Extracorporeal Stimulation of Sacral Nerve Roots for Observation of Pelvic Autonomic Nerve Integrity: Description of a Novel Methodological Setup.

INTRODUCTION: Neurophysiologic monitoring can improve autonomic nerve sparing during critical phases of rectal cancer surgery. OBJECTIVES: To develop a system for extracorporeal stimulation of sacral nerve roots. METHODS: Dedicated software controlled a ten-electrode stimulation array by switching between different electrode configurations and current levels. A built-in impedance and current level measurement assessed the effectiveness of current injection. Intra-anal surface electromyography (sEMG) informed on targeting the sacral nerve roots. All tests were performed on five pig specimens. RESULTS: During switching between electrode configurations, the system delivered 100% of the set current (25 mA, 30 Hz, 200 µs cathodic pulses) in 93% of 250 stimulation trains across all specimens. The impedance measured between single stimulation array contacts and corresponding anodes across all electrode configurations and specimens equaled 3.7 ± 2.5 kΩ. The intra-anal sEMG recorded a signal amplitude increase as previously observed in the literature. When the stimulation amplitude was tested in the range from 1 to 21 mA using the interconnected contacts of the stimulation array and the intra-anal anode, the impedance remained below 250 Ω and the system delivered 100% of the set current in all cases. Intra-anal sEMG showed an amplitude increase for current levels exceeding 6 mA. CONCLUSION: The system delivered stable electric current, which was proved by built-in impedance and current level measurements. Intra-anal sEMG confirmed the ability to target the branches of the autonomous nervous system originating from the sacral nerve roots. SIGNIFICANCE: Stimulation outside of the operative field during rectal cancer surgery is feasible and may improve the practicality of pelvic intraoperative neuromonitoring.

FGF23 expression in rodents is directly induced via erythropoietin after inhibition of hypoxia inducible factor proline hydroxylase.

Plasma levels of FGF23 are increased in patients with chronic kidney disease. Beside its role in phosphate homeostasis, iron deficiency and anemia are associated with increased FGF23 plasma levels. Recently, FGF23 plasma levels were shown to be increased in mice after treatment with hypoxia inducible factor-proline hydroxylase (HIF-PH) inhibitors which are strong inducers of erythropoietin and erythropoiesis and are known to modulate iron uptake and availability. Therefore we investigated a potential context between expression of FGF23 and stimulation of erythropoiesis using a HIF-PH inhibitor and erythropoietin in rats. FGF23 plasma levels are induced at peak levels 2 h after intravenous injection of recombinant human Erythropoietin (rhEPO). Likewise induction of endogenous EPO using a HIF-PH inhibitor (BAY 85-3934) is followed by an increase of FGF23 plasma levels. In contrast to rhEPO the HIF-PH inhibitor induces lower peak levels of FGF23 applying equivalent hematopoietic doses. Bone and bone marrow were identified as sources of EPO-induced FGF23. Immediate induction of FGF23 mRNA was also detected in EPO receptor positive murine hematopoietic BAF3 cells after treatment with rhEPO but not after treatment with the HIF-PH inhibitor. Pretreatment of mice with a neutralizing anti-EPO antibody abrogated FGF23 induction by the HIF-PH inhibitor. Thus, direct impact on FGF23 expression by HIF-PH inhibition in vivo via hypoxia mimicking and modulation of iron metabolism appears unlikely. Collectively, the findings point to an EPO dependent regulation pathway of FGF23 gene expression which might be important in the context of erythropoiesis stimulating therapies in patients with renal anemia.

Gas6 protein: its role in cardiovascular calcification.

BACKGROUND: Cardiovascular calcifications can be prevented by vitamin K and are accelerated by vitamin K antagonists. These effects are believed to be mainly mediated by the vitamin K-dependent matrix Gla protein. Another vitamin K-dependent protein, Gas6, is also expressed in vascular smooth muscle cells (VSMC). In vitro Gas6 expression was shown to be regulated in VSMC calcification and apoptotic processes. METHODS: We investigated the role of Gas6 in vitro using VSMC cultures and in vivo in young and old Gas6-deficient (Gas6(-/-)) and wildtype (WT) mice. In addition, Gas6(-/-) and WT mice were challenged by (a) warfarin administration, (b) uninephrectomy (UniNX) plus high phosphate diet, or (c) UniNX plus high phosphate plus electrocautery of the residual kidney. RESULTS: In vitro VSMC from WT and Gas6(-/-) mice exposed to warfarin showed increased apoptosis and calcified similarly. In vivo, aortic, cardiac and renal calcium content in all groups was similar, except for a lower cardiac calcium content in Gas6(-/-) mice (group a). Von Kossa staining revealed small vascular calcifications in both WT and Gas6(-/-) mice (groups a-c). In aging, non-manipulated mice, no significant differences in vascular calcification were identified between Gas6(-/-) and WT mice. Gas6(-/-) mice exhibited no upregulation of matrix Gla protein in any group. Cardiac output was similar in all treatment groups. CONCLUSIONS: Taken together, in our study Gas6 fails to aggravate calcification against the previous assumption.

Surface Electromyography Reliably Records Electrophysiologically Evoked Internal Anal Sphincter Activity: A More Minimally Invasive Approach for Monitoring Extrinsic Innervation.

BACKGROUND: Even in the case of minimally invasive pelvic surgery, sparing of the autonomic nerve supply is a prerequisite for maintaining anal sphincter function. Internal anal sphincter (IAS) innervation could be electrophysiologically identified based on processed electromyographic (EMG) recordings with conventional bipolar needle electrodes (NE). This experimental study aimed for the development of a minimally invasive approach via intra-anal surface EMG for recordings of evoked IAS activity. METHODS: Six male pigs underwent nerve-sparing low anterior rectal resection. Electric autonomic nerve stimulations were performed under online-processed EMG of the IAS. EMG recordings were simultaneously carried out with conventional bipolar NE as the reference method and newly developed intra-anal surface electrodes (SE) in different designs. RESULTS: In all experiments, the IAS activity could be continuously visualized via EMG recordings based on NE and SE. The median number of bipolar electric stimulations per animal was 27 (range 5-52). The neurostimulations resulted in significant EMG amplitude increases for both recording types [NE: median 3.0 µV (interquartile range, IQR 2.8-3.5) before stimulation vs. 7.1 µV (IQR 3.9-13.8) during stimulation, p < 0.001; SE: median 3.6 µV (IQR 3.1-4.3) before stimulation vs. 6.8 µV (IQR 4.8-10.3) during stimulation, p < 0.001]. CONCLUSIONS: Intra-anal SE enabled reliable EMG of electrophysiologically evoked IAS activity similar to the conventional recording via NE. The transfer of the method to access platforms for transanal total mesorectal excision or robotics may offer a practical more minimally invasive approach for monitoring extrinsic innervation.

Serum levels of C-peptide are associated with coronary artery calcification in patients with rheumatoid arthritis.

C-peptide has pro-atherogenic effects in animal models, and elevated C-peptide levels are associated with cardiovascular and all-cause mortality in patients undergoing coronary angiography. This cross-sectional study investigated the association between C-peptide serum levels and coronary artery calcification (CAC) in patients with rheumatoid arthritis (RA), a high-risk group for cardiovascular events. Fifty-four patients with RA were recruited from an arthritis outpatient department at the University Hospital in Aachen, Germany. CAC was measured by multi-slice CT scan, and blood samples were drawn from all patients for the analysis of C-peptide and other cardiovascular biomarkers. Mean serum levels of C-peptide (1.187 ± 0.771 vs 0.745 ± 0.481 nmol/L, p = 0.02), YKL-40, LDL cholesterol, and triglycerides were significantly higher in patients with CAC (n = 32, 59 %) compared to those without CAC (n = 22, 41 %). Univariate analysis revealed a significant association of C-peptide [OR 4.7, 95 % CI (1.1, 20.2)], YKL-40, triglycerides, hypertension, smoking, age, and male sex with the presence of CAC. After adjustment for body mass index, cholesterol, diabetes, adiponectin, calcium, and phosphate, C-peptide was still significantly associated with CAC in a multivariate logistic regression model. In conclusion, C-peptide serum levels are independently associated with the presence of CAC in patients with RA. These data suggest a potential role of C-peptide in cardiovascular disease in patients with RA.

Impaired vitamin K recycling in uremia is rescued by vitamin K supplementation.

In chronic kidney disease, vitamin K-dependent proteins, including the calcification inhibitor matrix Gla protein, are largely uncarboxylated indicating that functional vitamin K deficiency may contribute to uremic vascular calcification. Since the effects of uremia on the vitamin K cycle are unknown, we investigated the influence of uremia and vitamin K supplementation on the activity of the vitamin K cycle and extraosseous calcification. Uremia was induced in rats by an adenine-supplemented diet and vitamin K1 or K2 was administered over 4 and 7 weeks. After 4 weeks of adenine diet, the activity of the vitamin K cycle enzyme γ-carboxylase but not the activities of DT-diaphorase or vitamin K epoxide reductase were reduced. Serum levels of undercarboxylated matrix Gla protein increased, indicating functional vitamin K deficiency. There was no light microscopy-detectable calcification at this stage but chemically determined aortic and renal calcium content was increased. Vitamin K treatment reduced aortic and renal calcium content after 4 weeks. Seven weeks of uremia induced overt calcification in the aorta, heart, and kidneys; however, addition of vitamin K restored intrarenal γ-carboxylase activity and overstimulated it in the liver along with reducing heart and kidney calcification. Thus, uremic vitamin K deficiency may partially result from a reduction of the γ-carboxylase activity which possibly contributes to calcification. Pharmacological vitamin K supplementation restored the vitamin K cycle and slowed development of soft tissue calcification in experimental uremia.

Relationship between sclerostin and cardiovascular calcification in hemodialysis patients: a cross-sectional study.

BACKGROUND: Sclerostin is a Wnt pathway antagonist regulating osteoblast activity and bone turnover. Here, we assessed the potential association of sclerostin with the development of coronary artery (CAC) and aortic valve calcifications (AVC) in haemodialysis (HD) patients. METHODS: We conducted a cross-sectional multi-slice computed tomography (MS-CT) scanning study in 67 chronic HD patients (59.4 ± 14.8 yrs) for measurement of CAC and AVC. We tested established biomarkers as well as serum sclerostin (ELISA) regarding their association to the presence of calcification. Fifty-four adults without relevant renal disease served as controls for serum sclerostin levels. Additionally, sclerostin expression in explanted aortic valves from 15 dialysis patients was analysed ex vivo by immunohistochemistry and mRNA quantification (Qt-RT-PCR). RESULTS: CAC (Agatston score > 100) and any AVC were present in 65% and in 40% of the MS-CT patient group, respectively. Serum sclerostin levels (1.53 ± 0.81 vs 0.76 ± 0.31 ng/mL, p < 0.001) were significantly elevated in HD compared to controls and more so in HD patients with AVC versus those without AVC (1.78 ± 0.84 vs 1.35 ± 0.73 ng/mL, p = 0.02). Multivariable regression analysis for AVC revealed significant associations with higher serum sclerostin. Ex vivo analysis of uraemic calcified aortic valves (n = 10) revealed a strong sclerostin expression very close to calcified regions (no sclerostin staining in non-calcified valves). Correspondingly, we observed a highly significant upregulation of sclerostin mRNA in calcified valves compared to non-calcified control valves. CONCLUSION: We found a strong association of sclerostin with calcifying aortic heart valve disease in haemodialysis patients. Sclerostin is locally produced in aortic valve tissue adjacent to areas of calcification.

Mechanisms and treatment of extraosseous calcification in chronic kidney disease.

Strong and unidirectional associations exist between the severity of cardiovascular calcifications and mortality in patients with advanced chronic kidney disease. In the past 10 years, a wealth of experimental and clinical information has been published on the key pathophysiological events that contribute to the development and progression of vascular and soft-tissue calcifications. These processes involve a sensitive balance of calcification inhibition, induction and removal. The traditional view of regarding secondary hyperparathyroidism and elevated calcium × phosphate product as the pivotal risk factors for calcification has been challenged by data demonstrating a role for other, more subtle and complex pathomechanisms. These mechanisms include the loss of endogenous calcification inhibitors, deficient clearance of calcified debris, effects of vitamin K and vitamin D, and the action of calcification inducers as in osteogenic transdifferentiation. In this Review, we describe our current knowledge of the factors involved in the passive and active regulation of extraosseous calcification processes, with an assessment of their importance as targets for future diagnostic and therapeutic interventions.

Serum fetuin-A levels are associated with vascular calcifications and predict cardiovascular events in renal transplant recipients.

BACKGROUND AND OBJECTIVES: Vascular calcifications predict cardiovascular disease, the major cause of death in renal transplant recipients (RTRs). We studied the determinants of fetuin-A, a potent circulating calcification inhibitor encoded by the AHSG gene, and tested its association with vascular calcifications and long-term survival and cardiovascular events (CVEs) in RTRs. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: Two hundred seventy-seven prevalent RTRs from a single center were included. CVEs and deaths were prospectively recorded during a 5-year follow-up. RESULTS: Independent determinants of lower serum fetuin-A levels were lower plasma cholesterol, the AHSG rs4918 G allele, and history of smoking. Low serum fetuin-A level was a determinant of aortic calcifications (assessed using spiral CT). Low fetuin-A levels (≤0.47 g/L, first quintile) were independently associated with CVEs and deaths (hazard ratio=1.83; 95% confidence interval, 1.07 to 3.04). The association was confirmed for all-cause mortality, and the major adverse cardiovascular endpoints were analyzed separately. Patients with low fetuin-A and high high-sensitivity C-reactive protein (>4.36 mg/L, fourth quintile) levels had a 3.5-fold increased risk of all-cause mortality and CVEs. In the presence of inflammation, CVE-free survival was influenced by common variants in the AHSG gene. CONCLUSIONS: These data show that low fetuin-A levels are independently associated with aortic calcifications and a higher risk of CVEs and mortality. They support fetuin-A as a circulating biomarker able to identify RTRs at risk for vascular calcifications and CVEs.

Circulating nonphosphorylated carboxylated matrix gla protein predicts survival in ESRD.

The mechanisms for vascular calcification and its associated cardiovascular mortality in patients with ESRD are not completely understood. Dialysis patients exhibit profound vitamin K deficiency, which may impair carboxylation of the calcification inhibitor matrix gla protein (MGP). Here, we tested whether distinct circulating inactive vitamin K-dependent proteins associate with all-cause or cardiovascular mortality. We observed higher levels of both desphospho-uncarboxylated MGP (dp-ucMGP) and desphospho-carboxylated MGP (dp-cMGP) among 188 hemodialysis patients compared with 98 age-matched subjects with normal renal function. Levels of dp-ucMGP correlated with those of protein induced by vitamin K absence II (PIVKA-II; r = 0.62, P < 0.0001). We found increased PIVKA-II levels in 121 (64%) dialysis patients, indicating pronounced vitamin K deficiency. Kaplan-Meier analysis showed that patients with low levels of dp-cMGP had an increased risk for all-cause and cardiovascular mortality. Multivariable Cox regression confirmed that low levels of dp-cMGP increase mortality risk (all-cause: HR, 2.2; 95% CI, 1.1 to 4.3; cardiovascular: HR, 2.7; 95% CI, 1.2 to 6.2). Furthermore, patients with higher vascular calcification scores showed lower levels of dp-cMGP. In 17 hemodialysis patients, daily supplementation with vitamin K2 for 6 weeks reduced dp-ucMGP levels by 27% (P = 0.003) but did not affect dp-cMGP levels. In conclusion, the majority of dialysis patients exhibit pronounced vitamin K deficiency. Lower levels of circulating dp-cMGP may serve as a predictor of mortality in dialysis patients. Whether vitamin K supplementation improves outcomes requires further study.

Chronic kidney disease aggravates arteriovenous fistula damage in rats.

Neointimal hyperplasia (NIH) and impaired dilatation are important contributors to arteriovenous fistula (AVF) failure. It is unclear whether chronic kidney disease (CKD) itself causes adverse remodeling in arterialized veins. Here we determined if CKD specifically triggers adverse effects on vascular remodeling and assessed whether these changes affect the function of AVFs. For this purpose, we used rats on a normal diet or on an adenine-rich diet to induce CKD and created a fistula between the right femoral artery and vein. Fistula maturation was followed noninvasively by high-resolution ultrasound (US), and groups of rats were killed on 42 and 84 days after surgery for histological and immunohistochemical analyses of the AVFs and contralateral femoral vessels. In vivo US and ex vivo morphometric analyses confirmed a significant increase in NIH in the AVFs of both groups with CKD compared to those receiving a normal diet. Furthermore, we found using histological evaluation of the fistula veins in the rats with CKD that the media shrank and their calcification increased significantly. Afferent artery dilatation was significantly impaired in CKD and the downstream fistula vein had delayed dilation after surgery. These changes were accompanied by significantly increased peak systolic velocity at the site of the anastomosis, implying stenosis. Thus, CKD triggers adverse effects on vascular remodeling in AVFs, all of which contribute to anatomical and/or functional stenosis.

Ultrastructural analysis of vascular calcifications in uremia.

Accelerated intimal and medial calcification and sclerosis accompany the increased cardiovascular mortality of dialysis patients, but the pathomechanisms initiating microcalcifications of the media are largely unknown. In this study, we systematically investigated the ultrastructural properties of medial calcifications from patients with uremia. We collected iliac artery segments from 30 dialysis patients before kidney transplantation and studied them by radiography, microcomputed tomography, light microscopy, and transmission electron microscopy including electron energy loss spectrometry, energy dispersive spectroscopy, and electron diffraction. In addition, we performed synchrotron x-ray analyses and immunogold labeling to detect inhibitors of calcification. Von Kossa staining revealed calcification of 53% of the arteries. The diameter of these microcalcifications ranged from 20 to 500 nm, with a core-shell structure consisting of up to three layers (subshells). Many of the calcifications consisted of 2- to 10-nm nanocrystals and showed a hydroxyapatite and whitlockite crystalline structure and mineral phase. Immunogold labeling of calcification foci revealed the calcification inhibitors fetuin-A, osteopontin, and matrix gla protein. These observations suggest that uremic microcalcifications originate from nanocrystals, are chemically diverse, and intimately associate with proteinaceous inhibitors of calcification. Furthermore, considering the core-shell structure of the calcifications, apoptotic bodies or matrix vesicles may serve as a calcification nidus.

Analysis of calcifications in patients with coral reef aorta.

BACKGROUND: Coral reef aorta is a rare vascular disease with intraluminal calcifications of the dorsal part of the visceral aorta. The pathogenesis of this disease with its topographic and morphologic characteristics is unknown. The aim of our study was to investigate calcification inhibitors and the ultrastructure of calcifications in patients with coral reef aorta. METHODS: Ten patients with coral reef aorta were examined. Calcified specimens were investigated by immunohistochemical techniques for the expression of the calcification inhibitors matrix gla protein (MGP) and fetuin-A. Vessel walls were also assessed by electron microscopic techniques including electron energy-lost spectroscopy, electron dispersive spectroscopy, and electron diffraction. Sera of patients were analyzed for fetuin-A, uncarboxylated MGP (ucMGP), and osteoprotegerin. RESULTS: As assessed by immunohistochemistry, most MGP was detected in the vicinity of calcified regions. Serum levels of the calcification inhibitors ucMGP, fetuin-A, and osteoprotegerin were 370+/-107 nmol/L, 0.57+/-0.03 g/L, and 5.64+/-0.79 pmol/L, respectively. Ultrastructural analysis of calcified specimens showed a core-shell structure with multiple calcification nuclei. Calcifications displayed a fine-crystalline character, and elemental analysis revealed hydroxyl apatite as the chemical compound. CONCLUSION: The coral reef aorta represents an extreme exophytic growth of vascular calcification with multiple nuclei which resemble typical media calcification. Positive vascular immunostaining and low serum levels of both fetuin-A and ucMGP suggest a pathophysiologic role of these calcification inhibitors in the development of coral reef aorta.

Clinical features of hemodialysis patients with intimal versus medial vascular calcifications.

BACKGROUND: Vascular calcifications (VCs) contribute to the massive mortality in hemodialysis (HD) patients. We aimed to identify prevalence and risk factors for arterial medial calcifications (AMCs) versus intimal calcifications (AICs) in a single-center HD population. METHODS: This cross-sectional study included 134 patients, mean age 56.9 +/- 9.7 years, on HD for 8.2 +/- 5.0 years. VCs were scored based on plain radiographs and ultrasonography of the common carotid arteries. RESULTS: Patients were categorized into groups I (13% without VC), II (10% with an AMC pattern), III (24% with an AIC pattern) and IV (53% with a mixed pattern). AIC and mixed patterns were associated with older age (p=0.006 and p=0.004, respectively), and mixed pattern with longer dialysis vintage (p=0.001). Pulse pressure was significantly higher in patients from group III than group IV, and intima-media thickness (IMT) was higher in both groups with AIC. By multivariate analysis, risk factors for any VC were high serum Ca, phosphate, CaxP product, low total protein, high body mass index (BMI), systolic and diastolic blood pressure, IMT and history of smoking. Elevated calcium and/or phosphate predicted an AMC pattern, and high calcium, BMI and IMT an AIC pattern. Finally, high IMT, systolic blood pressure, BMI and older age were predictors of a mixed pattern. CONCLUSION: We observed a very high prevalence of VC, mostly with a mixed AIC+AMC pattern. Apart from well-known risk factors, the data stress the importance of smoking, an under-recognized cause of AMC, and systolic blood pressure for AIC+AMC.

Risk factors for cardiovascular calcifications in non-diabetic Caucasian haemodialysis patients.

BACKGROUND/AIMS: Dialysis patients display an increased mortality which is associated with cardiovascular calcifications. Diabetes mellitus and ethnicity are known factors that affect the extent of cardiovascular calcifications. However, most studies have investigated mixed cohorts with diabetics and/or mixed ethnicity. METHODS: Cardiovascular calcifications were assessed in non-diabetic Caucasian haemodialysis patients by the semiquantitative Adragao calcification score (X-ray pelvis and hands) and a novel composite calcification score encompassing the Adragao score as well as calcifications detected by X-ray of the fistula arm, echocardiography of heart valves and carotid ultrasound. RESULTS: Using multivariate analysis, age, male gender, dialysis vintage, lower Kt/V, calcium-phosphate product, smoking and high-sensitivity CRP were independent risk factors for cardiovascular calcifications as assessed by the Adragao or the composite score. Pulse wave velocity was independently related to both calcification scores. Body mass index, cholesterol, triglycerides, iPTH and serum levels of fetuin-A and uncarboxylated matrix Gla protein were not associated with cardiovascular calcifications. CONCLUSIONS: In our cohort of non-diabetic Caucasian haemodialysis patients, age, male gender, dialysis vintage, smoking, calcium-phosphate product, high-sensitivity CRP and lower Kt/V were independent risk factors for cardiovascular calcifications. Whether lowering the calcium-phosphate product and increasing dialysis efficiency can reduce cardiovascular calcifications in dialysis patients remains to be determined.

Fetuin-A protects against atherosclerotic calcification in CKD.

Reduced serum levels of the calcification inhibitor fetuin-A associate with increased cardiovascular mortality in dialysis patients. Fetuin-A-deficient mice display calcification of various tissues but notably not of the vasculature. This absence of vascular calcification may result from the protection of an intact endothelium, which becomes severely compromised in the setting of atherosclerosis. To test this hypothesis, we generated fetuin-A/apolipoprotein E (ApoE)-deficient mice and compared them with ApoE-deficient and wild-type mice with regard to atheroma formation and extraosseous calcification. We assigned mice to three treatment groups for 9 wk: (1) Standard diet, (2) high-phosphate diet, or (3) unilateral nephrectomy (causing chronic kidney disease [CKD]) plus high-phosphate diet. Serum urea, phosphate, and parathyroid hormone levels were similar in all genotypes after the interventions. Fetuin-A deficiency did not affect the extent of aortic lipid deposition, neointima formation, and coronary sclerosis observed with ApoE deficiency, but the combination of fetuin-A deficiency, hyperphosphatemia, and CKD led to a 15-fold increase in vascular calcification in this model of atherosclerosis. Fetuin-A deficiency almost exclusively promoted intimal rather than medial calcification of atheromatous lesions. High-phosphate diet and CKD also led to an increase in valvular calcification and aorta-associated apoptosis, with wild-type mice having the least, ApoE-deficient mice intermediate, and fetuin-A/ApoE-deficient mice the most. In addition, the combination of fetuin-A deficiency, high-phosphate diet, and CKD in ApoE-deficient mice greatly enhanced myocardial calcification, whereas the absence of fetuin-A did not affect the incidence of renal calcification. In conclusion, fetuin-A inhibits pathologic calcification in both the soft tissue and vasculature, even in the setting of atherosclerosis.

Vascular access calcification predicts mortality in hemodialysis patients.

Vascular calcification is a recognized risk factor for cardiovascular mortality in patients with end-stage renal disease. The aim of this study was to identify risk factors for vascular access calcification and to determine if patients with this disorder are at increased risk of death. Vascular access calcification was found in 49 of 212 hemodialysis patients as measured by plain X-ray (arteriovenous fistula or synthetic graft) in two dimensions. Male gender, diabetes mellitus, and length of time on dialysis were independent predictors for access calcification determined by logistic regression multivariate analysis. Serum parameters were not independently related to access calcification. Kaplan-Meier analysis showed an increased mortality risk, and Cox regression analysis confirmed that vascular access calcification was an independent mortality predictor. Our study suggests that detection of vascular access calcification is a cost-effective method to identify patients at increased mortality risk.

Predictors of low circulating endothelial progenitor cell numbers in haemodialysis patients.

BACKGROUND: End-stage renal disease (ESRD) patients exhibit increased cardiovascular mortality associated with cardiovascular calcifications and endothelial dysfunction. As circulating endothelial progenitor cells (EPCs) harbour vascular regenerative potential and are altered in uraemia, we examined clinical and biochemical factors influencing EPC levels as well as the relation between EPC numbers and function and uraemic cardiovascular calcifications. METHODS: Sixty-five haemodialysis patients were investigated. Cardiovascular calcifications were assessed by multi-slice spiral CT (MSCT, n = 44) with the calculation of coronary Agatston scores and indirectly by carotid-femoral pulse wave velocity (PWV, n = 61). EPCs were quantified in peripheral blood (CD34(+)/KDR(+)) and at day 7 after ex vivo cultivation (ac-LDL(+)/lectin(+)) by flow cytometry. In addition, colony-forming units (CFUs), migratory activity, adhesion and viability of isolated EPCs were analysed. RESULTS: EPC numbers were reduced (P < 0.001) compared to 27 healthy controls (-64%) or 81 patients with documented coronary artery disease and normal renal function (-58%). Coronary calcifications did not exhibit a significant association with the numbers of circulating CD34(+)/KDR(+) or isolated ac-LDL(+)/lectin(+) EPCs. No difference in EPC functions was observed between the 10 patients with the lowest Agatston scores (range 0-41) versus those with the highest scores (range 1181-3736). Multivariate analysis revealed low fetuin-A serum levels to be a positive predictor, while haematocrit and reticulocytes were negative predictors of reduced ac-LDL(+)/lectin(+) EPC numbers. CONCLUSIONS: EPC numbers and function did not correlate with the degree of coronary calcifications in haemodialysis patients. Rather they appear to be related to serum fetuin-A levels, haematocrit and reticulocytes.