Plasma hydrogenated cationic detonation nanodiamonds efficiently deliver to human cells in culture functional siRNA targeting the Ewing sarcoma junction oncogene

The expression of a defective gene can lead to major cell dysfunctions among which cell proliferation and tumor formation. One promising therapeutic strategy consists in silencing the defective gene using small interfering RNA (siRNA). In previous publications we showed that diamond nanocrystals (ND) of primary size 35 nm, rendered cationic by polyethyleneimine-coating, can efficiently deliver siRNA into cell, which further block the expression of EWS/FLI-1 oncogene in a Ewing sarcoma disease model. However, a therapeutic application of such nanodiamonds requires their elimination by the organism, particularly in urine, which is impossible for 35 nm particles. Here, we report that hydrogenated cationic nanodiamonds of primary size 7 nm (ND-H) have also a high affinity for siRNA and are capable of delivering them in cells. With siRNA/ND-H complexes, we measured a high inhibition efficacy of EWS/FLI-1 gene expression in Ewing sarcoma cell line. Electron microscopy investigations showed ND-H in endocytosis compartments, and especially in macropinosomes from which they can escape before siRNA degradation occurred. In addition, the association of EWS/FLI-1 silencing by the siRNA/ND-H complex with a vincristine treatment yielded a potentiation of the toxic effect of this chemotherapeutic drug. Therefore ND-H appears as a promising delivery agent in anti-tumoral gene therapy.     

Is the present cut-point to define type 2 diabetes appropriate in Latin-Americans?

The diagnosis of diabetes mellitus type 2 (DM2) is based either on increased plasma glucose or Glycated hemoglobin levels. Since these measures are the only means for diagnosis of DM2, they must be well adapted to each population according to their metabolic characteristics, given that these may vary in each population. The World Health Organization (WHO) determined the cut-points of plasma glucose levels for the diagnosis of DM2 by associating hyperglycemia with the risk of a specific microvascular complication-retinopathy. Cardiovascular diseases are however the principal causes of mortality in patients with DM2 and we reported that in the Colombo-Ecuadorian population impaired fasting glucose and impaired glucose tolerance are both risk markers for myocardial infarction. We propose that the current cut-points accepted by the WHO need to be revaluated in populations such as Latin America and that there should be lower cut points for glycaemia in this population, to reduce the prevalence of cardiovascular complications associated with DM2.     

Improved in vitro evaluation of novel antimicrobials: potential synergy between human plasma and antibacterial peptidomimetics,AMPs and antibiotics against human pathogenic bacteria

Stable peptidomimetics mimicking natural antimicrobial peptides (AMPs) have emerged as a promising class of potential novel antibiotics. In the present study, we aimed at determining whether the antibacterial activity of two α-peptide/β-peptoid peptidomimetics against a range of bacterial pathogens was affected by conditions mimicking in vivo settings. Their activity was enhanced to an unexpected degree in the presence of human blood plasma for thirteen pathogenic Gram-positive and Gram-negative bacteria. MIC values typically decreased 2- to 16-fold in the presence of a human plasma concentration that alone did not damage the cell membrane. Hence, MIC and MBC data collected in these settings appear to represent a more appropriate basis for in vivo experiments preceding clinical trials. In fact, concentrations of peptidomimetics and peptide antibiotics (e.g. polymyxin B) required for in vivo treatments might be lower than traditionally deduced from MICs determined in laboratory media. Thus, antibiotics previously considered too toxic could be developed into usable last-resort drugs, due to ensuing lowered risk of side effects. In contrast, the activity of the compounds was significantly decreased in heat-inactivated plasma. We hypothesize that synergistic interactions with complement proteins and/or clotting factors most likely are involved.     

Diagnosis and Management of Atypical Hemolytic Uremic Syndrome In Children: Single Centre Experience

Atypical hemolytic uremic syndrome (aHUS) although rare is the commonest cause of acute renal failure (ARF) in children and has poor prognosis. We present single centre experience of aHUS. Thirty six children (29 males, 7 females) with mean age, 7.9 years presented with ARF, 2 children also had tonic–clonic type convulsions. Their hematology examination revealed hemolytic anemia with s. creatinine (SCr), 5.54 mg/dl. Acute HUS was observed in 75 %, acute on chronic HUS in 19.4 % and patchy cortical necrosis (PCN) in 5.6 % biopsies. Mean 5.4 plasma exchanges (PE) were carried out. Supportive management of anti-hypertensives and prednisone was also given. Recovery end points were establishment of urine output, improvement of SCr and hematological profile. Hematology and renal function profile improved variably in all children, 5.6 % died, relapse was observed in 80.5 % over mean 70 days; 13.9 % children are doing well over mean follow-up of 268.8 days. Thus poor prognosis was observed in 86.1 % children. Children with acute on chronic HUS and PCN did not recover. Six children who recovered had acute HUS. aHUS in Indian children occurs at an older age of around 8 years and chronic/irreversible changes on histopathology examination are harbingers of poor prognosis. PE is life-saving however further research for developing strategies to improve long-term survival is needed.     

Rapid and sensitive analysis of cyclobenzaprine by LC–MS/MS: Application to a pharmacokinetic study of cyclobenzaprine in dog

A rapid and sensitive liquid chromatography–tandem mass spectrometry method was developed and validated for the quantification of cyclobenzaprine in dog plasma. After extracted with organic solvent, post-treatment samples were separated on an Agela C18 column interfaced with a triple quadrupole tandem mass spectrometer in positive electrospray ionization mode. Multiple reaction monitoring was performed using the transitions of m/z 276.2 → 216.1 and m/z 325.1 → 109.0 to quantify cyclobenzaprine and escitalopram (internal standard), respectively. The mobile phase consisted of acetonitrile: 5 mM ammonium acetate: formic acid (90:10:0.01, v/v/v) at a flow rate of 0.3 ml/min. The total analysis time was 2.4 min. The method was linear over the concentration range of 0.0200–10.0 ng/ml. The intra- and inter-day precision was within 12.8% in terms of relative standard deviation (RSD%) and the accuracy within 5.6% in terms of relative error. This method was successfully applied in a pharmacokinetic study of extended-release cyclobenzaprine in dogs.