Understanding the product-ion spectra of T-rich tetradeoxynucleotides is a starting point in the development of a mass spectrometric scheme to determine the mutagenicity of individual types of DNA damage. We obtained product-ion spectra for electrospray-produced ions that were activated in the ion source (electrospray ionization-source collision-activated-dissociation) and by high-energy collisions in the MS/MS mode of a four-sector instrument. We also activated singly and doubly charged ions by low-energy collisions in an ion-trap mass spectrometer and investigated post source decompositions of matrix-assisted laser desorbed ions in a time-of-flight mass spectrometer. The various methods of extracting structural information give remarkably consistent results. The difference in the relative abundances of wn and dn ions of the singly charged oligonucleotides and the formation of [a3-B3] ions, where B3 is the base on the third position, are effective for identification and distinction of pairs of isomeric tetranucleotides. A sufficient number of tetramers and pentamers were studied to enable us to propose a charge-remote mechanism for the formation of site-specific [an-Bn] ion.

To determine if cyclophilin (Cyp) inhibition can prevent renal IRI, and examine the specific contributions of CypA and CypD in promoting renal IRI. Methods: Groups of 10 mice underwent bilateral renal ischaemia and were killed 24 hours after reperfusion: (1) C57BL/6J mice were treated with 30mg/kg/BID CYPi or vehicle by oral gavage; (2) CypA-/-versus wild type (WT) mice on the 129 background. Controls were sham operated. (3) CypD-/-vs WT cultured proximal tubular epithelial cells (PTEC) were exposed to 0.5 mM of H 2 O 2 for 24hr, with or without CYPi. Results: Renal IRI caused acute renal failure in C57BL/6J mice (179.0AE19.1 vs 12.2AE1.4umol/L serum creatinine (sCr) in sham; P<0.0001). CYPi protected against renal failure (sCr 36.4AE6.7umol/L; P<0.0001). CYPi treatment reduced histologic tubular damage (P<0.0001). CYPi treatment also reduced apoptotic tubular cells (TUNEL+ cells), TNF-a mRNA levels and neutrophil and macrophage infiltration (all P<0.005 vs vehicle). Renal IRI induced acute renal failure in 129 mice (sCr 41AE13.76 vs 6.25AE1.66umol/L in sham; P<0.0001). CypA-/-mice were protected from renal dysfunction (sCr 20.7AE3.53umol/L; P<0.0001). CypA-/-mice had less histologic tubular damage (P<0.005) and lower KIM-1 mRNA levels (P<0.005). CypA-/mice also had less tubular cell death (TUNEL+ cells; P<0.001), inflammatory cytokines (TNF-a and IL-36-a PCR; P<0.05), and reduced neutrophil infiltration (P<0.001). In addition, cell culture studies showed that CYPi treatment significantly reduced H 2 O 2 induced cell death in WT PTEC (P<0.0001). CypD-/-PTEC also showed reduced H 2 O 2 induced cell death which was not further reduced by CYPi treatment, indicating that it is CypD and not the other cyclophilins that mediate oxidant-induced cell death. Conclusions: Pan-cyclophilin inhibition prevented anticipated IRIinduced acute renal failure by suppressing tubular cell death and inflammation. Our data suggest that CypA promotes leukocyte infiltration and inflammation leading to acute kidney injury following renal IRI, whilst CypD specifically contributes to cell death from the ischaemic/hypoxic insult.

Previous epidemiologic studies have established that there is a strong relationship between anterior cruciate ligament (ACL) disruption and the risk for subsequent development of osteoarthritis (OA). Though not normally considered a classical inflammatory arthropathy, OA is often associated with low-grade synovitis. In patients with OA, synovial inflammation is one factor associated with risk of progression of structural joint deterioration and symptoms. The aims of the study were to characterize the synovial features in patients undergoing ACL reconstruction after traumatic ACL rupture and to determine the relationship between inflammation and meniscal and cartilage abnormalities. Methods: The study was conducted in the context of the ACL registry at the Hospital for Special Surgery, which maintains extensive records of preclinical, intraoperative and post-operative data for 1200 patients who have undergone ACL reconstruction for traumatic ACL rupture. Synovial biopsies were collected from 40 patients during arthroscopic surgery and processed for histology and RNA extraction. Synovial features of inflammation, reaction, and degeneration were assessed using a combination histological synovial scoring system developed at our institution in which the following six features were graded: perivascular mononuclear cell infiltration, detritus, mucoid change, fibrosis, increased vascularity and hyperplasia of the synovial lining layer leading to a total combined score of 15. The following clinical data were also collected: age, sex, BMI, date of injury, time to surgery and the intraoperative presence or absence of meniscal and/or cartilage abnormalities. Suprapatellar synovial levels of mRNA for 4 chemokines (IL-8, CCL19, CCL21, and CCL5) and 1 chemokine receptor (CCR7), shown previously to be associated with synovial inflammation in patients undergoing meniscectomy by microarray pathway analysis (C Scanzello A&R 2011), were measured by real-time PCR. Results: Of the 40 patients undergoing synovial biopsy and analysis, 19 were female and 21 male, with mean ages of 29.1 and 38.4, respectively. 75% of the patients exhibited histological evidence of synovial inflammation in the suprapatellar area: 44.1% of patients scored 1 or 2, 44.1% scored 3 or 4, 5.8% scored 5 or 6, and 5.8% scored 7 or 8. Arthroscopically, meniscal abnormalities (mostly tears) were observed in 19/40 patients (47%), and cartilage defects were observed in 16/40 patients (40%). No association was observed between the synovitis score and BMI, date of injury, time to surgery, and frequency of cartilage defects or meniscal pathology. The patients were divided in two groups based on age: 19 of patients older than 30 years and 21 patients younger than 30 yr. Meniscal tears were associated with synovial inflammation in patients older than 30 yr. 16 synovial suprapatellar biopsies from patients > 30 yr were chosen for qPCR analysis, 8 each from patients without synovial inflammation (grade 0-2) and with synovial inflammation (grade 3-8). The levels of CCL5 and C...

No abstract