Cardiovascular Status of Children 5 Years After Kawasaki Disease

All participants will be examined in Medical University Warsaw Children's Hospital.

Children with history of KD will be recruited from 2 paediatric hospitals in Warsaw and via advertisement by Polish support group for parents of children with KD in social media. Diagnosis of KD will be verified according to current American Heart Association (AHA) guidelines.

All children after KD will be examined 5 years after IVIG treatment exact to 2 months.

CAA presence at the time of KD diagnosis will be determined on the basis of medical records, after specialist consultation, in accordance to AHA definition. Worst-ever echocardiographic picture of coronary arteries will be considered in analysis.

Healthy age- and sex-matched controls (HC) will be recruited from KD patients' siblings.

Informed consent will be obtained from the parents of all patients and all HC.

Assessment of cardiovascular status

All children included in the study will undergo following tests:

1. Laboratory tests

   Blood samples will be drawn after over-night fasting. A) 1.6 ml of blood will be collected in vacutainer tube with ethylenediaminetetraacetic acid (EDTA), B) 4.9 ml of blood will be collected in vacutainer tube with clot-activator, without separation gel (serum tube).

   Routine laboratory techniques will be used to measure lipid profile, glucose and complete blood count. 1 ml of whole blood will be used for circulating endothelial cells (CEC) isolation. CEC will be identified with CD146-immunomagnetic bead extraction based on an international consensus standardised protocol. 3 ml of blood will be centrifuged at room temperature within 2 h of collection and serum will be stored in separate tubes at -70°C until analyzed. Endothelial injury markers: galectin-3 and soluble VCAM levels will be measured using standardized ELISA assays.

2. Echocardiography

   Echocardiography (ECHO) will be performed with Philips Epiq 7 ultrasound equipment with appropriate transducers by a single specialist supervised by an experienced paediatric echocardiographer. All the standard anatomic and physiological imaging will be done. Multiple imaging planes and transducer positions will be used for optimal visualization of the coronary arteries in all major coronary segments - main stem of left coronary artery, anterior interventricular branch, circumflex branch and right coronary artery will be measured according to AHA guidelines - internal vessel diameter will be assessed from inner edge to inner edge of vessel. The number and location of aneurysms and the presence or absence of intraluminal thrombi and stenotic lesions will be evaluated.

   Another evaluation will include assessment of the left ventricular form and function (ejection fraction measured by Teicholz and Simpson's method, end-systolic and end-diastolic volumes, regional wall motion estimated by M-Mode and speckle tracking modes, evaluation of diastolic function in Tissue Doppler Imaging, both systolic and diastolic function measured as myocardial performance index - i.e. Tei index), aortic root imaging (possible dilatation), valvular function (especially mitral and aortic regurgitation assessed in pulsed and color doppler), presence of pericardial effusion. All of the parameters will be calculated as Z-scores assessed by the health professionals Cardio Z mobile application developed by the experienced Paediatric Cardiology Team at Evelina Children's Hospital in London.

3. Carotid intima media thickness (cIMT)

   cIMT will be evaluated in all subjects by a single experienced specialist using 13-megahertz (MHz) linear transducer, Aloka Prosound Alpha 6, Hitachi Aloka Medical, Mitaka, Japan.

   cIMT will be defined as the mean distance from the leading edge of the lumen-intima interface to the leading edge of the media adventitia interface of the far wall, approximately 1 cm proximal to the carotid bulb. Six determinations of cIMT [mm], three on the left and three on the right side, will be obtained and averaged.

4. Pulse Wave Analysis (PWA) and Pulse Wave Velocity (PWV)

Arterial pulse waveform and aortal pulse wave velocity will be evaluated by the same investigator using a Sphygmocor device, AtCor Medical Pty Ltd., Sydney, Australia. All pulse wave and velocity measurements will be performed in the sitting position in a quiet, temperature-controlled room (20 ± 5°C) after a 5 min rest.

Peripheral pressure waveforms will be recorded from the radial artery at the right wrist, using applanation tonometry. After 20 sequential waveforms had been acquired, a validated generalized transfer function will be used to generate the corresponding central aortic pressure waveform.

All examiners will be unaware of patients' clinical details.