Insulin Resistance in PCOS Patients Enhances Oxidative Stress and Leukocyte Adhesion: Role of Myeloperoxidase

Abstract

Cardiovascular diseases and oxidative stress are related to polycystic ovary syndrome (PCOS) and insulin resistance (IR). We have evaluated the relationship between myeloperoxidase (MPO) and leukocyte activation in PCOS patients according to homeostatic model assessment of IR (HOMA-IR), and have explored a possible correlation between these factors and endocrine and inflammatory parameters. This was a prospective controlled study conducted in an academic medical center. The study population consisted of 101 PCOS subjects and 105 control subjects. We divided PCOS subjects into PCOS non-IR (HOMA-IR<2.5) and PCOS IR (HOMA-IR>2.5). Metabolic and anthropometric parameters, total and mitochondrial reactive oxygen species (ROS) production, MPO levels, interactions between human umbilical vein endothelial cells and leukocytes, adhesion molecules (E-selectin, ICAM-1 and VCAM-1) and proinflammatory cytokines (IL-6 and TNF-α) were evaluated. Oxidative stress was observed in PCOS patients, in whom there was an increase in total and mitochondrial ROS production and MPO levels. Enhanced rolling flux and adhesion, and a decrease in polymorphonuclear cell rolling velocity were also detected in PCOS subjects. Increases in IL-6 and TNF-α and adhesion molecules (E-selectin, ICAM-1 and VCAM-1) were also observed, particularly in the PCOS IR group, providing evidence that inflammation and oxidative stress are related in PCOS patients. HOMA-IR was positively correlated with hsCRP (p<0.001, r = 0.304), ROS production (p<0.01, r = 0.593), leukocyte rolling flux (p<0.05, r = 0.446), E-selectin (p<0.01, r = 0.436) and IL-6 (p<0.001, r = 0.443). The results show an increase in the rate of ROS and MPO levels in PCOS patients in general, and particularly in those with IR. Inflammation in PCOS induces leukocyte-endothelium interactions and a simultaneous increase in IL-6, TNF-α, E-selectin, ICAM-1 and VCAM-1. These conditions are aggravated by the presence of IR.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Fig 1. Effects of PCOS on levels of ROS in PMN.

(A) Levels of DCFH-DA fluorescence measured by fluorimetry in controls and PCOS subjects; (B) Levels of DCFH fluorescence in controls and in PCOS non-IR and PCOS IR subjects; (C) Mean DCFH fluorescence assessed by static cytometry; % vs control. (D) Representative images of DCFH-DA fluorescence in PMNs assessed by fluorescence microscopy; nuclei: Hoechst 33342 signal (blue); ROS: DCFH-DA signal (green). *pcp<0.001 between PCOS non-IR and PCOS IR subjects.

Fig 2. Leukocyte mitochondrial ROS production in PCOS non-IR and IR patients and control women.

A) Mitochondrial ROS (Mitosox Red Fluorescence) in PCOS versus control subjects. B) Mitochondrial ROS (Mitosox Red Fluorescence) in PCOS non IR and IR patients versus control subjects. Values in the bar graphs were obtained by calculating the percentage of fluorescence intensity relative to the control. *p

Fig 3. Plasma MPO concentrations in PCOS and control subjects (A) and in control, PCOS non-IR and PCOS IR subjects (B).

*pap<0.05 between PCOS non-IR and PCOS IR subjects.

Fig 4. Leukocyte/endothelium interactions in PCOS and control subjects.

PMN rolling velocity (μsecond-1) (A), rolling flux (PMN per minute) (C), and PMN adhesion (PMN per square millimetre) (E); and PCOS non-IR and PCOS IR subjects: PMN rolling velocity (μsecond-1) (B), rolling flux (PMN per minute) (D), and PMN adhesion (PMN per square millimetre) (F). *p<0.05 and ***p<0.001 vs. Control. ap<0.05 and bp<0.01 between PCOS non-IR and PCOS IR subjects.

Fig 5. Adhesion molecules in the serum of PCOS and control subjects.

(A) E-selectin, (C) ICAM-1 and (E) VCAM-1; and PCOS non-IR, PCOS IR and control subjects. (B) E-selectin, (D) ICAM-1 and (F) VCAM-1. *pap<0.05 and bp<0.01 between PCOS non-IR and PCOS IR subjects.

Fig 6. Proinflammatory cytokines in the serum of PCOS and control subjects.

(A) IL-6, and (C) TNF-α; and PCOS non-IR, PCOS IR and control subjects. (B) IL-6, and (D) TNF-α. *pap<0.05 between PCOS non-IR and PCOS IR subjects.

Fig 7. Correlation studies of HOMA-IR and inflammation and adhesion parameters in PCOS women.