RESUMEN
Our research aimed to expand the knowledge of relationships between the structure of multifunctional cationic dicephalic surfactants with a labile linker-N,N-bis[3,3-(dimethylamine)propyl]alkylamide dihydrochlorides and N,N-bis[3,3-(trimethylammonio)propyl]alkylamide dibromides (alkyl: n-C9H19, n-C11H23, n-C13H27, n-C15H31)-and their possible mechanism of action on fungal cells using the model organism Saccharomyces cerevisiae. General studies performed on surfactants suggest that in most cases, their main mechanism of action is based on perforation of the cell membranes and cell disruption. Experiments carried out in this work with cationic dicephalic surfactants seem to modify our understanding of this issue. It was found that the investigated compounds did not cause perforation of the cell membrane and could only interact with it, increasing its permeability. The surfactants tested can probably penetrate inside the cells, causing numerous morphological changes, and contribute to disorders in the lipid metabolism of the cell resulting in the formation of lipid droplet aggregates. This research also showed that the compounds cause severe oxidative stress within the cells studied, including increased production of superoxide anion radicals and mitochondrial oxidative stress. Dicephalic cationic surfactants due to their biodegradability do not accumulate in the environment and in the future may be used as effective antifungal compounds in industry as well as medicine, which will be environmentally friendly. KEY POINTS: ⢠Dicephalic cationic surfactants do not induce disruption of the cell membrane. ⢠Surfactants could infiltrate into the cells and cause accumulation of lipids. ⢠Surfactants could cause acute oxidative stress in yeast cells. ⢠Compounds present multimodal mechanism of action. Graphical abstract.
Asunto(s)
Amidas , Antifúngicos , Antifúngicos/farmacología , Cationes , Membrana Celular , TensoactivosRESUMEN
Quorum sensing (QS) is a mechanism that enables microbial communication. It is based on the constant secretion of signaling molecules to the environment. The main role of QS is the regulation of vital processes in the cell such as virulence factor production or biofilm formation. Due to still growing bacterial resistance to antibiotics that have been overused, it is necessary to search for alternative antimicrobial therapies. One of them is quorum quenching (QQ) that disrupts microbial communication. QQ-driving molecules can decrease or even completely inhibit the production of virulence factors (including biofilm formation). There are few QQ strategies that comprise the use of the structural analogues of QS receptor autoinductors (AI). They may be found in nature or be designed and synthesized via chemical engineering. Many of the characterized QQ molecules are enzymes with the ability to degrade signaling molecules. They can also impede cellular signaling cascades. There are different techniques used for testing QS/QQ, including chromatography-mass spectroscopy, bioluminescence, chemiluminescence, fluorescence, electrochemistry, and colorimetry. They all enable qualitative and quantitative measurements of QS/QQ molecules. This article gathers the information about the mechanisms of QS and QQ, and their effect on microbial biofilm formation. Basic methods used to study QS/QQ, as well as the medical and biotechnological applications of QQ, are also described. Basis research methods are also described as well as medical and biotechnological application.
Asunto(s)
Biopelículas , Percepción de Quorum , Antibacterianos/farmacología , Bacterias/efectos de los fármacos , Bacterias/genética , Fenómenos Fisiológicos Bacterianos/efectos de los fármacos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Biopelículas/efectos de los fármacos , Percepción de Quorum/efectos de los fármacos , Factores de Virulencia/genética , Factores de Virulencia/metabolismoRESUMEN
Quaternary ammonium salts (QASs) are commonly used in medicine, agriculture and industry and their wide usage caused the development of microbial resistance, thus there is still a need for new effective antimicrobial agents. Present work describes the biological activity of alanine- (DMALM-n) and glycine-derived (DMGM-n) QASs against planktonic and biofilm forms of micro-organisms. The antimicrobial activity was dependent mainly on the hydrocarbon chain length and surfactants with 12-16 atoms of carbon in the alkyl chain were the most active ones. The lowest MIC value was determined for DMALM-14 against Rhodotorula rubra and Saccharomyces cerevisiae (2·5 µmol l- 1 ). Generally, alanine derivatives showed stronger effects against micro-organisms than glycine-derived QASs. Alanine-derived surfactants with 12-16 carbons in the alkyl chain had antiadhesive properties on the polystyrene surface, preventing cell attachment (about 70% of inhibition for C. albicans and 40% for S. epidermidis). Strong adhesion reduction was also observed on the stainless steel surface and the highest reduction was observed for C. albicans cells incubated on surface pretreated with DMGM-16. Moreover, DMGM-16 and DMALM-16 prevented C. albicans filamentation, one of the determinants of cell adhesion. Surfactants with C16 alkyl chain (DMGM-16 and DMALM-16) eradicated bacterial and yeast biofilm (from 60 to 90% of reduction observed after incubation of the previously grown biofilm in the presence of the highest tested concentration of the surfactant - 400 µmol l- 1 ) and reduced its viability. Strong antimicrobial activity as well as antiadhesive properties make alanine- and glycine-derived QASs the potential candidates for future application as disinfectants. SIGNIFICANCE AND IMPACT OF THE STUDY: Cationic surfactants are used in many fields, among others in medicine, cosmetic and pharmaceutical industry. Their usage on a large scale caused the development of microbial resistance mechanisms to such compounds. Thus, there is a need to synthesize new surfactants with potential application as effective disinfectants to combat both planktonic and biofilm forms of micro-organisms. Present work focuses on the antimicrobial activity of chosen quaternary ammonium salts.
Asunto(s)
Alanina/análogos & derivados , Antiinfecciosos/farmacología , Biopelículas/efectos de los fármacos , Glicina/análogos & derivados , Compuestos de Amonio Cuaternario/farmacología , Tensoactivos/farmacología , Candida albicans/efectos de los fármacos , Desinfectantes/farmacología , Pruebas de Sensibilidad Microbiana , Poliestirenos , Rhodotorula/efectos de los fármacos , Sales (Química)/farmacología , Acero Inoxidable , Staphylococcus epidermidis/efectos de los fármacosRESUMEN
The studies were aimed to contribute to the elucidation of the relationships between structure of the double-headed cationic surfactants - N,N-bis[3,3'-(dimethylamine)propyl]alkylamide dihydrochlorides and N,N-bis[3,3'-(trimethylammonio)propyl]alkylamide dibromides (alkyl: n-C9H19, n-C11H23, n-C13H27, n-C15H31), which are of particular interest, as they contain a labile amide group in the molecule and their antifungal activity. Therefore, the minimal inhibitory and fungicidal concentrations (MIC and MFC) of dicephalic surfactants against various fungi were tested using standardized methods. Most of the tested fungi were resistant to the Cn(TAPABr)2 compounds. The strongest growth inhibition was caused by Cn(DAPACl)2 series, which MICs ranged from 6.5 to 16⯵M. The influence of dicephalic surfactants on Candida albicans biofilm and adhesion to the various surfaces was investigated with crystal violet staining or colony counting. The reduction of fungal adhesion was also observed, especially to the glass surface. One of the compounds (C14(DAPACl)2) caused DNA leakage from C. albicans cells. Further studies showed the impact of dicephalic surfactants on ROS production, accumulation of lipid droplets and filament formation. This study points to the possibility of application of dicephalic surfactants as the surface-coating agents to prevent biofilm formation or as disinfectants. The results give an insight into the possible mechanism of action of newly synthesized dicephalic surfactants in yeast cells.
Asunto(s)
Antifúngicos/farmacología , Tensoactivos/farmacología , Biopelículas/efectos de los fármacos , Candida albicans/citología , Candida albicans/efectos de los fármacos , Cationes , Adhesión Celular/efectos de los fármacos , Gotas Lipídicas/metabolismo , Pruebas de Sensibilidad Microbiana , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Controlling relative daughter cell size is key during cytokinesis. Uncontrolled size asymmetries can lead to aneuploidy and division failure. At the same time, precisely regulated size asymmetries are of crucial importance in many divisions during embryonic development. Therefore, being able to monitor daughter cell size is important in cytokinesis studies. However, freely available tools allowing to effectively measure the size of daughter cells in three dimensions during cytokinesis are missing. Here, we describe an open-access plugin for ImageJ or Fiji based on an active contour surface representation of the cells. Our method provides a user-friendly and accurate way to monitor the size of the two daughter cells throughout cytokinesis.
Asunto(s)
Rastreo Celular/métodos , Citocinesis/genética , Imagen Molecular/métodos , Algoritmos , Tamaño de la Célula , Conformación MolecularRESUMEN
AIMS: The studies were aimed to contribute to the elucidation of the relationships between structure of the double-headed cationic surfactants-N,N-bis[3,3'-(dimethylamine)- propyl]alkylamide dihydrochlorides and N,N-bis[3,3'-(trimethylammonio)propyl]alkylamide dibromides (alkyl: n-C9 H19 , n-C11 H23 , n-C13 H27 , n-C15 H31 ) and their antibacterial and biofilm preventing activity. METHODS AND RESULTS: The minimal inhibitory and bactericidal concentrations (MIC and MBC) of dicephalic surfactants against Staphylococcus epidermidis and Pseudomonas aeruginosa were tested using standard methods. Pseudomonas aeruginosa was resistant to studied compounds but MBC values against Staph. epidermidis reached 0·48-0·01 mmol l(-1) . The influence of dicephalic surfactants on bacterial biofilm and adhesion to the various surfaces was investigated with crystal violet staining or colony counting. The reduction in bacterial adhesion was observed, especially in the case of glass and stainless steel. The condensation of the DNA was shown in the ethidium bromide intercalation assay. CONCLUSIONS: Dicephalic surfactants exhibited antibacterial activity against Staph. epidermidis. The activity of studied compounds depended on the hydrocarbon chain length and the counterion. Surfactants deposited on different materials reduced Staph. epidermidis adhesion, dependently on the surfactant structure and the substratum. Dicephalic surfactants showed the ability of DNA compaction. SIGNIFICANCE AND IMPACT OF THE STUDY: This study points the possibility of application of dicephalic surfactants as the surface-coating agents to prevent biofilm formation. These compounds efficiently condensed DNA and are potential candidates for further studies towards the transfection.
Asunto(s)
Amidas/farmacología , Biopelículas/efectos de los fármacos , Pseudomonas aeruginosa/efectos de los fármacos , Compuestos de Amonio Cuaternario/farmacología , Staphylococcus epidermidis/efectos de los fármacos , Tensoactivos/farmacología , Amidas/química , Antibacterianos/farmacología , Adhesión Bacteriana/efectos de los fármacos , ADN Bacteriano/química , Vidrio , Conformación de Ácido Nucleico/efectos de los fármacos , Poliestirenos , Pseudomonas aeruginosa/fisiología , Compuestos de Amonio Cuaternario/química , Siliconas , Acero Inoxidable , Infecciones Estafilocócicas/microbiología , Staphylococcus epidermidis/fisiología , Factores de Virulencia/metabolismoRESUMEN
Heat shock proteins (Hsp) are the group of proteins observed in both prokaryotic and eukaryotic cell types. Hsp synthesis takes place in response to many environmental conditions, including ultraviolet radiation, heavy metal ions, hypoxia and toxic agents. Many authors have suggested that Hsp can be used in immunoprophylaxis, yet Hsp70 proteins expressed in bovine leukocytes have not been fully characterized. Hence the aim of this study was to evaluate the expression of Hsp70 proteins in bovine leukocytes exposed to temp. 41degrees C. The material for the study consisted of bovine white blood cells incubated at 41 degrees C for 2 hours. SDS-Page electrophoresis, Western blotting, and two-dimensional electrophoresis (2D) were performed to estimate the proteins obtained. The results of the study confirmed the influence of the temperature of 41 degrees C on induction of Hsp70 in bovine leukocytes. These proteins were mainly localized within molecular mass 70kDa. Some of the proteins with molecular mass from 20 to 50 kDa also showed positive reactions in Western blotting with anti-Hsp70 antibodies. Analysis of 2D electrophoresis showed a change in the localization of these proteins in the pH gradient. It can be postulated that analysis of Hsp70 expression in bovine leukocytes can be a very helpful marker for evaluating an organism's adaptation to environmental heat stress. The proteins obtained also showed immunological reactivity with rabbit antibodies in Western blotting reactions, indicating that they can be used as protective factors in the pathogenesis of many diseases.
Asunto(s)
Proteínas HSP70 de Choque Térmico/metabolismo , Leucocitos/metabolismo , Temperatura , Animales , Bovinos , Células CultivadasRESUMEN
We undertook this trial to determine whether ipratropium bromide nasal spray 0.03% (IB) reduced the nasal hypersecretion associated with nonallergic perennial rhinitis (NAPR) without causing excessive dryness or irritation of the nasal mucosa. We compared two drug doses of IB (21 micrograms and 42 micrograms per nostril) to a placebo, administered as two sprays to each nostril twice daily. The study design consisted of a 1-week screening period without treatment, a 1-week single-blind placebo period, a 4-week double-blind treatment comparison period, and a 1-week follow-up period without medication to evaluate nasal rebound. One hundred fifty-two patients were entered and 140 completed the trial. Both doses of IB reduced the severity and duration of rhinorrhea compared with placebo (P = .05 and .03, respectively). Treatment differences were noticeable during the first week of therapy, continued to widen during the second week, and then remained stable throughout the next 2 weeks. There was no evidence of nasal rebound observed during the week after treatment. The drug was well tolerated with side effects limited to infrequent nasal adverse events of nasal dryness, blood-tinged mucus, and epistaxis occurring in 2% to 6% of patients. We conclude that IB is a safe and effective therapy for control of rhinorrhea associated with NAPR.
Asunto(s)
Ipratropio/administración & dosificación , Rinitis/tratamiento farmacológico , Administración Intranasal , Adolescente , Adulto , Anciano , Método Doble Ciego , Estudios de Evaluación como Asunto , Femenino , Humanos , Ipratropio/efectos adversos , Ipratropio/uso terapéutico , Masculino , Persona de Mediana Edad , Cooperación del Paciente , Calidad de Vida , Terfenadina/uso terapéutico , Factores de TiempoRESUMEN
Serum or plasma specimens were assayed in indirect immunfluorescence tests on cryostat sections of normal human skin for the presence and titer of antibodies reactive with human epidermal cytoplasmic antigens. A polyvalent fluorescein-labeled goat anti-human immunoglobulin antiserum was used in all tests. Three distinct staining patterns were noted: upper epidermal cytoplasmic fluorescence, U-CYT, produced by antibodies reactive with antigen present in cells of the upper and middle layers of the epidermis; general cytoplasmic fluorescence, G-CYT, produced by antibodies reactive with antigens present in cells throughout the epidermis; and basal cell cytoplasmic fluorescence, BCL, produced by antibodies reactive with components present only in basal cells. Sera from 8% of 52 normal blood donors produced the U-CYT pattern at dilutions greater than 1:10. The incidence of antibodies reactive with epidermal cytoplasmic antigens in patients with a clinical history of not more than 2 basal cell carcinomas of the skin was 5%, compared to an incidence of 89% in those individuals with 3 or more separate instances of skin neoplasms. There was no difference in the frequency with which cryosurgery was used in the treatment of skin neoplasms in either of these 2 groups. Antibodies to epidermal cytoplasmic antigens were also detected in 10% of patients with nondermatologic, nonpulmonary neoplasms, in 43% of patients with pulmonary neoplasms and in 1 of 11 patient with nonneoplastic diseases. Positive sera yielded titers ranging from 1:16 to 1:1024. The most common staining patterns noted in all of these cases were the U-CYT and G-CYT patterns; the BCL staining pattern was noted in only one instance.
Asunto(s)
Anticuerpos Antineoplásicos/análisis , Autoanticuerpos/análisis , Citoplasma/inmunología , Piel/citología , Antineoplásicos/uso terapéutico , Criocirugía , Técnica del Anticuerpo Fluorescente , Humanos , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/inmunología , Neoplasias Cutáneas/cirugíaRESUMEN
Lung-carcinoma-reactive antibodies have been previously isolated from tumor tissues and pleural effusions. To explore the immunoglobulin (Ig) content of bronchial secretions, bronchial washings of patients with inflammatory and neoplastic lung diseases were studied with respect to Ig levels and specificity. Expressed as Ig/potassium ratios, statistically significant increases in Ig levels were found in inflammatory diseases and even higher increases in lung carcinomas. The isolation of Ig from bronchial washings was achieved by dissociation of immune complexes at low pH, neutralization and subsequent purification by anion exchange chromatography. The isolated immunoglobulins were tested in indirect immunofluorescence against lung cancer cells of various histologic types in tissue cultures and fresh suspensions. Positive cytoplasmic fluorescence was obtained with cells of adeno carcinomas and squamous-cell carcinomas of the lung but not with cells of normal lung. The accessibility of bronchial washings makes the investigation of their lung-cancer-reactive immunoglobulins relatively easy and raises the possibility of its eventual conversion into a screening test.
Asunto(s)
Bronquios/inmunología , Inmunoglobulinas , Neoplasias Pulmonares/inmunología , Adenocarcinoma/inmunología , Complejo Antígeno-Anticuerpo , Bronquios/metabolismo , Carcinoma de Células Escamosas/inmunología , Línea Celular , Técnicas de Cultivo , Técnica del Anticuerpo Fluorescente , Humanos , Inmunoglobulina A/análisis , Inmunoglobulina G/análisis , Inmunoglobulina M/análisis , Inmunoglobulinas/análisisRESUMEN
Low pH elution techniques were used on lung cancer tissues and pleural effusions of lung cancer patients to dissociate antigen-antibody complexes. The immunoglobulins obtained were assayed by indirect immunofluorescence against tissue cultures and fresh cell suspensions of various target cells; they reacted positively, in significant titers, with cells of squamous cell carcinomas and adenocarcinomas of the lung but not with cells of normal adult and fetal lung or of nonpulmonary tumors. Immunoglobulins, similarly dissociated from tumor effusions of other organs, showed no reactivity in indirect immunofluorescence tests against lung carcinoma cells.
Asunto(s)
Anticuerpos Antineoplásicos/aislamiento & purificación , Neoplasias Pulmonares/inmunología , Derrame Pleural/inmunología , Adenocarcinoma/inmunología , Especificidad de Anticuerpos , Complejo Antígeno-Anticuerpo , Antígenos de Neoplasias , Carcinoma de Células Pequeñas/inmunología , Carcinoma de Células Escamosas/inmunología , Reacciones Cruzadas , Epítopos , Femenino , Humanos , Inmunoglobulina G , MasculinoRESUMEN
The local immune response to lung cancer was investigated by histologic and immunologic means. Distinctive patterns of stromal cellular reaction, characteristic for different histologic types of lung carcinoma, were recognized. The amount of cellular infiltration was highest in squamous cell carcinomas and lowest or nonexistent in oat cell carcinomas. Within the various histologic categories the well-differentiated tumors appeared to be accompanied by more reactive cells than the poorly differentiated ones; there was no relation between tumor necrosis and cellular infiltration. The plasma cells were distinctly associated with squamous cell carcinomas; their number in the stroma was proportionate to the degree of differentiation and the presence of keratin produced by the tumors. Eluates with a high content of immunoglobulins were recovered from pleural effusions and from solid lung carcinomas by dissociation of antigen-antibody complexes. These preparations reacted positively in indirect immunofluorescence tests with tissue cultures and with fresh suspensions of lung carcinoma cells, but not with tissue culture cells of most nonpulmonary tumors or with cell suspensions of normal adult and fetal lung. Similarly prepared fractions of noncarcinomatous pleural effusions did not react with lung cancer cells.