Osteopontin-stimulated expression of matrix metalloproteinase-9 causes cardiomyopathy in the mdx model of Duchenne muscular dystrophy

Duchenne muscular dystrophy (DMD), caused by mutations in the dystrophin gene, is a common and lethal form of muscular dystrophy. With progressive disease, most patients succumb to death from respiratory or heart failure, or both. However, the mechanisms, especially those governing cardiac inflammation and fibrosis in DMD, remain less understood. Matrix metalloproteinase (MMPs) are a group of extracellular matrix proteases involved in tissue remodeling in both physiologic and pathophysiologic conditions. Previous studies have shown that MMP-9 exacerbates myopathy in dystrophin-deficient mdx mice. However, the role and the mechanisms of action of MMP-9 in cardiac tissue and the biochemical mechanisms leading to increased levels of MMP-9 in mdx mice remain unknown. Our results demonstrate that the levels of MMP-9 are increased in the heart of mdx mice. Genetic ablation of MMP-9 attenuated cardiac injury, left ventricle dilation, and fibrosis in 1-y-old mdx mice. Echocardiography measurements showed improved heart function in Mmp9-deficient mdx mice. Deletion of the Mmp9 gene diminished the activation of ERK1/2 and Akt kinase in the heart of mdx mice. Ablation of MMP-9 also suppressed the expression of MMP-3 and MMP-12 in the heart of mdx mice. Finally, our experiments have revealed that osteopontin, an important immunomodulator, contributes to the increased amounts of MMP-9 in cardiac and skeletal muscle of mdx mice. This study provides a novel mechanism for development of cardiac dysfunction and suggests that MMP-9 and OPN are important therapeutic targets to mitigating cardiac abnormalities in patients with DMD.     

Diurnal rhythm of emergence from pupae in parasitic wasp Trichogramma evanescens Westw. (Insecta: Hymenoptera)

Abstract

Pupal eclosion of Trichogramma evanescens Westw. was studied in different conditions of light‐darkness and temperature fluctuations. The results revealed that under natural light cycles Trichogramma exhibits a distinct rhythm of emergence from pupae. Maximum emergence takes place in the morning. This rhythm persists in constant dim red light and temperature, so it is endogenous in nature. The rhythm can be entrained by artificial 24‐h temperature cycles or by day‐night cycles of light with a very low intensity of illumination (<0.01 lux). Nevertheless a single pulse of bright light or of high temperature is not able to reset the rhythm. The emergence rhythm was also absent if the culture was grown in constant darkness and temperature.     

Construction of amperometric uric acid biosensor based on uricase immobilized on PBNPs/cMWCNT/PANI/Au composite

A chitosan-glutaraldehyde crosslinked uricase was immobilized onto Prussian blue nanoparticles (PBNPs) absorbed onto carboxylated multiwalled carbon nanotube (c-MWCNT) and polyaniline (PANI) layer, electrochemically deposited on the surface of Au electrode. The nanohybrid-uricase electrode was characterized by scanning electron microscopic (SEM), Fourier transform infrared spectroscopy (FTIR) and cyclic voltammetry. An amperometric uric acid biosensor was fabricated using uricase/c-MWCNT/PBNPs/Au electrode as working electrode, Ag/AgCl as standard and Pt wire as auxiliary electrode connected through a potentiostat. The biosensor showed optimum response within 4 s at pH 7.5 and 40 °C, when operated at 0.4 V vs. Ag/AgCl. The linear working range for uric acid was 0.005-0.8 mM, with a detection limit of 5 μM. The sensor was evaluated with 96% recovery of added uric acid in sera and 4.6 and 5.4% within and between batch of coefficient of variation respectively and a good correlation (r = 0.99) with standard enzymic colorimetric method. This sensor measured uric acid in real serum samples. The sensor lost only 37% of its initial activity after its 400 uses over a period of 7 months, when stored at 4 °C.     

Breast cancer hypofractionated radiotherapy in 2-weeks with 2D technique: 5-year clinical outcomes of a phase 2 trial

BackgroundTo report clinical outcomes and late toxicities of a 2-week hypofractionated post-operative loco-regional radiotherapy in patients with breast cancer.Materials and methodsThis trial was approved by the Institutional Ethics Committee and registered with gov, no. {"type":"clinical-trial","attrs":{"text":"NCT02460744","term_id":"NCT02460744"}}NCT02460744. Between June 2013 and October 2014, 50 patients with breast cancer, post mastectomy or breast conserving surgery (BCS) were included in this study, of whom 10 had BCS. Patients were planned on a 2-dimentional (2D) simulator with 2 tangential fields and an incident supraclavicular field. Radiotherapy dose was 34 Gy/10#/2 weeks and a sequential boost of 10 Gy/5#/1 wk in BCS patients. The primary endpoint was the rate of acute skin toxicities previously reported. Here, we report the secondary end points of late toxicities, cosmesis, local recurrence, disease-free survival (DFS) and overall survival (OS). Late skin toxicities were recorded according to the Radiotherapy and Oncology Group (RTOG) scoring criteria. Cosmetic outcomes were assessed using the Harvard/National Surgical Adjuvant Breast and Bowel Project (NSABP)/RTOG breast cosmesis and the Late Effects Normal Tissue/Subjective Objective Management Analytic (LENT/SOMA) scales for the breast and chest wall, respectively. Kaplan-Meier estimates of DFS and OS were calculated, and 5-year DFS and OS rates (with approximate 95% CIs) were estimated.ResultsLate grade ≥ 2 chest wall induration, hypopigmentation and subcutaneous fibrosis were seen in 3 (6%), 3 (6%) and 1 (2%) patients, respectively. Chest wall cosmesis was excellent/good in 34 (72%) and fair/bad in 13 (28%) patients. In BCS patients, grade 2 skin induration, subcutaneous fibrosis and edema was observed in 1 patient (11%) each. Cosmesis was excellent/good in 7 (78%) and fair/bad in 2 (22%) patients. Late grade ≥ 2 arm edema, pain and shoulder stiffness were reported by 1 (2%), 2 (4%) and 2 (4%) patients, respectively. No local recurrences were observed. Five patients developed distant metastases (10%). Seven patients died (14%). The 5-year DFS and OS rate was 90% (95% CI: 77–96%) and 88% (95% CI: 75–94%), respectively.ConclusionHypofractionated radiotherapy in 2 weeks in patients with breast cancer was associated with minimal late toxicity, good cosmetic outcome and excellent local control. This trial may be of relevance for developing countries where resources are limited.     

The role of sphingomyelin synthetase and sphingomyelinase in 1,2-dimethylhydrazine-induced lipid alterations of rat colonic plasma membranes

Recently, our laboratory, utilizing the 1,2-dimethylhydrazine model of colonic adenocarcinoma, demonstrated alterations in the 'dynamic component' of fluidity in brush-border membranes prepared from distal colonocytes of rats administered this agent for 5, 10 and 15 weeks, i.e., before the development of colon cancer. Furthermore, changes in the sphingomyelin content and sphingomyelin/phosphatidylcholine molar ratio of these membranes appeared, at least partially, to be responsible for these fluidity alterations. In an attempt to elucidate the mechanism(s) involved in these dimethylhydrazine-induced lipid changes, in the present studies the activities of sphingomyelin synthetase and magnesium-dependent neutral sphingomyelinase, enzymes involved in the synthesis and degradation of this phospholipid, respectively, were examined and compared in distal colonic brush-border membranes prepared from rats after 5, 10 or 15 weeks administration of dimethylhydrazine or diluent. The results of these studies demonstrate that alterations in both these enzymatic activities can be detected after administration of dimethylhydrazine and appear to, at least in part, be responsible for the changes in membrane sphingomyelin composition noted previously. These results as well as a discussion of their possible serve as the basis for the present report.     

Dexamethasone-induced alterations in lipid composition and fluidity of rat proximal-small-intestinal brush-border membranes.

A series of experiments were conducted to examine the possible effects of subcutaneous administration of the synthetic glucocorticoid dexamethasone (100 micrograms/day per 100 g body wt.) on the lipid fluidity and lipid composition of rat proximal-small-intestinal brush-border membranes. After 4 days of treatment, membranes and their liposomes prepared from treated animals possessed a greater fluidity than did their control (diluent, 0.9% NaCl) counterparts, as assessed by steady-state fluorescence-polarization techniques using several different fluorophores. Examination of the effects of temperature on the anisotropy values of 1,6-diphenylhexa-1,3,5-triene, using Arrhenius plots, moreover, revealed that the mean break-point temperatures of the treated preparations were approx. 3-4 degrees C lower than those of their control-preparation counterparts. Changes in the sphingomyelin/phosphatidylcholine (PC) molar ratio as well as in certain of the fatty acids of the PC fraction of treated membranes, secondary to alterations in membrane PC levels and in lysophosphatidylcholine acyltransferase activities respectively, were also noted after dexamethasone administration. These compositional alterations appeared to be responsible, at least in part, for the differences in fluidity noted between treated and control plasma membranes. These results therefore demonstrate that dexamethasone administration can modulate the lipid fluidity and lipid composition of rat proximal-small-intestinal brush-border membranes.     

Hypofractionated radiotherapy in young versus older women with breast cancer: a retrospective study from India

BackgroundYoung women with breast cancer (BC) are not represented in the trials on hypofractionation. In this study we compared outcomes in young patients with BC to their older counterparts treated with hypofractionated radiotherapy (RT) in a regional cancer centre in India.Materials and methodsBetween January 1990 to December 2010, women with BC, treated with hypofractionated RT dose of 35–40 Gy/15#/3 weeks were divided into two groups, ≤ 35 years and > 35 years. Outcomes compared were locoregional recurrence rate (LRR), locoregional recurrence-free survival (LRRFS), disease-free survival (DFS), overall survival (OS) and toxicities. LRRFS, DFS and OS were estimated using the Kaplan-Meier method.ResultsOf total 2244 patients, 359 were ≤ 35 years of age and 1885 were > 35 years. Patient and disease characteristics were comparable between the two groups, except that comorbidities were significantly higher in the > 35 years age group, more patients aged ≤ 35 years had nodal N3 disease, received chemotherapy and RT to internal mammary nodes and more patients in the > 35 years group received hormonal therapy. Median follow up was 10 years (range 1–30 years). LRR and distant metastases were comparable between the two groups. However, synchronous LRR and distant metastases were significantly higher in the ≤ 35 years group 18 (5.1%) as compared to the > 35 years group 39 (2.1%) with p = 0.018. Estimated 10-year LRRFS, DFS and OS were 92% vs. 94% (p = 0.95), 68% vs. 73%(p = 0.058) and 78% vs. 76% (p = 0.10) in ≤ 35 years and > 35 years, respectively. OS for stage 1 was comparable between the two groups. However, for stage 2 and 3 it was 77% vs. 82% (p = 0.048) and 53% vs. 62% (p = 0.045) in the ≤ 35 years and > 35 years group, respectively. Acute and late toxicity were similar in the two groups.ConclusionYoung BC patients had higher LRR and distant metastases. LRRFS, DFS and toxicities were comparable between the two groups. However, OS was poorer in young BC patients with stage 2 and 3 disease.     

Immunogold localization of callose and other cell wall components in pea nodule transfer cells

Summary Transfer cells are located adjacent to xylem and phloem elements in pea nodule vascular tissues. The composition of the labyrinthine wall intrusions was investigated by immunogold labeling using specific antibody probes. Callose antigen was found at the base of newly formed cell wall intrusions and also in adjacent plasmodesmata. Sections through developed labyrinthine intrusions revealed that wall ingrowths had an internal structure with small domains of callose suggesting the presence of channels or vents. Xyloglucan and pectin antigens were uniformly distributed within the wall, but the distribution of extensin antigens was variable, with different antigens being detected in different regions of the wall ingrowth. A lectinlike glycoprotein, PsNLEC-1, was localized in intercellular spaces associated with nodule transfer cells. Previously, expression of this component was observed in other types of cells showing complex involution of the plasma membrane, namely root cortical cells harboring arbuscular mycorrhizae and nodule cells harboring nitrogen-fixing rhizobia.