Peripheral Blood Lymphocyte Subsets (CD4+, CD8+ T Cells, NK Cells) in Patients with Cardiovascular and Neurological Complications after Carotid Endarterectomy

Katarzyna Kotfis, Jowita Biernawska, Małgorzata Zegan-Barańska, Maciej Żukowski, Katarzyna Kotfis, Jowita Biernawska, Małgorzata Zegan-Barańska, Maciej Żukowski

Abstract

Background: The aim of the study was to evaluate the differences in the circulating immune cells' subgroups after the atherosclerotic plaque removal in patients presenting with postoperative complications as compared to the patients without complications after carotid endarterectomy (CEA).

Methods: Patients with significant carotid atherosclerosis (n=124, age range: 44 to 87 years) who underwent CEA were enrolled in a prospective study. The immunology study using flow cytometry was performed to determine the percentages of peripheral blood T cells (CD4+, CD8+, Treg-CD4+/CD25+) and NK (natural killer) cells before and after the procedure. The data were expressed as the percentage of total lymphocytes±the standard error of mean.

Results: The mean percentage of lymphocytes (61.54%±17.50% vs. 71.82%±9.68%, p=0.030) and CD4 T lymphocytes (T helper, 38.13%±13.78% vs. 48.39%±10.24%, p=0.027) was significantly lower six hours after CEA in patients with postoperative 30-day cardiovascular and neurological complications as compared to the group without complications. On the other hand the mean NK level in the group with complications was significantly higher (21.61%±9.00% vs. 15.80%±9.31%, p=0.048).

Conclusions: The results of this study suggest that after carotid endarterectomy the percentages of circulating immune cells subsets differ in patients with and without postoperative complications.

Figures

Figure 1
Figure 1
Gated lymphocyte sub-population based on SSC (Side-Scattered light) vs. FSC (Forward-Scattered light) measurement.
Figure 2
Figure 2
(a) Tube No. 1; Dot plot of MouseIgG1 fluorescein isothiocyanate (FITC)/MouseIg2a phycoerythrin (PE) with quadrant markers, showing negative control; (b) Tube No. 1; Gating tree showing gating strategy for Fluorescence-activated cell sorting (FACS) analysis as well as parent and total cell percentages within each of the gates for negative control.
Figure 3
Figure 3
(a) Tube No. 2; Dot plot of CD3 FITC/CD19 PE with quadrant markers, differentiating lymphocytes B (CD19+) from lymphocytes T (CD3+); (b) Tube No. 2; Gating tree showing gating strategy for FACS analysis as well as parent and total cell percentages within each of the gates for CD19+ and CD3+.
Figure 4
Figure 4
(a) Tube No. 3; Dot plot of CD3 FITC/CD8 PE with quadrant markers, differentiating lymphocytes T cytotoxic (CD3+/CD8+) from all lymphocytes; (b) Tube No. 3; Gating tree showing gating strategy for FACS analysis as well as parent and total cell percentages within each of the gates for CD8+ and CD3+.
Figure 4
Figure 4
(a) Tube No. 3; Dot plot of CD3 FITC/CD8 PE with quadrant markers, differentiating lymphocytes T cytotoxic (CD3+/CD8+) from all lymphocytes; (b) Tube No. 3; Gating tree showing gating strategy for FACS analysis as well as parent and total cell percentages within each of the gates for CD8+ and CD3+.
Figure 5
Figure 5
(a) Tube No. 4; Dot plot of CD3 FITC/CD4 PE with quadrant markers, differentiating lymphocytes T helper (CD3+/CD4+) from all lymphocytes; (b) Tube No. 4; Gating tree showing gating strategy for FACS analysis as well as parent and total cell percentages within each of the gates for CD4+ and CD3+.
Figure 6
Figure 6
(a) Tube No. 5; Dot plot of CD3 FITC/CD16CD56 PE with quadrant markers, differentiating NK cells (CD3+/CD56CD16+) from all lymphocytes; (b) Tube No. 5; Gating tree showing gating strategy for FACS analysis as well as parent and total cell percentages within each of the gates for CD56CD16+ and CD3+.
Figure 7
Figure 7
(a) Tube No. 7; Dot plot of CD25 FITC/CD4 PE with quadrant markers, showing Treg CD25+/CD4+ lymphocytes; (b) Tube No. 7; Gating tree showing gating strategy for FACS analysis as well as parent and total cell percentages within each of the gates for a representative CD4+CD25+ lymphocytes.
Figure 7
Figure 7
(a) Tube No. 7; Dot plot of CD25 FITC/CD4 PE with quadrant markers, showing Treg CD25+/CD4+ lymphocytes; (b) Tube No. 7; Gating tree showing gating strategy for FACS analysis as well as parent and total cell percentages within each of the gates for a representative CD4+CD25+ lymphocytes.

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