Thwarting resistance: MgrA inhibition with methylophiopogonanone a unveils a new battlefront against S. aureus.

Limitations in the clinical treatment of Staphylococcus aureus (S. aureus) infections have arisen due to the advent of antibiotic-resistant strains. Given the immense potential of therapeutic strategies targeting bacterial virulence, the role of MgrA as a pivotal virulence determinant in S. aureus-orchestrating resistance, adherence, and hundreds of virulence targets-becomes indispensable. In this investigation, leveraging advanced virtual screening and fluorescence anisotropy assays, we discerned methylophiopogonanone A (Mo-A), a flavonoid derivative, as a potent disruptor of the MgrA-DNA interaction nexus. Subsequent analysis revealed that Mo-A effectively inhibits the expression of virulence factors such as Hla and Pvl in S. aureus and markedly reduces its adhesion capability to fibrinogen. On a cellular landscape, Mo-A exerts a mitigating influence on the deleterious effects inflicted by S. aureus USA300 on A549 cells. Furthermore, our data indicate that Mo-A downregulates the transcription of genes associated with immune evasion, such as nucleases (nuc), Staphylococcal Chemotaxis Inhibitory Protein (chips), and Staphylococcal Complement Inhibitor (scin), thereby undermining immune escape and amplifying neutrophil chemotaxis. Upon application in an in vivo setting, Mo-A assumes a protective persona in a murine model of S. aureus USA300-induced pneumonia and demonstrates efficacy in the Galleria mellonella infection model. Of note, S. aureus displayed no swift acquisition of resistance to Mo-A, and the effect was synergistically enhanced when used in combination with vancomycin. Our findings add substantive weight to the expanding field of virulence-targeted therapeutic strategies and set the stage for more comprehensive exploration of Mo-A potential in combating antibiotic-resistant S. aureus.

Resveratrol protects mesangial cells under high glucose by regulating the miR-1231/IGF1/ERK pathway.

Diabetic nephropathy (DN) is one of the complications of diabetes mellitus and the main cause of end-stage renal disease (ESRD), which is a serious threat to human health. In DN, mesangial cells (MCs) are a critical target cell that perform a variety of key functions, and abnormal proliferation of MCs is a common and prominent pathological change in DN. In recent years, the investigation of Chinese medicine interventions for DN has increased significantly in recent years due to the many potential adverse effects and controversies associated with the treatment of DN with Western medicines. In this study, we evaluated the protective effect of resveratrol (RES), an active ingredient known as a natural antioxidant, on HMCs under high glucose and explored its possible mechanism of action. We found that RES inhibited the proliferation of human mesangial cell (HMC) under high glucose and blocked cell cycle progression. In the high glucose environment, RES upregulated miR-1231, reduced IGF1 expression, inhibited the activity of the extracellular signal-regulated kinase (ERK) signaling pathway and reduced levels of the inflammatory factors TNF-α and IL-6. In addition, we found that miR-1231 mimics were synergistically inhibited with RES, whereas miR-1231 inhibitor attenuated the protective effect of RES on HMCs. Thus, our results suggest that the protective effect of RES on HMCs under high glucose is achieved, at least in part, through modulation of the miR-1231/IGF1/ERK pathway. The discovery of this potential mechanism may provide a new molecular therapeutic target for the prevention and treatment of DN, and may also bring new ideas for the clinical research in DN.

Isovanillic acid protects mice against Staphylococcus aureus by targeting vWbp and Coa.

Aim: Our primary objective was to investigate the protective effects and mechanisms of isovanillic acid in mice infected with Staphylococcus aureus Newman. Methods: In vitro coagulation assays were used to validate vWbp and Coa as inhibitory targets of isovanillic acid. The binding mechanism of isovanillic acid to vWbp and Coa was investigated using molecular docking and point mutagenesis. Importantly, a lethal pneumonia mouse model was used to assess the effect of isovanillic acid on survival and pathological injury in mice. Results & Conclusion: Isovanillic acid reduced the virulence of S. aureus by directly binding to inhibit the clotting activity of vWbp and Coa, thereby reducing lung histopathological damage and improving the survival rate in mice with pneumonia.

Oroxylin a glucuronide as a novel class of reversible inhibitors of Sortase a, combats MRSA-induced infections.

AIMS: The main purpose of this study was to study the therapeutical effect of oroxylin A glucuronide (OAG) on methicillin-resistant Staphylococcus aureus (MRSA). METHODS AND RESULTS: By substrate peptide reaction-based fluorescence resonance energy transfer (FRET) screening, we identified that OAG was an efficient inhibitor of Sortase A (SrtA) with an IC50 of 45.61 µg mL-1, and achieved efficacy in the treatment of Staphylococcus aureus (S. aureus) infections. We further demonstrated that OAG inhibited the adhesion of the S. aureus to fibrinogen, the surface protein A anchoring and diminished biofilm formation. Results obtained from fluorescence quenching assay elucidated a direct interaction between OAG and SrtA. Employing molecular dynamics simulations, we proved that OAG binds to the binding sites of R197, G192, E105, and V168 in the SrtA. Notably, OAG exhibited a robust therapeutic effect in a MRSA-induced pneumonia model. CONCLUSIONS: We identified that OAG as a novel class of reversible inhibitors of SrtA, combats MRSA-induced Infections.

Hibifolin, a Natural Sortase A Inhibitor, Attenuates the Pathogenicity of Staphylococcus aureus and Enhances the Antibacterial Activity of Cefotaxime.

This study aimed to identify hibifolin as a sortase A (SrtA) inhibitor and to determine whether it could attenuate the virulence of methicillin-resistant Staphylococcus aureus (MRSA). We employed a fluorescence resonance energy transfer (FRET) assay to screen a library of natural molecules to identify compounds that inhibit SrtA activity. Fluorescence quenching assay and molecular docking were performed to verify the direct binding interaction between SrtA and hibifolin. The pneumonia model was established using C57BL/6J mice by MRAS nasal administration for evaluating the effect of hibifolin on the pathogenicity of MRSA. Herein, we found that hibifolin was able to inhibit SrtA activity with an IC50 of 31.20 µg/mL. Further assays showed that the capacity of adhesion of bacteria to the host cells and biofilm formation was decreased in hibifolin-treated USA300. Results obtained from fluorescence quenching assay and molecular docking indicated that hibifolin was capable of targeting SrtA protein directly. This interaction was further confirmed by the finding that the inhibition activities of hibifolin on mutant SrtA were substantially reduced after mutating the binding sites (TRP-194, ALA-104, THR-180, ARG-197, ASN-114). The in vivo study showed that hibifolin in combination with cefotaxime protected mice from USA300 infection-induced pneumonia, which was more potent than cefotaxime alone, and no significant cytotoxicity of hibifolin was observed. Taken together, we identified that hibifolin attenuated the pathogenicity of S. aureus by directly targeting SrtA, which may be utilized in the future as adjuvant therapy for S. aureus infections. IMPORTANCE We identified hibifolin as a sortase A (SrtA) inhibitor by screening the natural compounds library, which effectively inhibited the activity of SrtA with an IC50 value of 31.20 µg/mL. Hibifolin attenuated the pathogenic behavior of Staphylococcus aureus, including adhesion, invasion, and biofilm formation. Binding assays showed that hibifolin bound to SrtA protein directly. Hibifolin improved the survival of pneumonia induced by S. aureus USA300 in mice and alleviated the pathological damage. Moreover, hibifolin showed a synergistic antibacterial effect with cefotaxime in USA300-infected mice.

Solamargine Inhibits the Development of Hypopharyngeal Squamous Cell Carcinoma by Decreasing LncRNA HOXA11-As Expression.

Hypopharyngeal squamous cell carcinoma (HSCC) is one of the high mortality cancers with a poor prognosis, which is driving the development of new chemotherapeutic agents. We identified the anticancer effects of a natural compound, solamargine (SM), on FaDU cells and explored its mechanism in terms of non-coding RNA. It was observed that SM inhibited the proliferation of FaDU cells with an IC50 of 5.17 µM. High-throughput sequencing data revealed that lncRNA HOXA11-AS was significantly downregulated in cells co-incubated with SM. Further assays demonstrated that SM-induced downregulation of lncRNA HOXA11-AS showed important implications for apoptosis. Given the properties of HOXA11-AS as a miR-155 sponge, we further confirmed that SM upregulated the expression of miR-155 in FaDU cells. C-Myc is a transcription factor that regulates cell differentiation and apoptosis, whose mRNA is considered to be targeted by miR-155. We showed that c-Myc expression was downregulated by SM and accompanied by increased apoptosis, which was consistent with the findings of transcriptome sequencing. Furthermore, SM administration suppressed xenograft tumor growth in a xenograft mouse model in vivo. In the light of the aforementioned findings, our results suggested that SM downregulated the expression of HOXA11-AS, which in turn induces apoptosis by downregulating c-Myc in FaDU, providing evidence for the anticancer effect of SM on HSCC and uncovering the effect of SM on non-coding RNAs as, at least partly, a mechanism of action.

The global succinylation of SARS-CoV-2-infected host cells reveals drug targets.

SARS-CoV-2, the causative agent of the COVID-19 pandemic, undergoes continuous evolution, highlighting an urgent need for development of novel antiviral therapies. Here we show a quantitative mass spectrometry-based succinylproteomics analysis of SARS-CoV-2 infection in Caco-2 cells, revealing dramatic reshape of succinylation on host and viral proteins. SARS-CoV-2 infection promotes succinylation of several key enzymes in the TCA, leading to inhibition of cellular metabolic pathways. We demonstrated that host protein succinylation is regulated by viral nonstructural protein (NSP14) through interaction with sirtuin 5 (SIRT5); overexpressed SIRT5 can effectively inhibit virus replication. We found succinylation inhibitors possess significant antiviral effects. We also found that SARS-CoV-2 nucleocapsid and membrane proteins underwent succinylation modification, which was conserved in SARS-CoV-2 and its variants. Collectively, our results uncover a regulatory mechanism of host protein posttranslational modification and cellular pathways mediated by SARS-CoV-2, which may become antiviral drug targets against COVID-19.

Natural flavone hispidulin protects mice from Staphylococcus aureus pneumonia by inhibition of α-hemolysin production via targeting AgrAC.

The emergence of drug-resistant Staphylococcus aureus (S. aureus) has limited drug options for the clinical treatment of S. aureus infections. Considering recent reports, therapeutic strategies targeting bacterial virulence hold great promise, and alpha-hemolysin (encoded by hla), a critical virulence factor of S. aureus, plays a vital role during bacterial infection. Herein, we demonstrated that hispidulin effectively inhibited the hemolytic activity of S. aureus USA300 without suppressing bacterial growth, along with inhibiting hla transcription and expression in a dose-dependent manner. As heptamer formation is essential for hla-mediated invasion of cells, nevertheless, hispidulin did not affect the deoxycholate-induced oligomerization of hla, suggesting that hispidulin did not affect the protein activity of hla. In vitro assays illustrated that hispidulin bound to agrAC, a crucial protein in quorum sensing. Meanwhile, hispidulin alleviated A549 cell damage caused by S. aureus USA300 and reduced lactate dehydrogenase release. In vivo studies showed that hispidulin had a protective effect against pneumonia caused by S. aureus USA300 in mice. S. aureus did not develop resistance to hispidulin in the short term. Interestingly, our research indicated that hispidulin synergized with the antibacterial activity of cefoxitin. These results showed that hispidulin effectively inhibited α-hemolysin expression by inhibiting the agr quorum sensing of S. aureus. It has promise as an agent to treat S. aureus infection.

Punicalagin, an Inhibitor of Sortase A, Is a Promising Therapeutic Drug to Combat Methicillin-Resistant Staphylococcus aureus Infections.

Antimicrobial resistance (AMR) poses a major threat to human health globally. Staphylococcus aureus is recognized as a cause of disease worldwide, especially methicillin-resistant S. aureus (MRSA) and vancomycin-resistant S. aureus (VRSA). The enzyme sortase A (SrtA), present on the cell surface of S. aureus, plays a key role in bacterial virulence without affecting the bacterial viability, and SrtA-deficient S. aureus strains do not affect the growth of bacteria. Here, we found that punicalagin, a natural compound, was able to inhibit SrtA activity with a very low half maximal inhibitory concentration (IC50) value of 4.23 µg/mL, and punicalagin is a reversible inhibitor of SrtA. Moreover, punicalagin has no distinct cytotoxicity toward A549, HEK293T, or HepG2 cells at a much higher concentration than the IC50 detected by MTT [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide] assays. In addition, punicalagin visibly attenuated the virulence-related phenotype of SrtA in vitro by decreasing adhesion of S. aureus to fibrinogen, reducing the ability of protein A (SpA) displayed on the surface of the bacteria and biofilm formation. Fluorescence quenching elucidated the interaction between punicalagin and SrtA. Molecular docking further implied that the inhibitory activity lay in the bond between punicalagin and SrtA residues LYS190, TYR187, ALA104, and GLU106. In In vivo studies, we surprisingly found that punicalagin had a more effective curative effect combined with cefotaxime when mice were infected with pneumonia caused by MRSA. Essentially, punicalagin, a therapeutic compound targeting SrtA, demonstrates great potential for combating MRSA infections.

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.

Scutellarin potentiates vancomycin against lethal pneumonia caused by methicillin-resistant Staphylococcus aureus through dual inhibition of sortase A and caseinolytic peptidase P.

The strategy of targeting virulence factor has received great attention as it barely develops bacterial resistance. Sortase A (SrtA) and caseinolytic peptidase P (ClpP), as important virulence factors, are considered to be ideal pharmacological targets for methicillin-resistant Staphylococcus aureus (MRSA) infection. Through screening hundreds of compounds, we found scutellarin, a natural flavonoid, markedly inhibited SrtA and ClpP activities of MRSA strain USA300 with an IC50 of 53.64 µg/mL and 107.00 µg/mL, respectively. Subsequently, we observed that scutellarin could inhibit the SrtA-related virulence of MRSA. To demonstrate whether scutellarin directly binding to SrtA, fluorescence quenching assay and molecular docking were performed and the results indicated that scutellarin directly bonded to SrtA molecule with a KA value of 7.58 × 104 L/mol. In addition to direct SrtA inhibition, scutellarin could also inhibit hemolytic activity of S. aureus by inhibiting the expression of Hla in a SrtA-independent manner. Further assays confirmed that scutellarin inhibited hemolysis by inhibiting ClpP. The combination of scutellarin and vancomycin showed enhancing inhibition of USA300 in vitro and in vivo, evidenced by decreased MIC from 3 µg/mL to 0.5 µg/mL and increased survival and improvement of lung pathology in pneumonia mice. Taken together, these results suggest that scutellarin exhibited di-inhibitory effects on SrtA and ClpP of USA300. The di-inhibition of virulence factors by scutellarin combined with vancomycin to prevent MRSA invasion of A549 cells and pneumonia in mice, indicating that scutellarin is expected to be a potential adjuvant against MRSA in the future.

Kaempferol inhibits the expression of α-hemolysin and protects mice from methicillin-resistant Staphylococcus aureus-induced lethal pneumonia.

Staphylococcus aureus (S. aureus) is a common pathogenic bacterium that induces a variety of diseases in humans and animals. The significant pathogenicity of S. aureus is due to its expression of several virulence factors. Alpha-hemolysin (Hla) has attracted attention as a virulence factor in staphylococcal pathogenesis and has been the predominant focus of intense research. In this study, we found that kaempferol, a flavonoid compound, inhibited hemolysis at a low concentration (32 µg/mL) and exerted no effect on bacterial growth. Western blot and RT-qPCR assays further demonstrated that kaempferol downregulated the expression of Hla in S. aureus. We observed that kaempferol alleviated the damage from S. aureus Hla in A549 cells. More importantly, kaempferol showed a potent protective effect on mice pneumonia induced by MRSA, as evidenced by a significant improvement in the survival of mice, a reduction in the number of colonized colonies in lung tissue and a decrease in the pathological damage to lung tissues. In summary, the results demonstrate the protective effect of kaempferol on MRSA-induced lethal pneumonia in mice and indicate that kaempferol could be developed as a potential anti-MRSA drug.

Assessment of the reporting quality of randomized controlled trials on treatment of coronary heart disease with traditional chinese medicine from the chinese journal of integrated traditional and Western medicine: a systematic review.

BACKGROUND: Due to language limitations, little is known about the reporting quality of randomized clinical trials (RCTs) on the treatment of coronary heart disease (CHD) with traditional Chinese medicine (TCM) in Chinese Journal of Integrated Traditional and Western Medicine (CJITWM). OBJECTIVE: In this study, we utilized the CONSORT 2010 statement to understand the reporting quality of RCTs on CHD with TCM from the CJITWM. METHODS: The China National Knowledge Infrastructure (CNKI) electronic database was searched for CJITWM RCTs on the treatment of CHD with TCM, published between January 1, 2006 and December 31, 2011. We excluded articles reported as "animal studies," "topic review," "diagnostic test," "editorials," or "others." The CONSORT checklist was applied to evaluate the reporting quality of all eligible articles by two independent authors after extensive discussion. Each item was graded as either "yes" or "no" depending on whether the authors had reported it or not. RESULTS: We identified 21 articles meeting our inclusion criteria. The percentage of 11 of the 37 items was 4.8 ∼ 95.2%, 14 of the 37 items were reported in all included articles, while 12 items were not mentioned at all. The average reporting percentage for the "title and abstract" section was 52.4%, for the "introduction" section 100.0%, for the "methods" section 45.4%, for the "results" section 57.1%, for the "discussion" section 79.4%, and for the "other information" section 17.5%. CONCLUSION: In general, the reviewed RCTs were not consistent with the CONSORT 2010 statement. Authors should adhere to the CONSORT statement in reporting RCTs; editorial departments may consider the CONSORT statement as a guideline and should instruct authors to write manuscripts, and reviewers to judge them according to CONSORT statutes.

Assessment of the reporting quality of randomized controlled trials on the treatment of diabetes mellitus with traditional chinese medicine: a systematic review.

BACKGROUND: After the publication of the CONSORT 2010 statement, few studies have been conducted to assess the reporting quality of randomized clinical trials (RCTs) on treatment of diabetes mellitus with Traditional Chinese Medicine (TCM) published in Chinese journals. OBJECTIVE: To investigate the current situation of the reporting quality of RCTs in leading medical journals in China with the CONSORT 2010 statement as criteria. METHODS: The China National Knowledge Infrastructure (CNKI) electronic database was searched for RCTs on the treatment of diabetes mellitus with TCM published in the Journal of Traditional Chinese Medicine, Chinese Journal of Integrated Traditional & Western Medicine, and the China Journal of Chinese Materia Medica from January to December 2011. We excluded trials reported as "animal studies", "in vitro studies", "case studies", or "systematic reviews". The CONSORT checklist was applied by two independent raters to evaluate the reporting quality of all eligible trials after discussing and comprehending the items thoroughly. Each item in the checklist was graded as either "yes" or "no" depending on whether it had been reported by the authors. RESULTS: We identified 27 RCTs. According to the 37 items in the CONSORT checklist, the average reporting percentage was 45.0%, in which the average reporting percentage for the "title and abstract", the "introduction", the "methods", the "results", the "discussion" and the "other information" was 33.3%, 88.9%, 36.4%, 54.4%, 71.6% and 14.8%, respectively. In the Journal of Traditional Chinese Medicine, Chinese Journal of Integrated Traditional & Western Medicine, and the China Journal of Chinese Materia Medica the average reporting percentage was 42.2%, 56.8%, and 46.0%, respectively. CONCLUSIONS: The reporting quality of RCTs in these three journals was insufficient to allow readers to assess the validity of the trials. We recommend that editors require authors to use the CONSORT statement when reporting their trial results as a condition of publication.