An investigation into the mechanism of nobiletin's inhibition of papillary thyroid cancer using network pharmacology analysis and experimental pharmacology.

OBJECTIVE: Surgery and radioactive iodine therapy are the main treatments for papillary thyroid carcinoma (PTC), and effective drugs are lacking. As a promising natural product, nobiletin (NOB) has a wealth of pharmacological activities like anti-tumor, antivirus, and other effects. In this research, bioinformatics methods and cellular assays were combined to explore how NOB inhibited PTC. MATERIALS AND METHODS: Our NOB targets were derived from three databases, including the SwissTargetPrediction database, Traditional Chinese Medicine System Pharmacology Database, and the TargetNet server. Four databases were used to identify disease-related targets: GeneCards, PharmGkb, Online Mendelian Inheritance in Man, and DisGeNET. Finally, cross-targets of disease and drug were deemed as pharmacological targets, and they were used for GO and KEGG enrichment analysis. STRING and Cytoscape were applied for PPI Network and core Targets Ranking. Molecular docking analysis validated binding affinity values for NOB and core targets. By using cell proliferation and migration assays, NOB was assessed for its effects on PTC proliferation and migration phenotype. Western blot validated the downregulation of the PI3K/Akt pathway. RESULTS: (1) Preliminarily, 85 NOB targets were predicted for NOB intervention in PTC. (2) Our core target screening identified TNF, TP53, and EGFR, and our molecular docking results confirmed good binding between NOB and protein receptors. (3) NOB inhibited proliferation and migration of PTC cells. PI3K/AKT pathway target proteins were downregulated. CONCLUSIONS: (1) Bioinformatics analyses revealed that NOB may inhibit PTC by regulating TNF, TP53, EGFR and PI3K/AKT signalling pathway. (2) As evidenced by cell experiments, there was an inhibition of proliferating and migrating PTCs by NOB via the PI3K/AKT signalling pathway.

The analysis of the effects of Liuwei Dihuang decoction on aging-related metabolites and metabolic pathways in naturally aging mice by ultra-performance liquid chromatography quadruple time-of-light mass spectrometry.

Liuwei Dihuang decoction (LWDH) has been used to treat age-related diseases in Asia for thousands of years. Its effect is significant, but the mechanism is not entirely understood. The development and maturity of metabonomics technology have created a new way to understand its effects. In this study, 20-month-old naturally aging mice were used as the research subjects. Mice were randomly divided into the young group, elderly group, vitamin E group, and LWDH group, with 10 mice in each group. The UPLC-Q-TOF-MS technique analyzed the changes in plasma metabolism of these mice. Principal component analysis (PCA) and orthogonal partial least-squares discriminant analysis (OPLS-DA) were used to study metabolic markers and metabolic pathways. Compared with the elderly group, the organ index and weightbearing swimming time of the LWDH and vitamin E groups increased (P ≤ 0.01). The results show that LWDH can benignly regulate the expression level of 11 aging-associated metabolites in aged mice and alleviate the aging state of mice. These metabolites mainly affect the metabolism of glutathione, unsaturated fatty acid biosynthesis, pyrimidine, selenium, pentose phosphate, arginine, and proline.

Astragalus membranaceus root supplementation improves average daily gain, rumen fermentation, serum immunity and antioxidant indices of Tibetan sheep.

The use of antibiotics as supplements in animal feed is restricted due to possible health hazards associated with them. Consequently, there is increasing interest in exploiting natural products to improve health and production of livestock with no detrimental side effects. In this study, we examined the effect of Astragalus membranaceus root (AMT) supplementation on DM intake, growth performance, rumen fermentation and immunity of Tibetan sheep. Twenty-four male Tibetan sheep (31 ±â€¯1.4 kg; 9 months old) were assigned randomly to one of four dietary treatments with different levels of AMT: 0, 20, 50 and 80 g/kg DM (A0, A2, A5 and A8, respectively) in addition to their basal diets. A0 acted as a control group, and measurements were recorded over a 56-d feeding period. Sheep fed with AMT had a higher average daily gain and a lower feed:gain ratio than controls (P < 0.001). Rumen concentrations of NH3-N (P < 0.001), total volatile fatty acids (P = 0.028), acetate (P = 0.017) and propionate (P = 0.031) in A5 and A8 were higher than those in A0. The addition of AMT in the feed significantly increased serum antioxidant and immunity factors of the sheep and increased the concentrations of serum interleukin, immunoglobulin and tumour necrosis factor-α (P = 0.010). We concluded that AMT can be used as a feed additive to improve growth performance and rumen fermentation and enhance the immunity of Tibetan sheep. Some responses exhibited a dose-dependent response, whereas other did not exhibit a pattern, with an increase in AMT. The addition of 50 and 80 g/kg AMT of total DM intake showed the most promising results.

[Current practice patterns of preoperative bowel preparation in elective colorectal surgery: a nation-wide survey of Chinese surgeons].

Objective: To understand the current practice of preoperative bowel preparation in elective colorectal surgery in China. Methods: A cross-sectional questionnaire survey was conducted through wechat. The content of the questionnaire survey included professional title of the participants, the hospital class, dietary preparation and protocol, oral laxatives and specific types, oral antibiotics, gastric intubation, and mechanical enema before elective colorectal surgery. A stratified analysis based on hospital class was conducted to understand their current practice of preoperative bowel preparation in elective colorectal surgery. Result: A total of 600 questionnaires were issued, and 516 (86.00%) questionnaires of participants from different hospitals, engaged in colorectal surgery or general surgeons were recovered, of which 366 were from tertiary hospitals (70.93%) and 150 from secondary hospitals (29.07%). For diet preparation, the proportions of right hemicolic, left hemicolic and rectal surgery were 81.59% (421/516), 84.88% (438/516) and 84.88% (438/516) respectively. The average time of preoperative dietary preparation was 2.03 days. The study showed that 85.85% (443/516) of surgeons chose oral laxatives for bowel preparation in all colorectal surgery, while only 4.26% (22/516) of surgeons did not choose oral laxatives. For mechanical enema, the proportions of right hemicolic, left hemicolic and rectal surgery were 19.19% (99/516), 30.04% (155/516) and 32.75% (169/516) respectively. Preoperative oral antibiotics was used by 34.69% (179/516) of the respondents. 94.38% (487/516) of participants were satisfied with bowel preparation, and 55.43% (286/516) of participants believed that preoperative bowel preparation was well tolerated. In terms of preoperative oral laxatives, there was no statistically significant difference between different levels of hospitals [secondary hospitals vs. tertiary hospitals: 90.00% (135/150) vs. 84.15% (308/366), χ(2)=2.995, P=0.084]. Compared with the tertiary hospitals, the surgeons in the secondary hospitals accounted for higher proportions in diet preparation [87.33% (131/150) vs. 76.78% (281/366), χ(2)=7.369, P=0.007], gastric intubation [54.00% (81/150) vs. 36.33% (133/366), χ(2)=13.672, P<0.001], preoperative oral antibiotics [58.67% (88/150) vs. 24.86% (91/366), χ(2)=12.259, P<0.001] and enema [28.67% (43/150) vs. 15.30% (56/366), χ(2)=53.661, P<0.001]. Conclusion: Although the preoperative bowel preparation practice in elective colorectal surgery for most of surgeons in China is basically the same as the current international protocol, the proportions of mechanical enema and gastric intubation before surgery are still relatively high.

Long noncoding RNA SNHG14 promotes malignancy of prostate cancer by regulating with miR-5590-3p/YY1 axis.

OBJECTIVE: Studies have demonstrated that long non-coding RNAs (lncRNAs) are important in the development and prognosis of prostate cancer. The aim of this study was to investigate the functions and mechanism of lnc-SNHG14 in prostate cancer. PATIENTS AND METHODS: Quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) or Western blot (WB) were performed to detect mRNA expressions of SNHG14 and miR-5590-3p, and the protein levels of Yin Yang-1 (YY1) in prostate cancer tissues, adjacent tissues, and cancer cell lines. The correlation analysis was used to analyze the correlations between SNHG14, miR-5590-3p, and YY1. Kaplan-Meier survival analysis was used to analyze the overall survival for prostate cancer patients. Cell Counting Kit-8 (CCK-8) assay was performed to measure cell proliferation ability and flow cytometry assay was used to detect cell apoptotic rate. Besides, transwell assay was used to measure cell invasion ability. In addition, WB was performed to measure protein expressions in prostate cancer cell lines. Finally, Luciferase reporter assay was performed to verify the binding sites between SNHG14 and miR-5590-3p, miR-5590-3p, and YY1. RESULTS: The results showed that SNHG14 was significantly increased in prostate cancer tissues and prostate cancer cell lines, which were related with advanced stage and poor diagnosis for prostate cancer patients. MiR-5590-3p was reduced in prostate cancer tissues and cell lines, which were negatively correlated with SNHG14. YY1 was found to be increased in prostate cancer tissues, which was negatively correlated with miR-5590-3p and positively correlated with SNHG14. Furthermore, SNHG14 knockdown inhibited cell proliferation, invasion, and promoted cell apoptosis in DU145 cells. In addition, protein expressions of Cyclin D1, Bcl-2, and N-cadherin were repressed, and the levels of Bax, Cleaved Caspase-3, and E-cadherin were increased. Besides, miR-5590-3p inhibition promoted cell proliferation and invasion, and inhibited apoptosis in DU145 cells. Importantly, Luciferase reporter assay proved that SNHG14 could directly sponge with miR-5590-3p, which could bind with YY1 and regulate the functions of cancer cell. Finally, we proved that SNHG14 regulated cell proliferation, cell apoptosis, and invasion via miR-5590-3p/ YY1 axis in prostate cancer. CONCLUSIONS: Above all, we found that SNHG14 was increased in prostate cancer patients, which was related with future diagnosis for prostate cancer patients. Of note, we discovered that SNHG14 could promote cell proliferation, invasion, and repress cell apoptosis via miR-5590-3p/YY1 axis in prostate cancer, which might provide a new target for treating prostate cancer.

Neuroprotective effect of gastrodin in methamphetamine-induced apoptosis through regulating cAMP/PKA/CREB pathway in cortical neuron.

OBJECTIVE: Methamphetamine (MA) abuse induces neurotoxicity and causes neuronal cell apoptosis. Gastrodin is a traditional Chinese herbal medicine used for the treatment of nerve injuries, spinal cord injuries, and some central nervous system diseases as well. The present study investigated the neuroprotective effects of gastrodin against MA-induced neurotoxicity in neuronal cells and its potential protective mechanism. METHODS: The primary cortex neuronal culture was divided into four groups (control group, MA group, MA + gastrodin group, and MA + gastrodin + small interfering RNA group). The neurotoxicity of MA was assessed by detecting apoptotic cells by terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick-end labeling assay and cell viability by cell counting kit 8 (CCK-8) method, the Tuj1-positive cells and the average axonal length were detected by immunofluorescence, and the expressions of cyclic adenosine monophosphate (cAMP), protein kinase A (PKA), cAMP-response element-binding (CREB), and brain-derived neurotrophic factor (BDNF) proteins were detected by Western blot. RESULTS: The results of CCK-8 assay showed that 0.5 mM MA was an optimal concentration that induced neurotoxicity (p < 0.01). Pretreatment with 25 mg/L gastrodin exerted maximum protective effects on neuronal cells. The expression levels of cAMP, PKA, phosphorylated PKA, CREB, phosphorylated CREB, and BDNF proteins were decreased in the MA group, and pretreatment with gastrodin upregulated the expression levels of these proteins (p < 0.01). The expressions of PKA and CREB proteins showed no significant changes in the control group, MA group, and gastrodin group. Compared the MA + gastrodin + small interfering RNA group with MA + gastrodin group, the Tuj1-positive cells and the average axonal length were decreased significantly, while the number of apoptotic cells was increased (p < 0.05). CONCLUSION: Gastrodin has neuroprotective effects against MA-induced neurotoxicity, which exerts neuroprotective effects via regulation of cAMP/PKA/CREB signaling pathway and upregulates the expression of BDNF.

Increased activity of linezolid in combination with rifampicin in a murine pneumonia model due to MRSA.

Objectives: The chloramphenicol/florfenicol resistance gene cfr, which mediates cross-resistance to linezolid and other classes of antimicrobial agents, represents a global therapeutic challenge due to its dissemination among MDR nosocomial pathogens, including MRSA. This study aimed to compare the efficacy of the linezolid/rifampicin combination in a murine pneumonia model caused by cfr-positive and cfr-negative clinical MRSA strains. Methods: Synergistic activity between linezolid and rifampicin was evaluated by chequerboard and time-kill assays. Pharmacokinetic profiles in plasma and epithelial lining fluid (ELF) as well as the therapeutic efficacy of linezolid alone and in combination with rifampicin were investigated in a murine pneumonia model. The Emax Hill equation was used to model the dose-response relationship. Results: Increased susceptibility of the study MRSA strains to linezolid was observed with the rifampicin combination (MIC decreased 2- to 16-fold versus linezolid alone). The combination had synergistic activity (fractional inhibitory concentration index ≤0.5) against all cfr-positive MRSA isolates. Linezolid demonstrated excellent pulmonary penetration with an ELF/fplasma AUC ratio of 2.68 ±âŸ0.17. The addition of rifampicin significantly improved the efficacy of linezolid in the pneumonia model due to cfr-positive and cfr-negative MRSA strains. The fAUC/MIC targets of linezolid associated with stasis, 1 log10 kill and 2 log10 kill were 15.9, 38.8 and 175 in plasma, and 43.5, 108 and 415 in ELF, respectively. Importantly, the linezolid fAUC/MIC targets in both plasma and ELF were 2.4-6.7 times lower in combined linezolid/rifampicin therapy versus linezolid monotherapy (P < 0.005). Conclusions: Combination of linezolid with rifampicin significantly improved the efficacy of linezolid in the murine pneumonia model caused by MRSA strains in the presence and absence of the cfr gene.

[Hypoglossal nerve stimulation therapy for obstructive sleep apnea hypopnea syndrome].

Obstructive sleep apnea hypopnea syndrome(OSAHS) is a clinical and critical disease to health, which the accuratly pathogenesis is not very explicit. The mainstream research suggested that dysfunction of upper airway dilator muscles is one of the hazard factors. Genioglossus innervated by the hypoglossal nerve plays crucial roles. It could expand volume of pharyngeal cavity, and increase airflow rate when stimulate it.With the development of anatomy and physiology, hypoglossal nerve stimulation(HGNS) entered into clinical trials, and had made tremendous progress since 2001. Most of clinical trials show that, it could dramatically improve the smooth general peculiarity of the upper respiratory tract in patients with OSAHS in the sleeping state.

The protective effect of Shenfu injection against elderly severe pneumonia.

OBJECTIVE: The purpose of this study is to investigate the effect of Shenfu injection (SFI) on the tumor necrosis factor-α (TNF-α) and the interleukin (IL-6, IL-8, IL-10) of elderly patients who suffered from severe pneumonia. METHODS: From June 2012 to September 2014, we performed in our department 89 cases of elderly patients with severe pneumonia. These patients were randomly divided into two groups: the treatment group (45 cases) and the control group (44 cases). The control group was given the treatment of anti-infection, reducing sputum, and support therapy, while the treatment group was fed by SFI intravenously based on the control group. The TNF-α and the interleukin were detected by enzyme-linked immunosorbent assay (ELISA). Meanwhile, the changes in the inflammatory response indicators, the blood gas analysis, and the parameters of vital signs were measured and compared before and after therapy. RESULTS: Prior to treatment, there is no significant difference between the treatment group and the control group (p > 0.05); after the treatment for 7 days, the levels of TNF-α, IL-6, and IL-8 were significantly decreased, while the level of IL-10 was obviously increased. The APACHE II score was significantly decreased in comparison to that before the treatment (p < 0.05), and the time of mechanical ventilation, the duration of time in ICU, and the application time of vasoactive drugs were notably shortened. CONCLUSION: The application of Shenfu injection exhibited a positive and effective effect on removing the inflammation media during the treatment of elderly severe pneumonia.

Telavancin Is Active against Experimental Aortic Valve Endocarditis Caused by Daptomycin- and Methicillin-Resistant Staphylococcus aureus Strains.

We compared the efficacy of telavancin (TLV) and daptomycin (DAP) in an experimental rabbit endocarditis model caused by two clinically derived daptomycin-resistant (DAPr) methicillin-resistant Staphylococcus aureus (MRSA) strains. TLV treatment significantly reduced MRSA densities in all target tissues and increased the percentage of these organs rendered culture negative compared to those with the untreated control or DAP-treated animals. These results demonstrate that TLV has potent in vivo efficacy against DAPr MRSA isolates in this invasive endovascular infection model.

Comparative efficacy of telavancin and daptomycin in experimental endocarditis due to multi-clonotype MRSA strains.

BACKGROUND: MRSA strains of clonal complexes (CCs) 5, 8, 30 and 45 are leading causes of complicated endovascular infections associated with suboptimal clinical outcomes. Telavancin is a novel anti-MRSA agent that both inhibits bacterial cell wall synthesis and disrupts membranes by depolarization. METHODS: In this study, we compared the in vitro susceptibility and in vivo efficacy of telavancin versus daptomycin in an experimental rabbit infective endocarditis (IE) model caused by four MRSA strains representing each of the above CC types. RESULTS: All study strains were susceptible to telavancin (MICs of ≤0.12 mg/L) and daptomycin (MICs of ≤0.5 mg/L). In vitro time-kill analyses revealed that supra-MIC levels of telavancin were effective at preventing regrowth at 24 h of incubation. In the IE animal model for all CC types, treatment with telavancin produced significantly greater reductions in MRSA counts as compared with daptomycin-treated animals in all target tissues. Moreover, telavancin-treated animals had a significantly higher percentage of sterile tissue cultures versus daptomycin-treated animals (e.g. 78%-100% versus 0% sterile vegetations and 100% versus 0%-11% sterile kidneys and spleen, in the telavancin- and daptomycin-treated animals, respectively). CONCLUSIONS: These results suggest that telavancin exhibits significantly greater efficacies versus daptomycin in treating experimental IE caused by MRSA clinical isolates across four common CC types.

Impact of vancomycin on sarA-mediated biofilm formation: role in persistent endovascular infections due to methicillin-resistant Staphylococcus aureus.

BACKGROUND: Staphylococcus aureus is the most common cause of endovascular infections. The staphylococcal accessory regulator A locus (sarA) is a major virulence determinant that may potentially impact methicillin-resistant S. aureus (MRSA) persistence in such infections via its influence on biofilm formation. METHODS: Two healthcare-associated MRSA isolates from patients with persistent bacteremia and 2 prototypical community-acquired MRSA strains, as well as their respective isogenic sarA mutants, were studied for in vitro biofilm formation, fibronectin-binding capacity, autolysis, and protease and nuclease activities. These assays were done in the presence or absence of sub-minimum inhibitory concentrations (MICs) of vancomycin. In addition, these strain pairs were compared for intrinsic virulence and responses to vancomycin therapy in experimental infective endocarditis, a prototypical biofilm model. RESULTS: All sarA mutants displayed significantly reduced biofilm formation and binding to fibronectin but increased protease production in vitro, compared with their respective parental strains. Interestingly, exposure to sub-MICs of vancomycin significantly promoted biofilm formation and fibronectin-binding in parental strains but not in sarA mutants. In addition, all sarA mutants became exquisitely susceptible to vancomycin therapy, compared with their respective parental strains, in the infective endocarditis model. CONCLUSIONS: These observations suggest that sarA activation is important in persistent MRSA endovascular infection, potentially in the setting of biofilm formation.

Role of the LytSR two-component regulatory system in adaptation to cationic antimicrobial peptides in Staphylococcus aureus.

Many host defense cationic antimicrobial peptides (HDPs) perturb the staphylococcal cell membrane (CM) and alter transmembrane potential (ΔΨ) as key parts of their lethal mechanism. Thus, a sense-response system for detecting and mediating adaptive responses to such stresses could impact organism survival; the Staphylococcus aureus LytSR two-component regulatory system (TCRS) may serve as such a ΔΨ sensor. One well-known target of this system is the lrgAB operon, which, along with the related cidABC operon, has been shown to be a regulator in the control of programmed cell death and lysis. We used an isogenic set of S. aureus strains: (i) UAMS-1, (ii) its isogenic ΔlytS and ΔlrgAB mutants, and (iii) plasmid-complemented ΔlytSR and ΔlrgAB mutants. The ΔlytS strain displayed significantly increased in vitro susceptibilities to all HDPs tested (neutrophil-derived human neutrophil peptide 1 [hNP-1], platelet-derived thrombin-induced platelet microbicidal proteins [tPMPs], and the tPMP-mimetic peptide RP-1), as well as to calcium-daptomycin (DAP), a cationic antimicrobial peptide (CAP). In contrast, the ΔlrgAB strain exhibited no significant changes in susceptibilities to these cationic peptides, indicating that although lytSR positively regulates transcription of lrgAB, increased HDP/CAP susceptibilities in the ΔlytS mutant were lrgAB independent. Further, parental UAMS-1 (but not the ΔlytS mutant) became more resistant to hNP-1 and DAP following pretreatment with carbonyl cyanide m-chlorophenylhydrazone (CCCP) (a CM-depolarizing agent). Of note, lytSR-dependent survival against CAP/HDP killing was not associated with changes in either surface positive charge, expression of mprF and dlt, or CM fluidity. The ΔlytS strain (but not the ΔlrgAB mutant) displayed a significant reduction in target tissue survival in an endocarditis model during DAP treatment. Collectively, these results suggest that the lytSR TCRS plays an important role in adaptive responses of S. aureus to CM-perturbing HDPs/CAPs, likely by functioning as a sense-response system for detecting subtle changes in ΔΨ.

Reduced vancomycin susceptibility in an in vitro catheter-related biofilm model correlates with poor therapeutic outcomes in experimental endocarditis due to methicillin-resistant Staphylococcus aureus.

Staphylococcus aureus is the most common cause of endovascular infections, including catheter sepsis and infective endocarditis (IE). Vancomycin (VAN) is the primary choice for treatment of methicillin-resistant S. aureus (MRSA) infections. However, high rates of VAN treatment failure in MRSA infections caused by VAN-susceptible strains have been increasingly reported. Biofilm-associated MRSA infections are especially prone to clinical antibiotic failure. The present studies examined potential relationships between MRSA susceptibility to VAN in biofilms in vitro and nonsusceptibility to VAN in endovascular infection in vivo. Using 10 "VAN-susceptible" MRSA bloodstream isolates previously investigated for VAN responsiveness in experimental IE, we studied the mechanism(s) of such in vivo VAN resistance, including: (i) VAN binding to MRSA organisms; (ii) the impact of VAN on biofilm formation and biofilm composition; (iii) VAN efficacy in an in vitro catheter-related biofilm model; (iv) effects on cell wall thickness. As a group, the five strains previously categorized as VAN nonresponders (non-Rsp) in the experimental IE model differed from the five responders (Rsp) in terms of lower VAN binding, increased biofilm formation, higher survival in the presence of VAN within biofilms in the presence or absence of catheters, and greater biofilm reduction upon proteinase K treatment. Interestingly, sub-MICs of VAN significantly promoted biofilm formation only in the non-Rsp isolates. Cell wall thickness was similar among all MRSA strains. These results suggest that sublethal VAN levels that induce biofilm formation and reduce efficacy of VAN in the in vitro catheter-associated biofilms may contribute to suboptimal treatment outcomes for endovascular infections caused by "VAN-susceptible" MRSA strains.

Telavancin in therapy of experimental aortic valve endocarditis in rabbits due to daptomycin-nonsusceptible methicillin-resistant Staphylococcus aureus.

A number of cases of both methicillin-susceptible Staphylococcus aureus (MSSA) and methicillin-resistant S. aureus (MRSA) strains that have developed daptomycin resistance (DAP-R) have been reported. Telavancin (TLV) is a lipoglycopeptide agent with a dual mechanism of activity (cell wall synthesis inhibition plus depolarization of the bacterial cell membrane). Five recent daptomycin-susceptible (DAP-S)/DAP-R MRSA isogenic strain pairs were evaluated for in vitro TLV susceptibility. All five DAP-R strains (DAP MICs ranging from 2 to 4 µg/ml) were susceptible to TLV (MICs of ≤0.38 µg/ml). In vitro time-kill analyses also revealed that several TLV concentrations (1-, 2-, and 4-fold MICs) caused rapid killing against the DAP-R strains. Moreover, for 3 of 5 DAP-R strains (REF2145, A215, and B(2.0)), supra-MICs of TLV were effective at preventing regrowth at 24 h of incubation. Further, the combination of TLV plus oxacillin (at 0.25× or 0.50× MIC for each agent) increased killing of DAP-R MRSA strains REF2145 and A215 at 24 h (∼2-log and 5-log reductions versus TLV and oxacillin alone, respectively). Finally, using a rabbit model of aortic valve endocarditis caused by DAP-R strain REF2145, TLV therapy produced a mean reduction of >4.5 log(10) CFU/g in vegetations, kidneys, and spleen compared to untreated or DAP-treated rabbits. Moreover, TLV-treated rabbits had a significantly higher percentage of sterile tissue cultures (87% in vegetations and 100% in kidney and spleen) than all other treatment groups (P < 0.0001). Together, these results demonstrate that TLV has potent bactericidal activity in vitro and in vivo against DAP-R MRSA isolates.

Astragalus membranaceus lectin (AML) induces caspase-dependent apoptosis in human leukemia cells.

OBJECTIVES: Recently, plant lectins have attracted great interest due to their various biological activities such as anti-cancer, anti-fungal and anti-viral activities. We have reported earlier concerning anti-proliferation of human cancer cell lines by a galactose-binding lectin (AML), from a Chinese herb, ASTRAGALUS MEMBRANACEUS: In the present study, detailed investigations into the mechanism of such anti-proliferation properties have been carried out. MATERIALS AND METHODS: Mechanism of apoptosis initiation in K562 cells by AML was investigated by morphology, flow cytometry and western blot analysis. RESULTS: AML induced apoptosis in a caspase-dependent manner in the chronic myeloid leukemia cell line, K562. Furthermore, we observed that cytotoxicity and apoptosis of K562 cells induced by AML were completely abolished in presence of lactose or galactose. CONCLUSIONS: Our results suggest that AML could act as a potential anti-cancer drug.

Relationship of agr expression and function with virulence and vancomycin treatment outcomes in experimental endocarditis due to methicillin-resistant Staphylococcus aureus.

The accessory gene regulator (agr) locus has been shown to be important for virulence in several animal models of Staphylococcus aureus infection. However, the role of agr in human infections, and specifically in antibiotic treatment, is controversial. Interestingly, agr dysfunction has been associated with reduced vancomycin responses. To systematically investigate the role of agr in virulence and treatment outcome in the context of endovascular infection, 10 well-characterized vancomycin-susceptible methicillin-resistant S. aureus (MRSA) bloodstream isolates (5 agr-I [clonal complex 45, or CC45] and 5 agr-II [CC5]) were studied for (i) agr function, (ii) RNAIII transcriptional profiles, (iii) agr locus sequences, (iv) intrinsic virulence and responses to vancomycin therapy in an experimental infective endocarditis (IE) model, and (v) in vivo RNAIII expression. Significant differences in agr function (determined by delta-hemolysin activity) correlated with the time point of RNAIII transcription (earlier RNAIII onset equals increased agr function). Unexpectedly, four MRSA strains with strong delta-hemolysin activities exhibited significant resistance to vancomycin treatment in experimental IE. In contrast, five of six MRSA strains with weak or no delta-hemolysin activity were highly susceptible to vancomycin therapy in the IE model. agr sequence analyses showed no common single-nucleotide polymorphism predictive of agr functionality. In vivo RNAIII expression in cardiac vegetations did not correlate with virulence or vancomycin treatment outcomes in the IE model. Inactivation of agr in two strains with strong delta-hemolysin activity did not affect virulence or the in vivo efficacy of vancomycin. Our findings suggest that agr dysfunction does not correlate with vancomycin treatment failures in this experimental IE model in two distinct MRSA genetic backgrounds.

Efficacy of NZ2114, a novel plectasin-derived cationic antimicrobial peptide antibiotic, in experimental endocarditis due to methicillin-resistant Staphylococcus aureus.

Cationic antimicrobial peptides (CAPs) play important roles in host immune defenses. Plectasin is a defensin-like CAP isolated from the saprophytic fungus Pseudoplectania nigrella. NZ2114 is a novel variant of plectasin with potent activity against Gram-positive bacteria. In this study, we investigated (i) the in vivo pharmacokinetic and pharmacodynamic (PK/PD) characteristics of NZ2114 and (ii) the in vivo efficacy of NZ2114 in comparison with those of two conventional antibiotics, vancomycin or daptomycin, in an experimental rabbit infective endocarditis (IE) model due to a methicillin-resistant Staphylococcus aureus (MRSA) strain (ATCC 33591). All NZ2114 regimens (5, 10, and 20 mg/kg of body weight, intravenously [i.v.], twice daily for 3 days) significantly decreased MRSA densities in cardiac vegetations, kidneys, and spleen versus those in untreated controls, except in one scenario (5 mg/kg, splenic MRSA counts). The efficacy of NZ2114 was clearly dose dependent in all target tissues. At 20 mg/kg, NZ2114 showed a significantly greater efficacy than vancomycin (P < 0.001) and an efficacy similar to that of daptomycin. Of importance, only NZ2114 (in 10- and 20-mg/kg regimens) prevented posttherapy relapse in cardiac vegetations, kidneys, and spleen, while bacterial counts in these target tissues continued to increase in vancomycin- and daptomycin-treated animals. These in vivo efficacies were equivalent and significantly correlated with three PK indices investigated: fC(max)/MIC (the maximum concentration of the free, unbound fraction of a drug in serum divided by the MIC), fAUC/MIC (where AUC is the area under the concentration-time curve), and f%T(>MIC) (%T(>MIC) is the cumulative percentage of a 24-h period that the drug concentration exceeds the MIC under steady-state pharmacokinetic conditions), as analyzed by a sigmoid maximum-effect (E(max)) model (R(2) > 0.69). The superior efficacy of NZ2114 in this MRSA IE model suggests the potential for further development of this compound for treating serious MRSA infections.

Daptomycin-oxacillin combinations in treatment of experimental endocarditis caused by daptomycin-nonsusceptible strains of methicillin-resistant Staphylococcus aureus with evolving oxacillin susceptibility (the "seesaw effect").

In vivo development of daptomycin resistance (DAPr) among Staphylococcus aureus strains, especially methicillin-resistant S. aureus (MRSA) strains, in conjunction with clinical treatment failures, has emerged as a major problem. This has raised the question of DAP-based combination regimens to enhance efficacy against such strains. We studied five recent DAP-susceptible (DAPs)/DAPr clinical MRSA strain pairs obtained from patients who failed DAP monotherapy regimens, as well as one DAPs/DAPr MRSA strain pair in which the resistant strain was generated by in vitro passage in DAP. Of note, we identified a DAP-oxacillin (OX) "seesaw" phenomenon in vitro in which development of DAPr was accompanied by a concomitant fall in OX resistance, as demonstrated by 3- to 4-fold decreases in the OX MIC, a susceptibility shift by population analyses, and enhanced early killing by OX in time-kill assays. In addition, the combination of DAP and OX exerted modest improvement in in vitro bactericidal effects. Using an experimental model of infective endocarditis and two DAPs/DAPr strain pairs, we demonstrated that (i) OX monotherapy was ineffective at clearing DAPr strains from any target tissue in this model (heart valve, kidneys, or spleen) and (ii) DAP-OX combination therapy was highly effective in DAPr strain clearances from these organs. The mechanism(s) of the seesaw effect remains to be defined but does not appear to involve excision of the staphylococcal cassette chromosome mec (SCCmec) that carries mecA.