Mechanisms of Antibiotic Resistance and Developments in Therapeutic Strategies to Combat Klebsiella pneumoniae Infection.

Infections with drug-resistant bacteria have become one of the greatest public health challenges, and K. pneumoniae is among the top six drug-resistant bacteria. K. pneumoniae often causes nosocomial infections, leading to illnesses such as pneumonia, liver abscesses, soft tissue infections, urinary tract infections, bacteremia, and in some cases death. As the pathogen continues to evolve and its multidrug resistance increases, K. pneumoniae poses a direct threat to humans. Drug resistance in K. pneumoniae may occur due to the formation of biofilms, efflux pumps, and the production of ß-lactamases. In many cases, resistance is further enhanced by enzymatic modification and loss of porins. Drug resistance to K. pneumoniae has led to a decline in the effectiveness of conventional therapies against this pathogen. Therefore, there is an urgent need to accelerate the development of new antibiotics and explore new therapeutic approaches such as antimicrobial peptides, phages, traditional Chinese medicine, immunotherapy, Antimicrobial nanoparticle technology, antisense oligonucleotides and gene editing technologies. In this review, we discuss the mechanisms of drug resistance in K. pneumoniae and compare several new potential therapeutic strategies to overcome drug resistance in the treatment of K. pneumoniae infections.

Study on Synergistic Anti-Inflammatory Effect of Typical Functional Components of Extracts of Ginkgo Biloba Leaves.

There are some differences in the anti-inflammatory activities of four typical components in EGB (extracts of ginkgo biloba leaves), and there is also a synergistic relationship. The order of inhibiting the NO-release ability of single functional components is OA > GF > OPC > G. Ginkgolide (G), proanthocyanidins (OPC), and organic acids (OA) all have synergistic effects on ginkgo flavonoids (GF). GF:OA (1:9) is the lowest interaction index among all complexes, showing the strongest synergy. The anti-inflammatory mechanism of the compound affects the expression of p-JNK, p-P38, and p-ERK1/2 proteins by inhibiting the expression of iNOS and COX2 genes on NFKB and MAPK pathways. This also provides a research basis for the development of anti-inflammatory deep-processing products of EGB.

Network Pharmacology Analysis and Experimental Verification on Antiangiogenesis Mechanism of Hedyotis diffusa Willd in Liver Cancer.

Purpose: Hedyotis diffusa Willd (HDW) is one of the most well-known herbs used in the therapy of cancer. However, the potential mechanisms of its antiangiogenic effects have not been fully explored. Here, we applied a network pharmacology approach to explore the potential mechanisms of HDW against liver cancer angiogenesis (LCA) and used a mouse orthotopic liver cancer model for experimental verification accordingly. Methods: The effective components, primary active compounds, and possible targets in the therapy of LCA were predicted using network pharmacology and bioinformatics. In vivo testing of the pharmacodynamic foundation of HDW in the treatment of LCA was performed. Hepa1-6 cells were implanted in C57BL/6 mice to establish an orthotopic liver cancer model to evaluate the antitumor and antiangiogenesis effects of the drug. Furthermore, protein levels were evaluated by western blotting, immunofluorescence, and immunohistochemistry. Results: We firstly confirmed the therapeutic effect of HDW on LCA and subsequently screened 7 active compounds from HDW according to their pharmacokinetic properties. Network analysis and enrichment analysis indicated that these compounds exhibit antiangiogenic effect by acting on multiple targets and thereby regulating multiple pathways mainly involved in Akt1, IL-6, IL-1ß, IL-17, hypoxia inducible factor-1α (HIF-1α), and tumor necrosis factor-α (TNF-α). Importantly, we preliminarily verified the results of the network pharmacology analysis in vivo. Conclusion: Collectively, our work initially explored the therapeutic mechanism of HDW on tumor angiogenesis, which lays an experimental reference for further exploring its pharmacological action and its clinical application.

Study on Synergistic Antioxidant Effect of Typical Functional Components of Hydroethanolic Leaf Extract from Ginkgo Biloba In Vitro.

The predicted anti-oxidation is related to apoptosis, proliferation, lipid metabolism, cell differentiation, and immune response. There are some differences in the antioxidant capacity of the four typical components of ginkgo biloba extract (EGb) including ginkgo flavone (GF), ginkgolide (G), procyanidins (OPC), and organic acids (OA), and any two members of them can exhibit apparent synergistic effects. The order of DPPH scavenging ability was: OPC > GF > OA > G. The scavenging ability of procyanidins was close to that of VC; the scavenging capacity of ABTS was GF > OPC > OA > G. The GF:OPC (1:9) showed the best synergism in scavenging DPPH and ABTS radicals. The 193 kinds of small molecules reported in EGb were obtained by analyzing the properties of EGb. In order to construct a corresponding biological activity target set, molecular docking and the network pharmacology method were employed to build the molecular action mechanism network of a compound target, and the main biological functions and signaling pathways involved with their antioxidant activities were predicted. The results displayed that the top ten compounds which belonged to the two broad categories, ginkgo flavonoids and proanthocyanidins, could interact closely with several important target proteins (CASP3, SOD2, MAPK1, HSPA4, and NQO1). This would be expected to lay a theoretical foundation for the deep development of Ginkgo biloba extract.

Deoxyelephantopin decreases the release of inflammatory cytokines in macrophage associated with attenuation of aerobic glycolysis via modulation of PKM2.

Growing evidence suggests that activated immune cells undergo metabolic reprogramming in the regulation of the innate inflammatory response. Remarkably, macrophages activated by lipopolysaccharide (LPS) induce a switch from oxidative phosphorylation to aerobic glycolysis, and consequently results in release of proinflammatory cytokines. Pyruvate Kinase M2 (PKM2) plays a vital role in the process of macrophage activation, promoting the inflammatory response in sepsis and septic shock. Deoxyelephantopin (DET), a naturally occurring sesquiterpene lactone from Elephantopus scaber, has been shown to counteracts inflammation during fulminant hepatitis progression, but the underlying mechanism remains unclear. Here, we studied the function of the DET on macrophage activation and investigated the anti-inflammatory effects of DET associated with interfering with glycolysis in macrophage. Our results first demonstrated that DET attenuates LPS-induced interleukin-1ß (IL-1ß) and high-mobility group box 1 (HMGB1) release in vitro and in vivo and protected mice against lethal endotoxemia. Furthermore, DET decreased the expression of pyruvate dehydrogenase kinase 1 (PDK1), glucose transporter 1(GLUT1), lactate dehydrogenase A (LDHA), and reduced lactate production dose-dependently in macrophages. Moreover, we further revealed that DET attenuates aerobic glycolysis in macrophages associated with regulating the nuclear localization of PKM2. Our results provided a novel mechanism for DET suppression of macrophages activation implicated in anti-inflammatory therapy.

Enrichment and Purification of Total Ginkgo Flavonoid O-Glycosides from Ginkgo Biloba Extract with Macroporous Resin and Evaluation of Anti-Inflammation Activities In Vitro.

In the present study, the performance and separation characteristics of six macroporous resins for the enrichment and purification of total ginkgo flavonoid O-glycosides (TGFs) (quercetin (I), kaempferol (II), isorhamnetin (III)) from Ginkgo Biloba extracts (EGB) are evaluated. The adsorption and desorption properties of TGFs are studied on macroporous resins, including D101, D201, AB-8, HPD400, D301, and D311. Along with the results, AB-8 resin exhibits the best adsorption and desorption capacity for these three ginkgo flavonoid O-glycosides among the six resins. Adsorption isotherms are created on AB-8 resin and fit well to the Langmuir (R² > 0.96) and Freundlich (R² > 0.92, 0.3 < 1/n < 0.7) models. After the treatment with gradient elution on AB-8 resin packed chromatography column, the contents of the three main ginkgo flavonoid O-glycosides (I, II, and III) increase from 8.93%, 9.88%, and 6.11% in the extracts to 30.12%, 35.21%, and 14.14%, respectively, in the product. The recoveries of compounds I, II, and III are 88.76%, 93.78%, and 60.90%, respectively. Additionally, the anti-inflammatory effects of TGFs are evaluated in LPS-treated RAW 264.7 macrophages, and the result demonstrates that TGFs could significantly inhibit LPS-induced NO release in vitro in a dose-dependent manner compared with the control group. These findings suggest that TGFs could potentially be natural antioxidants and anti-inflammatory ingredients that could be used in pharmaceutical products and functional food additives.

Identification of Human Acetylcholinesterase Inhibitors from the Constituents of EGb761 by Modeling Docking and Molecular Dynamics Simulations.

AIM AND OBJECTIVE: EGb761, a standardized and well-defined product extract of Ginkgo biloba leaves, has beneficial role in the treatment of multiple diseases, particularly Alzheimer's disease (AD). Identification of natural acetylcholinesterase (AChE) inhibitors from EGb761 would provide a novel therapeutic approach against the Alzheimer's disease. MATERIAL AND METHOD: A series of 21 kinds of promising EGb761 compounds were selected, and subsequently evaluated for their potential ability to bind AChE enzyme by molecular docking and a deep analysis of protein surface pocket features. RESULTS: Docking results indicated that these compounds can bind tightly with the active site of human AChE, with favorable distinct interactions around several important residues Asp74, Leu289, Phe295, Ser293, Tyr341, Trp286 and Val294 in the active pocket. Most EGB761 compounds could form the hydrogen bond interactions with the negatively charged Asp74 and Phe295 residues. Among these compounds, diosmetin is the one with the best-predicted docking score while three key hydrogen bonds can be formed between small molecule and corresponding residues of the binding site. Besides, other three compounds luteolin, apigenin, and isorhamnetin have better predicted docking scores towards AChE than other serine proteases, i.e. Elastase, Tryptase, Factor XA, exhibiting specificity for AChE inhibition. The RMSD and MM-GBSA results from molecular dymamic simulations indicated that the docking pose of diosmetin-AChE complex displayed highly stable, which can be used for validating the accuracy of molecular docking study. Subsequently, the AChE inhibitory activities of these compounds were evaluated by the Ellman's colorimetric method. CONCLUSION: The obtained results revealed that all the four compounds exhibited modest AChE inhibitory activity, among which Diosmetin manifested remarkable anti-AChE activity, comparable with the reference compound, Physostigmine. It can be deduced that these EGB761 compounds can be regarded as a promising starting point for developing AChE inhibitors against AD.

[Isolation and characteristic of SmbHLH1 gene in Salvia miltiorrhiza].

OBJECTIVE: A novel bHLH-like gene, designated SmbHLH1, was isolated from Salvia miltiorrhiza, in order to identify a bHLH gene in related to danshinone biosysnthesis. METHOD: SmbHLH1 was isolated by RT-PCR,and Semi-quantitative RT-PCR was used to detect the gene expression level. RESULT: The full length of SmbHLH1 cDNA has an open reading frame of 999 bp. The deduced amino acid sequence of SmbHLH1 has 332 amino acid residues which forms a 36 kDa polypeptide with a calculated pI of 5.4. SmbHLH1 gene was expressed at high level in root, but low level in stem, leaf and flower of S. miltiorrhiza. The transcripts of SmbHLH1 was suppressed when the plants were treated with exogenous MeJA, Yeast + Ag+. The transcripts of SmbHLH1 constitutively accumulated in response to exogenous ABA and low concentration of salicylic acid. CONCLUSION: SmbHLH is a new member of the S. miltiorrhiza bHLH family, and its possible roles in brassinosteriods signaling responses.