Colistin interacts synergistically with echinocandins against Candida auris

Antifungal combination is an interesting approach for the treatment of several fungal infections but there is currently little evidence to support combined therapy in Candida auris infections. The antibacterial colistin has recently been shown to interact synergistically with antifungals against Candida spp., including azole-resistant isolates. The current study evaluated the in vitro interaction between colistin and either caspofungin or micafungin against 15 C. auris isolates by a checkerboard methodology based on the European Committee on Antimicrobial Susceptibility Testing (EUCAST) reference method. Results were analysed by two approaches: calculation of the fractional inhibitory concentration index (FICI) and response surface analysis based on the Bliss model. The minimum inhibitory concentration (MIC) range (geometric mean [Gmean]) of caspofungin and micafungin was 0.25 to 1 µg/mL (0.691 µg/mL) and 0.03 to 0.125 µg/mL (0.114 µg/mL), respectively. No activity was observed for colistin alone with MIC of >64 µg/mL for all the isolates. When colistin was combined with caspofungin, synergistic interactions were observed for all strains with FICI values of 0.08 to 0.14. In contrast, indifferent interactions were observed for the combination of colistin with micafungin with FICI values of 0.51 to 1.01. Synergy was also demonstrated using the Bliss model against all isolates for the colistin-caspofungin combination and in 60% of isolates for the colistin-micafungin combination. Antagonism was not observed for any combination.     

Colistin/daptomycin: an unconventional antimicrobial combination synergistic in vitro against multidrug-resistant Acinetobacter baumannii

The in vitro activity of the combination colistin/daptomycin was evaluated against multidrug-resistant Acinetobacter baumannii clinical isolates. Clonal relationships were assessed by pulsed-field gel electrophoresis. The following synergy studies were undertaken: (i) daptomycin MICs were determined by E-test on Mueller–Hinton agar plates supplemented with a subinhibitory concentration of colistin; and (ii) time–kill methodology using tubes containing an inoculum of 5 × 10⁵ CFU/mL and subinhibitory concentrations of each antibiotic alone or in combination subcultured at 0, 5 and 24 h for colony counting. Synergy was defined as ≥2 log₁₀ CFU/mL decrease of viable colonies compared with colistin alone. Ten colistin-susceptible and four colistin-resistant A. baumannii isolates were tested. Isolates were assigned to nine different clonal types. Enhanced in vitro activity of the combination was detected only against colistin-susceptible isolates; using plates supplemented with colistin, the daptomycin MIC was reduced by 4- to 128-fold. From a total of 30 isolate–concentration combinations in time–kill studies, a synergistic interaction was detected in 16 (53.3%). The combination exhibited synergy against 8 and 12 of these combinations at 5 h and 24 h, respectively. No antagonism was detected. Colistin alone was bactericidal against two colistin-susceptible isolates at 24 h, whereas the combination was bactericidal against 9 colistin-susceptible isolates at 24 h. Against all colistin-resistant isolates, the combination exhibited a static effect and indifference in time–kill studies. Potent in vitro synergistic interactions between colistin and daptomycin provide evidence that this unorthodox combination may be beneficial in the treatment of colistin-susceptible multidrug-resistant A. baumannii.     

Amelioration of melatonin on oxidative stress and genotoxic effects induced by cisplatin in vitro

In this study, we made an effort to evaluate the possible protective actions of melatonin on cisplatin-induced oxidative damage in mice brain homogenate and genotoxic effects in human lymphocytes under in vitro conditions. The tissue homogenate was divided into three parts. The first portion was kept as control treated with dimethyl sulphoxide (DMSO) (group 1) while the second and third portion were treated with 24 µg/g tissue cisplatin alone (group 2) and 24 µg/g tissue cisplatin in combination with 3?mM melatonin (group 3), respectively. We measured the oxidative stress biomarkers such as lipid peroxidation, 8-hydroxy 2′ deoxyguanosine (8-OHdG) and antioxidant parameters such as reduced glutathione, superoxide dismutase, glutathione peroxidase, and glutathione reductase in brain homogenate. Likewise peripheral venous blood was collected from healthy donors and human lymphocyte culture was done using karyotyping medium. Cultures were divided into three groups. Group 1 was the control i.e. lymphocytes treated with DMSO 5 µg/mL. In group 2, lymphocytes were treated with 2 µg/mL cisplatin and group 3 with a combination of 2 µg/mL cisplatin and 0.3?mM melatonin. Incubation of tissue homogenates with cisplatin elevated the malondialdehyde and 8-OHdG levels which were then reversed by melatonin. Reduction in antioxidant parameters with respect to corresponding controls were also restored by melatonin treatment. Furthermore, supplementation of melatonin was found to modulate the chromosome damage elicited by cisplatin which was determined using Giemsa (GTG) banding and karyotyping. These findings suggest that melatonin improves the cellular function and helps them to survive in the belligerent environment created by free radicals.     

Pharmacokinetics of colistin in cerebrospinal fluid after intraventricular administration alone in intracranial infections

The aim of this study was to investigate the pharmacokinetics of colistin in cerebrospinal fluid (CSF) after intraventricular (IVT) administration of colistin methanesulfonate (CMS) for central nervous system (CNS) infections caused by multidrug-resistant Gram-negative bacteria. Ten patients with CNS infection were treated with CMS (active substance colistin equivalent to 100 000 units, every 24 h) by IVT administration. After 3 days of treatment, the concentration of colistin in the CSF was determined by selective ultra-performance liquid chromatography (UPLC) at 2, 4, 6, 8, 12 and 24 h after CMS administration. A pharmacokinetic analysis was performed using Phoenix WinNonlin. Following IVT administration of CMS, the estimated colistin apparent CSF half-life (t_1/2) was 10.46 ± 6.98 h, the average peak colistin concentration (C_max) was 16.95 ± 7.39 μg/mL and the average time to peak concentration (T_max) was 4.6 ± 0.97 h. The measured trough concentration (C_min; colistin concentration in CSF at 24 h after administration of CMS) was 1.12–8.33 μg/mL and the average C_min was 2.91 ± 2.11 μg/mL. CSF concentrations of colistin were above the minimum inhibitory concentration (MIC) of 0.5 μg/mL at 24 h after IVT administration in all patients. Microbiological cure was observed in all patients. In conclusion, this is the first study of colistin pharmacokinetics in CSF after IVT administration alone in patients with CNS infection. It provides essential data for designing relatively safe and effective CMS dosing regimens.     

Characterisation of a Staphylococcus aureus strain with progressive loss of susceptibility to vancomycin and daptomycin during therapy

Following an initial response to vancomycin therapy, a patient with meticillin-resistant Staphylococcus aureus (MRSA) bacteraemia developed endocarditis, failed a second course of vancomycin and then failed daptomycin therapy. An increase in the vancomycin minimum inhibitory concentrations of four consecutive MRSA blood isolates from 2 μg/mL to 8 μg/mL was shown by Etest. Population analysis of four successive blood culture isolates recovered over the 10-week period showed that the MRSA strain became progressively less susceptible to both vancomycin and daptomycin. Retrospectively, the macro Etest method using teicoplanin indicated a decrease in vancomycin susceptibility in the second blood isolate. The patient improved after treatment with various courses of trimethoprim/sulfamethoxazole, quinupristin/dalfopristin and linezolid. Early detection of vancomycin-heteroresistant S. aureus isolates, which appeared to have clinical significance in this case, continues to be a challenge for the clinical laboratory. Development of suitable practical methods for this should be given priority. Concurrent development of resistance to vancomycin and daptomycin, whilst rare, must be considered in a patient who is unresponsive to daptomycin following vancomycin therapy.     

Quantitative determination of colistin A/B and colistin methanesulfonate in biological samples using hydrophilic interaction chromatography tandem mass spectrometry

Colistin (polymyxin E) is a polycation antibiotic which is increasingly used (administered as colistin methanesulfonate, CMS) as a salvage therapy in critically ill patients with multidrug resistant Gram‐negative infections. Even though colistin has been used for more than 50 years, its metabolic fate is poorly understood. One of the current challenges for studying the pharmacokinetics (PK) is the precise and accurate determination of colistin in in vitro and in vivo studies. In the present study, we developed and validated a series of sensitive and robust liquid chromatography tandem mass spectrometry (LC–MS/MS) methods for analysing biological samples obtained from in vitro and in vivo disposition assays. After a zinc acetate‐mediated precipitation, hydrophilic–lipophilic‐balanced solid phase extraction (HLB‐SPE) was used for the extraction of colistin. The compounds were retained on a hydrophilic interaction liquid chromatography (HILIC) column and were detected by MS/MS. CMS was quantified by determining the produced amount of colistin during acidic hydrolysis. The developed methods are sensitive with lower limits of quantification varying between 0.009 μg/mL and 0.071 μg/mL for colistin A, and 0.002 μg/mL to 0.013 μg/mL for colistin B. The intra‐ and inter‐day precision and accuracy were within ±15%. Calibration curves of colistin were linear (0.063 μg/mL to 8.00 μg/mL) within clinically relevant concentration ranges. Zinc acetate‐mediated precipitation and the use of a HILIC column were found to be essential. The developed methods are sensitive, accurate, precise, highly efficient and allow monitoring colistin and CMS in biological samples without the need for an internal standard.     

Efficacies of colistin and tigecycline in mice with experimental pneumonia due to NDM-1-producing strains of Klebsiella pneumoniae and Escherichia coli

New Delhi metallo-β-lactamase-1 (NDM-1)-producing Enterobacteriaceae have emerged as a global threat. The aim of this study was to assess the efficacies of colistin and tigecycline in an experimental model of pneumonia caused by NDM-1-producing Escherichia coli and Klebsiella pneumoniae. The susceptibilities of K. pneumoniae NDM, E. coli NDM and K. pneumoniae ATCC 29665 were determined using the broth microdilution technique. The pharmacokinetics of colistin and tigecycline in an experimental model of pneumonia were performed using immunocompetent C57BL/6 mice. Mice were treated with colistin (60 mg/kg/day) or tigecycline (10 mg/kg/day). Mortality, bacteraemia and lung bacterial concentrations were recorded. The strains were susceptible to colistin and tigecycline. The ratio of area under the concentration-time curve/minimum inhibitory concentration (AUC/MIC) for colistin was 158.5 (all three strains) and that for tigecycline was 18.5 (K. pneumoniae NDM) and 37 (K. pneumoniae ATCC 29665 and E. coli NDM). In vivo, colistin decreased bacterial lung concentrations of K. pneumoniae NDM and K. pneumoniae ATCC 29665 by 1.16 log colony-forming units (CFU)/g and 2.23 logCFU/g, respectively, compared with controls (not significant). Tigecycline reduced K. pneumoniae NDM and K. pneumoniae ATCC 29665 load by 2.67 logCFU/g and 4.62 logCFU/g (P<0.05). Colistin and tigecycline decreased lung concentrations of E. coli NDM by 2.27 logCFU/g and 4.15 logCFU/g (P<0.05), respectively, compared with controls, and was more active than colistin (P<0.05). In conclusion, these results suggest that colistin is inappropriate for treating pneumonia due to NDM-1-producing K. pneumoniae and its efficacy was suboptimal against NDM-1-producing E. coli. A high tigecycline dose was efficacious for treating experimental pneumonia due to NDM-1-producing E. coli and K. pneumoniae.     

妇安消疹洗液对特应性皮炎模型小鼠的疗效实验

目的探讨妇安消疹洗液对特应性皮炎模型小鼠的疗效。方法将48只小鼠分为正常组(A组),模型组(B组),对照组(C组,卤米松乳膏),妇安消疹洗液低、中、高剂量组(D_1组、D_2组、D_3组,生药0.334,0.672,1.344 g/mL),各8只。采用二硝基氯苯丙酮溶液建立特应性皮炎小鼠模型。A组、B组小鼠患处涂抹生理盐水,C组小鼠患处涂抹卤米松乳膏,D_1组、D_2组、D_3组小鼠患处分别涂抹不同浓度的妇安消疹洗液,每日1次,给药7 d。结果与A组比较,B组小鼠血清干扰素γ(IFN-γ)水平显著降低,白细胞介素4(IL-4)、白细胞介素5(IL-5)、免疫球蛋白E(IgE)水平、耳肿胀度、胸腺指数、脾脏指数、皮损厚度均显著升高(P<0.05);与B组比较,D_2组、D_3组小鼠表皮剥脱、出血、渗出、红斑、结痂变浅显著更优,血清IFN-γ水平显著升高,IL-4,IL-5,IgE水平均显著降低耳肿胀度、胸腺指数、脾脏指数、皮损厚度均显著降低(P <0.05)。结论妇安消疹洗液具有消炎、抗变态反应的作用,对特应性皮炎模型小鼠疗效显著。     

Effects of silver nanoparticles on rat hepatic cytochrome P450 enzyme activity

Silver nanoparticles (AgNPs) are increasingly used in various products and consequentially the potential adverse effects associated with exposure to them are of concern. This study investigated the effects of AgNPs on the hepatic drug-metabolizing enzymes of the cytochrome P450 (CYP) families 1, 2 and 3, using both in vitro and in vivo biological assays. AgNPs were orally administered to Sprague-Dawley rats at various concentrations (0–1000?mg/kg body weight/day) for 2 weeks. No effect was found on the plasma levels of ALT, AST and ALP in all treated rat groups, and no significant change in the activities of CYP1A, CYP2C, CYP2D, CYP2E1 and CYP3A was observed for all tested AgNP doses. The results correlated with the observation that no AgNPs were detected in the liver sections of the tested rats. However, the in vitro system using rat liver microsomes demonstrated a strong inhibition of CYP2C (IC₅₀?=?28 µg/mL) and CYP2D (IC₅₀?=?23 µg/mL) activities, but not of CYP1A, CYP2E1 and CYP3A activities (IC₅₀ > 100 µg/mL) at concentrations up to 100 µg/mL of AgNPs. The inhibitory effect of AgNPs on these CYPs indicates the possibility of the AgNP-drug interaction when co-administered with some medicines and this may cause adverse effects to patients.     

Anti-Candida albicans effectiveness of citral and investigation of mode of action

Context: Candidiasis is a mycosis caused by Candida species, which is of clinical importance due to the increase in resistant yeasts. Candida infection has been a serious health problem due to the inappropriate use of antibiotics. Therefore, it is necessary to study molecules with an antifungal action. Citral is a monoterpene with known pharmacological properties, including antimicrobial action.

Objective: The aim of this work was to determine the minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) of citral and the probable mode of action.

Materials and methods: The MIC of citral was determined by the broth microdilution method using Sabouraud dextrose medium. Additionally, the interference of citral in cell wall (sorbitol assay) and the binding of citral to ergosterol and cholesterol were studied, carried out by broth microdilution method.

Results: The MIC and MFC of citral were 512 and 1024 µg/mL, respectively. The MIC of amphotericin B was 1 µg/mL. The mechanism of action did not involve either the cell wall or ergosterol. However, the presence of cholesterol increased the MIC of citral to 1024 µg/mL, indicating there is some interaction between citral and cholesterol. Amphotericin B was used as the positive control, and it showed a high MIC in the presence of ergosterol (32 µg/mL), while in the presence of cholesterol MIC increased to 4 µg/mL.

Conclusion: Citral inhibits the growth of C. albicans. The probable mechanism of action did not involve the cell wall or ergosterol. Citral is able to interact with cholesterol. More studies are necessary to describe their effects completely.     

Antioxidant and cytotoxic potential of a new thienyl derivative from Tagetes erecta roots

Context: The search for newer compounds against pathogenic species continues unabated due to drug resistance. Traditionally, Tagetes erecta Linn. (Compositae) has been used for the treatment of various parasitic and microbial diseases.

Objective: To evaluate the antioxidant activity of the ethanol extract of Tagetes erecta roots and its cytotoxicity against prostate and HeLa cancer cell lines followed by activity-guided isolation.

Materials and Methods: The antioxidant screening was carried out using diphenylpicrylhydrazyl (DPPH) radical scavenging assay with serial concentrations ranging from 2 to 100 µg/mL, and cytotoxicity was evaluated against prostate (PC-3) and HeLa cell lines using microculture tetrazolium test (MTT) assay with concentrations ranging from 500 to 1.89 µg/mL. Isolation of the ethanol extract was carried out using column chromatography whereby 21 isolates were obtained (T₁-T₂₁), and the most active isolate was subjected for characterization using ultraviolet (UV), infrared (IR), nuclear magnetic resonance (NMR), and mass spectroscopic techniques.

Results: The ethanol extract scavenged DPPH free radicals thereby exhibiting antioxidant activity with an IC₅₀ of 35.9 µg/mL. In addition, the extract conferred noticeable cytotoxicity against the HeLa (LD₅₀ of 164.28 µg/mL) and PC-3 cell lines (LD₅₀ of 407.3 µg/mL). Among all the isolates, T₃ showed antioxidant activity with IC₅₀ of 11.56 µg/mL and cytotoxicity with LD₅₀ of 12.5 µg/mL against HeLa and 30.25 µg/mL against PC-3 cell lines and was characterized as 2-ethynyl-5-(thiophen-2-yl) thiophene.

Discussion: The new thienyl compound (T₃) exhibited profound antioxidant activity and cytotoxicity at relatively lower concentrations than the extract.

Conclusion: The observations provide support for the ethnobotanical use of the plant.     

Novel anticancer alkene lactone from Persea americana

Abstract

Context: Persea americana Mill (Lauraceae) root bark is used in ethnomedicine for a variety of diseases including cancer.

Objective: To isolate and characterize the chemical constituent in P. americana, and also to determine the anticancer property of a new alkene lactone from the root bark of P. americana.

Materials and methods: The MCF-7 cells were treated with different concentrations of the pure compound for 48?h. The percentage of cells in the various phases, online monitoring of metabolic changes and integrin receptor expression determined by flow cytometry.

Results: One novel alkene lactone (4-hydroxy-5-methylene-3-undecyclidenedihydrofuran-2 (3H)-one) (1) was isolated and characterized using 1D-NMR, 2D-NMR, infrared, UV and MS. At a concentration of 10?µg/mL, significant reduction of proliferation of MCF-7 was induced while MCF-12?A cell was significantly stimulated by 10?µg/mL. The IC₅₀ value for MCF-7 cells is 20.48?µg/mL. Lower concentration of 1 harbor no significant effect on either MCF-7 or MCF-12A. The apoptotic rates of MCF-7 cells were increased significantly. At the final concentration 10?µg/mL, up to 80% of all breast cancer cells were dead. On the non-tumorigenic cell line MCF-12A, the same concentrations (1 and 10?µg/mL) of compound 1 caused significant enhanced apoptotic rates. A total of 1?µg/mL of 1 caused a decrease of α4-, α6-, β1- and β3-integrin expression.

Conclusions: The compound caused a stimulatory effect on non-tumorigenic MCF-12A cells with respect to cell adhesion while tumorigenic MCF-7 cells detached continuously. This is the first report on the anticancer effects of this class of compound.     

Colistin offers prolonged survival in experimental infection by multidrug-resistant Acinetobacter baumannii: the significance of co-administration of rifampicin

The effect of colistin on bacterial eradication and survival was tested in experimental infection by multidrug-resistant Acinetobacter baumannii. The thigh infection model was applied in 86 neutropenic Wistar rats. Six rats were used for the induction of neutropenia and for the selection of the dose regimen of colistin; the remainder was equally divided into four groups: A, controls; B, rifampicin; C, colistin; and D, both agents. Therapy was administered 5 h after bacterial challenge; 5mg/kg of rifampicin was administered intravenously and 3mg/kg of colistin intramuscularly. Survival was recorded in 10 animals of each group. The remaining 10 rats per group were killed 4h after therapy; blood and tissue samples were sampled. Median survival of animals of groups A, B, C and D was 2.00, 2.50, 4.00 and 4.00 days, respectively (P=0.0048 between A and C and P=0.0012 between A and D. Mortality rates after 6 days of follow-up were 100, 100, 100 and 70%, respectively (P=0.018 between groups). Statistically significant decreases of bacteria were found in blood, liver, lung and spleen of group B compared with A; in lung of group C compared with A; and in blood and liver of group D compared with A. Colistin was effective in prolonging survival in an experimental thigh infection by multidrug-resistant A. baumannii in neutropenic rats. Its activity was enhanced after co-administration with rifampicin. These results mandate the application of colistin in the event of infections by multidrug-resistant pathogens and the need for its co-administration with rifampicin.     

Wnt通路对胰岛素样生长因子-1抑制小鼠毛细胞凋亡的作用

目的探讨胰岛素样生长因子-1(IGF-1)抑制小鼠毛细胞凋亡作用的调控机制。方法体外培养新生小鼠耳蜗基底膜。在分别含有正常对照培养液(A组)、0.5 mmol/L庆大霉素(GM)(B组)、0.5 mmol/L GM+1 ng/mL IGF(C组)、0.5 mmol/L GM+1 ng/mL IGF+5μg/mL wif(D组)、0.5 mmol/L GM+1 ng/mL IGF+10μg/mLwif(E组)、0.5 mmol/L GM+1 ng/mL IGF+15μg/mL wif(F组)的成分中培养24 h后收集细胞及固定。应用TUNEL法观察各组耳蜗毛细胞的凋亡情况。应用Western blot及Real-time PCR法观察Caspase-3的表达情况。结果 B组耳蜗毛细胞经刺激后,细胞凋亡数量及Caspase-3 mRNA和蛋白表达较A组明显上升(P<0.01),C组耳蜗毛细胞经IGF刺激后,细胞凋亡数量及Caspase-3mRNA和蛋白表达较B组明显下降(P<0.01),应用Wnt阻断剂(wif)后D、E及F组细胞凋亡数量及Caspase-3 mRNA和蛋白表达较C组升高,其中F组升高显著(P<0.01)。结论 IGF可能通过激活Wnt通路抑制毛细胞凋亡。     

水合氯醛在C57BIM6小鼠麻醉中的应用研究

目的探索不同浓度的水合氯醛对C57BL／6小鼠的麻醉效果。方法将60只C57BL／6小鼠随机分为6组,A（2．5％、0．06mL／10g）、B（2．5％、0．1mL／10g）、C（5％、0．06mL／10g）、D（5％、0．1mL／10g）、E（10％、0．06mL/10g）、F（10％、0．1mL／10g）,分别进行腹腔注射水合氯醛麻醉实验,观察临床表现、诱导时间反维持时间。结果B、c、D和E组取得不同的麻醉效果,分别在3．34±0．48min、14．93±3．59rain、4．72±3．02min、9．20±1．51min进入麻醉状态,维持时间分别为45．74±1．94rain、44．18±8．64rain、179．25±3．99min、100．05±22．33rain。结论根据不同实验需要,选择水合氯醛恰当的浓度与剂量行腹腔注射麻醉,是安全有效的。     

Antifungal activity of ribavirin used alone or in combination with fluconazole against Candida albicans is mediated by reduced virulence

The incidence of fungal infections has increased continuously in recent years, and drug resistance, especially resistance to fluconazole (FLC), has emerged. To overcome this challenge, research on the antifungal activities of non-antifungal agents has gained more attention. In this study, we determined the anti-Candida activity of ribavirin (RBV), an antiviral drug commonly used in the clinic, and found that RBV displayed potent antifungal activity when used alone or in combination with FLC in vitro and in vivo. In vitro, the MIC₈₀ values of RBV were 2–4 µg/mL for FLC-susceptible Candida albicans and 8 µg/mL for FLC-resistant C. albicans. When RBV at a dose of 1 µg/mL was combined with FLC, significant synergistic effects were exhibited against FLC-resistant C. albicans, and the MICs of FLC decreased from >512 µg/mL to 0.25–1 µg/mL. Synergism was also exhibited against C. albicans biofilms. In vivo, RBV plus FLC significantly improved the survival of infected Galleria mellonella larvae compared with the FLC-treated group over a 4-day period and attenuated the damage of FLC-resistant C. albicans to G. mellonella larvae tissue. Furthermore, mechanistic studies indicated that the antifungal effects of RBV used alone or in combination with FLC might be associated with inhibition of biofilm formation, reduced extracellular phospholipase activity and inhibition of hyphal growth, but is not related to promotion of FLC uptake and inhibition of FLC efflux. These results provide a promising direction for overcoming drug resistance and for expanding the clinical application of existing drugs.     

Genotoxic effects of chlorophenoxy herbicide diclofop-methyl in mice in vivo and in human lymphocytes in vitro

Diclofop-methyl (DM) is a chlorophenoxy derivative used in large quantities for the control of annual grasses in grain and vegetable crops. In this study, the genotoxic effects of DM were investigated by measuring chromosomal aberrations (CAs) in mouse bone-marrow cells and CA and the comet assay in human peripheral lymphocytes. Mice were treated with 15.63, 31.25, 62.5, and 125?mg/kg body weight of DM intraperitoneally for 24 hours, and 15.63-, 31.25-, 62.5-, 125-, and 250-µg/mL concentrations were applied to human lymphocytes for both 24 and 48 hours. In in vivo treatments, DM significantly, but not dose dependently, increased the total chromosome aberrations, compared to both negative and solvent controls. Cell proliferation was significantly, but not dose dependently, affected by all doses. In in vitro treatments, DM (except 15.63 µg/mL) significantly and dose dependently increased the frequency of chromosome aberrations. Also, 250 µg/mL of 48-hour treatment was found to be toxic. Cell proliferation was significantly and dose dependently affected by DM applications, when compared to negative control. In in vitro treatments, DM significantly decreased the mitotic index only at the highest concentration for 24 hours, and 62.5- and 125-µg/mL concentrations for 48 hours. In the comet assay, a significant and dose-dependent increase in comet-tail intensity was observed at 62.5-, 125-, and 250-µg/mL concentrations. The mean comet-tail length was significantly increased in all concentrations. Our results demonstrate that DM is genotoxic in mammalian cells in vivo and in vitro.     

The α-glucosidase inhibitory activity of avicularin and 4-O-methyl gallic acid isolated from Syzygium myrtifolium leaves

Diabetes Mellitus is the main cause of death on a global scale. In 2019, there were 463 million people with diabetes, and WHO predicts that by 2030, there will be 578 million. As an antidiabetic agent, α-glucosidase inhibitors are one of the methods employed to reduce the prevalence of diabetes. Diabetes is traditionally treated with Syzygium as a primary material, medicine, fruit, ornamental plant, and source of carpentry. This investigation aimed to examine the inhibitory effect of seven species of Syzygium against α-glucosidase enzyme using an in vitro assay and isolate active substances and ascertain their concentrations in each sample. As a solvent, ethanol was used in maceration to extract the substance. Afterward, the extract underwent a series of fractionation techniques, including liquid–liquid extraction, vacuum liquid chromatography, column chromatography, and preparative Thin Layer Chromatography (TLC) for purification and isolation. The compound's structures were elucidated using TLC, UV–Visible spectrophotometry, and nuclear magnetic resonance (NMR) spectroscopy. Based on concentrations of 100 and 200 µg/mL, Syzygium myrtifolium exhibited the most significant inhibitory effect, followed by other species of Syzygium. The proportion of ethyl acetate had the strongest activity (IC₅₀ 0.40 ± 0.02 µg/mL) contrasted to positive control acarbose (IC₅₀ 55.39 ± 0.67 g/mL) and quercitrin (IC₅₀ 6.47 ± 0.40 µg/mL). Avicularin and 4-O-methyl gallic acid were discovered in the ethyl acetate fraction of Syzygium myrtifolium with IC₅₀ values of 17.05 ± 0.75 µg/mL and 25.19 ± 0.21 µg/mL, respectively. As α-glucosidase inhibitory, the results of this study indicate Syzygium myrtifolium can be used as a dietary supplement to manage hyperglycemia.     

The effects of amphotericin B on angiogenesis in chick chorioallantoic membrane

Objective: Amphotericin B (AmB) is widely used as a mainstay in the treatment of sight-threatening fungal endophthalmitis. From the time that itraconazole was discovered to have a previously unknown anti-angiogenic activity, we have suspected that AmB may have possible effects on ocular angiogenesis. The purpose of this study was to evaluate the in vivo anti-angiogenic effect of AmB in the chick chorioallantoic membrane (CAM) model.

Materials and methods: Atak-S type fertilized eggs obtained from the Poultry Institution were used. The eggs were kept under 37?°C at 85–90% relative humidity throughout the experiment. Amphotericin B was prepared in two different concentrations (AmB 125?μg/1?mL and 0.125?μg/1?mL). The CAMs treated with sterile distilled water was specified as controls. About 0.1?mL of each containing 12.5 and 0.0125?µg of AmB, respectively, were dropped to CAM surface. Thirteen eggs were used for each group. The results were evaluated at the 48th hour of the administration of the drugs and recorded by digital camera.

Results: A reduction of angiogenesis in CAM area which treated with 125?μg/1?mL of AmB was appreciable macroscopically. The affected areas showed impaired radial arrangement of small vessels with the presence of avascular zone at periphery. The dose of 0.125?μg/1?mL AmB did not show any visible anti-angiogenic effect. Numerous blood vessels with a radially arranged pattern developed toward the periphery after 48?h of treatment. In the CAMs that treated with distilled water, physiological angiogenesis was observed in allantoic vessels. Vessel formation seems to be similar in CAMs treated with 0.125?μg/1?mL AmB with the presence of visibly non-malformed alive embryos.

Conclusions: The present study gives the impression that AmB has the capacity to serve as an anti-angiogenic treatment. As it is a preliminary CAM study only, further studies on both animals and humans are required.     

1，25-二羟维生素D3减轻严重烫伤小鼠应激反应的作用

目的：探讨1,25-二羟维生素D3对严重烫伤小鼠应激状态下胰岛素抵抗和炎症反应的影响。方法： 130只小鼠随机分为：健康组10只,实验组Ⅰ、实验组Ⅱ、对照组各40只。实验组Ⅰ、实验组Ⅱ和对照组小鼠制成30%全身体表面积（TBSA）Ⅲ°烫伤模型。每隔1日晨8时,实验组Ⅰ小鼠1,25-(0H)2VitD3 1μg?kg-1+0.6mL花生油灌胃;实验组Ⅱ小鼠1,25-(0H)2VitD3 4μg?kg-1+0.6mL花生油灌胃;对照组小鼠0.6mL花生油灌胃。分别于伤后1、3、7、14 天测空腹血糖（FBG）、空腹胰岛素（FIns）、血清TNF-α浓度和创面组织NF-κB阳性率。结果：（1）实验组Ⅰ、实验组Ⅱ和对照组各时相点胰岛素抵抗指数（HOMA-IR）、血清TNF-α含量和创面组织NF-κB表达阳性率均数均高于健康组水平。（2）实验组Ⅰ和实验组Ⅱ小鼠伤后同一时相点胰岛素抵抗指数（HOMA-IR）均数均低于对照组水平,且实验组Ⅱ低于实验组Ⅰ（P<0.05）;同一组内不同时相点实验小鼠伤后第3天HOMA-IR均数最高,以后逐渐降低（P<0.05）。（3）实验组Ⅰ和实验组Ⅱ小鼠伤后同一时相点血清TNF-α含量、创面组织NF-κB表达阳性率均数均低于对照组水平,且实验组Ⅱ低于实验组Ⅰ（P<0.05）;同一组内不同时相点血清TNF-α含量、创面组织NF-κB表达阳性率均数均逐渐降低（P<0.05）。结论：1,25-二羟维生素D3能够减轻严重烫伤小鼠应激状态下胰岛素抵抗作用和炎症反应。