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Optimized Protocol for the Extraction of Proteins from the Human Mitral Valve.

ABSTRACT: Analysis of the cellular proteome can help to elucidate the molecular mechanisms underlying diseases due to the development of technologies that permit the large-scale identification and quantification of the proteins present in complex biological systems.The knowledge gained from a proteomic approach can potentially lead to a better understanding of the pathogenic mechanisms underlying diseases, allowing for the identification of novel diagnostic and prognostic disease markers, and, hopefully, of therapeutic targets. However, the cardiac mitral valve represents a very challenging sample for proteomic analysis because of the low cellularity in proteoglycan and collagen-enriched extracellular matrix. This makes it challenging to extract proteins for a global proteomic analysis. This work describes a protocol that is compatible with subsequent protein analysis, such as quantitative proteomics and immunoblotting. This can allow for the correlation of data concerning protein expression with data on quantitative mRNA expression and non-quantitative immunohistochemical analysis. Indeed, these approaches, when performed together, will lead to a more comprehensive understanding of the molecular mechanisms underlying diseases, from mRNA to post-translational protein modification. Thus, this method can be relevant to researchers interested in the study of cardiac valve physiopathology.

SUBMITTER: Banfi C

PROVIDER: S-EPMC5608423 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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Optimized Protocol for the Extraction of Proteins from the Human Mitral Valve.

Banfi Cristina C Guarino Anna A Brioschi Maura M Brioschi Maura M Ghilardi Stefania S Mastrullo Valeria V Tremoli Elena E Polvani Gianluca G

Journal of visualized experiments : JoVE 20170614 124

Analysis of the cellular proteome can help to elucidate the molecular mechanisms underlying diseases due to the development of technologies that permit the large-scale identification and quantification of the proteins present in complex biological systems.The knowledge gained from a proteomic approach can potentially lead to a better understanding of the pathogenic mechanisms underlying diseases, allowing for the identification of novel diagnostic and prognostic disease markers, and, hopefully, ...[more]

PMID: 28654069

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Project description:ObjectiveIntraoperative predictors of functional mitral valve (MV) stenosis after surgical repair of mitral regurgitation (MR) caused by prolapse remain poorly characterised. This study evaluated the effect of annuloplasty size on postoperative MV haemodynamics during exercise and evaluated predictors of MV hemodynamics.Methods104 patients were randomly assigned to leaflet resection or preservation for surgical repair of MR in the Canadian Mitral Research Alliance CardioLink-2 study. In this post hoc analysis, we compared MV haemodynamics between the two surgical groups and examined the relationship between annuloplasty size and MV haemodynamics 1 year after repair in the combined groups. Echocardiograms were performed at baseline and intraoperatively. Exercise transthoracic echocardiography was performed 1 year postoperatively. Multivariable linear regression analysis was used to identify predictors of exercise MV gradients at follow-up.ResultsMean age of participants was 65±10 years, and 83% were male. Median annuloplasty size was 34 (IQR 32-36). Dividing by the median, 48 (46%) had annuloplasty size of <34 mm and 56 (54%) had ≥34 mm. Mean and peak exercise gradients at 1 year were 11±5 mm Hg and 22±9 mm Hg in <34, and 6±3 mm Hg and 14±5 mm Hg in ≥34 (p<0.001). Rate of residual MR was similar in both groups. In multivariable analyses, annuloplasty size of ≥34 mm was associated with lower mean and peak exercise gradients at 12 months, after adjustment for repair type, age, sex, heart rate and body surface area (β -4.1, 95% CI -6 to -3, p<0.001, and β -7 95% CI -10 to -4, p<0.001, respectively). Intraoperative mean and peak MV gradients by transesophageal echocardiography independently predicted mean and peak resting and exercise gradients at follow-up (p<0.001). Similar results were obtained in both leaflet resection and preservation.ConclusionAnnuloplasty size of ≥34 mm is associated with a 4 and 7 mm Hg reduction in mean and peak exercise MV gradients, respectively, 1 year post MV repair regardless of the repair strategy used. Intraoperative TEE MV gradients predict exercise MV gradients 1 year post repair.Trial registration numberNCT02552771.

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Optimized Protocol for the Extraction of Proteins from the Human Mitral Valve.

Publications

Optimized Protocol for the Extraction of Proteins from the Human Mitral Valve.

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