Antibacterial activity of t-cinnamaldehyde: An approach to its mechanistic principle towards enterohemorrhagic Escherichia coli (EHEC).

BACKGROUND: Compounds of natural origin are potent source of drugs with unique mechanisms of action. Among phytochemicals, trans-cinnamaldehyde (t-CA) exhibits a wide range of biological activity, thus has been used for centuries to fight bacterial and fungal infections. However, the molecular basis of these properties has not been fully covered. Considering that difficult-to-control infections are becoming a rising global problem, there is a need to elucidate the molecular potential of t-CA. PURPOSE: To evaluate the antibacterial activity of t-CA against Shiga-toxigenic E. coli strains and elucidate its mechanism of action based on the inhibition of the virulence factor expression. METHODS: The antimicrobial potential of t-CA was assessed with two-fold microdilution and time-kill assays. Further evaluation included bioluminescence suppression assays, quantification of reactive oxygen species (ROS) and assessment of NAD+/NADH ratios. Morphological changes post t-CA exposure were examined using transmission electron microscopy. RNA sequencing and radiolabeling of nucleotides elucidated the metabolic alterations induced by t-CA. Toxin expression level was monitored through the application of fusion proteins, monitoring of bacteriophage development, and fluorescence microscopy studies. Lastly, the therapeutic efficacy in vivo was assessed using Galleria mellonella infection model. RESULTS: A comprehensive study of t-CA's bioactivity showed unique properties affecting bacterial metabolism and morphology, resulting in significant bacterial cell deformation and effective virulence inhibition. Elucidation of the underlying mechanisms indicated that t-CA activates the global regulatory system, the stringent response, manifested by its alarmone, (p)ppGpp, overproduction mediated by the RelA enzyme, thereby inhibiting bacterial proliferation. Intriguingly, t-CA effectively downregulates Shiga toxin gene expression via alarmone molecules, indicating its potential for therapeutic effect. In vivo validation demonstrated a significant improvement in larval survival rates post- t-CA treatment with 50 mg/kg (p < 0.05), akin to the efficacy observed with azithromycin, thus indicating its effectiveness against EHEC infections (p < 0.05). CONCLUSIONS: Collectively, these results reveal the robust antibacterial capabilities of t-CA, warranting its further exploration as a viable anti-infective agent.

[Self-made metal freezing plate for cryopreservation of human epididymal sperm: An experimental study].

OBJECTIVE: To study the effect of a new human sperm freezing method on the sperm recovery rate and search for an optimal method for cryopreservation of human epididymal sperm. METHODS: We collected semen samples from 76 men with obstructive azoospermia by percutaneous epididymal sperm aspiration and divided each sample into two parts to be cryopreserved with a self-made metal freezing plate (the experimental group) or by slow freezing (the control group), respectively. We measured the percentage of progressively motile sperm (PMS) with the computer-assisted semen analysis system and compared the membrane function, DNA fragmentation index (DFI), acrosin activity and morphological abnormality of the sperm between the two groups before and after cryopreservation. RESULTS: After thawing, both the percentages of PMS and hypotonically swollen sperm were significantly higher in the experimental than in the control group (ï¼»12.0 ± 7.5ï¼½% vs ï¼»8.0 ± 5.1ï¼½%, P < 0.05; ï¼»22.0 ± 17.5ï¼½% vs ï¼»18.0 ± 20.5ï¼½%, P < 0.05), though both decreased in comparison with the pre-freezing parameters (ï¼»20.7 ± 8.8ï¼½% and ï¼»30.0 ± 13.5ï¼½%) (P < 0.05). The sperm acrosin activity was remarkably higher in the experimental than in the control group after thawing (ï¼»75.2 ± 9.5ï¼½ vs ï¼»55.7 ± 8.3ï¼½ µIU/106sperm, P < 0.05), though decreased as compared with the baseline (ï¼»120.0 ± 10.5ï¼½ µIU/106 sperm, P < 0.05). No statistically significant differences were observed between the experimental and the control groups after thawing in the percentage of morphologically abnormal sperm (ï¼»98.7 ± 8.8ï¼½% vs ï¼»98.5±9.2ï¼½%, P > 0.05) or sperm DFI ï¼»38.2 ± 8.5ï¼½% vs ï¼»39.5 ± 10.2ï¼½%, P > 0.05), though both markedly elevated in comparison with the pre-freezing parameters (ï¼»97.2 ± 9.5ï¼½% and ï¼»30.8 ± 9.7ï¼½%) (P < 0.05). The post-thaw recovery rate of sperm was significantly higher in the experimental than in the control group (ï¼»65.2 ± 12.0ï¼½% vs ï¼»52.3 ± 18.0ï¼½%, P < 0.05). CONCLUSIONS: The self-made metal freezing plate, with its advantages of low cost, high efficiency, and easy operation, can be used as an effective method for cryopreservation of human sperm to achieve a high post-thaw sperm recovery rate, progressive sperm motility, and sperm acrosin activity.

Mangiferin induces apoptosis by regulating Bcl-2 and Bax expression in the CNE2 nasopharyngeal carcinoma cell line.

To investigate the anti-proliferative mechanism of mangiferin in a human nasopharyngeal carcinoma cell line, CNE2 cells were incubated with different concentrations of mangiferin (12.5, 25, 50, 100, 150 and 200 µM) or with PBS as a control for 72 hours. Analyses were made of the cell cycle and apoptosis with measurement of mRNA and protein levels of two apoptosis-related genes, Bcl-2 and Bax. Flow cytometry assays showed mangiferin could inhibit CNE2 cell proliferation via G2/M arrest and induction of early apoptosis. Real time PCR and Western blotting showed the mRNA and protein level of Bcl-2 to be down-regulated, while those of Bax were up-regulated, when CNE2 cells were treated with mangiferin. This investigation indicated anti-proliferation effects of mangiferin through induction of cell apoptosis regulated by Bcl-2 and Bax expression.

Gentiana manshurica Kitagawa prevents acetaminophen-induced acute hepatic injury in mice via inhibiting JNK/ERK MAPK pathway.

AIM: To investigate the in vivo hepatoprotective effects and mechanisms of Gentiana manshurica Kitagawa (GM) in acetaminophen (APAP)-induced liver injury in mice. METHODS: GM (200, 150 or 50 mg/kg body weight) or N-acetyl-L-cysteine (NAC; 300 mg/kg body weight) was administrated orally with a single dose 2 h prior to APAP (300 mg/kg body weight) injection in mice. RESULTS: APAP treatment significantly depleted hepatic glutathione (GSH), increased serum aspartate aminotransferase (AST), alanine aminotransferase (ALT) and malonyldialdehyde (MDA) and 4-hydroxynonenal levels, and decreased hepatic activity of glutathione peroxidase (GSH-px) and superoxide dismutase (SOD). However, the pretreatment of GM significantly alleviated APAP-induced oxidative stress by increasing GSH content, decreasing serum ALT, AST and MDA, and retaining the activity of GSH-px and SOD in the liver. Furthermore, GM pretreatment can inhibit caspase-3 activation and phosphorylation of c-Jun-NH2-terminal protein kinase 2 (JNK1/2) and extracellular signal-regulated kinase (ERK). GM also remarkably attenuated hepatocyte apoptosis confirmed by the terminal deoxynucleotidyl transferase mediated dUTP nick end-labeling method. CONCLUSION: Hepatoprotective effects of GM against APAP-induced acute toxicity are mediated either by preventing the decline of hepatic antioxidant status or its direct anti-apoptosis capacity. These results support that GM is a potent hepatoprotective agent.