Jung-Hwa Ryu

Clopidogrel Effectively Suppresses Endothelial Microparticle Generation Induced by Indoxyl Sulfate via Inhibition of the p38 Mitogen-Activated Protein Kinase Pathway

Background/Aims: Endothelial microparticles (EMPs) are closely associated with vascular dysfunction. We investigated the effects of several drugs on EMP generation in human umbilical vein endothelial cells (HUVECs), and the involvement of the mitogen-activated protein kinase (MAPK) in EMP generation. Methods: CD31+CD42-EMP counts were measured by flow cytometry in supernatants of HUVECs incubated with indoxyl sulfate. The EMP responses to losartan, lovastatin, clopidogrel, and mesoglycan were examined. We then measured the effects of MAPK inhibitors on EMPs. Results: (1) Indoxyl sulfate induced EMP release in HUVECs in a dose-dependent fashion; (2) all drugs (10–50 µM) inhibited EMP generation induced by indoxyl sulfate, with clopidogrel being the most effective; (3) the p38 MAPK inhibitor suppressed EMP generation induced by indoxyl sulfate, and (4) clopidogrel significantly suppressed MAPK signaling activated by indoxyl sulfate, with the most potency on p38. Conclusion: The p38 signaling involves EMP generation induced by indoxyl sulfate and is effectively suppressed by clopidogrel.

Non-Dipper Status and Left Ventricular Hypertrophy as Predictors of Incident Chronic Kidney Disease

We have hypothesized that non-dipper status and left ventricular hypertrophy (LVH) are associated with the development of chronic kidney disease (CKD) in non-diabetic hypertensive patients. This study included 102 patients with an estimated glomerular filtration rate (eGFR) ≥ 60 mL/min/1.73 m2. Ambulatory blood pressure monitoring and echocardiography were performed at the beginning of the study, and the serum creatinine levels were followed. During the average follow-up period of 51 months, CKD developed in 11 patients. There was a significant difference in the incidence of CKD between dippers and non-dippers (5.0% vs 19.0%, P < 0.05). Compared to patients without CKD, patients with incident CKD had a higher urine albumin/creatinine ratio (52.3 ± 58.6 mg/g vs 17.8 ± 29.3 mg/g, P < 0.01), non-dipper status (72.7% vs 37.4%, P < 0.05), the presence of LVH (27.3% vs 5.5%, P < 0.05), and a lower serum HDL-cholesterol level (41.7 ± 8.3 mg/dL vs 50.4 ± 12.4 mg/dL, P < 0.05). Based on multivariate Cox regression analysis, non-dipper status and the presence of LVH were independent predictors of incident CKD. These findings suggest that non-dipper status and LVH may be the therapeutic targets for preventing the development of CKD in non-diabetic hypertensive patients.

Interleukin-6 -634 C/G and -174 G/C Polymorphisms in Korean Patients Undergoing Hemodialysis

Background/Aims Chronic inflammatory status is a possible risk factor for vascular access dysfunction in hemodialysis (HD) patients, but susceptibility differences appear among individuals. Interleukin (IL)-6 is a well-known inflammatory cytokine with various polymorphisms. We examined whether IL-6 polymorphisms are associated with vascular access dysfunction in HD patients. Methods A total of 80 HD patients (including 42 diabetic patients) were enrolled. Polymorphisms in the IL-6 gene promoter (-634 C/G and -174 G/C) were studied using restriction length polymorphism polymerase chain reaction analysis. Vascular access patency was compared between the patient groups with respect to IL-6 polymorphisms. An additional 89 healthy individuals were enrolled in the control group. Plasma IL-6 levels were de termined by enzyme-linked immunosorbent assay. Results The GG genotype and G allele at position -634 in the IL-6 promoter were more frequently observed in HD patients than in controls. Furthermore, the distribution of the -634 polymorphism differed according to vascular access patency in non-diabetic HD patients. However, the G allele was not a significant risk factor for early access failure. No significant association appeared between the IL-6 -634 C/G polymorphism and plasma IL-6 levels. The C allele of the IL-6 -174 G/C polymorphism was not detected in our study population. Conclusions The IL-6 -634 G allele appears with greater frequently in patients with end-stage renal disease and may be associated with vascular access dysfunction in non-diabetic HD patients.

Voice Change in End-Stage Renal Disease Patients After Hemodialysis: Correlation of Subjective Hoarseness and Objective Acoustic Parameters

INTRODUCTION Patients with end-stage renal disease (ESRD) who are treated with hemodialysis (HD) frequently complain about hoarseness after completion of each HD session. The HD treatment affects laryngeal volume and muscle function. This study attempted to evaluate the vocal effect of HD by acoustic and aerodynamic analysis and to determine the difference between voice change group (VCG) and nonvoice change group (NVCG). MATERIALS AND METHODS A total of 55 patients (34 females and 21 males) diagnosed with ESRD and undergoing outpatient HD were enrolled. The subjects were divided into the VCG (n=13) and NVCG (n=42) by the change of the Korean Voice Handicap Index score. Patients underwent weighing and acoustic, aerodynamic analysis before and after the HD. Fundamental frequency (F0), jitter, shimmer, noise-to-harmonics ratio (NHR), pitch range, habitual pitch, voice energy, and maximal phonation time (MPT) were obtained. The pre- and post-HD data were compared using paired t test. The results were compared after dividing the total group into the VCG and NVCG categories. Correlation between the change of the weight and change of the voice analysis result was certified by Pearson correlation coefficient. RESULTS The F0 and habitual pitch increased in all subjects. The NHR and MPT parameters significantly decreased (P<0.05). In the NVCG group, all the results were same as the total group. In the VCG group, the NHR result differed from the total group. All acoustic parameters showed no statistically significant differences between the two groups. There was no correlation between the weight change (%) and the change of acoustic parameter results. CONCLUSIONS The NVCG group of patient displayed improvement in NHR, whereas the VCG group showed no change. Weight change did not significantly correlate with the voice analysis results.

A case of membranoproliferative glomerulonephritis associated with metastatic colon cancer

To the Editor, Glomerular diseases frequently develop in patients with malignancy. It has been reported that 11% of adult patients with nephrotic syndrome also have malignant tumors [1]. Previous data show an excess incidence of de novo cancer (5.2%) after diagnosis of renal diseases in patients with biopsy proven glomerulopathy, compared to the general population [2]. Membranous glomerulonephritis (MGN) is the most common malignancy related glomerular disease [3]. Although a few reports have described membranoproliferative glomerulonephritis (MPGN) in association with lung cancer [3], MPGN related to colon cancer has to our knowledge not yet been reported. Here, we present a case of rapidly progressive MPGN associated with metastatic colon cancer. A 58-year-old male patient was admitted to our hospital for his first chemotherapy session to treat metastatic sigmoid colon cancer. The patient had been diagnosed with sigmoid colon cancer with hepatic metastasis (stage IV, T4aN1bM1) 50 days earlier. Other past medical history was unremarkable. Sigmoidectomy with partial hepatectomy had been performed 40 days earlier. The pathological type was characterized as adenocarcinoma. He was referred to the nephrology department due to abruptly increased levels of blood urea nitrogen (BUN; 68 mg/dL) and serum creatinine (sCr; 2.2 mg/dL). His laboratory results were within the normal range 50 days earlier: BUN, 11 mg/dL; sCr, 1.0 mg/dL; and there was no proteinuria or hematuria on urinalysis. On day 1 of admission, he complained of nausea, vomiting, and poor oral intake. Blood pressure was 140/70 mmHg, pulse rate was 70 beats/min, and body temperature was 37.4℃. Bilateral pretibial pitting edema and moderate ascites were noted on physical examination. Laboratory studies revealed white blood cell (WBC) count 9,400/mm3, hemoglobin 10.5 g/dL, hematocrit 31.0%, platelet count 153,000/mm3, potassium 5.2 mEq/L, total protein 5.4 g/dL, albumin 2.5 g/dL, phosphorus 3.9 mg/dL, and total cholesterol 178 mg/dL. Urinalysis with microscopic examination showed urine protein (3+), a red blood cell count of 21 to 30/high power field (HPF) and a WBC count of 6 to 10/HPF. The spot urine protein/creatinine ratio was 19.9 and the fractional excretion of sodium was 0.62%. Twenty-four hour urine collection could not be performed due to a decreased urine output and poor patient cooperation. Urine protein electrophoresis showed nonselective glomerular proteinuria. Immunological studies showed no specific abnormalities. Antineutrophilic cytoplasmic antibody, antinuclear antibody, and antiglomerular basement membrane antibody were negative. Serologic markers of hepatitis B and hepatitis C were negative. The carcinoembryonic antigen level was decreased compared with the value at diagnosis (5.7 ng/mL vs. 163.9 ng/mL). Urine cytology revealed negative results. Abdominal sonography showed normal sized kidneys with appropriate echogenicity. Ascitic fluid analysis revealed transudate with no abnormal cells. On day 4 of admission, a kidney biopsy was performed because his renal function did not improve. Upon light microscopic examination, glomerular enlargement due to diffuse endocapillary proliferation, diffuse thickening of capillary loop with double contour, subendothelial and mesangial fuchsinophilic deposition, and glomerular neutrophil infiltration were found. Cellular crescents were found in four of 24 glomeruli. Mild tubular atrophy, interstitial edema, fibrosis, and mild neutrophilic and mononuclear cellular infiltrations were found. Immunofluorescence microscopy showed immunoglobulin G (IgG), IgA, C1q, dominant C3, fibrinogen, and mild IgM, as well as C4 deposition in both the mesangium and capillary loops. These findings were compatible with type 1 MPGN (Fig. 1). Chemotherapy was delayed due to the poor general condition of the patient. On day 12 of admission, hemodialysis was initiated because BUN and sCr levels were elevated to 57 and 4.2 mg/dL, respectively. His uremic symptoms, including nausea, and vomiting were aggravated, and daily urine output was < 500 mL. On day 20 of admission, the first chemotherapy session was performed with FOLFOX (oxaliplatin, leucovorin, and 5-fluorouracil). After that, four more cycles of FOLFOX chemotherapy were performed at intervals of 2 weeks. Follow-up abdominopelvic computed tomography at 3 months showed no evidence of recurrence of primary cancer. The patient is currently undergoing maintenance hemodialysis three times per week. Figure 1 (A) A glomerulus showing a membranoproliferative pattern of injury. There was hypercellularity due to diffuse mesangial and endocapillary proliferation with neutrophilic infiltration (periodic acid-methenamine silver stain, × 400). Capillar y ... Various glomerular diseases such as MGN and minimal change disease frequently present as a paraneoplastic syndrome in different solid or hematological malignancies. Although there are some case reports of MPGN associated lung cancer or renal cell carcinoma [3], MPGN associated solid organ malignancy has rarely been reported. The mechanisms that underlie how malignancy induces renal diseases remain unresolved. Three hypotheses have been suggested to explain the pathogenesis of tumor associated glomerulonephritis: 1) an undiagnosed malignancy associated with antigen deposition may have caused glomerular disease-like paraneoplastic syndrome; 2) immunosuppressive therapy used in glomerular disease may trigger tumorigenesis; and 3) viral infection may have induced both glomerulopathy and cancer by: intrinsic viral oncogenic activity; disrupted renal clearance of biological mediators associated with oncogenesis; or both [2]. The treatment of MPGN depends in part upon the underlying cause because most cases appear to have an identifiable chronic disease with circulating immune complexes [4]. However, there are limitations in proving the effectiveness of immunosuppressive drugs in progressive MPGN. Furthermore, the development of severe nephrotic syndrome, renal insufficiency, hypertension at presentation, and negative findings of renal biopsy (formation of crescents and tubulointerstitial disease) are known as poor prognostic factors [5]. We did not administer any immunosuppressive agents to our patient because he was considered to have a low probability of renal recovery, despite treatment of MPGN itself, and he was at higher risk due to his poor general condition. In this patient, amelioration of renal function after cancer excision and chemotherapy was not observed. Based on this finding, we cannot rule out a coincidental association of MPGN with colon cancer. However, remnant metastasis may have been present, although the macroscopic masses were removed by surgery. Nevertheless, cytotoxic therapy was not performed due to age, poor general condition, drug toxicity, and low life expectancy. Other glomerular diseases such as crescentic glomerulonephropathy or acute tubular necrosis (ATN) were possible causes of the aggravated renal function. However, the pathology showed that the crescentic lesions comprised < 50% of the total and no findings compatible with ATN were identified. If ATN was the cause of the renal dysfunction, renal function should have recovered after conservative treatment. However, his renal function remains exacerbated. These findings suggest that renal failure in this patient was caused by rapid progress of glomerulopathy associated malignancy. In addition, there were no additional etiologies to explain the development of MPGN. Therefore, we believe that colon cancer may have contributed to the development of MPGN in our patient, in part, because the development of idiopathic MPGN in a relatively elderly individual is uncommon. We report here a case of rapidly progressive MPGN associated with metastatic colon cancer.

Association between vascular access failure and microparticles in hemodialysis patients

Background Vascular access failure, a major cause of morbidity in hemodialysis (HD) patients, occurs mainly at stenotic endothelium following an acute thrombotic event. Microparticles (MPs) are fragments derived from injured cell membrane and are closely associated with coagulation and vascular inflammatory responses. Methods We investigated the relationship between levels of circulating MPs and vascular access patency in HD patients. A total of 82 HD patients and 28 healthy patients were enrolled. We used flow cytometry to measure endothelial MPs (EMPs) identified by CD31+CD42− or CD51+ and platelet-derived MPs (PMPs) identified by CD31+CD42+ in plasma samples of participants. Vascular access patency was defined as an interval from the time of access formation to the time of first access stenosis in each patient. MP counts were compared according to access patent duration. Results The levels of EMP (both CD31+CD42− and CD51+) and CD31+CD42+PMP were significantly higher in patients than in healthy participants. Levels of CD31+CD42−EMP and CD31+CD42+PMP showed a positive correlation. In non-diabetic HD patients, CD31+CD42−EMPs and CD31+CD42+PMPs were more elevated in the shorter access survival group (access survival <1 year) than in the longer survival group (access survival ≥ 4 years). Conclusion Elevated circulating EMP or PMP counts are influenced by end-stage renal disease and increased levels of EMP and PMP may be associated with vascular access failure in HD patients.

The effects of indoxyl sulfate-induced endothelial microparticles on neointimal hyperplasia formation in an ex vivo model

Purpose Neointimal hyperplasia (NH) is considered to be one of the main causes of vascular access occlusion in patients receiving hemodialysis. Endothelial injury and TGF-β-mediated proliferation of vascular smooth muscle cells (VSMCs) induce NH. Endothelial microparticles (EMPs) are also increased by endothelial injury. We aimed to investigate the effects of EMPs and TGF-β expression on VSMC proliferation and their contributions to NH formation in an ex vivo model. Methods EMPs were collected from the culture media of human umbilical vein endothelial cells treated with indoxyl sulfate (IS, 250 µg/mL) after ultracentrifugation at 100,000 × g. Porcine internal jugular veins were isolated and treated with EMPs (2 × 106 /mL) or left untreated for 12 days and subsequently compared with TGF-β (10 ng/mL)-treated venous tissue. Intima-media thickness and NH area were assessed using a digital program. Massons trichrome staining and immunohistochemistry (IHC) analysis for α-smooth muscle actin, phosphorylated Akt, ERK1/2, p38 mitogen-activated protein kinase (MAPK), and Smad3 were performed on each vein sample. Results NH and VSMC proliferation developed to a significantly greater degree in EMP-treated veins compared to controls, with similar patterns seen in TGF-β-stimulated samples. IHC analysis demonstrated that EMPs markedly increased phosphorylation of Akt, ERK1/2, p38 MAPK, and Smad3 in areas of venous NH formation. Conclusion Our results showed that IS-induced EMPs provoked massive VSMC proliferation and NH formation via activation of the TGF-β signaling pathways. Further investigation is needed to elucidate the precise mechanism of EMP activity on vascular access stenosis in vivo.