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A long hike in the country can give you many things. A refreshed, relaxed mind to clear away the city cobwebs and a pleasant drink in the hostelry at the end of the walk spring to mind. So do the potential discomforts, including stiff legs, sore feet and blisters. There is nothing quite like the pain of a fat, weeping blister to weaken that resolve about getting out more and taking more exercise. In the medical world, however, blisters can be a source of useful information. Not the blisters inflicted by walking or by a poorly fitting pair of shoes, but artificial ones induced on the skin by suction. The fluid drawn into them can be used to measure endogenous metabolites or the pharmacokinetic profiles of drugs in the body. And now the world of proteomics has latched on to the potential of blister fluid for revealing biomarkers of disease. Blisters are produced on the skin, so might be a repository for proteins or peptides located in the interstitial fluid or the underlying tissue during skin diseases. One study published in 2006 identified a total of 670 proteins in artificial blister fluid and found that nine proteins were differentially expressed in a patient with plaque psoriasis compared with a control subject. A more general study of blister fluid has now been undertaken by European researchers who have compared the proteomic profile of blister fluid with that of serum determined under identical procedures. Albert Heck from Utrecht University with colleagues from The Netherlands and Norway explained that blister fluid obtained by suction, although invasive, is far less so than a skin biopsy and could provide a valid alternative for studying skin disorders. A skin suction device was used to create the blisters. It had a suction chamber fitted with seven holes (5 mm diameter each) through which the skin could expand. When placed on the skin, a pump withdrew air from the chamber, which was left in place until blisters containing sufficient fluid had formed, typically up to 2.5 hours. The blisters each contained about 30-100 µL fluid which was withdrawn by syringe. Serum was collected from a different set of volunteers. Before the blister fluid or serum were analysed, the top six or top twelve most abundant proteins were removed to facilitate detection of the less abundant proteins. The largest enrichment factor was achieved with the twelve-protein depletion. It removed about 95% of the protein content from both fluids so was adopted for the main studies. The proteins from both fluids were compared by two-dimensional gel electrophoresis and, after digestion with trypsin, by dual column HPLC/MS/MS on a high resolution FT/ICR mass spectrometer. The 30 fractions obtained in the first separation were each analysed by reversed-phase chromatography in the second dimension and the data were searched against the standard SwissProt database for protein identification. The overall 2D gel profiles from both media were fairly similar both before and after protein depletion. The total concentration of proteins in the blister fluid was about 5-fold lower than that in serum. Nevertheless, using stringent criteria that gave a false positive rate of less than 1%, 410 proteins were identified in blister fluid, compared with only 240 in serum using identical protocols. Virtually all of the serum proteins were also found in blister fluid. Conversely, 34 proteins were identified in blister fluid that were absent from serum. The team noted that several classes of blister fluid proteins were related to cell-mediated processes such as metabolism, protein synthesis, oxidation state and cell shape. So, it appears likely that they "result from cell leakage processes during blister formation in addition to blood related proteins that diffuse via extracellular spaces into the blisters." The proteins unique to blister fluid included some which have previously been recognised as biomarkers, such as cystatin A and ezrin (skin tumours), cystatin C (renal function), C-reactive protein (psoriasis) and interleukin-6 signal transducer, also known as gp130 (scleroderma). Although no new biomarkers were found, that was not one of the aims of the study. The preliminary data show that known biomarkers can be measured in blister fluid, especially following abundant protein depletion, suggesting that it should be considered as an alternative body fluid for biomarker detection, especially for diseases of the skin. Related links:
The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.
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