Prevention and modulation of aminoglycoside ototoxicity (Review)

More than 60 years after their isolation and characterization, aminoglycoside (AG) antibiotics remain powerful agents in the treatment of severe gram-negative, enterococcal or mycobacterial infections. However, the clinical use of AGs is hampered by nephrotoxicity and ototoxicity, which often develop as a consequence of prolonged courses of therapy, or of administration of increased doses of these drugs. The discovery of non-ototoxic antibacterial agents, showing a wider spectrum of activity, has gradually decreased the use of AGs as first line antibiotics for many systemic infections. However, AGs are now undergoing an unexpected revival, being increasingly indicated for the treatment of severe emerging infections caused by organisms showing resistance to most first-line agents (e.g., multidrug-resistant tuberculosis, complicated nosocomially-acquired acute urinary tract infections). Increasing adoption of aminoglycosides poses again to scientists and physicians the problem of toxicity directed to the kidneys and to the inner ear. In particular, aminoglycoside-induced deafness can be profound and irreversible, especially in genetically predisposed patients. For this reason, an impressive amount of molecular strategies have been developed in the last decade to counteract the ototoxic effect of aminoglycosides. The present article overviews: i) the molecular mechanisms by which aminoglycosides exert their bactericidal activity, ii) the mechanisms whereby AGs exert their ototoxic activity in genetically-predisposed patients, iii) the drugs and compounds that have so far proven to prevent or modulate AG ototoxicity at the preclinical and/or clinical level, and iv) the dosage regimens that have so far been suggested to decrease the incidence of episodes of AG-induced ototoxicity

Resistance of uropathogens to antibacterial agents: Emerging threats, trends and treatments

Urinary tract infections are among the most common infectious diseases in humans. Today, resistance to nearly all antimicrobial classes is dramatically growing, and extremely drug-resistant or even pan-drug resistant pathogens are increasingly isolated around the world. It is foreseen that in the next decades the world will be facing a major medical emergency generated by the rapid spread of pathogens carrying resistance determinants of unprecedented power. Carbapenemase-producing Enterobacteriaceae, multidrug-resistant Enterococci and fluoroquinolone resistance determinants in both Gram-negative and Gram-positive uropathogens are among the greatest emergencies. In this article, the major emerging threats of particular interest to urologists are reviewed, worldwide resistance trends are illustrated, and novel and older-but still active-recommended drugs are summarized

Characterization of the c.190T>C missense mutation in BRCA1 codon 64 (Cys64Arg).

In the Milan area (Northern Italy), we identified a family characterized by a high prevalence of ovarian and breast cancer cases (5 out of 6 subjects, over 3 generations), and a predominant prevalence of ovarian lesions (4 out of 5 patients). Analysis of BRCA1 and BRCA2 genes allowed the identification of the missense c.190T>C mutation in codon 64 (Cys64Arg) of BRCA1. The aims of the present investigation were to characterize the functional implications of the c.190T>C mutation at the molecular level, and to search whether additional polymorphisms might be linked to the peculiar phenotypic features observed in the Italian pedigree. Molecular modelling studies suggested that substitution of the cysteine 64 with an arginine likely disrupts the architecture of the BRCA1 RING finger domain, responsible for the interaction with BARD1, essential for the tumor-suppressor activity of the BRCA1-BARD1 complex. By splicing site information analysis, exonic splicing enhancer site characterization, and analysis of transcript fragment length and sequence, we showed that the c.190T>C mutation was able to modulate the splicing of exon 5 in a fashion opposite to the c.190T>G transversion, responsible for the functionally-related Cys64Gly amino acid substitution. Genotyping of BRCA1 and BRCA2 in the Italian family revealed the presence of two significant polymorphisms: the cancer-associated c.2612C>T SNP in BRCA1, and the c.-26G>A SNP in the BRCA2 gene, acting as an ovarian cancer risk modifier in carriers of deleterious BRCA1 mutations. Analysis of these SNPs in a genotypically-unrelated Polish family, characterized by prevalent breast neoplasms in carriers of the c.190T>C mutation, revealed a genetic profile consistent with the hypothetic role of both polymorphisms

Infusion Micro-Pump Development Using MEMS Technology

International audienceDiabetes is a chronic condition that occurs when the pancreas does not produce enough insulin or when the body cannot effectively use the insulin it produces. People having type 1 diabetes require insulin (10% of all diabetics). People with type 2 diabetes can be treated with oral medication, but may also require insulin; 10% of all type 2 diabetics require insulin. Among the actual different methods to administer insulin (syringes, pens and conventional infusion pumps) a possibility to increase infuser performances is offered by the utilization of silicon based MEMS pumps (Micro- Electro Mechanical Systems). The main two pump families are classified as mechanical and non-mechanical pumps. The former contains check-valve, peristaltic, rectification without valves and rotary ones (“Displacement Pumps”) or Ultrasonic and Centrifugal (“Dynamic Pumps”); the latter consists in Pressure, Concentration, Electrical Potential gradients and Magnetic Potential micro-pumps. The micro-pump described here is an electro-mechanical device actuated with a piezoelectric-element and based on MEMS technology, able to minimize size and costs, offering a high precision pharmacological dispense. Three slices are bonded to reach the final results: top and bottom caps and an intermediate SOI. In case of anodic bonding, top and bottom caps are constituted of micromachined borophosphosilicate wafers, whereas in case of metallic bonding three silicon slices are used. The paper deals with the fabrication evolution of the device according to the different items that had to be faced during development: design, fluidic, mechanical and electrical simulations and characterization, safety requirements and final testing. Built-in reliability is ensured by two inner sensors able to detect any occlusion or malfunctioning and informing so the patient. The result is a compact, core pump chip that can deliver from 0.02 Units of insulin up to 3.6 Units per minute with accuracy better than 5%

Semen analysis in chronic bacterial prostatitis: diagnostic and therapeutic implications

The significance and diagnostic value of semen analysis in chronic bacterial prostatitis has been extensively debated and remains controversial. To investigate the diagnostic relevance of semen culture in the bacteriological workup of prostatitis patients, we retrospectively analyzed a clinical database of 696 symptomatic patients. All patients were routinely subjected to a four-glass test, followed by semen culture and analysis. This allowed to dissect from the database three different diagnostic scenarios, and to compare the \u2018two-glass\u2019 pre-/post- massage test and the standard \u2018four-glass\u2019 test with a \u2018five-glass\u2019 test (four-glass plus post-VB3 semen culture). The \u2018five-glass\u2019 test showed 3.6- or 6.5-fold increases in relative sensitivity and lesser reductions (\u201313.2% or \u201314.7%) in relative specificity for traditional uropathogens (TUs) compared with the four-glass or two-glass test, respectively. The area under the ROC curve and Jouden\u2019s index were increased, whereas positive and negative likelihood ratios were lower than comparators, indicating that the \u2018five-glass\u2019 assay may be superior in confirming the negative outcome of both standard tests. The five-, four-, and two-glass tests detected TUs (Enterobacteriaceae, Enterococci, etc.) in 120, 33, and 20 patients and unusual pathogens (Streptococci, other Gram-positive species, Mycoplasmata, and others) in 130, 56, and 45 patients, respectively. When patients were subjected to pharmacological treatment, including a combination of a fluoroquinolone and a macrolide, no differences in eradication rates were observed between groups diagnosed with different tests, irrespective of pathogen category. Eradication was associated with long-term sign/symptom remission; no significant intergroup differences in sign/symptom scores were observed throughout a 24-month off-therapy follow-up period. In conclusion, our data support the usefulness of semen analysis in the diagnostic workup of prostatitis patients when this test is used to complement thefour-glass Meares and Stamey test. Improvement of microbiological assays conveys important diagnostic and therapeutic implications

Tumor Suppression by RNA from C/EBPβ 3′UTR through the Inhibition of Protein Kinase Cε Activity

BACKGROUND: Since the end of last century, RNAs from the 3'untranslated region (3'UTR) of several eukaryotic mRNAs have been found to exert tumor suppression activity when introduced into malignant cells independent of their whole mRNAs. In this study, we sought to determine the molecular mechanism of the tumor suppression activity of a short RNA from 3'UTR of C/EBPβ mRΝΑ (C/EBPβ 3'UTR RNA) in human hepatocarcinoma cells SMMC-7721. METHODOLOGY/PRINCIPAL FINDINGS: By using Western blotting, immunocytochemistry, molecular beacon, confocal microscopy, protein kinase inhibitors and in vitro kinase assays, we found that, in the C/EBPβ 3'UTR-transfectant cells of SMMC-7721, the overexpressed C/EBPβ 3'UTR RNA induced reorganization of keratin 18 by binding to this keratin; that the C/EBPβ 3'UTR RNA also reduced phosphorylation and expression of keratin 18; and that the enzyme responsible for phosphorylating keratin 18 is protein kinase Cε. We then found that the C/EBPβ 3'UTR RNA directly inhibited the phosphorylating activity of protein kinase Cε; and that C/EBPβ 3'UTR RNA specifically bound with the protein kinase Cε-keratin 18 conjugate. CONCLUSION/SIGNIFICANCE: Together, these facts suggest that the tumor suppression in SMMC-7721 by C/EBPβ 3'UTR RNA is due to the inhibition of protein kinase Cε activity through direct physical interaction between C/EBPβ 3'UTR RNA and protein kinase Cε. These facts indicate that the 3'UTR of some eukaryotic mRNAs may function as regulators for genes other than their own

Increased chromosomal radiosensitivity in asymptomatic carriers of a heterozygous BRCA1 mutation

Background: Breast cancer risk increases drastically in individuals carrying a germline BRCA1 mutation. The exposure to ionizing radiation for diagnostic or therapeutic purposes of BRCA1 mutation carriers is counterintuitive, since BRCA1 is active in the DNA damage response pathway. The aim of this study was to investigate whether healthy BRCA1 mutations carriers demonstrate an increased radiosensitivity compared with healthy individuals. Methods: We defined a novel radiosensitivity indicator (RIND) based on two endpoints measured by the G2 micronucleus assay, reflecting defects in DNA repair and G2 arrest capacity after exposure to doses of 2 or 4 Gy. We investigated if a correlation between the RIND score and nonsense-mediated decay (NMD) could be established. Results: We found significantly increased radiosensitivity in the cohort of healthy BRCA1 mutation carriers compared with healthy controls. In addition, our analysis showed a significantly different distribution over the RIND scores (p = 0.034, Fisher’s exact test) for healthy BRCA1 mutation carriers compared with non-carriers: 72 % of mutation carriers showed a radiosensitive phenotype (RIND score 1–4), whereas 72 % of the healthy volunteers showed no radiosensitivity (RIND score 0). Furthermore, 28 % of BRCA1 mutation carriers had a RIND score of 3 or 4 (not observed in control subjects). The radiosensitive phenotype was similar for relatives within several families, but not for unrelated individuals carrying the same mutation. The median RIND score was higher in patients with a mutation leading to a premature termination codon (PTC) located in the central part of the gene than in patients with a germline mutation in the 5′ end of the gene. Conclusions: We show that BRCA1 mutations are associated with a radiosensitive phenotype related to a compromised DNA repair and G2 arrest capacity after exposure to either 2 or 4 Gy. Our study confirms that haploinsufficiency is the mechanism involved in radiosensitivity in patients with a PTC allele, but it suggests that further research is needed to evaluate alternative mechanisms for mutations not subjected to NMD