Development and characterization of vancomycin-loaded levan-based microparticular system for drug delivery.

Encapsulation of vancomycin (VANCO) into biodegradable levan microparticles was achieved using a simple preparation technique. Microparticles were prepared by using levan polysaccharide produced by a halophilic bacterium Halomonas smyrnensis AAD6T. To optimize efficiency of encapsulation process by precipitation method, three parameters were studied: drug and polymer concentrations and preparation rotating speed. The particles were characterized in vitro. The size of levan microparticles was changed between 0.404 µm and 1.276 µm. The surface charge was detected between +4.1 mV and +6.5 mV. The highest drug encapsulation capacity of the system was 74.7% and was depending on the polymer concentration. In dissolution studies, initial burst effect around 10-20% from all the formulations was observed and then the release was slowed down and continued at a constant level. In vitro antibiotic release from the microparticles was controlled with the drug carrier system and release fit to Higuchi kinetic model. All the released samples collected at different time intervals during dissolution studies have exhibited intrinsic bactericidal activity against Bacillus subtilis ATCC 6633. WST-1 cell proliferation and viability studies showed that VANCO-loaded levan microparticles at concentrations between 100 µg/mL and 1000 µg/mL were nontoxic to L929 cells. As conclusion, levan microparticulate system could be a potential carrier of antibiotic drugs such as VANCO.

Effect of Antibiotics on Polymorphonuclear Leukocyte Functions and Myeloperoxidase Activity, Glutathione and Malondialdehyde Levels in Allergic Asthma.

We investigated the effect of ciprofloxacin, rifampicine and doxycycline on myeloperoxidase (MPO) activity, glutathione (GSH) and malondialdehyde (MDA) levels in allergic asthma patients and healthy volunteers. Polymorphonuclear leukocytes (PMNs) were isolated with ficoll-hypaque gradient centrifugation method. MPO activity was assayed with modified o-dianisidine, GSH by Ellman's and MDA levels by Beuge's method. PMN functions and MDA levels of patients significantly decreased when compared with healthy volunteers. Ciprofloxacin significantly increased PMN functions, MPO activity and MDA levels of both groups. We have demonstrated that ciprofloxacin has beneficial effects on MPO activity and PMN functions in allergic asthma patients and healthy volunteers.

Potential adjuvant effects of Nigella sativa seeds to improve specific immunotherapy in allergic rhinitis patients.

OBJECTIVE: To investigate the effects of Nigella sativa seed supplementation on symptom levels, polymorphonuclear leukocyte (PMN) functions, lymphocyte subsets and hematological parameters of allergic rhinitis. SUBJECTS AND METHODS: Twenty-four patients randomly selected from an experimental group of 31 (mean age 34 years) sensitive to house dust mites with allergic rhinitis and a control group of 8 healthy volunteers (mean age 23 years) were treated with allergen-specific immunotherapy in conventional doses for 30 days. After a month of immunotherapy, 12 of the 24 patients and the 8 healthy volunteers were given N. sativa seed supplementation (2 g/day orally) for 30 days. The remaining 12 patients continued only on immunotherapy during the same period. The other 7 patients were given 0.1 ml saline solution subcutaneously once a week as a placebo. The symptom scores, PMN functions, lymphocyte subsets and other hematological parameters were evaluated before and after all treatment periods. RESULTS: There was a statistically significant increase in the phagocytic and intracellular killing activities of PMNs of patients receiving specific immunotherapy, especially after the addition of N. sativa seed. The CD8 counts of patients receiving specific immunotherapy plus N. sativa seed supplementation significantly increased compared to patients receiving only specific immunotherapy. PMN functions of healthy volunteers significantly increased after N. sativa seed supplementation compared to baseline. CONCLUSION: N. sativa seed supplementation during specific immunotherapy of allergic rhinitis may be considered a potential adjuvant therapy.

The effects of some antibiotics on polymorphonuclear leukocyte functions of elderly patients in vitro before and after zinc supplementation.

The effects of ciprofloxacin, cefodizime, rifampicine, doxycycline and cefodizime + rifampicine combination on polymorphonuclear leukocyte (PMN) functions (phagocytosis and intracellular killing activity) were investigated in vitro in elderly patients and compared with those of healthy young volunteers before and after zinc supplementation. PMNs of 13 elderly hypertensive patients and 10 healthy young volunteers were isolated by Ficoll-Hypaque gradient centrifugation method from venous blood with EDTA. The subjects were given 22 mg/daily/oral zinc supplementation for 1 month. Serum zinc levels before and after supplementation were measured by flame atomic absorbtion spectrophotometer and the effects of each drug on PMN functions at therapeutic concentrations were investigated. Ciprofloxacin significantly increased the PMN's phagocytic activity of elderly patients (p = 0.002) before zinc supplementation and significantly increased both PMN functions of elderly patients (p = 0.002) after zinc supplementation. The same antibiotic significantly increased both PMN functions of healthy young volunteers (p = 0.005 and p<0.05, respectively) before and after zinc supplementation when compared with the control (drug-free). Cefodizime significantly increased the PMN's phagocytic activity of elderly patients (p = 0.003, p = 0.002) before and after zinc supplementation when compared with the control (drug-free). It also significantly increased both PMN functions of healthy young volunteers (p = 0.005 and p<0.05, respectively) before and after zinc supplementation when compared with the control (drug-free). Doxycycline significantly increased PMN's intracellular killing activity of healthy young volunteers before zinc supplementation (p<0.05) when compared with the control (drug-free) values. Rifampicine significantly decreased PMN's phagocytic activity of elderly patients (p<0.05) after zinc supplementation. Cefodizime+rifampicine combination significantly increased PMN's phagocytic activity at therapeutic concentrations of healthy young volunteers (p = 0.005) before zinc supplementation and PMN's phagocytic activity of elderly patients (p<0.05) after zinc supplementation when compared with the control (drug-free). Consequently, in the present study from the antibiotics ciprofloxacin, cefodizime and cefodizime + rifampicine combination, which are accepted as biological response modifiers have demonstrated stimulatory effects by significantly increasing polymorphonuclear leucocyte functions (phagocytosis and/or intracellular killing activity) of elderly patients and healthy young volunteers in vitro before and after zinc supplementation. Additionally zinc supplementation has more immunostimulatory effects on PMN functions of healthy young volunteers than elderly patients.

The effects of allergen-specific immunotherapy on polymorphonuclear leukocyte functions in patients with seasonal allergic rhinitis.

Immunotherapy plays an important role in the therapy of allergic rhinitis and bronchial asthma. However, there is not much information about the effects of allergen-specific immunotherapy (SIT) on the polymorphonuclear leukocyte functions. The aim of this study was to investigate the effects of specific immunotherapy on phagocytic and intracellular killing activities of polymorphonuclear leukocytes (PMN) derived from patients with seasonal allergic rhinitis. Twenty-four patients with seasonal allergic rhinitis documented to be sensitive to grass pollen were included in this study. Patients were divided into 3 groups. Group 1 (n=7) received conventional immunotherapy whereas patients in Group 2 (n=7) were treated with short-term immunotherapy and the third group (n=10) were given placebo during the study process. Both phagocytic and intracellular killing activities were significantly increased (p=0.002, p<0.0001, respectively) by conventional immunotherapy when compared to the first determination. In the short-term immunotherapy group, phagocytic activity was increased very significantly (p=0.0001), whereas intracellular killing activity was not affected (p=0.252). There were no changes in these parameters in the placebo group. These results suggest that allergen-specific immunotherapy has an enhancing effect on PMNs functions in the patients with seasonal allergic rhinitis. It should be clarified by further studies whether this enhancement might be considered as another beneficial effect of the immunotherapy.

Effect of antituberculous drugs on human polymorphonuclear leukocyte functions in vitro.

The aim of the study was to investigate antituberculous drugs effects on polymorphonuclear leukocyte (PMN) functions (phagocytic activity and intracellular killing activity) in vitro. PMNs obtained from healthy volunteers were incubated with antituberculous drugs (isoniazid [INH], rifampin [RIF], pyrazinamide [PZA], ethambutol [EMB], streptomycin [S], amikacin [A], ofloxacin [OFLX], prothionamide [PTH] and cycloserine [CyC]) and different combinations at therapeutic serum concentrations. Phagocytic activity of PMNs was significantly increased when compared with controls by PTH (p<0.001), A (p<0.001), OFLX (p<0.001), INH+RIF+S combination (p<0.01), A+OFLX combination (p<0.05), A+OFLX+CyC combination (p<0.01) and A+OFLX+CyC+PTH+EMB combination (p<0.01). Intracellular killing activity of PMNs was significantly increased by OFLX when compared with the control (p<0.05). No significant difference was observed in functions of PMN for other drugs when compared with control (p>0.05). Functions of PMN were significantly increased by OFLX when compared with A+OFLX combination (p<0.05). Phagocytic activity of PMNs was significantly increased by A+OFLX+CyC combination and A+OFLX+CyC+PTH+EMB combination when compared with A+OFLX+CyC+PTH combination and A+OFLX+CyC+PTH+PZA combination (p<0.05). No significant difference was found in functions of PMN between the other groups (p>0.05). In conclusion, some antituberculous drugs alone or in combination enhanced PMN functions, although in combination no additive or synergistic effects were detected. Moreover, none of the antituberculous drugs alone or in combination significantly decreased PMN functions. The drugs having adverse effects on immune functions would better be replaced with equally effective drugs or drug combinations having positive effects on PMN functions.

The effects of rifampicin and fluoroquinolones on tubercle bacilli within human macrophages.

The objective of this study was to determine the activities of rifampicin, ciprofloxacin and sparfloxacin against M. tuberculosis H37Rv in human macrophages. The minimal inhibitory concentrations (MIC) were determined by agar macrodilution and broth microdilution methods and were 0.5, 1 and 0.125 mg/l, respectively. Concentrations of rifampicin at 0.5 and 2.5 microg (P<0.001), ciprofloxacin at 4 and 8 microg (P<0.001) and sparfloxacin at 0.125 microg (P<0.05), 0.625 microg (P<0.001) and 1.25 microg (P<0.001) were found to be effective against intracellular bacteria. Ciprofloxacin and especially sparfloxacin were effective in macrophages and may be useful in the treatment of tuberculosis particularly infections caused by multiply drug resistant strains.

Effect of montelukast on polymorphonuclear leukocyte functions in asthmatic patients.

Leukotriene receptor antagonists are being used widely in the treatment of bronchial asthma. They have been shown to possess anti-inflammatory properties, but there is no sufficient data about their effects on polymorphonuclear leukocyte functions. The aim of this study was to investigate the effects of montelukast, a specific cysteinyl leukotriene-1 receptor antagonist, on human polymorphonuclear leukocyte (PMN) functions (phagocytic and intracellular killing activity) in asthmatic patients. Fifteen mild to moderate asthmatic patients were included in the study. They were treated with montelukast (10 mg/day per os) in addition to their previous medications for 2 weeks. Whole blood samples of patients were taken before and after this treatment period. Phagocytic activities and intracellular killing activities of polymorphonuclear leukocytes isolated from whole blood samples were tested by using appropriate technics. Phagocytic and intracellular killing activities of PMNs were significantly increased (p<0.001, p<0.05) by montelukast compared to those before treatment. These results show that montelukast has an enhancing effect on PMN functions in asthmatic patients.

Effect of antipyretics on polymorphonuclear leukocyte functions in children.

The aim of this study was to investigate whether fever and antipyretic drugs had an adverse effect on human polymorphonuclear leukocyte (PMN) functions (phagocytic and intracellular killing activity). Twenty febrile children with an axillary temperature of 39-40 degrees C and 20 healthy children without fever were included. Polymorphonuclear leukocytes were isolated. The effects of in vitro addition of antipyretic drugs (acetaminophen, metamizole sodium, nimesulid and ibuprofen) on PMN functions were tested. Phagocytic activity was assayed by the ingestion of yeast cells by PMNs and intracellular killing activity by the ingestion of yeast cells (stained blue) killed by PMNs. PMNs derived from febrile children exhibited better phagocytic activity when ibuprofen was added. In contrast, phagocytic activity was enhanced when acetaminophen, metamizole sodium or nimesulid was added in children without fever. Intracellular killing activity was enhanced when ibuprofen or metamizole sodium was added in children without fever. We conclude the antipyretic drugs at safely achievable concentrations do not suppress PMN function in vitro.

Phagocytic activity of neutrophils improves over the course of therapy of diabetic foot infections.

AIMS: The aim of this study was to investigate changes in phagocytic activity of neutrophils of type 2 diabetic patients with foot infections over short treatment courses. The potential utility of the phagocytic index in determining the efficacy of treatment modalities and it's relationship with metabolic control parameters were evaluated. METHODS: The phagocytic activity of neutrophils was determined in blood samples of 38 type 2 diabetic patients with foot infections (14 women and 24 men). Mean age and mean duration of diabetes were 66.3+/-9.4 and 19.1+/-11.2 (yrs), respectively. All patients received standard treatment (intensive insulin therapy, antibiotherapy, hyperbaric oxygen therapy and surgical debridement). Phagocytic activity of neutrophils was determined by a standard method. Phagocytic activity of neutrophils, acute phase proteins (C-reactive protein) and glycosylated haemoglobin was determined before therapy and two weeks later. RESULTS: The phagocytic index before and after therapy were 47.7+/-11.4 and 62.5+/-15.6, respectively (p<0.05). There was a significant correlation between phagocytic index and both CRP and HbA1c (r=0.52, p<0.05 and r=-0.41, p<0.05, respectively). CONCLUSIONS: Derangement of carbohydrate metabolism may underlie the impairment of bactericidal activity of neutrophils of poorly controlled diabetic patients. These data reveal that phagocytic activity improves during short-course standard therapy and might enable monitoring of efficacy of treatment modalities in diabetic patients with foot infections.

Comparison of polymorphonuclear leukocyte functions in elderly patients and healthy young volunteers.

OBJECTIVE: To compare the polymorphonuclear leukocyte (PMN) functions (phagocytosis and intracellular killing activity) of elderly patients with healthy young volunteers. SUBJECTS AND METHODS: Fifty-nine elderly patients who had various diseases (cancer, hypertension and diabetes mellitus, DM) and 10 healthy young volunteers were included in this study. Ficoll-Hypaque gradient centrifugation was used to isolate PMNs from venous blood containing EDTA (0.1 g/ml). Phagocytosis and intracellular killing activity of neutrophils were assayed using a modification of Alexander's method, in which serum opsonins, number of neutrophils and number of microorganisms are standardized in order to detect both increases and decreases in phagocytosis and intracellular killing as well as combined abnormalities of these two functions. The least significant difference test was used to compare the results in the two groups. RESULTS: Phagocytic activity of PMNs from patients with cancer was significantly higher than that of healthy young volunteers (p < 0.05) and elderly patients with hypertension and DM (p < 0.05). There was no statistical difference between the phagocytic activity of PMNs from elderly patients with hypertension and DM and healthy young volunteers (p > 0.05). The intracellular killing activity of PMNs from elderly patients with hypertension, DM and cancer was significantly lower than that of healthy young volunteers (p = 0.001, p < 0.0001, p = 0.003, respectively). CONCLUSIONS: The intracellular killing activity of PMNs from elderly patients was significantly decreased when compared with that of healthy young volunteers. Ageing, chronic diseases and drugs used in the treatment of these elderly patients may be the cause for decreased intracellular killing activity.