Role of Endothelial Progenitor Cells Dysregulation and Inflammation in the Pathophysiology of Cardiovascular Complications of Type 2 Diabetes

Weill Cornell Medical College in Qatar90 enrolled

Overview

This study aims to isolate endothelial progenitor cells (EPCs) from participants with type 2 diabetes (T2D) and cardiovascular complications and to comprehensively characterize EPC dysfunction. Specifically, the study will evaluate maladaptive angiocrine signaling, calcium signaling pathways, and the role of inflammation in EPC function and the progression of atherosclerosis during T2D development. A sub-study will assess EPC functionality by examining endothelial nitric oxide synthase (eNOS) expression and activity, as well as the effectiveness of in vitro eNOS gene enhancement.

Type 2 diabetes (T2D) is associated with damage to blood vessels, which can lead to serious complications such as heart disease, stroke, and other vascular problems. Endothelial progenitor cells (EPCs) help maintain healthy blood vessels by repairing vascular injury. In people with T2D, the number and function of these cells are reduced, which may contribute to poor blood vessel repair and increased cardiovascular risk. The mechanisms responsible for this dysfunction are not fully understood. This study aims to examine how changes in cell signaling and chronic low-grade inflammation in T2D affect EPC function. EPCs will be isolated from patients with T2D, with and without cardiovascular complications, to assess their signaling properties, function, and ability to mature into vascular cells. An in vitro sub-study will evaluate a potential therapeutic strategy to improve EPC function by increasing the activity of endothelial nitric oxide synthase (eNOS), a protein that plays a key role in maintaining healthy blood vessels. Reduced eNOS activity is an important contributor to vascular dysfunction in diabetes. Enhancing eNOS expression and function in EPCs may improve their regenerative capacity and help prevent or treat diabetic vascular complications.

Study Type

OBSERVATIONAL

Enrollment

Conditions

Diabete Type 2 Cardio Vascular Disease

Outcomes

Primary Outcomes

Characterization of EPCs dysregulation in Type 2 diabetes

Functional characterization of endothelial progenitor cells (EPCs) isolated from participants will be performed by assessing cellular signaling and functional pathways. Measurements will be compared among EPCs isolated from participants with type 2 diabetes and cardiovascular diseases, participants with type 2 diabetes without cardiovascular diseases, and healthy volunteers without diabetes or cardiovascular diseases.

Time frame: Following the EPC isolation (15 - 20 days).

Functional Analysis of inflammatory responses in Endothelial Progenitor Cells in type 2 Diabetes with cardiovascular complications

Time frame: Following the EPC isolation (15 - 20 days)

Secondary Outcomes

Expression levels of angiocrine factor genes in EPCs

Expression levels of angiocrine related genes in EPCs measured by RNA sequencing and reported as normalized transcript counts.

Time frame: Following the EPC isolation (15-20 days)

Cytosolic calcium concetration in EPCs

Cytosolic calcium concentration and expression of calcium signaling transcripts and proteins in EPCs assessed using functional calcium assays and molecular analyses.

Time frame: Following the EPC isolation (15-20 days)

Quantification of Mitochondrial reactive oxygen species (ROS) levels in EPCs

Mitochondrial ROS production in EPCs measured using pre-designed fluorescence-based assay kits and reported as relative fluorescence units. Comparisons will be made among the three study groups.

Time frame: Following the EPC isolation (15-20 days).

Protein Expression of Inflammatory Transcription Factors in Endothelial Progenitor Cells

Protein expression levels in lysates are quantified using western blot analysis and normalized to housekeeping proteins.

Time frame: Following the EPC isolation (15 - 20 days).

Angiogenic Transcription Factor Expression in EPCs

Protein expression levels in lysates are quantified using western blot analysis and normalized to housekeeping proteins.

Time frame: Time Frame: Following the EPC isolation (15 - 20 days).

Proportion of EPCs differentiating into endothelial cells in vitro

Differentiation of EPCs into adherent endothelial cells following culture in VEGF-, FGF-2-, IGF-1-, and heparin-supplemented media, assessed by endothelial morphology and marker expression.

Time frame: 14 days after initiation of in vitro differentiation.

Measurement of eNOS(endothelial nitric oxide synthase)-Dependent Nitric Oxide Production in Endothelial Progenitor Cells

eNOS(endothelial nitric oxide synthase) mRNA expression measured by RT-PCR and eNOS protein expression measured by western blotting in EPCs isolated from participants with type 2 diabetes and healthy volunteers.

Time frame: Following EPC isolation (15-20 days) and 72 hours post-gene transfection.

Functional Outcomes of Genetically Enhanced Endothelial Progenitor Cells

eNOS mRNA expression measured by RT-PCR and eNOS protein expression measured by western blotting in EPCs isolated from participants with type 2 diabetes and healthy volunteers.

Time frame: Following EPC isolation (15-20 days) and 72 hours post-gene transfection