New planar substrates for the in situ synthesis of peptide arrays

The performance of new cellulose membranes, aminofunctionalized via a PEG spacer, as a solid support in the synthesis of peptide arrays is described. The new membranes are stable to trifluoroacetic acid (TFA) and strong aqueous base for days. These properties extend the scope of synthesis considerably, e.g., more efficient side chain cleavage protocols can be applied which yielded peptides of improved purity. For the first time, cellulose membranes with a loading as high as 5 micromol/cm2 were accessible. Additionally, newly developed polypropylene membranes with hydroxy- or amino functionalities were successfully employed for the SPOT synthesis of peptides and phosphopeptides. The membranes are compatible with antibody binding as well as enzymatic phosphorylation assays.     

Sonoporation-mediated transduction of siRNA ameliorated experimental arthritis using 3 MHz pulsed ultrasound

The goal of this feasibility study was to examine whether sonoporation assisted transduction of siRNA could be used to ameliorate arthritis locally. If successful, such approach could provide an alternative treatment for the patients that have or gradually develop adverse response to chemical drugs. Tumor necrosis factor alpha (TNF-α) produced by synovial fibroblasts has an important role in the pathology of rheumatoid arthritis, inducing inflammation and bone destruction. In this study, we injected a mixture of microbubbles and siRNA targeting TNF-α (siTNF) into the articular joints of rats, and transduced siTNF into synovial tissue by exposure to a collimated ultrasound beam, applied through a probe 6 mm in diameter with an input frequency of 3.0 MHz, an output intensity of 2.0 W/cm² (spatial average temporary peak; SATP), a pulse duty ratio of 50%, and a duration of 1 min. Sonoporation increased skin temperature from 26.8 °C to 27.3 °C, but there were no adverse effect such as burns. The mean level of TNF-α expression in siTNF-treated knee joints was 55% of those in controls. Delivery of siTNF into the knee joints every 3 days (i.e., 7, 10, 13, and 16 days after immunization) by in vivo sonoporation significantly reduced paw swelling on days 20–23 after immunization. Radiographic scores in the siTNF group were 56% of those in the CIA group and 61% of those in the siNeg group. Histological examination showed that the number of TNF-α positive cells was significantly lower in areas of pannus invasion into the ankle joints of siTNF- than of siNeg-treated rats. These results indicate that transduction of siTNF into articular synovium using sonoporation may be an effective local therapy for arthritis.