Van Eyk, Dunn - Proteomic and Genomic Analysis of Cardiovascular Disease - 2003 (522919), страница 103
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Recent studies in our laboratory have also characterized novel changes in protein expression in rat neonatal cardiomyocytes made hypertrophic following endothelin treatment [52]. In rats with coronary ligation-induced heart failure we observed alterations in protein profiles that were significant, time-dependent (i.e., 1 day–16 weeks of ligation) and correlated with cardiacdysfunction as measured by echocardiography [53]. Proteome maps of rat cardiacsarcoplasmic reticulum and sarcolemma were also developed, given the importance that these membranes play in mediating beat-to-beat cardiac functionthrough Ca+2 regulatory proteins like phospholamban [54]. These studies havebeen extended to the assessment of protein expression changes in mouse heartsmade hyperdynamic through transgenic knockout of phospholamban, the key regulator of sarcoplasmic reticulum Ca2+-ATPase activity [55].Hemostasis or thrombotic disorders are of particular clinical interest and a major focus for several of the large pharmaceutical companies.
A primary goal hasbeen to better understand the exact molecular mechanisms in blood platelet activation and aggregation that lead to the formation of thrombi and vascular plaques.While there have been many proteomic studies performed using platelets, it isonly recently that the techniques have been robust and sensitive enough to provide meaningful identification and functionality of proteins [56–58]. Researchersat Bayer established a non-activated human blood platelet protein map, and inparticular identified tyrosine-phosphorylated proteins in the cytosolic fraction, typically a major challenge for protein analysis [56]. A more comprehensive analysisof the platelet proteome was developed by Watson’s group that revealed at least 5new proteins (along with a host of transcription factors and signaling molecules)and has helped build a basis for future identification of new drug targets andtherapeutic approaches [58].
Another study that focused on protein changes fol-21.3 Target Identificationlowing activation of platelets with thrombin was associated with the translocationof novel candidate proteins to the cytoskeletal actin scaffold [57]. Our own proteomic efforts towards characterizing the platelet proteome ± pharmacological treatment (i.e., activators like thrombin, ADP, collagen, and inhibitors such a glycoprotein IIb/IIIa antagonists, the P2Y12 receptor antagonist clopidogrel, aspirin andthromboxane inhibitors) utilizing 2-DE and ICAT have confirmed and extendedthe aforementioned studies (unpublished results).
Thus, the application of proteomics to platelets is increasingly leading to a more useful molecular understandingof the role this blood cell type may play in health and disease.The utility of proteomic analysis in identifying differentially expressed proteinsunder conditions related to hypertension has been demonstrated in only a few instances [59–63]. An early study by Kohane et al. noted an absence of HSP induction using an in vivo deoxycorticosterone-salt-treated rat model of hypertension orfollowing treatment of aortic smooth muscle cells with either norepinephrine orangiotensin II [60].
Abnormal vascular smooth muscle cell (VSMC) growth following treatment with either hypertrophic (angiontensin II) or hyperplastic (plateletderived growth factor) agents was accompanied by differential expression of agroup of proteins involved in protein synthesis and folding [59]. These had notpreviously been recognized as being regulated by growth factors in VSMCs noridentified by RNA expression profiling. The investigators speculated that the transition from hyperplasia to hypertrophy may be a critical event regulated by growthfactor signaling events as identified by proteomic analysis, and differentially-regulated proteins may serve as putative sites for therapeutic intervention.
Pleibner etal. demonstrated that despite significant myocardial hypertrophy, the expressionpattern of the most abundant myocardial proteins was not altered in hypertrophic,non-failing hearts in renovascular hypertension [61]. Berk’s group studied the secretion of stress-induced oxidative factors (SOXF) from VSMC in response toLY83583, a generator of reactive oxygen species, and showed that the secreted proteins stimulated ERK1/2 signaling [62].
The functional role of SOXF candidateproteins remains to be elucidated, but these proteins are speculated to be potential therapeutic targets under pathophysiological conditions of oxidative stresssuch as hypertension, ischemia/reperfusion or atherosclerosis. Lastly, Shusta et al.utilized a novel technique that combines a tissue-specific polyclonal antiserumwith a cDNA library expression cloning system in an attempt to analyze differential protein expression in the brain microvasculature [63].
This subtractive expression cloning strategy revealed changes in a number of tissue-specific membraneproteins that could either be tracked as marker proteins or be useful in identifying molecular transporters for non-invasive drug delivery to the brain. Thus, hypertension as a disease is by and large lacking in a suitable understanding of itsetiology as assessed by protein profiling. By the same token, it is prime area forproteomic study, especially by pharmaceutical researchers, as there is a paucity ofviable and novel molecular targets suitable for drug discovery.
This is particularlyimportant given the many patients who are non-responders to the stable of available anti-hypertensive therapies.37137221 Proteomics in Pharmaceutical R & D21.3.2CancerThe direct comparison of proteomic profiles of normal vs malignant tissues is rapidly becoming a fertile area of drug research and has revealed a number of potential therapeutic targets.
Petricoin’s group has found that the known negative regulator of Ras associated small GTP-binding proteins, RhoGDI, over-expresses in invasive ovarian cancer [64]. This protein has also been associated with chemoresistance in several cancer cell lines [65], suggesting that it is potentially a useful therapeutic target. A separate study revealed decreased expression of the molecularchaperone 14-3-3 sigma in breast cancer cells [66]. The researchers suggested thatrestoring higher levels of the protein in those cells might lead to their decreasedproliferation since this protein directly associates with cyclin-dependent kinases tonegatively regulate cell growth.
Pearl et al. investigated the signaling network of fibroblast growth factor FGF-2, a pleiotropic polypeptide known to be involved inmany forms of cancer cell growth and metastasis [67]. Differential protein expression was noted in MCF-7 cells by studying both the rapid changes in intracellularsignaling and modifications in protein synthesis induced by FGF-2. In addition tothe expected induction of tyrosine phosphorylation, expression of four proteinswas upregulated within the first 12 hours of FGF-2 stimulation and these included proliferating cell nuclear antigen (PCNA), HSP90 and HSP70, and thetranscriptionally-controlled tumor protein.
Geldanamycin, an inhibitor of HSP90activity, completely blocked the FGF-2 induced proliferation and growth of breastcancer cells. The data suggested that expression of this protein is causally linkedto breast epithelial cell tumorigenesis, and thus constitutes a potential drug discovery target.
In the largest published study of its kind, researchers at Large ScaleBiology, in concert with collaborators at the NCI, tested over 60,000<b>Текст обрезан, так как является слишком большим</b>.
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