Shear stress induces cell apoptosis via a c-Src-phospholipase D-mTOR signaling pathway in cultured podocytes

The glomerular capillary wall, composed of endothelial cells, the glomerular basement membrane and the podocytes, is continually subjected to hemodynamic force arising from tractional stress due to blood pressure and shear stress due to blood flow. Exposure of glomeruli to abnormal hemodynamic force such as hyperfiltration is associated with glomerular injury and progressive renal disease, and the conversion of mechanical stimuli to chemical signals in the regulation of the process is poorly understood in podocytes. By examining DNA fragmentation, apoptotic nuclear changes and cytochrome c release, we found that shear stress induced cell apoptosis in cultured podocytes. Meanwhile, podocytes exposed to shear stress also stimulated c-Src phosphorylation, phospholipase D (PLD) activation and mammalian target of rapamycin (mTOR) signaling. Using the antibodies against c-Src, PLD(1), and PLD(2) to perform reciprocal co-immunoprecipitations and in vitro PLD activity assay, our data indicated that c-Src interacted with and activated PLD(1) but not PLD(2). The inhibition of shear stress-induced c-Src phosphorylation by PP(2) (a specific inhibitor of c-Src kinase) resulted in reduced PLD activity. Phosphatidic acid, produced by shear stress-induced PLD activation, stimulated mTOR signaling, and caused podocyte hypertrophy and apoptosis

Automated fluorometer/photometer system for homogeneous immunoassays.

Abstract A fully automated bench-top clinical analyzer (OPTIMATE TM; Ames/Gilford) performs homogeneous fluorescent immunoassays, colorimetric immunoassays, and determinations of routine blood analytes; drugs, enzymes, metabolites, specific proteins, and hormones in serum. Unique features include a combination fluorescence/absorbance aspirating thermocuvette, a photon-counting fluorometer/photometer, a multi-reagent distribution valve to dispense as many as three reagents plus buffer, and a user-replaceable programmable memory cartridge for software updates. We have evaluated the performance of OPTIMATE substrate-labeled fluorescent immunoassays for gentamicin, tobramycin, amikacin, theophylline, phenytoin, phenobarbital, primidone, carbamazepine, and quinidine with this automated system. A sample throughput of 92 samples per hour is achieved by reading fixed-point fluorescence results every 39 s after an initial 4-min reaction period. Precision studies indicate typical CVs of less than or equal to 6% for mid-range controls. Standard curves can be reused for as long as two weeks before recalibration. With clinical samples, results by the OPTIMATE procedure correlated well (r greater than or equal to 0.97) with those by a reference method.</jats:p

Towards standardisation of evidence-based clinical care process specifications.

There is a strong push towards standardisation of treatment approaches, care processes and documentation of clinical practice. However, confusion persists regarding terminology and description of many clinical care process specifications which this research seeks to resolve by developing a taxonomic characterisation of clinical care process specifications. Literature on clinical care process specifications was analysed, creating the starting point for identifying common characteristics and how each is constructed and used in the clinical setting. A taxonomy for clinical care process specifications is presented. The De Bleser approach to limited clinical care process specifications characterisation was extended and each clinical care process specification is successfully characterised in terms of purpose, core elements and relationship to the other clinical care process specification types. A case study on the diagnosis and treatment of Type 2 Diabetes in the United Kingdom was used to evaluate the taxonomy and demonstrate how the characterisation framework applies. Standardising clinical care process specifications ensures that the format and content are consistent with expectations, can be read more quickly and high-quality information can be recorded about the patient. Standardisation also enables computer interpretability, which is important in integrating Learning Health Systems into the modern clinical environment. The approach presented allows terminologies for clinical care process specifications that were widely used interchangeably to be easily distinguished, thus, eliminating the existing confusion