Streamlined design of a self-inactivating feline immunodeficiency virus vector for transducing ex vivo dendritic cells and T lymphocytes.

BACKGROUND: Safe and efficient vector systems for delivering antigens or immunomodulatory molecules to dendritic cells (DCs), T lymphocytes or both are considered effective means of eliciting adaptive immune responses and modulating their type, extent, and duration. As a possible tool toward this end, we have developed a self-inactivating vector derived from feline immunodeficiency virus (FIV) showing performance characteristics similar to human immunodeficiency virus-derived vectors but devoid of the safety concerns these vectors have raised. METHODS: The pseudotyped FIV particles were generated with a three-plasmid system consisting of: the packaging construct, providing Gag, Pol and the accessory proteins; the vector(s), basically containing FIV packaging signal (psi), Rev responsive element, R-U5 region at both ends, and the green fluorescent protein as reporter gene; and the Env plasmid, encoding the G protein of vesicular stomatitis virus (VSV-G) or the chimeric RD114 protein. Both packaging and vector constructs were derived from p34TF10, a replication competent molecular clone of FIV. The pseudotyped particles were produced by transient transfection in the Crandell feline fibroblast kidney (CrFK) or the human epithelial (293T) cell line. RESULTS: To broaden its species tropism, the final vector construct was achieved through a series of intermediate constructs bearing a longer psi, the FIV central polypurin tract sequence (cPPT), or the woodchuck hepatitis post-regulatory element (WPRE). These constructs were compared for efficiency and duration of transduction in CrFK or 293T cells and in the murine fibroblast cell line NIH-3T3. Whereas psi elongation and cPPT addition did not bring any obvious benefit, insertion of WPRE downstream GFP greatly improved vector performances. To maximize the efficiency of transduction for ex-vivo murine DCs and T-lymphocytes, this construct was tested with VSV-G or RD114 and using different transduction protocols. The results indicated that the FIV construct derived herein stably transduced both cell types, provided that appropriate vector makeup and transduction protocol were used. Further, transduced DCs underwent changes suggestive of an induced maturation. CONCLUSION: In contrast to previously described FIV vectors that were poorly efficient in delivering genetic material to DCs and T lymphocytes, the vector developed herein has potential for use in experimental immunization strategies

Osteopontin plasma levels and accelerated atherosclerosis in patients with CAD undergoing PCI: a prospective clinical study.

OBJECTIVES: Growing evidence supports the role played by inflammation in atherosclerosis. Identifying sensitive biomarkers is useful in predicting accelerated atherosclerosis. We investigated prospectively the relationship between plasma levels of inflammatory biomarkers [osteopontin, C-reactive protein (CRP), interleukin-6 (IL-6)] and instent restenosis, and rapid coronary plaque progression in patients with coronary artery disease (CAD) undergoing percutaneous coronary intervention (PCI). METHODS: We studied 77 patients with CAD: 45 affected by unstable angina/non-ST elevation myocardial infarction [acute coronary syndrome (ACS)], and 32 by chronic coronary syndrome (CCS). Plasma osteopontin, IL-6, and CRP levels were measured before intervention in all patients; measurements were carried out on the basis of the following time course at 1,15, 30, 90, and 180 days follow-up in a subgroup of 39 consenting patients. Clinical and biohumoral data were correlated with baseline and 6-month PCI follow-up angiography. RESULTS: Osteopontin, IL-6, and CRP were higher in patients with ACS than in those with CCS (analysis of variance: P<0.001, 0.05, and 0.05, respectively). Baseline osteopontin levels proved to be associated with rapid coronary plaque progression (P=0.005) and instent restenosis (P=0.05). The highest osteopontin levels were found in patients with CAD with both rapid plaque progression and instent restenosis (P=0.003). PCI increased inflammatory markers acutely, and osteopontin remained elevated in patients with ACS. Patients with ACS showed a higher percentage (74%) of rapid plaque progression than those with CCS (26%) (P<0.05). CONCLUSION: The study prospectively shows the link between inflammatory status and accelerated atherosclerosis in patients with CAD undergoing PCI. The baseline and persistent rise of osteopontin is an expression of its contribution to the accelerated plaque progression, and therefore osteopontin may be a useful prognostic biomarker

Rituximab in Children with Steroid-Dependent Nephrotic Syndrome: A Multicenter, Open-Label, Noninferiority, Randomized Controlled Trial.

Steroid-dependent nephrotic syndrome (SDNS) carries a high risk of toxicity from steroids or steroid-sparing agents. This open-label, noninferiority, randomized controlled trial at four sites in Italy tested whether rituximab is noninferior to steroids in maintaining remission in juvenile SDNS. We enrolled children age 1-16 years who had developed SDNS in the previous 6-12 months and were maintained in remission with high prednisone doses ( 650.7 mg/kg per day). We randomly assigned participants to continue prednisone alone for 1 month (control) or to add a single intravenous infusion of rituximab (375 mg/m(2); intervention). Prednisone was tapered in both groups after 1 month. For noninferiority, rituximab had to permit steroid withdrawal and maintain 3-month proteinuria (mg/m(2) per day) within a prespecified noninferiority margin of three times the levels among controls (primary outcome). We followed participants for 651 year to compare risk of relapse (secondary outcome). Fifteen children per group (21 boys; mean age, 7 years [range, 2.6-13.5 years]) were enrolled and followed for 6460 months (median, 22 months). Three-month proteinuria was 42% lower in the rituximab group (geometric mean ratio, 0.58; 95% confidence interval, 0.18 to 1.95 [i.e., within the noninferiority margin of three times the levels in controls]). All but one child in the control group relapsed within 6 months; median time to relapse in the rituximab group was 18 months (95% confidence interval, 9 to 32 months). In the rituximab group, nausea and skin rash during infusion were common; transient acute arthritis occurred in one child. In conclusion, rituximab was noninferior to steroids for the treatment of juvenile SDNS

Renal outcome in patients with congenital anomalies of the kidney and urinary tract.

15openopenSanna-Cherchi S; Ravani P; Corbani V; Parodi S; Haupt R; Piaggio G; Innocenti ML; Somenzi D; Trivelli A; Caridi G; Izzi C; Scolari F; Mattioli G; Allegri L; Ghiggeri GM.Sanna Cherchi, S; Ravani, P; Corbani, V; Parodi, S; Haupt, R; Piaggio, G; Innocenti, Ml; Somenzi, D; Trivelli, A; Caridi, G; Izzi, C; Scolari, Francesco; Mattioli, G; Allegri, L; Ghiggeri, G. M