Efficient new cationic liposome formulation for systemic delivery of small interfering RNA silencing tumor necrosis factor alpha in experimental arthritis.

International audienceOBJECTIVE: Tumor necrosis factor alpha (TNFalpha) is among the most prominent cytokines in rheumatoid arthritis (RA) and is secreted mainly by macrophages. A direct method for restoring the immunologic balance in RA is use of small interfering RNA (siRNA) for silencing the TNFalpha transcript. The aim of this study was to determine the therapeutic effect of systemic administration of TNFalpha siRNA in an experimental model of RA, optimizing its delivery using new liposome formulations. METHODS: Murine macrophages were transfected with siRNA targeting TNFalpha, and expression was measured. The therapeutic effect in collagen-induced arthritis (CIA) was assessed after intravenous delivery of TNFalpha siRNA. Delivery was optimized using a carrier DNA for complexation with the cationic liposome RPR209120/DOPE. Levels of TNFalpha and other cytokines were measured in sera and joint tissue-conditioned media. Biodistribution was determined using a fluorescent siRNA. RESULTS: In vitro, TNFalpha siRNA efficiently and specifically modulated the expression of TNFalpha at both the messenger RNA and protein levels. In vivo, complete cure of CIA was observed when TNFalpha siRNA was administered weekly, complexed with the liposome and combined with carrier DNA. Inhibition (50-70%) of articular and systemic TNFalpha secretion was detected in the siRNA-injected groups, which correlated with a decrease in the levels of interleukin-6 and monocyte chemotactic protein 1. The main organs targeted by siRNA were the liver and spleen; the addition of liposome RPR209120 and carrier DNA significantly increased organ uptake. CONCLUSION: We demonstrated the efficiency of systemic delivery of siRNA designed to silence TNFalpha in CIA, using a liposome carrier system as a way to address the methodologic limitations in vivo

Diagnosis of vascular Ehlers-Danlos syndrome in Italy: clinical findings and novel COL3A1 mutations.

In this study, we describe the clinical features, the behaviour of dermal fibroblasts type III collagen (COLLIII) and COL3A1 gene mutations in 22 Italian vEDS patients

Diagnosis of vascular Ehlers-Danlos syndrome in Italy: clinical findings and novel COL3A1 mutations.

In this study, we describe the clinical features, the behaviour of dermal fibroblasts type III collagen (COLLIII) and COL3A1 gene mutations in 22 Italian vEDS patients. All subjects provided written, informed consent and authorised the processing of their personal data according to Italian bioethics laws. Control and patient skin fibroblasts were established from skin biopsies and were metabolically labelled. Pepsin-purified collagens (COLLs) from the culture medium and cell layer were run in a 6% SDS/PAGE, as previously described [9]. COLLIII from the medium was evaluated based on the a1(III)3/a1(I) + a2(I) COLLs chains ratio; the control ratio range (0.09\u20130.15) was established by analysis of 10 control fibroblast strains. Direct sequencing of cDNA and/or gDNA was used to search for mutations in the COL3A1 gene by standard procedures. All of the mutations identified by cDNA analysis were verified by gDNA sequencing and were investigated in the patients\u2019 parents; novel mutations were confirmed by analysing the alleles of 400 healthy donors