Manuka honey chelates iron and impacts iron regulation in key bacterial pathogens.

AIM: The aim of this study was to test the hypothesis that Manuka honey (MH) chelates iron and promotes an iron-limiting environment, which contributes to its antimicrobial activity. METHODS AND RESULTS: Employing a ferrozine-based assay, we observed that MH is an iron chelator that depletes iron from solution. Siderophores are small molecules that bind ferric iron (III) with high affinity and their levels are upregulated by bacteria under iron-limiting conditions. We demonstrated by quantitating siderophore production that Escherichia coli and Pseudomonas aeruginosa treated with MH sub-minimum inhibitory concentrations (sub-MIC) experience an iron-limiting environment and increase siderophore production. In addition, supplementation with ferrous iron (II) significantly increased growth of E. coli, Staphylococcus aureus and P. aeruginosa cultured at their MH MIC above that observed in nonsupplemented controls. By contrast, supplementation with ferric iron (III) significantly increased growth for only E. coli and P. aeruginosa, above their nonsupplemented controls. CONCLUSIONS: Manuka honey chelates iron, thereby generating an iron-limiting environment for E. coli and P. aeruginosa, and to a lesser extent S. aureus, which contributes to its antimicrobial properties. SIGNIFICANCE AND IMPACT OF THE STUDY: Our work demonstrates that MH-induced iron chelation is an antimicrobial mechanism that differentially impacts the bacterial species tested here. Iron chelation affects multiple diverse physiological processes in bacteria and would contribute to the lack of bacterial resistance to MH. Iron metabolism is tightly regulated; bacteria require this essential nutrient for survival, but in excess it is toxic. Additional exploration of MH's iron chelation mechanism will facilitate its future use in mainstream medicine.

Efficient mRNA electroporation of peripheral blood mononuclear cells to detect memory T cell responses for immunomonitoring purposes.

To date, the number of immunotherapy vaccines in clinical trials increases exponentially. To evaluate the efficacy of these clinical vaccination trials, we fervently need cellular immunomonitoring tools. In this study, we present a newly developed short-time assay which allows direct ex vivo analysis of multi-epitope antigen-specific T cell immune responses. This novel method is based on mRNA electroporation of isolated peripheral blood mononuclear cells (PBMC). Fresh and cryopreserved PBMC of both healthy volunteers as well as of allogeneic stem cell transplanted patients enrolled in a cytomegalovirus (CMV) dendritic cell vaccination trial were electroporated with CMV pp65-encoding mRNA. Using a direct IFN-gamma EliSPOT and intracellular cytokine flow cytometry we detected significantly higher numbers of CMV pp65-specific IFN-gamma-secreting T cells as compared to the assay with non-treated PBMC and as compared to PBMC electroporated with mRNA coding for an irrelevant protein. Compared to conventional methods to evaluate T cell-mediated immune responses, this method is time-saving and less labor-intensive because it obviates the need for in vitro cultured antigen-presenting cells and because an overnight incubation is sufficient for activation of T cells. Moreover, the use of CMV pp65-encoding mRNA will broaden the immune response because it covers every epitope with potential relevance. This is a major advantage compared to the recognition of a single epitope covered by a HLA-restricted peptides. In summary, we developed a highly efficient mRNA electroporation protocol for fresh and cryopreserved PBMC. This novel method is a rapid and elegant tool and will be convenient for monitoring the cellular immune status of patients in a clinical vaccination setting.

Survivability and immune responses after challenge with Edwardsiella ictaluri in susceptible and resistant families of channel catfish, Ictalurus punctatus.

Diseases in catfish farming are prevalent and costly, particularly the bacterial disease Enteric Septicemia of Catfish. Considerable research has focused on different aspects of this disease, including the biology of the causative agent, Edwardsiella ictaluri. However, no satisfactory treatment or preventive has resulted from these efforts. One solution is to increase the natural disease resistance of the fish through genetic selection. Recent research has demonstrated that genetic factors influence resistance to infection in mammals as well as fish. Selective breeding for disease resistance in channel catfish is ongoing, however differences in defence mechanisms among E. ictaluri challenged strains and families are only now being investigated. Antigen-specific as well as non-specific immune responses of full-sib families of channel catfish to laboratory challenge with E. ictaluri have been investigated. Both resistant and sensitive families produce a humoral response as specific antibody, but there were no differences found in the level of specific antibody produced. The sensitive family produced a slightly higher percentage of B lymphocytes in mononuclear cell preparations from peripheral blood, while the resistant family had a higher percentage of T lymphocytes in those preparations. The most significant observation was that the resistant family produced more macrophage aggregations in the spleen and posterior kidney throughout the infection than the sensitive family. Neither family produced stress-associated amounts of cortisol.

The assessment and treatment of concerns and anxiety in patients undergoing presurgical monitoring for epilepsy.

PURPOSE: This study investigated the impact of a treatment information package on patients being monitored for possible surgical treatment for temporal lobe epilepsy. METHODS: One hundred patients were randomly assigned to either a high- or low-information preparation condition. Levels of anxiety were tested soon after admission by using the Hospital Anxiety and Depression Scale (HADS), The State-Trait Anxiety Inventory (STAI), and a newly devised questionnaire to assess specific concerns and anxieties of epilepsy patients presenting for monitoring and surgery, the Concerns About Epilepsy Monitoring Questionnaire (CAEMQ). Dispositional desire for information was assessed by the Miller Behavioural Style Scale (MBSS) to investigate whether coping disposition affected coping styles in the hospital setting. Patients assigned to the high-information condition were exposed to an intervention package, which included viewing a video depicting two separate interviews with patients who had undergone surgery as well as an information package, which described the various tests that the patient would undergo in the course of the monitoring procedure. Patients assigned to the low-information group were given information that the hospital provided to all patients in their care. All subjects were then retested on anxiety levels a few days later. RESULTS: Those in the high-information group showed a significant decrease in anxiety and depression levels compared with those in the low-information group. CONCLUSIONS: Within the main findings, an effect of dispositional style was found. Identification as either a monitor or blunter on the MBSS showed different coping strategies on arrival in hospital as measured by the CAEMQ, indicating that the level of information given to patients with epilepsy on arrival needs to be mediated by awareness of these two dispositional styles so that they obtain maximal benefit from the information to which patients with epilepsy are exposed.

Seasonal values of selected blood parameters of farm-raised channel catfish (Ictalurus punctatus) in the Mississippi Delta.

Hematocrit, sodium, chloride, potassium, calcium, glucose, and pH were measured in whole blood of 1,522 channel catfish collected from 3 commercial food-fish ponds in the Mississippi Delta. Samples were collected from March 1995 to March 1996 to monitor seasonal fluctuations. A total of 10-20 fish were arbitrarily collected with snag lines from each pond on each sample day. The mean monthly hematocrits fluctuated seasonally from a low of 14.5% in midwinter to a high of 25.7% in midsummer (annual x = 21%, SE = 0.15). Sodium levels were consistent throughout the year with a mean (SE) of 134 (0.13) mM/liter. Mean chloride values for the year were 120 (0.14) mM/liter but increased to 132 mM/liter in midwinter. By March 1996, the chloride levels had returned to levels observed during spring 1995. Potassium and glucose levels varied throughout the year with means of 4.43 (0.06) mM/liter and 26.9 (0.46) mg/dl, respectively, and coefficients of variation of 51.8% and 63.3%, respectively. Calcium and pH values were fairly stable with means of 1.31 (0.004) mM/liter and 7.13 (0.004), respectively. All parameters except glucose and potassium may be adequately evaluated with a sample size of 25 or less. These data were collected to provide baseline information for ongoing pond health studies.

Parent information binder: individualizing education for parents of preterm infants.

To assist the parents of preterm infants at Mount Sinai Hospital, Toronto, Canada, we have developed a unique and creative method of individualizing parent information: The parent information binder (PIB). This method has supported a shift from nurse-directed education to parents as partners in their own education. Parents and health care professionals utilize the PIB collaboratively to facilitate organization and collection of information, which is individualized according to the infant's level of wellness and the parents' readiness to receive the information. This approach empowers parents to be active and integral participants in decisions regarding their infants' hospital care.

Transurethral microwave thermotherapy in the treatment of chronic abacterial prostatitis: a 2 years follow-up.

OBJECTIVE: Transurethral microwave thermotherapy (TUMT) is a minimally invasive, outpatient treatment, applied as a single session of 1-hour duration. SUBJECTS AND METHODS: A group of 11 patients with chronic abacterial prostatitis, who failed to respond to a variety of conventional treatments, underwent this therapy in our centre. RESULTS: The results after two years of follow-up are encouraging: 88% remain free of symptoms. CONCLUSION: Thermotherapy seems to add a new alternative for this hard to manage disease.

Molecular adaptations of GLUT1 and GLUT2 in renal proximal tubules of diabetic rats.

The renal reabsorption of glucose is mediated by two major classes of transporters. Initially, luminal glucose is concentrated in tubules by Na(+)-glucose cotransporters (Na(+)-GLUT). Afterwards, glucose reaches the blood space through facilitative glucose transporters, low-Michaelis constant (Km) GLUT1 and high-Km GLUT2. Hence, the transtubular flux of glucose could be impaired in hyperglycemia because the outwardly directed glucose gradient, from tubule to blood, is potentially lowered. However, in diabetic rats, transtubular glucose flux is not reduced but increased. In this work the molecular mechanism underlying this adaptation was examined. We tested the hypothesis that upregulation of renal tubular high-Km GLUT2 gene may compensate for the decrease in the tubule to blood glucose gradient. In rat tubules, GLUT1 protein and mRNA steady-state levels were reduced, and GLUT2 protein and mRNA levels were increased in rats after 2, 3, and 4 wk of uncontrolled streptozotocin-induced diabetes. These molecular adaptations were associated with augmented facilitative glucose flux. In summary, changes in GLUT1 and GLUT2 gene expression are important to the preservation of renal glucose reabsorption in hyperglycemia.

[Sterilization and the mentally handicapped: who decides?]. / Sterilisatie en mentaal gehandicapten: wie beslist?

At the moment sterilization of mentally handicapped persons is carried out in a medical and a legal vacuum in Belgium as well as abroad. To give an adequate answer to the problem of sterilization of mentally handicapped adolescents in medical practice, Dr. J. Denekens undertook an extensive literature review and an empirical investigation comprising case-studies of 52 adolescents, their parents and family doctors. The adolescents, between the ages of 16 and 21, were selected at random from students attending schools for mentally handicapped persons in the province of Antwerp during the school year 1989-1990. This study and investigation resulted in the formulation of medical and ethical criteria to assist decision makers confronted with a request for sterilization from or on the behalf of a mentally handicapped person who may be unable to decide this issue alone. These criteria have been ordered in a useful and practical algorithm that enables a motivated, logical and ethically responsible decision to be taken. A proposal of procedure for such a sterilization request has been elaborated. The medical-ethical criteria, the algorithm and the procedure proposal come out right in time as the European Parliament is discussing a motion that deals with the legal part of the subject.

Glucose transporters of rat proximal tubule: differential expression and subcellular distribution.

In the late proximal tubule, glucose reabsorption progressively lowers the concentration of luminal glucose, and concentrative glucose influx increases to ensure complete glucose reabsorption. The change in glucose influx is effected by luminal Na(+)-dependent glucose transporters (Na(+)-GLUT), which exhibit higher Na(+)-to-glucose stoichiometric ratios in the late proximal tubule. In this work, the corresponding changes in the axial distribution of basolateral glucose efflux transporters (GLUTs) were examined. mRNAs encoding high-affinity facilitative basolateral transporter GLUT1, low-affinity GLUT2, and apical Na(+)-GLUT were identified in mixed populations of proximal convoluted and straight tubules. The organization of the cognate proteins was also appraised on Western blots. GLUT1 was present in glomeruli, proximal convoluted, and straight tubules, GLUT2 was only expressed in the proximal convoluted tubule, and Na(+)-GLUT was present in both proximal convoluted and straight segments. GLUT1 and GLUT2 were confined to the basolateral membrane, whereas Na(+)-GLUT was preferentially localized to the brush-border membrane. These data are consistent with the idea that glucose influx in early and late proximal tubule is achieved through Na(+)-GLUT, that GLUT1 and GLUT2 are responsible for glucose efflux in the early proximal tubule, and that in the late proximal tubule, where transcellular glucose flux is lower, only GLUT1 mediates glucose efflux.

Role of penile nocturnal tumescence and rigidity measurement in the diagnosis of erectile impotence.

Reviewing a group of 100 patients undergoing nocturnal penile tumescence and rigidity measurement (NPTR), we tried to define normal values and compared these results with those of other authors. We also tried to answer two important questions: Is an abnormal NPTR registration always a proof of biogenic impotence? Is a normal NPTR always a proof of psychogenic impotence?

Potency and selectivity of the aromatase inhibitor R 76,713. A study in human ovarian, adipose stromal, testicular and adrenal cells.

The effects of R 76,713 on steroidogenesis were studied in primary cultures of four different human cell types, i.e. ovarian granulosa cells, adipose stromal cells, testicular cells and adrenal cells. In human granulosa cells aromatization of [1 beta, 2 beta-3H]androstenedione (as measured by the release of tritiated water) showed a Km (Michaelis constant) of 78 nM. R 76,713 competitively inhibited aromatization with a Ki (dissociation constant of the enzyme-inhibitor complex) of 1.6 nM. In human adipose stromal cells aromatization was measured by following the conversion of androstenedione to estrone and 17 beta-estradiol. In this system a Km for aromatization of androstenedione of 10.8 nM was found. R 76,713 again showed competitive kinetics with a Ki-value of 0.14 nM. In human testicular cells the synthesis of the androgens testosterone, androstenedione and dehydroepiandrosterone was only inhibited by drug concentrations exceeding 10(-6) M. At 10(-5) M of R 76,713, steroid concentrations were lowered to 56, 64 and 81% of the control for testosterone, androstenedione and dehydroepiandrosterone respectively. Concomitantly, a slight increase in the levels of pregnenolone (138% of the control) and progesterone (133% of the control) was seen. In human adrenal cells the synthesis of cortisol and aldosterone was slightly affected by R 76,713 also at concentrations exceeding 10(-6) M. At 10(-5) M of R 76,713 the concentrations of cortisol and aldosterone were lowered to respectively 59 and 51% of the control. At the same drug concentration the precursors 11-deoxycortisol and 11-deoxycorticosterone rose to 189 and 147% of the control. These results show that in primary cultures of human cells, R 76,713 is a very potent aromatase inhibitor with a selectivity of at least 1000-fold compared to other steps in steroidogenesis.

Bilateral ureteral inverted papillomas in association with bladder carcinoma: a case report and review of the literature.

A case of multiple inverted papillomas of both ureters, in association with a bladder carcinoma, is presented. Inverted papilloma is a rare benign urothelial tumor. Because of its curability by conservative surgery, it should be considered in the differential diagnosis of a ureteral filling defect and ureteral obstruction. Inverted papilloma may be associated with carcinomas of the urinary tract.

Effect of omega-3 fatty acids in diet of type I diabetic subjects on lipid values and hemorheological parameters.

Twelve type I (insulin-dependent) diabetic subjects in stable metabolic control for at least 3 mo received a controlled diet containing 50% carbohydrate, 35% fat, and 15% protein. Calorie intake varied from 1800 to 2200 calories, depending on individual needs. Part of the polyunsaturated omega-6 fatty acids (omega 6FAs) were isocalorically exchanged with omega 3FAs (2.7 g/day provided by fish oil concentrates) for 10 wk. Subject selection was based on the fact that the atherogenic index (total cholesterol/high-density lipoprotein cholesterol [HDL-chol]) remained greater than 5. Total cholesterol did not change, but HDL-chol (P less than .05) increased significantly, and the mean +/- SD atherogenic index decreased from 5.9 +/- 1.1 to 5.1 +/- 1.3. Plasma triglyceride levels also decreased (P less than .05). There was a small (approximately 2%) but significant (P less than .05) decrease of whole-blood viscosity at low shear rate because of a similarly small (approximately 2% decrease (P less than .05) of plasma viscosity. Erythrocyte viscosity values and the erythrocyte transit time, measured with the St. George's filtrometer, remained unchanged during fish oil intake. Four weeks after stopping the omega 3FA administration, the triglyceride level was again increased (P less than .05) and was even higher than the starting value (P less than .05). Plasma and whole-blood viscosity also increased to the starting levels, demonstrating that lipid alterations are accompanied with blood viscosity changes in the presence of a stable metabolic control.

Blood viscosity parameters in coronary heart disease: effect of fish oil supplementation.

We have evaluated blood viscosity parameters in 20 men suffering from coronary heart disease (CHD) and in 15 control subjects. Whole blood viscosity at a standardized haematocrit of 45% was significantly increased in the CHD-patients, both at low (p less than 0.001) and high (p less than 0.05) shear rate. The increased whole blood viscosity in these patients was explained by an increased plasma viscosity (p less than 0.01), while the erythrocyte suspension viscosity values at a standardized haematocrit of 70%, reflecting erythrocyte deformability, were within the normal range. We have studied the effects of a daily supplementation of 1.5 g omega-3 polyunsaturated fatty acids on blood viscosity in the CHD patients. After 6 weeks of treatment whole blood viscosity, at low and high shear rate, and plasma viscosity were significantly improved (all p less than 0.05), although not normalized. There was no effect of the fish oil on erythrocyte deformability.

Comparative effects of ketoconazole on rat, dog and human testicular steroidogenesis.

Ketoconazole is an antifungal azole derivative which also inhibits the cytochrome P-450(17)alpha, catalyzing the conversion of progestins into androgens. The effects of ketoconazole on human, dog and rat testosterone biosynthesis were compared using short term incubations of dispersed testicular cells. The results showed that ketoconazole inhibited androgen biosynthesis at lower concentrations in dispersed human testicular cells (IC50: 0.08 mumol/l) than in canine (IC50: 0.1 mumol/l) and rat cells (IC50 greater than or equal to 0.2 mumol/l). Furthermore, they demonstrated that ketoconazole first inhibited the 17,20-lyase activity and then the 17-hydroxylation in rat and dog cells whereas only the 17-hydroxylation was affected in human cells.