly reduced the heat stress-induced apoptosis rate, mediated by enhanced levels of antioxidant substances (superoxide dismutase (SOD), total antioxidant capacity (TAC), and GSH) and reduced levels of oxidative substances (reactive oxygen species (ROS) and malondialdehyde (MDA)). Glutamate addition to Sertoli cells before heat stress upregulated the levels of glutamate-cysteine ligase, modifier subunit (Gclm), glutathione synthetase (Gss), thioredoxin (Trx1) and B-cell leukemia/lymphoma 2 (Bcl-2), and the ratio of phosphorylated Akt (protein kinase B)/total Akt. However, it decreased the levels of Bcl2-associated X protein (Bax) and cleaved-caspase 3. Addition of the inhibitor of glutaminase (Gls1), Bptes (Bis-2-(5-phenylacetamido-1,3,4-thiadiazol-2-yl)ethyl sulfide, 30 µM)to Sertoli cells before heat stress reversed these effects. These results inferred that glutamate rescued heat stress-induced apoptosis in Sertoli cells by enhancing activity of antioxidant enzymes and activating the Trx1-Akt pathway. Thus, glutamate supplementation might represent a novel strategy to alleviate the negative effect of heat stress.

Vicious Cycle-Breaking Lipid Nanoparticles Remodeling Multicellular Crosstalk to Reverse Liver Fibrosis.

During liver fibrogenesis, the reciprocal crosstalk among capillarized liver sinusoidal endothelial cells (LSECs), activated hepatic stellate cells (HSCs), and dysfunctional hepatocytes constructs a self-amplifying vicious cycle, greatly exacerbating the disease condition and weakening therapeutic effect. Limited by the malignant cellular interactions, the previous single-cell centric treatment approaches show unsatisfactory efficacy and fail to meet clinical demand. Herein, a vicious cycle-breaking strategy is proposed to target and repair pathological cells separately to terminate the malignant progression of liver fibrosis. Chondroitin sulfate-modified and vismodegib-loaded nanoparticles (CS-NPs/VDG) are designed to efficiently normalize the fenestrae phenotype of LSECs and restore HSCs to quiescent state by inhibiting Hedgehog signaling pathway. In addition, glycyrrhetinic acid-modified and silybin-loaded nanoparticles (GA-NPs/SIB) are prepared to restore hepatocytes function by relieving oxidative stress. The results show successful interruption of vicious cycle as well as distinct fibrosis resolution in two animal models through multiregulation of the pathological cells. This work not only highlights the significance of modulating cellular crosstalk but also provides a promising avenue for developing antifibrotic regimens.

Meroterpenoids from Ganoderma petchii inhibiting migration of triple negative breast cancer cells.

Ganoderpetchoids A-E and (-)-dayaolingzhiol H, six undescribed meroterpenoids, were isolated from Ganoderma petchii. Their structures including the relative configurations were identified by means of spectroscopic methods and 13C NMR calculations. Chiral separation of the new racemics was performed to afford their respective enantiomers. The absolute configurations of the new isolates were clarified by computational approaches, CD comparisons and X-ray diffraction analysis. Biological studies toward triple negative breast cancer indicated that (+)-6 and (-)-6 significantly inhibit the migration of MDA-MB-231 cell line.

Isolation and Characterization of trans-p-Hydroxycinnamoyl Meroterpenoids from Ganoderma sinensis.

Zizhines V, W, Y, Z, (±)-zizhines X, and Z1-Z3, and (±)-ganosinensol L, thirteen new compounds including four pairs of enantiomers and a known compound (-)-ganosinensol L, were isolated from the fruiting bodies of Ganoderma sinensis. Their structures were identified by spectroscopic, computational methods, and CD (circular dichroism spectroscopy) comparisons. Zizhines V-Z and Z1-Z3 are meroterpenoids consisting of the phenolic and the terpenoidal parts. All the compounds except zizhine Z3 bear a common trans-p-hydroxycinnamoyl group. Biological evaluation shows that (-)-zizhine Z1 inhibits cell migration in the MDA-MB-231 cell lines. The present study discloses the chemical profiling of G. sinensis and paves the way for its development as functional products to benefit chronic disorders.

Resistance-driven innovations in the discovery of bactericides: novel triclosan derivatives decorating isopropanolamine moiety as promising anti-biofilm agents against destructive plant bacterial diseases.

BACKGROUND: Controlling bacterial infections in plants is a major challenge owing to the appearance of resistant strains. As a physical barrier, the bacterial biofilm helps bacterial infections acquire drug resistance by enabling bacteria to accommodate complex and volatile environmental conditions and avoid bactericidal effects. Thus, developing new antibacterial agents with antibiofilm potency is imperative. RESULTS: A series of simple triclosan derivatives containing isopropanolamine moiety were elaborately designed and assessed for their antibacterial behavior. Bioassay results showed that some title compounds had excellent bioactivity against three destructive bacteria Xanthomonas oryzae pv. oryzae (Xoo), Xanthomonas axonopodis pv. citri (Xac) and Pseudomonas syringae pv. actinidiae (Psa). Notably, compound C8 displayed high bioactivities toward Xoo and Xac, with EC50 values were 0.34 and 2.11 µg mL-1 , respectively. In vivo trials revealed that compound C8 exhibited excellent protective activities against rice bacterial blight and citrus bacterial canker at 200 µg mL-1 , with control effectivenesses of 49.57% and 85.60%, respectively. Compound A4 had remarkably inhibitory activity toward Psa, with an EC50 value of 2.63 µg mL-1 , and demonstrated outstanding protective activity with a value of 77.23% against Psa in vivo. Antibacterial mechanisms indicated that compound C8 dose-dependently prevented biofilm formation and extracellular polysaccharide production. C8 also significantly weakened the motility and pathogenicity of Xoo. CONCLUSION: This study contributes to the development and excavation of novel bactericidal candidates with broad-spectrum antibacterial activity by targeting bacterial biofilm to control refractory plant bacterial diseases. © 2023 Society of Chemical Industry.

Ectopic expression of a combination of 5 genes detects high risk forms of T-cell acute lymphoblastic leukemia.

BACKGROUND: T cell acute lymphoblastic leukemia (T-ALL) defines a group of hematological malignancies with heterogeneous aggressiveness and highly variable outcome, making therapeutic decisions a challenging task. We tried to discover new predictive model for T-ALL before treatment by using a specific pipeline designed to discover aberrantly active gene. RESULTS: The expression of 18 genes was significantly associated with shorter survival, including ACTRT2, GOT1L1, SPATA45, TOPAZ1 and ZPBP (5-GEC), which were used as a basis to design a prognostic classifier for T-ALL patients. The molecular characterization of the 5-GEC positive T-ALL unveiled specific characteristics inherent to the most aggressive T leukemic cells, including a drastic shut-down of genes located on the mitochondrial genome and an upregulation of histone genes, the latter characterizing high risk forms in adult patients. These cases fail to respond to the induction treatment, since 5-GEC either predicted positive minimal residual disease (MRD) or a short-term relapse in MRD negative patients. CONCLUSION: Overall, our investigations led to the discovery of a homogenous group of leukemic cells with profound alterations of their biology. It also resulted in an accurate predictive tool that could significantly improve the management of T-ALL patients.

Meroterpenoid Dimers from Ganoderma Mushrooms and Their Biological Activities Against Triple Negative Breast Cancer Cells.

(±)-Dimercochlearlactones A-J (1-10), ten pairs of novel meroterpenoid dimers and one known spirocochlealactone A (11), were isolated from Ganoderma mushrooms. The structural elucidation of new compounds, including their absolute configurations, depends on spectroscopic analysis and electronic circular dichroism (ECD) calculations. Biological studies showed that (+)- and (-)-2, (-)-3, and (+)- and (-)-11 are cytotoxic toward human triple negative breast cancer (TNBC) cells (MDA-MB-231) with IC50 values of 28.18, 25.65, 11.16, 8.18, and 13.02 µM, respectively. Wound healing assay revealed that five pairs of meroterpenoids (±)-5-(±)-8 and (±)-10 could significantly inhibit cell mobility at 20 µM in MDA-MB-231 cells. The results provide a new insight into the biological role of Ganoderma meroterpenoids in TNBC.

[Electroacupuncture improves long-term survival of myocardial infarction mice by promoting myocardial angiogenesis and inhibiting ventricular remodeling].

OBJECTIVE: To investigate the mechanism of electroacupuncture (EA) in improving the long-term survival rate of mice after myocardial infarction by promoting angiogenesis and inhibiting ventricular remodeling. METHODS: A total of 102 male C57BL/6 mice were randomly divided into sham operation, model and EA groups (n=34 /group). The myocardial infarction model was established by permanent ligation of the anterior descending branch of the left coronary artery. Beginning from the 3rd day after ligation, EA (2 Hz/20 Hz) was applied to bilateral "Neiguan" (PC6) and "Ximen" (PC4) for 30 min, once a day for 28 days. The survival rate in 140 d was recorded and the left ventricular ejection fraction (EF) calculated by using echocardiography after the treatment. The left cardiac ventricular tissue was cut into sections to be stained with Masson's trichrome, wheat germ agglutinin (WGA) or α-smooth muscle actin (α-SMA) immunohistochemistry method, followed by measuring the collagen area in the marginal region of myocardial infarction and calculating the collagen volume fraction (for assessing the severity of myocardial fibrosis), measuring the sectional area of cardiomyocytes (for assessing the degree of myocardial hypertrophy), and ob-serving the newborn blood vessels and calculating the ratio of neovascularization area (for assessing the state of angiogenesis). The expression of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor (HIF)-1α protein in the boundary area of myocardial infarction were detected by Western blot. RESULTS: After modeling, the survival rate, EF, and the thickness of the left cardiac ventricle were significantly decreased (P<0.01), whereas the percentage of collagen deposition area, sectional area of cardiomyocyte, percentage of angiogenesis area, and the expression levels of VEGF and HIF-1α proteins were significantly increased (P<0.01) in the model group relevant to the sham operation group. Compared with the model group, the survival rate, EF, the thickness of the left cardiac ventricle, the percentage of angiogenesis area, and the expression levels of VEGF and HIF-1α proteins were significantly increased (P<0.05, P<0.01), while the percentage of collagen deposition area and the sectional area of the cardiomyocyte were considerably decreased in the EA group (P<0.01, P<0.05). CONCLUSION: EA of PC6 and PC4 can significantly improve the cardiac function and long-term survival rate in mice with myocardial infarction, which may be related to its functions in up-regulating the expression of HIF-1α and VEGF to promote angiogenesis and in inhibiting ventricular remodeling.

Optimized outcome prediction of oncogenetic mutations in non-early T-cell precursor acute lymphoblastic leukemia.

BACKGROUND: Early biomarkers allowing effective treatment stratification in adult T-cell acute lymphoblastic leukemia (T-ALL) patients remain elusive. MATERIALS AND METHODS: The mutation spectrum of 116T-ALL adult patients enrolled in the Shanghai Institute of Hematology (SIH)-based hospital network or Multicenter Hematology-Oncology Protocols Evaluation System (M-HOPES) in China were studied by using RNA-sequencing or targeted next generation sequencing. A comprehensive survival analysis based on clinical characteristics, immunophenotype and oncogenetic classifier was performed. RESULTS: Early T-cell precursor acute lymphoblastic leukemia (ETP-ALL) has higher mutation rates of N/K-RAS and lower mutation rates of FBXW7 compared to non-ETP ALL, but the survival probability of ETP-ALL patients is similar to that of non-ETP ALL patients. T-ALLs with a NOTCH1/FBXW7 (N/F) mutation in the absence of RAS or PTEN abnormalities (NFRP class I) show a more favorable outcome compared to T-ALLs with no N/F mutation and/or with the presence of RAS/PTEN alterations (NFRP class II). A survival analysis of T-ALL, taking into account both the ETP-ALL/non-ETP T-ALL groups and the NFRP oncogenetic classifier, demonstrates that, within the non-ETP T-ALL subtype, NFRP class II identifies a group with poor prognosis and significant decreases of both OS (14.8% versus 50.9%, P = 0.019) and EFS (11.4% versus 42.4%, P = 0.001). In contrast, no survival difference is observed within ETP-ALL between the NFRP class I or class II (OS: 37.9% versus 33%, P = 0.876; EFS: 39.8% versus 33.7%, P = 0.969). CONCLUSION: In summary, the oncogenetic classifier based on the NFRP classes is particularly useful to improve the stratification of non-ETP ALL.

17ß-estradiol rescues the damage of thiazolidinedione on chicken Sertoli cell proliferation via adiponectin.

Thiazolidinedione (TZD) is an oral anti-diabetic drug that exhibits some side effects on the male reproductive system by interfering with the steroidogenesis and androgenic activity and also shows anti-proliferative effect on several cell types. This study investigated the effect of TZD on immature chicken Sertoli cell (SC) proliferation and the potential mechanism by which 17ß-estradiol regulated this process. Chicken SC viability was investigated under different treatment concentration and time of TZD. 17ß-estradiol (0.001 µM, 24 h) was added to analyze its effects on TZD-mediated cell viability, cell metabolic activity, cell growth, cell cycle progression, reactive oxygen species (ROS) level, antioxidant enzyme activity, mitochondria activity, oxygen consumption rate, adenosine triphosphate (ATP) level, and mitochondrial respiratory chain enzyme activity, adiponectin expression and several cell proliferation-related genes mRNA and protein levels. We performed the microRNA (miRNA) array to find TZD-induced differentially expressed miRNAs and validated whether miR-1577 can target on adiponectin via the dual luciferase reporter assay, as well as verified the effect of adiponectin addition with different concentrations on the SC viability. Further, SCs were transfected with miR-1577 agomir (a double-stranded synthetic miRNA mimic) in the presence or absence of TZD and antagomir (a single-stranded synthetic miRNA inhibitor) in the presence or absence of 17ß-estradiol to analyze whether miR-1577 was involved in TZD-mediated SC proliferation and whether 17ß-estradiol regulated this process. Results showed that TZD significantly inhibited SC viability, cell metabolic activity, cell growth, and cell cycle progression, while increased adiponectin level and ROS generation. TZD-treated SCs presented decreases of antioxidant enzyme activity, mitochondria activity, basal and maximal respiration, ATP production and level, mitochondrial respiratory chain enzyme activity, and mRNA and protein expressions of several cell proliferation-related genes, as well as the significant alteration of miRNA expressions (a total number of 55 miRNAs were up-regulated whereas 53 miRNAs down-regulated). Whereas, 17ß-estradiol played a positive role in chicken SC proliferation and rescued the damage of TZD on SC proliferation by up-regulating miR-1577 expression whose target gene was validated to be the adiponectin. In addition, exogenous adiponectin (more than 1 µg/ml) treatment exhibited a significant inhibition on the SC viability. Transfection of miR-1577 agomir promoted the SC proliferation via down-expressed adiponectin, and increased the mitochondrial function and cell proliferation-related gene expression, while TZD weakened the positive effect of miR-1577 agomir on SCs. On the other hand, transfection of miR-1577 antagomir inhibited SC proliferation by producing the opposite effects on above parameters, while 17ß-estradiol attenuated the negative effect of miR-1577 antagomir on SCs. These findings suggest down-expressed miR-1577 is involved in the regulation of TZD-inhibited SC proliferation through increasing adiponectin level, and this damage of TZD on the immature chicken SC proliferation can be ameliorated by appropriate dose of exogenous 17ß-estradiol treatment. This study provides an insight into the cytoprotective effect of 17ß-estradiol on TZD-damaged SC proliferation and may suggest a potential strategy for reducing the risk of SC dysfunction caused by the abuse of TZD.

Meroterpenoids From Ganoderma lucidum Mushrooms and Their Biological Roles in Insulin Resistance and Triple-Negative Breast Cancer.

Ganoderma fungi as popular raw materials of numerous functional foods have been extensively investigated. In this study, five pairs of meroterpenoid enantiomers beyond well-known triterpenoids and polysaccharides, dayaolingzhiols I-M (1-5), were characterized from Ganoderma lucidum. Their structures were identified using spectroscopic and computational methods. Structurally, compound 1 features a novel dioxabicyclo[2.2.2]octan-3-one motif in the side chain. Ethnoknowledge-derived biological evaluation found that (+)-5 could activate Akt and AMPK phosphorylation in insulin-stimulated C2C12 cells, and (+)-5 could activate glucose uptake dose dependently in C2C12 cells. Furthermore, we found that (+)-1 (+)-4, and (-)-4 could significantly inhibit cell migration of the MDA-MB-231 cell line, of which (+)-4 showed significant inhibitory effects against cell migration of the MDA-MB-231 cell line in a dose-dependent manner. These findings revealed the meroterpenoidal composition of G. lucidum and its roles in the prevention of chronic diseases such as diabetes mellitus and triple-negative breast cancer.

Estradiol ameliorates metformin-inhibited Sertoli cell proliferation via AMPK/TSC2/mTOR signaling pathway.

Metformin is a commonly used for treating type 2 diabetes and it acts on a variety of organs including the male reproductive system. 17ß-estradiol plays an important role in Sertoli cell (SC) proliferation which determines the germ cell development and spermatogenesis. The aim of this study is to investigate the effect of metformin on immature chicken SC proliferation and the potential mechanisms by which 17ß-estradiol regulate this process. Results showed that metformin significantly inhibited SC proliferation, whereas 17ß-estradiol weakened the inhibitory effects of metformin on SC viability, cell growth, and cell cycle progression. SC proliferation-inhibiting effect of metformin exposure was regulated by decreasing adenosine triphosphate level and respiratory enzyme activity in the mitochondria; this process was possibly mediated by the adenosine monophosphate-activated protein kinase (AMPK)/tuberous sclerosis complex 2 (TSC2)/mammalian target of rapamycin (mTOR) signaling pathway, which was regulated by the down-expressed miR-1764 and by the decreased antioxidant enzyme activity and excessive reactive oxygen species generation. In addition, SCs transfected with the miR-1764 agomir led to an improvement of proliferation capacity through down-regulating AMPKα2 level, which further decreased TSC2 expression and induced mTOR activation. However, the anti-proliferative effect of miR-1764 antagomir can be alleviated by 17ß-estradiol treatment via the up-expression of miR-1764 in transfected SCs. Our findings suggest appropriate dose of exogenous 17ß-estradiol treatment can ameliorate the inhibitory effect of metformin on SC proliferation via the regulation of AMPK/TSC2/mTOR signaling pathway, this might reduce the risk of poor male fertility caused by the abuse of anti-diabetic agents.

Identification and function prediction of novel microRNAs in adenosine monophosphate activated protein kinase-activated Sertoli cells of immature boar.

This study was carried out with the objective to identify function prediction of novel microRNAs (miRNAs) in immature boar Sertoli cells (SCs) treated with 5-aminoimidazole-4-carboxamide-1-ß-D-ribofuranoside (AICAR), which is an agonist of adenosine monophosphate-activated protein kinase (AMPK) for regulating cellular energy homeostasis. Two small RNA libraries (control and AICAR treatment) prepared from immature boar SCs were constructed and sequenced by the Illumina small RNA deep sequencing. We identified 77 novel miRNAs and predicted 177 potential target genes for 26 differential novel miRNAs (four miRNAs up-regulation and 22 miRNAs down-regulation) in AICAR-treated SCs. Gene Ontology enrichment and Kyoto Encyclopedia of Genes and Genomes pathway suggested that target genes of differential novel miRNAs were implicated in many biological processes and metabolic pathways. Our findings provided useful information for the functional regulation of novel miRNAs and target mRNAs on AMPK-activated immature boar SCs.

Clinical outcomes and 5-year follow-up results of keratosis pilaris treated by a high concentration of glycolic acid.

BACKGROUND: Keratosis pilaris is a hereditary abnormal keratosis of the hair follicle orifice. Gray-brown keratotic plugs in the pores and dark red keratotic papules at the openings of hair follicles can be seen, which contain coiled hair and are often accompanied by perifollicular erythema and pigmentation. Glycolic acid can correct the abnormalities of hair follicular duct keratosis and eliminate excessive accumulation of keratinocytes. It also promotes skin metabolism and accelerates the melanin metabolism. The therapeutic effect is related to the glycolic acid concentration. AIM: To evaluate the efficacy and safety of a high concentration of glycolic acid in the treatment of keratosis pilaris, and to observe the outcomes at 5-year of follow-up. METHODS: Twenty-five participants were recruited and areas with typical keratosis pilaris were selected as testing sites. High concentrations of glycolic acid (50% or 70%) were applied to a circular area (d = 8 cm, S = 50 cm2) and repeated four times, on days 0, 20, 40 and 60. Before each treatment and 20 d after the last treatment, on days 0, 20, 40, 60, and 80 and at a 5-year follow-up, The number of follicular keratotic papules were counted and the extent of perifollicular erythema and pigmentation was determined. At the same time, the participants provided subjective evaluations of treatment efficacy and safety. RESULTS: Treatment effectiveness was indicated by the percentage of keratotic papules in the test site, on days 20, 40, 60 and 80, which were 8%, 12%, 36%, and 60%, respectively. Compared with day 0, each difference was significant (P < 0.05). Compared with day 0, differences in melanin content (M) in the skin and skin lightness (L) on days 40, 60 and 80, the were statistically significant (P < 0.05); skin hemoglobin content (E) on days 60 and 80 was statistically different as compared with before treatment (P < 0.05). There were no significant differences in the number of keratotic papules, M, L, and E in 9 participants at the 5-year follow-up compared with before treatment (P > 0.05%). CONCLUSION: A high concentration of glycolic acid significantly improved skin roughness as well as follicular hyperpigmentation of patients with keratosis pilaris. The treatment was relatively safe, but there was no significant difference at the 5-year follow-up compared to before treatment.

Lucidumones B-H, racemic meroterpenoids that inhibit tumor cell migration from Ganoderma lucidum.

Seven new meroterpenoids, lucidumones B-H (1 and 3-8), along with one known meroterpenoid (2), were isolated from the fruiting bodies of Ganoderma lucidum. The structures of the new compounds were assigned by spectroscopic and computational methods. All the isolated compounds were tested for their inhibition on human cancer cell migration. It was found that compounds (-)-1, (+)-2, (-)-4, (+)-6, and (+)-8 could significantly inhibit cell migration in KYSE30 cell line. Further examination disclosed that cell migration inhibition of (+)-6 and (+)-8 might be related with downregulation of N-cadherin.

Meroterpenoids and alkaloids from Ganoderma australe.

Two new mertoterpenoids, australins A (1) and B (2), and a new alkaloid, australine (4), together with five known compounds (3, 5-8) were isolated from the fruiting bodies of Ganoderma australe. Their structures including absolute configurations were assigned by using spectroscopic methods and electronic circular dichroism (ECD) calculations. Racemic australin A was further purified by chiral HPLC. Biological assessments reveal that compounds (+)-1 and 7 could significantly protect SH-SY5Y cells from glutamate-induced neural excitotoxicity.

MiR-8-3p regulates hyperthermia-induced lactate secretion by targeting PPP2R5B in boar Sertoli cells.

Lactate produced by glycolysis in Sertoli cells (SCs) is the main energy substrate for developing germ cells and plays a vital role in spermatogenesis. MicroRNAs (miRNAs) function as posttranscriptional regulators of gene expression in biological processes. We have previously shown that hyperthermia (43°C, 30 min) promotes lactate secretion by inhibiting phosphorylation of adenosine monophosphate-activated protein kinase (AMPK) in cultured immature boar SCs. However, it is unclear whether miRNAs are involved in AMPK-modulated glycolysis in SCs. In the present study, we identified 349 miRNAs (227 upregulated and 122 downregulated) in hyperthermia-treated boar SCs by next-generation high-throughput RNA sequencing. MiR-8-3p, which was found to be a novel upregulated miRNA in hyperthermia-treated SCs, suppressed the expression of AMPK upstream genes (protein phosphatase 2 subunit B, PPP2R5B), and further downregulated the expression of p-AMPK. The miR-8-3p mimic upregulated expression of glucose transporter 3, lactate dehydrogenase A and monocarboxylate transporter 1, and increased lactic acid dehydrogenase activity, lactate secretion, and ATP depletion in SCs; the miR-8-3p inhibitor had the opposite effects on these parameters. Our findings indicate that miR-8-3p acts as a novel regulator of AMPK-modulated lactate secretion by targeting PPP2R5B in hyperthermic boar SCs.

Optimization of Ultrasound-Assisted Extraction of Antioxidant Polyphenols from the Seed Coats of Red Sword Bean (Canavalia gladiate (Jacq.) DC.).

The seed coat of red sword bean (Canavalia gladiata (Jacq.) DC.) is rich in antioxidant polyphenols. It is often discarded as a byproduct with the consumption of red sword bean, since it is very thick and not consumed by people. The aim of this study was to develop an ultrasound-assisted extraction method to extract natural antioxidants from the seed coats. The extraction process was optimized by using response surface methodology. After the single-factor experiments, three key factors, including ethanol concentration, liquid/solid ratio, and extraction time, were selected and their interactions were studied using a central composite design. The optimal extraction condition was 60.2% hydroethanol, a liquid/solid ratio of 29.3 mL/g, an extraction time of 18.4 min, an extraction temperature of 50 °C, and ultrasound power of 400 W. Under the optimal conditions, antioxidant activity of the extract was 755.98 ± 10.23 µmol Trolox/g dry weight (DW), much higher than that from maceration (558.77 ± 14.42 µmol Trolox/g DW) or Soxhlet extraction (479.81 ± 12.75 µmol Trolox/g DW). In addition, the main antioxidant compounds in the extract were identified and quantified by high-performance liquid chromatography-diode array detection-tandem mass spectrometry (HPLC-DAD-MS/MS). The concentrations of digalloyl hexoside, methyl gallate, gallic acid, trigalloyl hexoside, and digallic acid were 15.30 ± 0.98, 8.85 ± 0.51, 8.76 ± 0.36, 4.27 ± 0.21, and 2.89 ± 0.13 mg/g DW. This study provides an efficient and green extraction method for the extraction of natural antioxidants from the bean coat of red sword bean. The extract of antioxidants might be added into functional foods or nutraceuticals with potential beneficial functions.

Demethylation and microRNA differential expression regulate plasma-induced improvement of chicken sperm quality.

The sperm quality is a vital economical requisite of poultry production. Our previous study found non-thermal dielectric barrier discharge plasma exposure on fertilized eggs could increase the chicken growth and the male reproduction. However, it is unclear how plasma treatment regulates the reproductive capacity in male chickens. In this study, we used the optimal plasma treatment condition (2.81 W for 2 min) which has been applied on 3.5-day-incubated fertilized eggs in the previous work and investigated the reproductive performance in male chickens aged at 20 and 40 weeks. The results showed that plasma exposure increased sperm count, motility, fertility rate, and fertilization period of male chickens. The sperm quality-promoting effect of plasma treatment was regulated by the significant improvements of adenosine triphosphate production and testosterone level, and by the modulation of reactive oxygen species balance and adenosine monophosphate-activated protein kinase and mammalian target of rapamycin pathway in the spermatozoa. Additionally, the plasma effect suggested that DNA demethylation and microRNA differential expression (a total number of 39 microRNAs were up-regulated whereas 53 microRNAs down-regulated in the testis) regulated the increases of adenosine triphosphate synthesis and testosterone level for promoting the chicken sperm quality. This finding might be beneficial to elevate the fertilization rate and embryo quality for the next generation in poultry breeding.