3,3'-Diindolylmethane (DIM) is a natural component of cruciferous plants. It has strong antioxidant and anti-angiogenic effects and promotes the apoptosis of a variety of tumor cells. However, little is known about the critical role of DIM on cardiac hypertrophy. In the present study, we investigated the effects of DIM on cardiac hypertrophy.

Testosterone is overwhelmingly important in regulating erectile physiology. However, the associated molecular mechanisms are poorly understood. The purpose of this study was to explore the effects and mechanisms of testosterone in erectile dysfunction (ED) in castrated rats. Forty male Sprague-Dawley rats were randomized to four groups (control, sham-operated, castration and castration-with-testosterone-replacement). Reactive oxygen species (ROS) production was measured by dihydroethidium (DHE) staining. Erectile function was assessed by the recording of intracavernous pressure (ICP) and mean arterial blood pressure (MAP). Protein expression levels were examined by western blotting. We found that castration reduced erectile function and that testosterone restored it. Nitric oxide synthase (NOS) activity was decrease in the castrated rats, and testosterone administration attenuated this decrease (each p < 0.05). The testosterone, dihydrotestosterone, cyclic guanosine monophosphate (cGMP) and cyclic adenosine monophosphate (cAMP) concentrations were lower in the castrated rats, and testosterone restored these levels (each p < 0.05). Furthermore, the cyclooxygenase-2 (COX-2) and prostacyclin synthase (PTGIS) expression levels and phospho-endothelial nitric oxide synthase (p-eNOS, Ser1177)/endothelial nitric oxide synthase (eNOS) ratio were reduced in the castrated rats compared with the controls (each p < 0.05). In addition, the p40(phox) and p67(phox) expression levels were increased in the castrated rats, and testosterone reversed these changes (each p < 0.05). Overall, our results demonstrate that testosterone ameliorates ED after castration by reducing ROS production and increasing the activity of the eNOS/cGMP and COX-2/PTGIS/cAMP signaling pathways.

Cryopreservation induces sperm cryoinjuries, including physiological and functional changes. However, the molecular mechanisms of sperm cryoinjury and cryoresistance are still unknown. Cryoresistance or the freeze tolerance of sperm varies across species, and boar sperm is more susceptible to cold stress. Contrary to boar sperm, giant panda sperm appears to be strongly freeze-tolerant and is capable of surviving repeated cycles of freeze-thawing. In this study, differentially expressed (DE) PIWI-interacting RNAs (piRNAs) of fresh and frozen-thawed sperm with different freeze tolerance capacity from giant panda and boar were evaluated. The results showed that 1,160 (22 downregulated and 1,138 upregulated) and 384 (110 upregulated and 274 downregulated) DE piRNAs were identified in giant panda and boar sperm, respectively. Gene ontology (GO) enrichment analysis revealed that the target DE messenger RNAs (mRNAs) of DE piRNAs were mainly enriched in biological regulation, cellular, and metabolic processes in giant panda and boar sperm. Moreover, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that the target DE mRNAs of DE piRNAs were only distributed in DNA replication and the cyclic adenosine monophosphate (cAMP) signaling pathway in giant panda, but the cAMP, cyclic guanosine monophosphate (cGMP), and mitogen-activated protein kinase (MAPK) signaling pathways in boar sperm were considered as part of the olfactory transduction pathway. In conclusion, we speculated that the difference in the piRNA profiles and the DE piRNAs involved in the cAMP signaling pathway in boar and giant panda may have contributed to the different freeze tolerance capacities between giant panda and boar sperm, which helps to elucidate the molecular mechanism behind sperm cryoinjury and cryoresistance.

The stimulator of interferon genes (STING) protein is an important and promising innate immune target for tumor therapy. However, the instability of the agonists of STING and their tendency to cause systemic immune activation is a hurdle. The STING activator, cyclic di-adenosine monophosphate (CDA), produced by the modified Escherichia coli Nissle 1917, shows high antitumor activity and effectively reduces the systemic effects of the "off-target" caused by the activation of the STING pathway. In this study, we used synthetic biological approaches to optimize the translation levels of the diadenylate cyclase that catalyzes CDA synthesis in vitro. We developed 2 engineered strains, CIBT4523 and CIBT4712, for producing high levels of CDA while keeping their concentrations within a range that did not compromise the growth. Although CIBT4712 exhibited stronger induction of the STING pathway corresponding to in vitro CDA levels, it had lower antitumor activity than CIBT4523 in an allograft tumor model, which might be related to the stability of the surviving bacteria in the tumor tissue. CIBT4523 exhibited complete tumor regression, prolonged survival of mice, and rejection of rechallenged tumors, thus, offering new possibilities for more effective tumor therapy. We showed that the appropriate production of CDA in engineered bacterial strains is essential for balancing antitumor efficacy and self-toxicity.

Corydalis yanhusuo W. T. Wang (C. yanhusuo) has been traditionally used for drug addiction and pain relief in China. In our previous study, we showed that the extract of C. yanhusuo blocks dopamine receptors, demonstrating that its pharmacological activities are mostly due to the antagonistic effects of some of its components at dopamine receptors. As part of our ongoing project on C. yanhusuo, the aim of the present study is to establish a high-throughput and low-cost screening assay system and test the abilities of the isolated alkaloids from C. yanhusuo to inhibit dopamine-induced dopamine D₁ receptor activity. By using our established cyclic adenosine monophosphate (cAMP)-response element (CRE)-luciferase reporter gene assay system, we identified eight alkaloids from C. yanhusuo with D₁ receptor antagonistic activities. We next validated the activities of these compounds using fluorometric imaging plate reader (FLIPR) assay by measuring the intracellular Ca2+ change. Six out of eight compounds, including tetrahydropalmatine, corydaline, 13-methyldehydrocorydalmine, dehydrocorybubine, dehydrocorydaline, and columbamine, can be confirmed for their inhibitory activities. The dopamine-receptor-antagonistic effects of four compounds, including 13-methyldehydrocorydalmine, dehydrocorydaline, columbamine, and corydaline, are reported for the first time. The present study provides an important pharmacological basis to support the traditional use of C. yanhusuo in China.

Fungal growth is closely related to virulence. Finding the key genes and pathways that regulate growth can help elucidate the regulatory mechanisms of fungal growth and virulence in efforts to locate new drug targets. Fusarium oxysporum is an important plant pathogen and human opportunistic pathogen that has research value in agricultural and medicinal fields. A mutant of F. oxysporum with reduced growth was obtained by Agrobacterium tumefaciens-mediated transformation, the transferred DNA (T-DNA) interrupted gene in this mutant coded a hypothetical protein that we named FoDbp40. FoDbp40 has an unknown function, but we chose to explore its possible functions as it may play a role in fungal growth regulatory mechanisms. Results showed that F. oxysporum growth and virulence decreased after FoDbp40 deletion. FOXG_05529 (NCBI Gene ID, isocitrate lyase, ICL) was identified as a key gene that involved in the reduced growth of this mutant. Deletion of FoDbp40 results in a decrease of more than 80% in ICL expression and activity, succinate level, and energy level, plus a decrease in phosphorylated mammalian target of rapamycin level and an increase in phosphorylated 5'-adenosine monophosphate activated protein kinase level. In summary, our study found that the FoDbp40 regulates the expression of ICL at a transcriptional level and affects energy levels and downstream related pathways, thereby regulating the growth and virulence of F. oxysporum.

Activation of adenosine monophosphate-activated protein kinase (AMPK) is able to produce significant anti-non-small cell lung cancer (NSCLC) cell activity. ASP4132 is an orally active and highly effective AMPK activator. The current study tested its activity against NSCLC cells. In primary NSCLC cells and established cell lines (A549 and NCI-H1944) ASP4132 potently inhibited cell growth, proliferation and cell cycle progression as well as cell migration and invasion. Robust apoptosis activation was detected in ASP4132-treated NSCLC cells. Furthermore, ASP4132 treatment in NSCLC cells induced programmed necrosis, causing mitochondrial p53-cyclophilin D (CyPD)-adenine nucleotide translocase 1 (ANT1) association, mitochondrial depolarization and medium lactate dehydrogenase release. In NSCLC cells ASP4132 activated AMPK signaling, induced AMPKα1-ACC phosphorylation and increased AMPK activity. Furthermore, AMPK downstream events, including mTORC1 inhibition, receptor tyrosine kinases (PDGFRα and EGFR) degradation, Akt inhibition and autophagy induction, were detected in ASP4132-treated NSCLC cells. Importantly, AMPK inactivation by AMPKα1 shRNA, knockout (using CRISPR/Cas9 strategy) or dominant negative mutation (T172A) almost reversed ASP4132-induced anti-NSCLC cell activity. Conversely, a constitutively active AMPKα1 (T172D) mimicked and abolished ASP4132-induced actions in NSCLC cells. In vivo, oral administration of a single dose of ASP4132 largely inhibited NSCLC xenograft growth in SCID mice. AMPK activation, mTORC1 inhibition and EGFR-PDGFRα degradation as well as Akt inhibition and autophagy induction were detected in ASP4132-treated NSCLC xenograft tumor tissues. Together, activation of AMPK by ASP4132 potently inhibits NSCLC cell growth in vitro and in vivo.

The effects of maternal conjugated linoleic acid (CLA) on embryonic development and hepatic lipid metabolism were investigated in chick embryos. A total of 180 Arbor Acres female broiler breeders (36 wk old) were randomly divided into the following 3 dietary treatment groups: a basic diet (control), a basic diet containing 0.5% CLA (CLA1), and a basic diet containing 1.0% CLA (CLA2). The females were fed for 8 wk, and the eggs from each group were collected and hatched during the last 2 wk. The results showed that the addition of dietary CLA increased the broken egg rate and reduced the fertilization rate and the egg hatchability (P < 0.05). CLA enrichment decreased the polyunsaturated and monounsaturated fatty acids and increased the saturated fatty acids in the yolk sac (P < 0.05). The yolk sac weight, body weight, and body length had a linear decrease with CLA supplementation (P < 0.05). In the developing chick embryo (at E14) and newly hatched chick (D0), the serum triglyceride concentration decreased with maternal CLA supplementation and was accompanied by a reduction in subcutaneous adipose tissue deposition. In addition, maternal CLA supplementation mediated the hepatic lipid metabolism by decreasing the mRNA expression of sterol regulatory element-binding proteins-1c (SREBP-1c), fatty acid synthase and acetyl-CoA carboxylase, and increasing the mRNA expression of adenosine 5'-monophosphate-activated protein kinase α (AMPKα), peroxisome proliferator-activated receptors α (PPARα), liver fatty acid-binding protein, adipose triglyceride lipase and carnitine palmitoyltransferase in embryonic chick livers (P < 0.05). A drop in SREBP-1c protein expression and an increase in the protein expression of p-AMPKα and PPARα were also observed in the liver of chick embryo (P < 0.05). In conclusion, maternal CLA supplementation regulated the fatty acid composition in the yolk sac, and mediated embryonic chick development and hepatic lipometabolism, and these effects may be related to the AMPK pathway. These findings suggest the potential ability of maternal CLA supplementation to reduce fat deposition in chick embryos.