Local vibration induced vascular pathological structural changes and abnormal levels of vascular damage indicators.

BACKGROUND: The vascular component of the hand-arm-vibration syndrome (HAVS) is often characterized by vibration-induced white fingers (VWF). Active substances secreted by the vascular endothelial cells (VEC) maintain a dynamic balance but damage to the blood vessels may occur when the equilibrium is altered, thus forming an important pathological basis for VWF. This study was aimed at investigating vascular damage indicators as a basis for an early detection of disorders caused by vibration, using the rat tail model. METHODS AND RESULTS: Experiments were conducted using a control group of rats not exposed to vibration while two exposed groups having different exposure durations of 7 and 14 days were randomly formed. Following exposure, the structural changes of tail tissue samples in anesthetized rats were observed. Enzyme-linked immunosorbent assay (ELISA) was used for analyzing four vascular damage indicators myosin regulatory light chain (MLC2), endothelin-1 (ET-1), vinculin (VCL) and 5-hydroxytryptamine (5-HT) in tail blood samples. We found that both vascular smooth muscle and endothelial cells displayed changes in morphology characterized by vacuolization and swelling in the vibration-exposed group. The levels of vascular damage indicators were altered under the vibration. CONCLUSION: The degree of vascular pathology increased with the longer duration exposure. Furthermore, the levels of MLC2, ET-1 and 5-HT in rat plasma were associated with vascular injury caused by local vibration.

Identification of Vinculin as a Potential Diagnostic Biomarker for Acute Aortic Dissection Using Label-Free Proteomics.

Acute aortic dissection (AAD) is an emergent vascular disease. Currently, its diagnosis depends on clinical and radiological investigations but lacking of serum biomarkers. In this study, we aimed to identify potential serum biomarkers for AAD using label-free proteomics approach. A total of 90 serum samples were collected from three groups: patients with acute aortic dissection (AAD, n = 30), patients with acute myocardial infarction (AMI, n = 30), and healthy controls (n = 30), and the first four samples from each group were selected for label-free proteomics analysis. Using label-free approach, a total of 22 differentially expressed proteins were identified in the serum samples of the AAD group, of which 15 were upregulated and 7 were downregulated as compared to the AMI and healthy control groups. The most prominent increased protein was vinculin, which was selected to validate in total samples. The level of vinculin was significantly elevated in AAD patients (15.8 ng/ml, IQR: 9.3-19.9 ng/ml) than that in AMI patients (8.6 ng/ml, IQR:5.3-11.4 ng/ml) and healthy volunteers (5.3 ng/ml, IQR:2.8-7.6 ng/ml), P < 0.0001. Furthermore, the concentration of vinculin both increased in type A and B dissection. At the early stage of AAD, vinculin maintained a high level to 48 hours compared with that of AMI. Our study demonstrated that vinculin may play a role in the early diagnosis of AAD.

Second-Generation Biomarker Testing for Irritable Bowel Syndrome Using Plasma Anti-CdtB and Anti-Vinculin Levels.

BACKGROUND: ELISA testing for anti-CdtB and anti-vinculin can discriminate patients with irritable bowel syndrome with diarrhea (IBS-D) from those with inflammatory bowel disease (IBD). However, recent findings suggest the antigens can suffer from epitope instability. AIM: This study aimed to assess effects of incorporating epitope stabilization on test characteristics for distinguishing IBS-D from IBD subjects. METHODS: Plasma samples from IBS-D subjects from a large-scale clinical trial and subjects with endoscopically active IBD without concurrent immunomodulator therapy were used. After epitope stabilization, CdtB and vinculin were used in ELISA testing. Optical density readings were compared between IBS-D and IBD subjects. RESULTS: Samples from 100 IBS-D and 31 IBD (22 UC and 9 CD) subjects were tested. IBS-D subjects had higher anti-CdtB titers (P = 0.0001) and higher anti-vinculin titers (P = 0.004) than IBD subjects. The specificities of anti-CdtB and anti-vinculin to differentiate IBS-D from IBD were 93.5% and 90.9%, respectively, with sensitivities of 43.0% and 52.2%, respectively. The positive likelihood ratios of identifying IBS-D with anti-CdtB and anti-vinculin were 6.7 and 5.7, respectively. Assuming a pretest probability of 57% for diagnosis of IBS-D in patients with abdominal pain and change in bowel habits, testing positive for both antibodies resulted in a posttest probability of > 98%. CONCLUSIONS: Performing epitope stabilization for CdtB and vinculin enhances the test characteristics of ELISAs for anti-CdtB and anti-vinculin in discriminating IBS-D from IBD. Measurement of anti-CdtB and anti-vinculin with this second-generation methodology may further advance our understanding of the role of immunity in functional bowel diseases.

Identification of vinculin as a potential plasma marker for age-related macular degeneration.

PURPOSE: To identify plasma protein biomarkers for age-related macular degeneration (AMD) using a large-scale quantitative proteomic discovery procedure. METHODS: Plasma proteomes from 20 exudative AMD patients and 20 healthy control patients were comparatively profiled by four-dimensional liquid chromatography-tandem mass spectrometry (LC-MS/MS). Proteins existing at statistically different levels were validated by enzyme-linked immunosorbent assay (ELISA) and Western blotting in 233 case-controlled samples. Newly discovered plasma biomarkers were further confirmed using in vivo and in vitro experiments. RESULTS: Out of 320 proteins identified, vinculin, protein S100A9, triosephosphate isomerase, protein S100A8, protein Z-dependent protease inhibitor, C-X-C motif chemokine 7, and tenascin X showed significantly differential expression in AMD patient plasma compared to control plasma. Among these, the area under the curve (AUC) for vinculin was 0.871 for discriminating between exudative AMD and controls (n = 201) and 0.879 for discriminating between AMD and controls (n = 233). A proteogenomic combination model using vinculin and two known risk genotypes in ARMS2 and CFH genes additionally provided excellent discrimination of AMD from controls (AUC = 0.916). The plasma level of vinculin was not associated with any confounding clinical variables, such as age, smoking, and other comorbidities. Additionally, vinculin was strongly expressed in retinal pigment epithelial cells of human eyes, and its expression was elevated when exposed to oxidative stress in vitro. CONCLUSIONS: Vinculin was identified as a potential plasma biomarker for AMD. The early detection of AMD using novel plasma biomarkers with genetic modeling may enable timely treatment and vision preservation in the elderly.

On the role of the platelet membrane skeleton in mediating signal transduction. Association of GP IIb-IIIa, pp60c-src, pp62c-yes, and the p21ras GTPase-activating protein with the membrane skeleton.

The platelet plasma membrane is lined with a membrane skeleton composed of short actin filaments, actin-binding protein, spectrin, vinculin, and other unidentified proteins. It is connected to the outside of the cell through association with the cytoplasmic domains of transmembrane receptors. In detergent-lysed platelets, cytoplasmic actin filaments are sedimented by centrifugation at 15,600 x g, but the sedimentation of membrane skeleton fragments requires higher g-forces (100,000 x g). In the present study, we show that the major platelet integrin, glycoprotein (GP) IIb-IIIa, sediments from detergent-lysed platelets at 100,000 x g together with fragments of the membrane skeleton that contain the cytoskeletal proteins spectrin, vinculin, and talin. In addition, this cell fraction contained the tyrosine kinases pp60c-src and pp62c-yes and the p21ras GTPase-activating protein (GAP). After thrombin-induced platelet aggregation mediated by fibrinogen binding to GP IIb-IIIa on adjacent platelets, we detected a redistribution of spectrin, talin, vinculin, pp60c-src, and pp62c-yes to the fraction that sediments at 15,600 x g. The redistribution of these proteins from the high-speed detergent-insoluble fraction to the low-speed fraction correlated with the extent of aggregation and was not detected in aggregation-defective thrombasthenic platelets (which lack the GP IIb-IIIa complex). In addition, many of the proteins phosphorylated on tyrosine in activated platelets were present in detergent-insoluble fractions. These results are consistent with the possibilities that 1) GP IIb-IIIa, pp60c-src, pp62c-yes, and GAP associate with a membrane skeleton fraction that contains spectrin, vinculin, and talin, 2) the association of GP IIb-IIIa with adhesive ligand in a platelet aggregate causes components of the membrane skeleton to undergo altered association with cytoplasmic actin filaments, and 3) many of the proteins phosphorylated on tyrosine residues in activated platelets are components of the cytoskeleton. The results imply that the membrane skeleton may play an important role in binding signaling molecules at sites of integrin-cytoskeleton interactions and in mediating signal transduction events in platelets. Further, GP IIb-IIIa-induced redistribution of components of the membrane skeleton and associated signaling molecules may represent an important step in regulating integrin-induced motile events in platelets.