Exploring the mechanisms of action of human secretory RNase 3 and RNase 7 against Candida albicans

Human antimicrobial RNases, which belong to the vertebrate RNase A superfamily and are secreted upon infection, display a wide spectrum of antipathogen activities. In this work, we examined the antifungal activity of the eosinophil RNase 3 and the skin-derived RNase 7, two proteins expressed by innate cell types that are directly involved in the host defense against fungal infection. Candida albicans has been selected as a suitable working model for testing RNase activities toward a eukaryotic pathogen. We explored the distinct levels of action of both RNases on yeast by combining cell viability and membrane model assays together with protein labeling and confocal microscopy. Site-directed mutagenesis was applied to ablate either the protein active site or the key anchoring region for cell binding. This is the first integrated study that highlights the RNases' dual mechanism of action. Along with an overall membrane-destabilization process, the RNases could internalize and target cellular RNA. The data support the contribution of the enzymatic activity for the antipathogen action of both antimicrobial proteins, which can be envisaged as suitable templates for the development of novel antifungal drugs. We suggest that both human RNases work as multitasking antimicrobial proteins that provide a first line immune barrier

Magnetite Nanoparticles Functionalized with RNases against Intracellular Infection of Pseudomonas aeruginosa

Altres ajuts: Fundació La Marató de TV3/20180310Current treatments against bacterial infections have severe limitations, mainly due to the emergence of resistance to conventional antibiotics. In the specific case of Pseudomonas aeruginosa strains, they have shown a number of resistance mechanisms to counter most antibiotics. Human secretory RNases from the RNase A superfamily are proteins involved in a wide variety of biological functions, including antimicrobial activity. The objective of this work was to explore the intracellular antimicrobial action of an RNase 3/1 hybrid protein that combines RNase 1 high catalytic and RNase 3 bactericidal activities. To achieve this, we immobilized the RNase 3/1 hybrid on Polyetheramine (PEA)-modified magnetite nanoparticles (MNPs). The obtained nanobioconjugates were tested in macrophage-derived THP-1 cells infected with Pseudomonas aeruginosa PAO1. The obtained results show high antimicrobial activity of the functionalized hybrid protein (MNP-RNase 3/1) against the intracellular growth of P. aeruginosa of the functionalized hybrid protein. Moreover, the immobilization of RNase 3/1 enhances its antimicrobial and cell-penetrating activities without generating any significant cell damage. Considering the observed antibacterial activity, the immobilization of the RNase A superfamily and derived proteins represents an innovative approach for the development of new strategies using nanoparticles to deliver antimicrobials that counteract P. aeruginosa intracellular infection

"Review: Application of Bioequivalence Testing of Medicines in Peru"

"This is a review of the current status of drug bioequivalence studies in Peru. A bibliographic search was conducted in PubMed (Medline database) for bioequivalence studies in Peru. Generic drugs constitute the basis of pharmacological requests in health care systems in Latin American countries. Peru has enacted laws and regulations that require bioequivalence studies of high health risk drugs and exemptions, based on international legislation, to be conducted in research centers accredited by the authority of Health. There is a list of 19 drugs that must demonstrate their therapeutic equivalence through in vivo or in vitro studies, of which 13 have shown bioequivalence in vivo, and 8 of those have shown bioequivalence in vitro. There is a challenge for health authorities to enforce the current legislation and an even greater challenge for pharmaceutical laboratories to demonstrate bioequivalence of multi-source drugs with the reference drug.

The development of highly dense highly protected surfactant ionizable lipid RNA loaded nanoparticles

The long quest for efficient drug administration has been looking for a universal carrier that can precisely transport traditional drugs, new genomic and proteic therapeutic agents. Today, researchers have found conditions to overcome the two main drug delivery dilemmas. On the one side, the versatility of the vehicle to efficiently load, protect and transport the drug and then release it at the target place. On the other hand, the questions related to the degree of PEGylation which are needed to avoid nanoparticle (NP) aggregation and opsonization while preventing cellular uptake. The development of different kinds of lipidic drug delivery vehicles and particles has resulted in the development of ionizable lipid nanoparticles (iLNPs), which can overcome most of the typical drug delivery problems. Proof of their success is the late approval and massive administration as the prophylactic vaccine for SARS-CoV-2. These ILNPs are built by electrostatic aggregation of surfactants, the therapeutic agent, and lipids that self-segregate from an aqueous solution, forming nanoparticles stabilized with lipid polymers, such as PEG. These vehicles overcome previous limitations such as low loading and high toxicity, likely thanks to low charge at the working pH and reduced size, and their entry into the cells via endocytosis rather than membrane perforation or fusion, always associated with higher toxicity. We herein revise their primary features, synthetic methods to prepare and characterize them, pharmacokinetic (administration, distribution, metabolization and excretion) aspects, and biodistribution and fate. Owing to their advantages, iLNPs are potential drug delivery systems to improve the management of various diseases and widely available for clinical use

Identification of CFTR variants in Latino patients with cystic fibrosis from the Dominican Republic and Puerto Rico

BackgroundIn cystic fibrosis (CF), the spectrum and frequency of CFTR variants differ by geography and race/ethnicity. CFTR variants in White patients are wellâ described compared with Latino patients. No studies of CFTR variants have been done in patients with CF in the Dominican Republic or Puerto Rico.MethodsCFTR was sequenced in 61 Dominican Republican patients and 21 Puerto Rican patients with CF and greater than â â â â 60â mmol/L sweat chloride. The spectrum of CFTR variants was identified and the proportion of patients with 0, 1, or 2 CFTR variants identified was determined. The functional effects of identified CFTR variants were investigated using clinical annotation databases and computational prediction tools.ResultsOur study found 10% of Dominican patients had two CFTR variants identified compared with 81% of Puerto Rican patients. No CFTR variants were identified in 69% of Dominican patients and 10% of Puerto Rican patients. In Dominican patients, there were 19 identified CFTR variants, accounting for 25 out of 122 disease alleles (20%). In Puerto Rican patients, there were 16 identified CFTR variants, accounting for 36 out of 42 disease alleles (86%) in Puerto Rican patients. Thirty CFTR variants were identified overall. The most frequent variants for Dominican patients were p.Phe508del and p.Ala559Thr and for Puerto Rican patients were p.Phe508del, p.Arg1066Cys, p.Arg334Trp, and p.I507del.ConclusionsIn this first description of the CFTR variants in patients with CF from the Dominican Republic and Puerto Rico, there was a low detection rate of two CFTR variants after full sequencing with the majority of patients from the Dominican Republic without identified variants.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/153634/1/ppul24549.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/153634/2/ppul24549_am.pd

PCSK6 and Survival in Idiopathic Pulmonary Fibrosis

Rationale: Idiopathic pulmonary fibrosis (IPF) is a devastating disease characterized by limited treatment options and high mortality. A better understanding of the molecular drivers of IPF progression is needed. Objectives: To identify and validate molecular determinants of IPF survival. Methods: A staged genome-wide association study was performed using paired genomic and survival data. Stage I cases were drawn from centers across the United States and Europe and stage II cases from Vanderbilt University. Cox proportional hazards regression was used to identify gene variants associated with differential transplantation-free survival (TFS). Stage I variants with nominal significance (P < 5 x 10(-5)) were advanced for stage II testing and meta-analyzed to identify those reaching genome-wide significance (P < 5 x 10(-8)). Downstream analyses were performed for genes and proteins associated with variants reaching genome-wide significance. Measurements and Main Results: After quality controls, 1,481 stage I cases and 397 stage II cases were included in the analysis. After filtering, 9,075,629 variants were tested in stage I, with 158 meeting advancement criteria. Four variants associated with TFS with consistent effect direction were identified in stage II, including one in an intron of PCSK6 (proprotein convertase subtilisin/kexin type 6) reaching genome-wide significance (hazard ratio, 4.11 [95% confidence interval, 2.54-6.67]; P = 9.45 x 10(-9)). PCSK6 protein was highly expressed in IPF lung parenchyma. PCSK6 lung staining intensity, peripheral blood gene expression, and plasma concentration were associated with reduced TFS. Conclusions: We identified four novel variants associated with IPF survival, including one in PCSK6 that reached genome-wide significance. Downstream analyses suggested that PCSK6 protein plays a potentially important role in IPF progression

Pharmacogenetics of OATP Transporters Reveals That SLCO1B1 c.388A>G Variant Is Determinant of Increased Atorvastatin Response

Aims: The relationship between variants in SLCO1B1 and SLCO2B1 genes and lipid-lowering response to atorvastatin was investigated. Material and Methods: One-hundred-thirty-six unrelated individuals with hypercholesterolemia were selected and treated with atorvastatin (10 mg/day/4 weeks). They were genotyped with a panel of ancestry informative markers for individual African component of ancestry (ACA) estimation by SNaPshot® and SLCO1B1 (c.388A&gt;G, c.463C&gt;A and c.521T&gt;C) and SLCO2B1 (−71T&gt;C) gene polymorphisms were identified by TaqMan® Real-time PCR. Results: Subjects carrying SLCO1B1 c.388GG genotype exhibited significantly high low-density lipoprotein (LDL) cholesterol reduction relative to c.388AA+c.388AG carriers (41 vs. 37%, p = 0.034). Haplotype analysis revealed that homozygous of SLCO1B1*15 (c.521C and c.388G) variant had similar response to statin relative to heterozygous and non-carriers. A multivariate logistic regression analysis confirmed that c.388GG genotype was associated with higher LDL cholesterol reduction in the study population (OR: 3.2, CI95%:1.3–8.0, p &lt; 0.05). Conclusion: SLCO1B1 c.388A&gt;G polymorphism causes significant increase in atorvastatin response and may be an important marker for predicting efficacy of lipid-lowering therapy

Clonal chromosomal mosaicism and loss of chromosome Y in elderly men increase vulnerability for SARS-CoV-2

The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2, COVID-19) had an estimated overall case fatality ratio of 1.38% (pre-vaccination), being 53% higher in males and increasing exponentially with age. Among 9578 individuals diagnosed with COVID-19 in the SCOURGE study, we found 133 cases (1.42%) with detectable clonal mosaicism for chromosome alterations (mCA) and 226 males (5.08%) with acquired loss of chromosome Y (LOY). Individuals with clonal mosaic events (mCA and/or LOY) showed a 54% increase in the risk of COVID-19 lethality. LOY is associated with transcriptomic biomarkers of immune dysfunction, pro-coagulation activity and cardiovascular risk. Interferon-induced genes involved in the initial immune response to SARS-CoV-2 are also down-regulated in LOY. Thus, mCA and LOY underlie at least part of the sex-biased severity and mortality of COVID-19 in aging patients. Given its potential therapeutic and prognostic relevance, evaluation of clonal mosaicism should be implemented as biomarker of COVID-19 severity in elderly people. Among 9578 individuals diagnosed with COVID-19 in the SCOURGE study, individuals with clonal mosaic events (clonal mosaicism for chromosome alterations and/or loss of chromosome Y) showed an increased risk of COVID-19 lethality

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Penilaian Kinerja Keuangan Koperasi di Kabupaten Pelalawan

This paper describe development and financial performance of cooperative in District Pelalawan among 2007 - 2008. Studies on primary and secondary cooperative in 12 sub-districts. Method in this stady use performance measuring of productivity, efficiency, growth, liquidity, and solvability of cooperative. Productivity of cooperative in Pelalawan was highly but efficiency still low. Profit and income were highly, even liquidity of cooperative very high, and solvability was good