Ayanin, a natural flavonoid inhibitor of Caseinolytic protease, is a promising therapeutic agent to combat methicillin-resistant Staphylococcus aureus infections.

Antimicrobial resistance (AMR) is a global health threat. The dramatic increase of Methicillin-resistant Staphylococcus aureus (MRSA) infections emphasizes the need to find new anti-infective agents with a novel mode of action. The Caseinolytic protease (ClpP) is a central virulence factor in stress survival, virulence, and antibiotic resistance of MRSA. Here, we found ayanin, a flavonoid isolated from Callicarpa nudiflora, was an inhibitor of MRSA ClpP with an IC50 of 19.63 µM. Using quantitative real-time PCR, ayanin reduced the virulence of Staphylococcus aureus (S. aureus) by down-regulating the level of some important virulence factors, including agrA, RNAⅢ, hla, pvl, psmα and spa. The results of cellular thermal shift assay and thermal shift assay revealed a binding between ayanin and ClpP. Molecular docking showed that ASP-168, ASN-173 and ARG-171 were the potential binding sites for ClpP binding to ayanin. ClpP mutagenesis study further indicated that ARG-171 and ASN-173 were the main active sites of ClpP. The affinity constant (KD) value of ayanin with ClpP was 3.15 × 10-5 M measured by surface plasmon resonance. In addition, ayanin exhibited a significant therapeutic effect on pneumonia infection induced by S. aureus in mice in vivo, especially in combination with vancomycin. This is the first report of ayanin with in vivo and in vitro efficacy against S. aureus infection. In conclusion, ayanin is a promising therapeutic agent to combat MRSA infections by targeting ClpP.

Nepetin reduces virulence factors expression by targeting ClpP against MRSA-induced pneumonia infection.

The resistance of Staphylococcus aureus (S. aureus) to various antibiotics has increased dramatically due to the misuse of antibiotics, and thus the development of new anti-infective drugs with new targets is urgently needed to combat resistance. Caseinolytic peptidase P is a case in hydrolase that regulates the virulence level of S. aureus. Here, we found that nepetin, a small-molecule compound from traditional Chinese herbal flavonoids, effectively inhibits ClpP activity. Nepetin suppressed the virulence of S. aureus and effectively combated the lethal pneumonia caused by MRSA. The results of cellular thermal shift assay showed that nepetin could bind to ClpP and reduce the thermal stability of ClpP, and the KD value of 602 nM between them was determined using localized surface plasmon resonance. The binding mode of nepetin and ClpP was further investigated by molecular docking, and it was found that Ser-22 and Gln-47 of ClpP residues were found to be involved in the binding of nepetin to ClpP. In conclusion, we determined that nepetin is a ClpP inhibitor and an effective lead compound for the development of a virulence factor-based treatment for MRSA infection.

The Clinical Application Value of Multiple Combination Food Intolerance Testing.

BACKGROUND: We aimed to investigate the clinical value of detecting 14 food intolerances. METHODS: A total of 312 patients with food intolerance enrolled in the Third Affiliated Hospital of Qiqihar Medical University (Qiqihar, China) from Feb 2016 to Feb 2017 were selected. ELISA was used to detect intolerance specific IgG antibodies for 14 foods (pork, chicken, beef, shrimp, fish, crab, egg white/yolk, tomato, mushroom, milk, corn, rice, soybean, wheat). RESULTS: The highest average positive rate of the patients was 42.31% for the crab, followed by shrimp 21.15%, egg white/yolk 18.27% and milk 16.99%. The positive rate from high to low was crab, shrimp, egg white/yolk, milk, fish, corn, soybeans, tomatoes, rice, mushrooms, wheat, pork, beef, chicken. There were significant differences in the specific IgG antibody positive rates between shrimp, soybean and wheat in the skin symptoms group, gastrointestinal symptoms group, respiratory symptom group and nervous system symptom group (P<0.05). There was a significant difference in the positive rate of specific IgG antibodies between shrimp, crab and egg white/yolk in the adolescent group, the middle-aged group and the elderly group (P<0.05). CONCLUSION: The detection of food-specific IgG antibodies can help to determine which food intolerance caused the disease, and then adopt a fasting or diet method to avoid eating unsuitable foods and continually damaging the body, thus maintaining good health. The detection method provides a new idea for the diagnosis and prevention of diseases.

Danhong injection attenuates isoproterenol-induced cardiac hypertrophy by regulating p38 and NF-κb pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Danhong injection (DHI), derived from Rhizoma Salviae Miltiorrhizae (Salvia miltiorrhiza Bge., Labiatae, Danshen in Chinese) and Flos Carthami (Carthamus tinctorius L., Compositae, Salvia militiorrhiza Bunge), is an extensively-used Chinese material standardized clinical product for treatment of cardiovascular diseases. AIM OF THE STUDY: Cardiac hypertrophy (CH) is an adaptive response of cardiomyocytes. Long-lasting cardiac hypertrophy results in the loss of compensation by cardiomyocytes which could ultimately develop into heart failure. In the present study, we aimed to investigate the effect and exact mechanisms of DHI on isoproterenol (ISO)-induced CH. MATERIALS AND METHODS: H9c2 cells and male Wistar rats were stimulated by ISO in the present study to establish CH models in vitro and in vivo. CCk-8 assay, Western blot, real time-polymerase chain reaction (RT-PCR), electrophoretic mobility shift assay (EMSA) and Echocardiography were used in the present study. RESULTS: DHI significantly attenuated ISO-induced CH of H9c2 cells (p<0.01). DHI decreased ISO-induced atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) elevation both at the mRNA and protein levels (p<0.05 and p<0.01, respectively). Western blot showed that DHI down-regulated the phosphorylation of p38. Furthermore, we found that DHI inhibited the nuclear translocation and activation of NF-κb. Echocardiography from ISO-induced CH rats showed that DHI significantly decreased left ventricle (LV) mass, the thickness of the LV end-systolic posterior wall (LVPWs), and the LV end-diastolic posterior wall (LVPWd) elevated by ISO (p<0.01 and p<0.05, respectively). CONCLUSION: These data demonstrate that DHI might exert anti-cardiac hypertrophic effects by regulating p38 and NF-κb pathway.