| Food proteins were considered to be absorbed into the body after being digested to amino acids, dipeptides and tripeptides. However, there are studies indicating that some proteins can pass through the intestinal epithelium under normal physiological conditions, perhaps not in sufficient quantities to be of nutritional importance, but in quantities that may be antigenically or biologically active. In the present study, rat intestinal lymph samples were collected using a modified lymph fistula rat model in fasting and cow’s milk postprandial states. Low molecular weight proteins were enriched by ultrafiltration and differential solubilization, separated by1D-SDS-PAGE, digested in-gel based on molecular weight, and identified using nano-LCMS/MS. In the postprandial rat intestinal lymph, nine bovine-specific proteins (false discovery rate≤1%) were identified in different molecular weight regions. Most proteins identified in lymph were high-abundant proteins in the milk, such as β-lactoglobulin and caseins. Seven of the nine identified bovine-specific proteins are allergens in milk. This strategy can be used to search for proteins that can enter the intestinal lymph and analyze their common features. Understanding the common features of these proteins might help to develop protein drugs taken orally, so that therapeutic proteins might embody fusion domains for cross-barrier transport or translocation.Biomarker is the measurable change associated with a physiological or pathophysiological process. Unlike blood which has mechanisms to keep the internal environment homeostatic, urine is more likely to reflect changes of the body. As a result, urine is likely to be a better biomarker source than blood. However, since the urinary proteome is affected by many factors, including age, gender, medications, exercises, etc. Careful evaluation of those effects is necessary if urinary proteomics is used for biomarker discovery. Here, we evaluated the effects of three commonly-used diuretics (furosemide, F; hydrochlorothiazide, H; and spirolactone, S) on the urinary proteome in rats. Urine samples were collected before and after intragastric administration of diuretics at therapeutic doses and the proteomes were analyzed using label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS). Based on the criteria of P≤ 0.05, a fold change≥2, a spectral count≥5and false positive rate (FDR)<1%,14proteins (seven for F, five for H, and two for S) were identified by Progenesis LC-MS. The human orthologs of most of these14proteins are stable in the healthy human urinary proteome, and ten of them are reported as disease biomarkers. Thus, our results suggest that the effects of diuretics deserve more attention in future urinary protein biomarker studies. Moreover, the distinct effects of diuretics on the urinary proteome may provide clues to the mechanisms of diuretics.The kidney shows a quantifiable decrease in function with age. The glomerular filtration rate declines with20-25%from40to80years of age. Furthermore, the capacity of the kidney to concentrate urine reduces progressively with age. Urine samples from9young rats (2months) and9old rats (20months) were analyzed by label-free quantitative proteomics.37proteins were significantly changed between the two groups,17proteins were up-regulated in old group, and20proteins were down-regulated. Most of these proteins were found in the human core urinary proteome. Compared these proteins with human protein atlas database, they are highly or strongly expressed in40different tissues. Among these tissues, kidney has the most highly or strongly expressed proteins, which may due to its natural relationship with urine. The number of highly or strongly expressed proteins in brain, lung and intestines is relatively high, implying that urinary proteins may reflect the status of these organs.Renal cell carcinoma (RCC) is the most common neoplasm in adult kidney, making up3%of all adult malignancies. Histopathologically, about85%of RCC is clear-cell carcinoma. Early diagnosis of kidney-localized RCC is associated with a quite favorable prognosis with a five-year survival rate of about89%. Unfortunately, patients often present with few signs, symptoms, or laboratory abnormalities, and are frequently (-30%) diagnosed at the metastatic stage, when the prospects for cure are dismal (9%five-year survival rate).The clinical diagnosis of RCC is often confirmed by imaging studies, including ultrasonic, MRI and CT. There are currently no biomarkers available for the diagnosis of RCC. Because of the natural relationship of urine and kidney, urine is particularly suitable as a source of kidney disease biomarkers. We have identified4candidate markers by label-free quantitative proteomics from urine samples of4ccRCC patients. And then, the dis-regulated proteins are validated by ELISA in a large scale.We expect to find some usefull biomarkers of ccRCC for early diagnosis. |
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