Tuning the Loading and Release Properties of MicroRNA-Silencing Porous Silicon Nanoparticles by Using Chemically Diverse Peptide Nucleic Acid Payloads

Peptide nucleic acids (PNAs) are a class of artificial oligonucleotide mimics that have garnered much attention as precision biotherapeutics for their efficient hybridization properties and their exceptional biological and chemical stability. However, the poor cellular uptake of PNA is a limiting factor to its more extensive use in biomedicine; encapsulation in nanoparticle carriers has therefore emerged as a strategy for internalization and delivery of PNA in cells. In this study, we demonstrate that PNA can be readily loaded into porous silicon nanoparticles (pSiNPs) following a simple salt-based trapping procedure thus far employed only for negatively charged synthetic oligonucleotides. We show that the ease and versatility of PNA chemistry also allows for producing PNAs with different net charge, from positive to negative, and that the use of differently charged PNAs enables optimization of loading into pSiNPs. Differently charged PNA payloads determine different release kinetics and allow modulation of the temporal profile of the delivery process. In vitro silencing of a set of specific microRNAs using a pSiNP-PNA delivery platform demonstrates the potential for biomedical applications

Efficient Delivery of MicroRNA and AntimiRNA Molecules Using an Argininocalix[4]arene Macrocycle

MicroRNAs (miRNAs) are short non-coding RNA molecules acting as gene regulators by repressing translation or by inducing degradation of the target RNA transcripts. Altered expression of miRNAs may be involved in the pathogenesis of many severe human diseases, opening new avenues in the field of therapeutic strategies, i.e., miRNA targeting or miRNA mimicking. In this context, the efficient and non-toxic delivery of premiRNA and antimiRNA molecules might be of great interest. The aim of the present paper is to determine whether an argininocalix[4]arene is able to efficiently deliver miRNA, premiRNA, and antimiRNA molecules to target cells, preserving their biological activity. This study points out that (1) the toxicity of argininocalix[4]arene 1 is low, and it can be proposed for long-term treatment of target cells, being that this feature is a pre-requisite for the development of therapeutic protocols; (2) the delivery of premiRNA and antimiRNA molecules is efficient, being higher when compared with reference gold standards available; and (3) the biological activity of the premiRNAs and antimiRNAs is maintained. This was demonstrated using the argininocalix[4]arene 1 in miRNA therapeutic approaches performed on three well-described experimental model systems: (1) the induction of apoptosis by antimiR-221 in glioma U251 cells; (2) the induction of apoptosis by premiR-124 in U251 cells; and (3) the inhibition of pro-inflammatory IL-8 and IL-6 genes in cystic fibrosis IB3-1 cells. Our results demonstrate that the argininocalix[4]arene 1 should be considered a very useful delivery system for efficient transfer to target cells of both premiRNA and antimiRNA molecules, preserving their biological activity

A peptide-nucleic acid targeting miR-335-5p enhances expression of cystic fibrosis transmembrane conductance regulator (CFTR) gene with the possible involvement of the CFTR scaffolding protein NHERF1

(1) Background: Up-regulation of the Cystic Fibrosis Transmembrane Conductance Regulator gene (CFTR) might be of great relevance for the development of therapeutic protocols for cystic fibrosis (CF). MicroRNAs are deeply involved in the regulation of CFTR and scaffolding proteins (such as NHERF1, NHERF2 and Ezrin). (2) Methods: Content of miRNAs and mRNAs was analyzed by RT-qPCR, while the CFTR and NHERF1 production was analyzed by Western blotting. (3) Results: The results here described show that the CFTR scaffolding protein NHERF1 can be upregulated in bronchial epithelial Calu-3 cells by a peptide-nucleic acid (PNA) targeting miR-335-5p, predicted to bind to the 3′-UTR sequence of the NHERF1 mRNA. Treatment of Calu-3 cells with this PNA (R8-PNA-a335) causes also up-regulation of CFTR. (4) Conclusions: We propose miR-335-5p targeting as a strategy to increase CFTR. While the efficiency of PNA-based targeting of miR-3355p should be verified as a therapeutic strategy in CF caused by stop-codon mutation of the CFTR gene, this approach might give appreciable results in CF cells carrying other mutations impairing the processing or stability of CFTR protein, supporting its application in personalized therapy for precision medicine

Enzymatic spermine metabolites induce apoptosis associated with increase of p53, caspase-3 and mir-34a in both neuroblastoma cells, SJNKP and the N-Myc-amplified form IMR5

Neuroblastoma (NB) is a common malignant solid tumor in children and accounts for 15% of childhood cancer mortality. Amplification of the N-Myc oncogene is a well-established poor prognostic marker in NB patients and strongly correlates with higher tumor aggression and resistance to treatment. New therapies for patients with N-Myc-amplified NB need to be developed. After treating NB cells with BSAO/SPM, the detection of apoptosis was determined after annexin V-FITC labeling and DNA staining with propidium iodide. The mitochondrial membrane potential activity was checked, labeling cells with the probe JC-1 dye. We analyzed, by real-time RT-PCR, the transcript of genes involved in the apoptotic process, to determine possible down-or upregulation of mRNAs after the treatment on SJNKP and the N-Myc-amplified IMR5 cell lines with BSAO/SPM. The experiments were carried out considering the proapoptotic genes Tp53 and caspase-3. After treatment with BSAO/SPM, both cell lines displayed increased mRNA levels for all these proapoptotic genes. Western blotting analysis with PARP and caspase-3 antibody support that BSAO/SPM treatment induces high levels of apoptosis in cells. The major conclusion is that BSAO/SPM treatment leads to antiproliferative and cytotoxic activity of both NB cell lines, associated with activation of apoptosis

Purification and functional characterisation of rhiminopeptidase A, a novel aminopeptidase from the venom of Bitis gabonica rhinoceros

This study describes the discovery and characterisation of a novel aminopeptidase A from the venom of B. g. rhinoceros and highlights its potential biological importance. Similar to mammalian aminopeptidases, rhiminopeptidase A might be capable of playing roles in altering the blood pressure and brain function of victims. Furthermore, it could have additional effects on the biological functions of other host proteins by cleaving their N-terminal amino acids. This study points towards the importance of complete analysis of individual components of snake venom in order to develop effective therapies for snake bites

Matéria seca e nutrientes em plantas de pinhão manso (Jatropha curcas) em áreas de Cambissolo de Rio das Flores - RJ.

O Pinhão manso (Jatropha Curcas L.) é uma cultura de potencial econômico para a produção de biodiesel no Estado do Rio de Janeiro. O estudo foi realizado no município de Rio das Flores, Rio de Janeiro RJ, com o objetivo avaliar a matéria seca acumulada na parte aérea e nas raízes de plantas de pinhão manso amostradas em áreas sob regime hídrico distinto. Os teores de C, P, K, Ca e Mn nesses tecidos foram também avaliados. Doze plantas em cada área foram amostradas e seus tecidos pesados no campo. Sub-amostras de cada compartimento foram analisadas quimicamente. As plantas amostradas no ambiente mais úmido apresentaram teores de todos os nutrientes (exceto C) mais altos, embora a fertilidade das áreas não tenha apresentado diferenças marcantes. O pinhão Para tanto, foi realizada a coleta aleatória de quatro amostras de plantas na área total. Os materiais vegetais amostrados foram medidos, pesados, postos a secar e submetidos ás análises de carbono total e foi feitas coletas de amostras de solo. A partir dos resultados, os teores de carbono não tiveram grandes variações, portanto, somente o caule, próximo ao rio obteve uma concentração do teor de carbono com uma média de 351,9, nas observações de matéria seca, o experimento próximo ao rio, obteve uma distribuição de biomassa em todo os compartimentos do vegetal, ao contrário do experimento distante do rio. Já as análises de crescimento sofreu uma interrupção, decorrente a uma chuva de granizo, portanto, após este evento as plantas tiveram um crescimento reduzido

Health related quality of life measure in systemic pediatric rheumatic diseases and its translation to different languages: an international collaboration

Background: Rheumatic diseases in children are associated with significant morbidity and poor health-related quality of life (HRQOL). There is no health-related quality of life (HRQOL) scale available specifically for children with less common rheumatic diseases. These diseases share several features with systemic lupus erythematosus (SLE) such as their chronic episodic nature, multi-systemic involvement, and the need for immunosuppressive medications. HRQOL scale developed for pediatric SLE will likely be applicable to children with systemic inflammatory diseases.Findings: We adapted Simple Measure of Impact of Lupus Erythematosus in Youngsters (SMILEY (c)) to Simple Measure of Impact of Illness in Youngsters (SMILY (c)-Illness) and had it reviewed by pediatric rheumatologists for its appropriateness and cultural suitability. We tested SMILY (c)-Illness in patients with inflammatory rheumatic diseases and then translated it into 28 languages. Nineteen children (79% female, n= 15) and 17 parents participated. the mean age was 12 +/- 4 years, with median disease duration of 21 months (1-172 months). We translated SMILY (c)-Illness into the following 28 languages: Danish, Dutch, French (France), English (UK), German (Germany), German (Austria), German (Switzerland), Hebrew, Italian, Portuguese (Brazil), Slovene, Spanish (USA and Puerto Rico), Spanish (Spain), Spanish (Argentina), Spanish (Mexico), Spanish (Venezuela), Turkish, Afrikaans, Arabic (Saudi Arabia), Arabic (Egypt), Czech, Greek, Hindi, Hungarian, Japanese, Romanian, Serbian and Xhosa.Conclusion: SMILY (c)-Illness is a brief, easy to administer and score HRQOL scale for children with systemic rheumatic diseases. It is suitable for use across different age groups and literacy levels. SMILY (c)-Illness with its available translations may be used as useful adjuncts to clinical practice and research.Rutgers State Univ, Robert Wood Johnson Med Sch, New Brunswick, NJ 08903 USARutgers State Univ, Child Hlth Inst New Jersey, New Brunswick, NJ 08901 USAHosp Special Surg, New York, NY 10021 USAUniv Michigan, Ann Arbor, MI 48109 USARed Cross War Mem Childrens Hosp, Cape Town, South AfricaAin Shams Univ, Pediat Allergy Immunol & Rheumatol Unit, Cairo, EgyptAin Shams Univ, Pediat Rheumatol Pediat Allergy Immunol & Rheum, Cairo, EgyptKing Faisal Specialist Hosp & Res Ctr, Riyadh 11211, Saudi ArabiaCharles Univ Prague, Prague, Czech RepublicGen Univ Hosp, Prague, Czech RepublicUniv Hosp Motol, Dept Pediat, Prague, Czech RepublicAarhus Univ, Hosp Skejby, Aarhus, DenmarkRigshosp, Juliane Marie Ctr, DK-2100 Copenhagen, DenmarkUniv Med Ctr, Dept Pediat Immunol, Utrecht, NetherlandsWilhelmina Childrens Hosp, Utrecht, NetherlandsGreat Ormond St Hosp Sick Children, Children NHS Fdn Trust, Renal Unit, London, EnglandLyon Univ, Hosp Civils Lyon, Rheumatol & Dermatol Dept, Lyon, FranceMed Univ Innsbruck, A-6020 Innsbruck, AustriaPrim Univ Doz, Bregenz, AustriaHamburg Ctr Pediat & Adolescence Rheumatol, Hamburg, GermanyAsklepios Clin Sankt, Augustin, GermanyUniv Zurich, Childrens Hosp, Zurich, SwitzerlandAristotle Univ Thessaloniki, Pediat Immunol & Rheumatol Referral Ctr, GR-54006 Thessaloniki, GreeceIsrael Meir Hosp, Kefar Sava, IsraelSanjay Gandhi Postgrad Inst Med Sci, Lucknow, Uttar Pradesh, IndiaSemmelweis Univ, H-1085 Budapest, HungaryAnna Meyer Hosp, Florence, ItalyUniv Siena, Res Ctr System Autoimmune & Autoinflammatory Dis, I-53100 Siena, ItalyUniv Florence, Florence, ItalyOsped Pediat Bambino Gesu, IRCCS, Pediat Rheumatol Unit, Rome, ItalyUniv Genoa Pediat II Reumatol, Ist G Gaslini EULAR, Ctr Excellence Rheumatol, Genoa, ItalyUniv Cattolica Sacro Cuore, Inst Pediat, Rome, ItalyUniv Padua, Dept Pediat, Pediat Rheumatol Unit, Padua, ItalyYokohama City Univ, Sch Med, Yokohama, Kanagawa 232, JapanUniv Estadual Paulista, UNESP, Botucatu, SP, BrazilUniversidade Federal de São Paulo, Dept Pediat, São Paulo, BrazilUniv Estadual Campinas, Dept Med, Campinas, SP, BrazilUniv Fed Rio de Janeiro, Dept Pediat, Rio de Janeiro, BrazilUniv Estado do, Adolescent Hlth Care Unit, Div Pediat Rheumatol, Rio de Janeiro, BrazilUniv São Paulo, Fac Med, Childrens Inst, Dept Pediat,Pediat Rheumatol Unit, São Paulo, BrazilChildrens Inst, Pediat Rheumatol Unit, São Paulo, BrazilClin Pediat I, Cluj Napoca, RomaniaInst Rheumatol, Belgrade, SerbiaUniv Childrens Hosp, Univ Med Ctr Ljubljana, Ljubljana, SloveniaHead Rheumatol Hosp Pedro Elizalde, Buenos Aires, DF, ArgentinaHosp Gen Mexico City, Mexico City, DF, MexicoHosp Infantil Mexico Fed Gomez, Mexico City, DF, MexicoHosp San Juan Dios, Barcelona, SpainHosp Univ Valle Hebron, Barcelona, SpainMt Sinai Med Ctr, New York, NY 10029 USAMt Sinai Med Ctr, Miami Beach, FL 33140 USAComplejo Hosp Univ Ruiz & Paez, Bolivar, VenezuelaHacettepe Univ, Dept Pediat, Ankara, TurkeyIstanbul Univ, Cerrahpasa Med Sch, Istanbul, TurkeyFMF Arthrit Vasculitis & Orphan Dis Res Ctr, Inst Hlth Sci, Ankara, TurkeyUniv Calgary, Dept Pediat, Alberta Childrens Hosp, Res Inst, Calgary, AB T2N 1N4, CanadaUniversidade Federal de São Paulo, Dept Pediat, São Paulo, BrazilWeb of Scienc

A transcriptomic analysis of gene expression in the venom gland of the snake Bothrops alternatus (urutu)

<p>Abstract</p> <p>Background</p> <p>The genus <it>Bothrops </it>is widespread throughout Central and South America and is the principal cause of snakebite in these regions. Transcriptomic and proteomic studies have examined the venom composition of several species in this genus, but many others remain to be studied. In this work, we used a transcriptomic approach to examine the venom gland genes of <it>Bothrops alternatus</it>, a clinically important species found in southeastern and southern Brazil, Uruguay, northern Argentina and eastern Paraguay.</p> <p>Results</p> <p>A cDNA library of 5,350 expressed sequence tags (ESTs) was produced and assembled into 838 contigs and 4512 singletons. BLAST searches of relevant databases showed 30% hits and 70% no-hits, with toxin-related transcripts accounting for 23% and 78% of the total transcripts and hits, respectively. Gene ontology analysis identified non-toxin genes related to general metabolism, transcription and translation, processing and sorting, (polypeptide) degradation, structural functions and cell regulation. The major groups of toxin transcripts identified were metalloproteinases (81%), bradykinin-potentiating peptides/C-type natriuretic peptides (8.8%), phospholipases A₂(5.6%), serine proteinases (1.9%) and C-type lectins (1.5%). Metalloproteinases were almost exclusively type PIII proteins, with few type PII and no type PI proteins. Phospholipases A₂were essentially acidic; no basic PLA₂were detected. Minor toxin transcripts were related to L-amino acid oxidase, cysteine-rich secretory proteins, dipeptidylpeptidase IV, hyaluronidase, three-finger toxins and ohanin. Two non-toxic proteins, thioredoxin and double-specificity phosphatase Dusp6, showed high sequence identity to similar proteins from other snakes. In addition to the above features, single-nucleotide polymorphisms, microsatellites, transposable elements and inverted repeats that could contribute to toxin diversity were observed.</p> <p>Conclusions</p> <p><it>Bothrops alternatus </it>venom gland contains the major toxin classes described for other <it>Bothrops </it>venoms based on trancriptomic and proteomic studies. The predominance of type PIII metalloproteinases agrees with the well-known hemorrhagic activity of this venom, whereas the lower content of serine proteases and C-type lectins could contribute to less marked coagulopathy following envenoming by this species. The lack of basic PLA₂agrees with the lower myotoxicity of this venom compared to other <it>Bothrops </it>species with these toxins. Together, these results contribute to our understanding of the physiopathology of envenoming by this species.</p

American College of Rheumatology Provisional Criteria for Clinically Relevant Improvement in Children and Adolescents With Childhood-Onset Systemic Lupus Erythematosus

10.1002/acr.23834ARTHRITIS CARE & RESEARCH715579-59