The native membrane fusion machinery in cells.

Recombinant SNAREs have been demonstrated as the minimal membrane fusion machinery. The participation of additional proteins in the regulation of membrane fusion has been suggested. In this study we provide nanometer-resolution images of native NSF oligomers and SNARE complexes isolated from neurons and the pancreas. Our study reveals the presence of new coiled rod-like structures in association with the SNARE complex only in neuronal tissue. Neuronal SNAREs were found coiled and super-coiled with these structures. The existence of NSF as pentamers in its native state is also demonstrated. The extent of coiling and super-coiling of SNAREs may regulate the potency and efficacy of membrane fusion in cells.     

Indianisches sprachgut im Gebrauch bei den europäern

Field materials from North Bali are presented to question conventional anthropological conceptions of culture and common practices in its analysis. The author argues that there is a need for anthropology to reshape Us assumptions, particularly in response to recent reßexive and deconstructionist critiques. A revised set of assumptions is presented with regard to cultural meanings, sharing, positioning and function; and its fruitfulness in the analysis of cultural reproduction in Bali is explored.     

Age-related responses of hepatic ATPases to starvation and cold stress in male garden lizards

Hepatic Na⁺-K⁺-ATPase and Mg²⁺-ATPase activities of male green lizards declined during the maturation phase (juvenile to 1-year-old) and stabilized thereafter. On the other hand, the Ca²⁺-ATPase activity of the liver declined during the later half of the life span (1-year-old to 2–4-year-old). Starvation stress induced a decline in hepatic Na⁺-K⁺-ATPase and Mg²⁺-ATPase activities of juvenile lizards and caused an increase in 1-year-old and 2–4-year-old counterparts. The Ca²⁺-ATPase activity declined only in starved 1-year-old lizards. Following cold stress, the hepatic Na⁺-K⁺-ATPase activity of juvenile lizards showed a higher degree of decline than 2–4-year-old counterparts. The Mg²⁺-ATPase activity declined in cold-stressed juvenile lizards, but the parameter was elevated in similarly treated 1-year-old lizards. On the other hand, the increase in Ca²⁺-ATPase activity in response to cold stress was confined only to 2–4-year-old lizards.     

Multiplex PCR and Next Generation Sequencing for the Non-Invasive Detection of Bladder Cancer

Background

Highly sensitive and specific urine-based tests to detect either primary or recurrent bladder cancer have proved elusive to date. Our ever increasing knowledge of the genomic aberrations in bladder cancer should enable the development of such tests based on urinary DNA.

Methods

DNA was extracted from urine cell pellets and PCR used to amplify the regions of the TERT promoter and coding regions of FGFR3, PIK3CA, TP53, HRAS, KDM6A and RXRA which are frequently mutated in bladder cancer. The PCR products were barcoded, pooled and paired-end 2 x 250 bp sequencing performed on an Illumina MiSeq. Urinary DNA was analysed from 20 non-cancer controls, 120 primary bladder cancer patients (41 pTa, 40 pT1, 39 pT2+) and 91 bladder cancer patients post-TURBT (89 cancer-free).

Results

Despite the small quantities of DNA extracted from some urine cell pellets, 96% of the samples yielded mean read depths >500. Analysing only previously reported point mutations, TERT mutations were found in 55% of patients with bladder cancer (independent of stage), FGFR3 mutations in 30% of patients with bladder cancer, PIK3CA in 14% and TP53 mutations in 12% of patients with bladder cancer. Overall, these previously reported bladder cancer mutations were detected in 86 out of 122 bladder cancer patients (70% sensitivity) and in only 3 out of 109 patients with no detectable bladder cancer (97% specificity).

Conclusion

This simple, cost-effective approach could be used for the non-invasive surveillance of patients with non-muscle-invasive bladder cancers harbouring these mutations. The method has a low DNA input requirement and can detect low levels of mutant DNA in a large excess of normal DNA. These genes represent a minimal biomarker panel to which extra markers could be added to develop a highly sensitive diagnostic test for bladder cancer.     

The Crohn’s Disease Risk Factor IRGM Limits NLRP3 Inflammasome Activation by Impeding Its Assembly and by Mediating Its Selective Autophagy

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Sodium Butyrate Ameliorates l‐Arginine‐Induced Pancreatitis and Associated Fibrosis in Wistar Rat: Role of Inflammation and Nitrosative Stress

Several reports indicated that histone deacetylases (HDACs) play a crucial role in inflammation and fibrogenesis. Sodium butyrate (SB) is a short‐chain fatty acid having HDAC inhibition potential. The present study aimed to evaluate the protective effect of SB against l ‐arginine (l ‐Arg)‐induced pancreatic fibrosis in Wistar rats. Pancreatic fibrosis was induced by twice intraperitoneal (i.p.) injections of 20% l ‐Arg (250 mg/100 g) at 2‐h interval on day 1, 4, 7, and 10, whereas SB (800 mg/kg/day) was administrated for 10 days. At the end of the study, biochemical estimations, histological alterations, DNA damage, and the expression of various proteins were evaluated. Posttreatment of SB decreased l ‐Arg‐induced oxidative and nitrosative stress, DNA damage, histological alterations, and fibrosis. Interestingly, posttreatment of SB significantly decreased the expression of α‐smooth muscle actin, interleukin‐1β, inducible nitric oxide synthase, and 3‐nitrotyrosine. The present study demonstrated that posttreatment of SB alleviates l ‐Arg‐induced pancreatic damage and fibrosis in rat.     

3D organization and function of the cell: Golgi budding and vesicle biogenesis to docking at the porosome complex

Insights into the three-dimensional (3D) organization and function of intracellular structures at nanometer resolution, holds the key to our understanding of the molecular underpinnings of cellular structure-function. Besides this fundamental understanding of the cell at the molecular level, such insights hold great promise in identifying the disease processes by their altered molecular profiles, and help determine precise therapeutic treatments. To achieve this objective, previous studies have employed electron microscopy (EM) tomography with reasonable success. However, a major hurdle in the use of EM tomography is the tedious procedures involved in fixing, high-pressure freezing, staining, serial sectioning, imaging, and finally compiling the EM images to obtain a 3D profile of sub-cellular structures. In contrast, the resolution limit of EM tomography is several nanometers, as compared to just a single or even sub-nanometer using the atomic force microscope (AFM). Although AFM has been hugely successful in 3D imaging studies at nanometer resolution and in real time involving isolated live cellular and isolated organelles, it has had limited success in similar studies involving 3D imaging at nm resolution of intracellular structure-function in situ. In the current study, using both AFM and EM on aldehyde-fixed and semi-dry mouse pancreatic acinar cells, new insights on a number of intracellular structure-function relationships and interactions were achieved. Golgi complexes, some exhibiting vesicles in the process of budding were observed, and small vesicles were caught in the act of fusing with larger vesicles, possibly representing either secretory vesicle biogenesis or vesicle refilling following discharge, or both. These results demonstrate the power and scope of the combined engagement of EM and AFM imaging of fixed semi-dry cells, capable of providing a wealth of new information on cellular structure-function and interactions.