Novel strategies for inhibition of bacterial biofilm in chronic rhinosinusitis

An important role has been recently reported for bacterial biofilm in the pathophysiology of chronic diseases, such as chronic rhinosinusitis (CRS). CRS, affecting sinonasal mucosa, is a persistent inflammatory condition with a high prevalence around the world. Although the exact pathological mechanism of this disease has not been elicited yet, biofilm formation is known to lead to a more significant symptom burden and major objective clinical indicators. The high prevalence of multidrug-resistant bacteria has severely restricted the application of antibiotics in recent years. Furthermore, systemic antibiotic therapy, on top of its insufficient concentration to eradicate bacteria in the sinonasal biofilm, often causes toxicity, antibiotic resistance, and an effect on the natural microbiota, in patients. Thus, coming up with alternative therapeutic options instead of systemic antibiotic therapy is emphasized in the treatment of bacterial biofilm in CRS patients. The use of topical antibiotic therapy and antibiotic eluting sinus stents that induce higher antibiotic concentration, and decrease side effects could be helpful. Besides, recent research recognized that various natural products, nitric oxide, and bacteriophage therapy, in addition to the hindered biofilm formation, could degrade the established bacterial biofilm. However, despite these improvements, new antibacterial agents and CRS biofilm interactions are complicated and need extensive research. Finally, most studies were performed in vitro, and more preclinical animal models and human studies are required to confirm the collected data. The present review is specifically discussing potential therapeutic strategies for the treatment of bacterial biofilm in CRS patients.     

Structure-activity relationship of pyrazolo pyrimidine derivatives as inhibitors of mitotic kinesin Eg5 and anticancer agents

Human kinesin Eg5 is a potential inhibiting site for cancer chemotherapy. Blocking metaphase by binding foreign inhibitors with Eg5 eventually leads to apoptotic cell death. Here, we report the pyrazolopyrimidine derivates as potent inhibitors of Eg5 that prevents mitotic kinesin progression. IC₅₀ values were evaluated against the motor domain of Eg5 using steady-state ATPase assay. To better understanding, we have performed molecular docking simulation. It reveals that the interactions of the proposed inhibitors with both the allosteric sites (helices α2, α3 and loopL5, and helices α4 & α6). Out of fifteen pyrazolopyrimidine derivates, three compounds (12, 25, and 27) have shown significant inhibition of Eg5. The synthesized compounds (12, 25, and 27) were tested for their in-vitro anticancer activity against cervical cancer cell line (HeLa).     

Synthesis of (1,3,4-thiadiazol-2-yl)-acrylamide derivatives as potential antitumor agents against acute leukemia cells

A lead compound with the (1,3,4-thiadiazol-2-yl)-acrylamide scaffold was discovered to have significant cytotoxicity on several tumor cell lines in an in-house cell-based screening. A total of 60 derivative compounds were then synthesized and tested in a CCK-8 cell viability assay. Some of them exhibited improved cytotoxic activities. The most potent compounds had IC₅₀ values of 1–5 μM on two acute leukemia tumor cell lines, i.e. RS4;11 and HL-60. Flow cytometry analysis of several active compounds and detection of caspase activation indicated that they induced caspase-dependent apoptosis. It was also encouraging to observe that these compounds did not have obvious cytotoxicity on normal cells, i.e. IC₅₀ > 50 μM on HEK-293T cells. Although the molecular targets of this class of compound are yet to be revealed, our current results suggest that this class of compound represents a new possibility for developing drug candidates against acute leukemia.     

Efficacy of antifungal agents in tissue conditioners in treating candidiasis

Chronic atrophic candidiasis is prevalent in up to 72% of institutionalized geriatric populations and is causally associated with Candida albicans. Topical antifungal treatments are difficult to implement in some geriatric patients due to cognitive impairment, reduced motor dexterity and memory loss. Objective: This in vitro study incorporated antifungal agents into tissue conditioners to investigate the effectiveness of this method of drug delivery. Design: Combinations of nystatin, fluconazole, itraconazole and Coe Soft, Viscogel, Fitt were tested at 1, 3, 5, 7, 9 and 11wt/wt%, with and without sterilized saliva. 6 mm diameter cores were punched in Sabouraud plates pre-grown with standardized C. albicans. Antifungal agents plus tissue conditioner mixtures were injected into each core. Inhibition diameters were measured for 14 days. Results: Cores with only tissue conditioners acted as negative control and showed no significant inhibition activity (ANOVA, p>0.05). Peak activity was between 65 to 89 hours; followed by a plateau. Itraconazole had greater fungicidal activity than fluconazole; while nystatin was found to have the least fungicidal activity (ANOVA. p<0.05). The most effective concentration for nearly all combinations was 5%wt/wt (ANOVA, p<0.05). Specimens with saliva showed greater antifungal activity than those without (t-test. p<0.001). Itraconazole altered the physical properties of Viscogel hence this combination is not recommended for clinical use. Conclusion: The treatment of chronic atrophic candidiasis by incorporation of antifungal drugs into tissue conditioners is efficacious. 5% wt/wt itraconazole mixed with Coe Soft or Fitt is recommended for clinical study where compliance of patient or care giver cannot be relied upon. Peak antifungal activity at 3 days suggests that mixtures prepared for clinical study may be replaced soon after this time for maximum effectiveness.     

Reflective properties of different eyespot types in dinoflagellates

The reflective properties of different types of dinoflagellate eyespots were investigated using confocal laser scanning microscopy in the epireflection contrast mode. Although the eyespots studied differed with respect to localization (cytosol or plastid) and organization of the globule layer(s), all types effectively absorbed and reflected blue-green laser light (principal lines of 488/514 nm). The relative orientation of the eyespot surface towards the light source strongly influenced the reflective properties. Maximal reflection occurred when the eyespot surface was approximately perpendicular to the light source and rapidly decreased at increasing angles of light incidence. Horizontal and vertical optical sectioning of live and fixed cells resolved differences in the reflection patterns. Focusing of reflected light on the basal portion of the longitudinal flagellum was observed for the cytosolic eyespot of Glenodinium sp. and the triple membrane-bounded eyespot of Peridinium foliaceum, presumably a vestige of a host plastid. This flagellum is thought to be mainly involved in mediating orientational movement responses. In contrast, the reflection patterns obtained from the eyespot of Woloszynskia pascheri, which represents the third and most commonly observed dinoflagellate eyespot type within a plastid, point to only minor focusing. Reflection signals could be followed a considerable distance into the sulcus in all cases, indicating that in dinoflagellate eyespots, irrespective of the presumed receptor location (plasma membrane overlying the eyespot and/or the basal part of the longitudinal flagellum), back reflection of non-absorbed light can enhance the excitation probability of the photoreceptor(s). Such a combined reflection/absorption screen allows maximal contrast modulation and will, in conjunction with the specialized geometry of the dinoflagellate eyespots, increase the directionality of these eyespot aparatuses considerably.     

A dual approach in the study of poly (ADP-ribose) polymerase: In vitro random mutagenesis and generation of deficient mice

A dual approach to the study of poly (ADP-ribose)polymerase (PARP) in terms of its structure and function has been developed in our laboratory. Random mutagenesis of the DNA binding domain and catalytic domain of the human PARP, has allowed us to identify residues that are crucial for its enzymatic activity.In parallel PARP knock-out mice were generated by inactivation of both alleles by gene targeting. We showed that: (i) they are exquisitely sensitive to -irradiation, (ii) they died rapidly from acute radiation toxicity to the small intestine, (iii) they displayed a high genomic instability to -irradiation and MNU injection and, (iv) bone marrow cells rapidly underwent apoptosis following MNU treatment, demonstrating that PARP is a survival factor playing an essential and positive role during DNA damage recovery and survival.     

Microbiological Characteristics and Susceptibility Patterns of Strains of Rhodotorula Isolated from Clinical Samples

The members of the genus Rhodotorula show a marked ubiquity. In man, they have been isolated from faeces, nails, skin, sputum, digestive tract and adenoids, forming part of the normal human flora, although in recent years cases have been reported of both local and systemic infection by this yeast. There are virtually no studies in the literature on the sensitivity of this genus to the antifungal agents in common clinical use. Therefore, it is considered of interest to study the microbiological characteristics and the susceptibility patterns of Rhodotorula isolated from clinical samples. A total of 35 different strains of Rhodotorula were studied. In vitro susceptibility testing to 5-fluorocytosine, amphotericin B, ketoconazole, fluconazole and itraconazole was performed. All the strains were considered sensitive to 5-fluorocytosine, amphotericin B, ketoconazole and itraconazole and resistant to fluconazole. As a conclusion, we can state that all the antifungal agents tested, except fluconazole, are useful medicaments for the treatment of infections by the Rhodotorula genus. This revised version was published online in June 2006 with corrections to the Cover Date.     

New antifungal agents that inhibit the growth of Candida Species: Dichlorinated 8-quinolinols

Five dichlorinated 8-quinolinols (2,5- 5,6-, 3,5-, 3,7-, and 4,5-dichloro-8-quinolinol) were tested against Candida albicans and C. Tropicalis in Sabouraud dextrosebroth with and without bovine serum. The 5,6-, 3,5-, and 3,7-dichloro-8-quinolinols proved to be more effective than the control, 5-fluorocytosine. In cytotoxicity tests employing baby hamster kidney (BHK) cells, all test agents proved to be more cytotoxic than the control. However, the minimum inhibitory concentration (MIC) of 3,5-dichloro-8-quinolinol to both fungi was only one tenth the cytotoxic dose,suggesting that the compound may be useful as a topical or systemic antifungal agent.This revised version was published online in October 2005 with corrections to the Cover Date.     

Elongated mouse chromosomes suitable for enhanced molecular cytogenetics

Characterization of genetic disorders in humans and animal models requires identification of chromosomal aberrations. However, identifying fine deletions or insertion in metaphase chromosomes has been always a challenge due to limitations of resolution. In this study we developed a rapid method for chromosome elongation using two different intercalating agents: ethidium bromide and 5-bromo-2′-deoxyuridine (BrdU), together with a short-term mitotic block using colcemid. About 70% of the chromosomes from cells that underwent this elongation procedure reached three times longer than those prepared from control cells. FISH experiments using elongated chromosomes revealed a duplicated region of chromosome 11 that was not visible in cells prepared with conventional methods.