Unma, Japan) [27]. The serum levels of intact FGF23 were determined using a commercial sandwich ELISA kit (Kainos Laboratories, Inc., Tokyo, Japan). The serum levels of total protein, albumin, creatinine, calcium, inorganic phosphate and glucose, as well as the urinary levels of albumin, creatinine, calcium and inorganic phosphate, were measured in all patients.using a high resolution real-time scanner with a 7.5 MHz transducer, as previously Title Loaded From File described [40]. The examination was performed with the subject in the supine position, and the carotid bifurcation, as well as the common carotid artery, were scanned on both sides. The maximum IMT value was measured as follows. The carotid artery was scanned in the longitudinal and transverse directions. The site of the most advanced atherosclerotic lesion that showed the greatest distance between the lumen-intima interface and the media-adventitia interface was located in both the right and left carotid arteries. When plaque was detected on ultrasonography, it was observed as localized thickening rather than a circumferential change in the vessel wall. The greatest thickness of the intima-media complex (including plaque) was used for the maximum IMT value. We identified patients having atherosclerosis based on atheromatous plaques of focal increases in IMT 1.1 mm in accordance with a prior study that showed the normal limit of IMT to be #1.0 mm [69].Measurement of ankle-brachial pulse wave velocity (baPWV). Pulse wave velocity (PWV) measurements wereobtained at the bedside of each subject using a volume plethysmographic apparatus (FORM/ABI; Colin, Komaki, Japan) after the subject had rested in the supine position for at least five minutes, as previously described [40]. This instrument allows simultaneous recording of the baPWV and the brachial and ankle BPs on both sides, in addition to recording an electrocardiogram and heart sounds. We defined patients having arterial stiffness as those with baPWV 1400 since a baPWV 1400 cm/sec is an independent variable of the risk stratification according to theSoluble Klotho and Arterial Stiffness in CKDFramingham score and for the discrimination of patients with atherosclerotic cardiovascular disease [70].Measurement and calculation of the aortic calcification index (ACI). The ACI was determined as previously described[42,43]. A non-contrast CT scan of the abdominal aorta was performed. Calcification of the abdominal aorta above the bifurcation of the common iliac arteries was evaluated semiquantitatively in 10 CT slices at 1 cm intervals. Calcification was considered to be present if an area 1 mm2 displayed a density 130 Hounsfield units. The 1317923 cross-section of the abdominal aorta on each slice was divided into 12 Title Loaded From File segments radially. A segment containing an aortic wall with calcification in any section was defined as having aortic calcification. The number of calcified segments was counted in each slice and divided by 12. The values thus obtained for the 10 slices were added together, divided by 10 (the number of slices inspected) and then multiplied by 100 to express the result as a percentage: ACI ( ) = (total score for calcification in all slices)/(12 [number of segments in each slice]610 [number of slices])6100. The ACI was used as a marker for the extent of aortic calcification. We defined CKD patients having abdominal calcification as those with ACI.0 , as described previously [42,43].Correlation between the serum Klotho levels (pg/mL) and the other m.Unma, Japan) [27]. The serum levels of intact FGF23 were determined using a commercial sandwich ELISA kit (Kainos Laboratories, Inc., Tokyo, Japan). The serum levels of total protein, albumin, creatinine, calcium, inorganic phosphate and glucose, as well as the urinary levels of albumin, creatinine, calcium and inorganic phosphate, were measured in all patients.using a high resolution real-time scanner with a 7.5 MHz transducer, as previously described [40]. The examination was performed with the subject in the supine position, and the carotid bifurcation, as well as the common carotid artery, were scanned on both sides. The maximum IMT value was measured as follows. The carotid artery was scanned in the longitudinal and transverse directions. The site of the most advanced atherosclerotic lesion that showed the greatest distance between the lumen-intima interface and the media-adventitia interface was located in both the right and left carotid arteries. When plaque was detected on ultrasonography, it was observed as localized thickening rather than a circumferential change in the vessel wall. The greatest thickness of the intima-media complex (including plaque) was used for the maximum IMT value. We identified patients having atherosclerosis based on atheromatous plaques of focal increases in IMT 1.1 mm in accordance with a prior study that showed the normal limit of IMT to be #1.0 mm [69].Measurement of ankle-brachial pulse wave velocity (baPWV). Pulse wave velocity (PWV) measurements wereobtained at the bedside of each subject using a volume plethysmographic apparatus (FORM/ABI; Colin, Komaki, Japan) after the subject had rested in the supine position for at least five minutes, as previously described [40]. This instrument allows simultaneous recording of the baPWV and the brachial and ankle BPs on both sides, in addition to recording an electrocardiogram and heart sounds. We defined patients having arterial stiffness as those with baPWV 1400 since a baPWV 1400 cm/sec is an independent variable of the risk stratification according to theSoluble Klotho and Arterial Stiffness in CKDFramingham score and for the discrimination of patients with atherosclerotic cardiovascular disease [70].Measurement and calculation of the aortic calcification index (ACI). The ACI was determined as previously described[42,43]. A non-contrast CT scan of the abdominal aorta was performed. Calcification of the abdominal aorta above the bifurcation of the common iliac arteries was evaluated semiquantitatively in 10 CT slices at 1 cm intervals. Calcification was considered to be present if an area 1 mm2 displayed a density 130 Hounsfield units. The 1317923 cross-section of the abdominal aorta on each slice was divided into 12 segments radially. A segment containing an aortic wall with calcification in any section was defined as having aortic calcification. The number of calcified segments was counted in each slice and divided by 12. The values thus obtained for the 10 slices were added together, divided by 10 (the number of slices inspected) and then multiplied by 100 to express the result as a percentage: ACI ( ) = (total score for calcification in all slices)/(12 [number of segments in each slice]610 [number of slices])6100. The ACI was used as a marker for the extent of aortic calcification. We defined CKD patients having abdominal calcification as those with ACI.0 , as described previously [42,43].Correlation between the serum Klotho levels (pg/mL) and the other m.
