Urinary excretion of thyroid hormone in CKD patients: a proof-of-concept of nephrogenic hypothyroidism.

PURPOSE: Patients with chronic kidney disease (CKD) complicated by hypothyroidism exhibit a higher prevalence of urine protein than that in the general population. This study was aimed at investigating thyroid hormones and thyroid hormone-binding proteins excreted in urine to elucidate the urine protein-associated underlying mechanisms of hypothyroidism. METHODS: Between November 2016 and August 2018, thyroid function (serum free T3 [sFT3], free T4 [sFT4], and thyroid-stimulating hormone [sTSH]), kidney function (estimated glomerular filtration rate [eGFR]), thyroid antibodies and albumin (Alb) were evaluated in 99 Japanese CKD patients with proteinuria at our outpatient clinic. A urine examination was also performed to assess the following parameters: total T3, total T4, TSH, Alb, preAlb, thyroid-binding globulin, and protein. RESULTS: The median patient age at study recruitment was 60 years; 50 patients (50.5%) were male. The median eGFR and Alb level were 20.3 ml/min/1.73 m2 and 3.8 g/dL, respectively. 21 patients (21.2%) were diagnosed with nephrotic syndrome (NS). The median sFT3, sFT4, and sTSH levels were within normal limits. Approximately 70% of the patients had thyroid dysfunction and 51.5% had overt or subclinical hypothyroidism without predominantly antibody positive. Regarding NS and non-NS patients, age and Alb were significantly different between these groups, while sex and eGFR were not significant, but the urinary T4 and TSH levels were higher in the NS group; thus, more severe hypothyroid. CONCLUSION: We found a significant association between hypothyroidism and NS regardless of sex and antibodies. Urinary loss of thyroid hormones must be a factor influencing hypothyroidism independent of autoimmunity.

Molecular and biochemical characterization of novel PAM-like MBL variants, PAM-2 and PAM-3, from clinical isolates of Pseudomonas tohonis.

BACKGROUND: There is no comprehensive study on PAM-like MBLs. OBJECTIVES: Our aim was to characterize novel B3 MBL variants, PAM-2 and PAM-3, from Pseudomonas tohonis clinical isolates. METHODS: We evaluated the antimicrobial susceptibility and the MBL gene composition of three novel P. tohonis clinical isolates identified at a Japanese hospital, using the broth microdilution method and WGS, respectively. We characterized the PAM-2 and PAM-3 proteins using recombinant protein expression and biochemical evaluations. RESULTS: Low carbapenem MICs (meropenem MIC = 0.125-1 mg/L) were observed for all three P. tohonis isolates; however, the isolates produced MBLs. We identified blaPAM-2 and blaPAM-3 as potential genes, belonging to a novel subclass of B3 MBLs. Their genomic sequence was similar to that of blaPAM-1 from Pseudomonas alcaligenes. PAM-2 and PAM-3 comprised 287 amino acids and exhibited 90% amino acid identity with PAM-1, 73% identity with POM-1 from Pseudomonas otitidis and 61% identity with L1 from Stenotrophomonas maltophilia. Biochemical evaluations of recombinant PAM-2 and PAM-3 revealed similar kcat/Km ratios and demonstrated catalytic activity against all the tested ß-lactams, except for aztreonam. In addition, the kcat/Km ratio for imipenem was 40-fold lower than that for meropenem. CONCLUSIONS: P. tohonis harbours a species-specific PAM-family MBL gene. This enzyme has higher hydrolytic activity against meropenem compared with that against imipenem.

Comparative analysis on characteristics of two activated partial thromboplastin time reagents.

BACKGROUND: For the lack of standardized activated partial thromboplastin time (APTT), it has been pointed out that there are differences in values among several reagents. Recently, we have performed a parallel measurement on two reagents, Thrombocheck APTT-SLA and Coagpia APTT-n, and resulted with some dissociated samples. The purpose of this study is to clarify the possible factors related to ΔAPTT, the difference in measured values between the two reagents. MATERIALS AND METHODS: In order to clarify the factors related to ΔAPTT, multiple regression analysis was performed on 8324 samples, using clinical laboratory data of all test items requested simultaneously with APTT. To confirm the items extracted from the multiple regression analysis, the target substance was spiked to pooled plasma and measured with two APTT reagents. Additionally, by spiking phospholipids, the effect on APTT measurement system was assessed. RESULT: Multiple regression analysis detected albumin-globulin ratio (AGR), C-reactive protein (CRP), hematocrit, and prothrombin time as factors related to ΔAPTT (p < 0.001). Results revealed no significant differences when albumin was added to change the AGR. Whereas with the addition of CRP, prolongation of APTT was observed in Coagpia APTT-n compared to Thrombocheck APTT-SLA (p < 0.001). This prolongation was canceled by the addition of phospholipids, suggesting the interaction of CRP with phospholipids leads to the pseudo-prolongation. CONCLUSION: It is considered that the pseudo-prolongation of APTT is triggered by the interaction of CRP on the phospholipid in Coagpia APTT-n, which contributed to the APTT dissociation.

Quantitative evaluation of insulin-induced abdominal subcutaneous dystrophic tissue using shear wave elastography.

AIMS/INTRODUCTION: Subcutaneous dystrophic tissue (DT) produced by insulin injection causes dysglycemia owing to inadequate absorption of insulin. However, precise techniques for measuring DT have not been established. Shear wave elastography (SWE) is an imaging technology that can quantify tissue stiffness. In this study, insulin injection-induced DT was quantified using SWE to generate whole-abdominal wall subcutaneous tissue by three-dimensional (3D) imaging in patients with type 2 diabetes who were treated with multiple insulin injections. MATERIALS AND METHODS: Seven patients with type 2 diabetes were recruited who received long-standing multiple insulin injections. Using SWE, the shear wave velocity (SWV) of DT and control (normal subcutaneous tissue) was measured. Furthermore, two of seven patients underwent whole-abdominal SWE examination to calculate the proportion of DT. A subcutaneous insulin tolerance test was also performed in both the DT and control tissues. RESULTS: The SWV in DT was significantly higher than that in the control tissue (2.87 [2.66-2.98] vs 1.29 [1.23-1.44] m/s, P < 0.01). The proportion of the DT volume was 0.67% and 5.21% for two individuals from the entire abdominal subcutaneous tissue volume. The area under the curve for the subcutaneously injected insulin aspart concentration at the DT sites was lower than that of the control tissue (75.0 [52.1-111] vs 116 [86.9-152.5] h*mU/L, P = 0.1). CONCLUSIONS: SWE can be useful in quantifying abdominal subcutaneous insulin-induced DT, especially the 3D volume of insulin injection-induced DT from the entire abdominal subcutaneous tissue. This study is the first to examine the volume and distribution of abdominal subcutaneous DT using SWE.

Pseudomonas tohonis sp. nov., isolated from the skin of a patient with burn wounds in Japan.

Strain TUM18999T was isolated from the skin of a patient with burn wounds in Japan. The strain was successfully cultured at 20-42 °C (optimum, 30-35 °C) in 1.0-4.0% NaCl (w/v) and at pH 5.5-9.5, optimum pH 5.5-8.5. The phylogenetic tree reconstructed using 16S rRNA, gyrB, rpoB and rpoD gene sequences indicated that strain TUM18999T is closely related to Pseudomonas otitidis MCC10330T. Although the partial 16S rRNA gene sequence (1412 bp) of TUM18999T exhibits high similarity to those of Pseudomonas alcaligenes NBRC 14159T (99.08 %) and Pseudomonas otitidis MCC10330T (98.51 %), multi-locus sequence analysis using 16S rRNA, gyrB, rpoB and rpoD genes reveals a clear distinction between TUM18999T and other Pseudomonas species. In addition, an average nucleotide identity >90 % was not observed in the P. aeruginosa group. Moreover, TUM18999T and P. otitidis can be distinguished based on the minimum inhibitory concentration for carbapenem. Meanwhile, the cellular fatty acids are enriched with C18 : 1 ω7c/C18 : 1 ω6c (34.35 %), C16 : 1 ω7c/C16 : 1 ω6c (24.22 %), C16 : 0 (19.79 %) and C12 : 0 (8.25 %). Based on this evidence, strain TUM18999T can be defined as representing a novel Pseudomonas species, with the proposed name Pseudomonas tohonis sp. nov. The type strain is TUM18999T (GTC 22698T=NCTC 14580T).

Evaluation of the simplified carbapenem inactivation method as a phenotypic detection method for carbapenemase-producing Enterobacterales.

Carbapenemase-producing Enterobacterales (CPE) have become a global health concern. Current molecular detection methods require special equipment and reagents. Thus, there is an urgent need for a highly sensitive, specific, and simple method for phenotypic detection of CPE in clinical microbiology laboratories. A simplified carbapenem inactivation method (sCIM) was recently reported. However, its utility for CPE detection has not been sufficiently evaluated to date. We evaluated the sCIM and compared it with the modified CIM (mCIM), using 133 CPE strains (producing IMP, 92; NDM, 11; NDM and OXA-48-like, 1; KPC, 13; OXA-48-like, 12; GES-24, 3; Nmc-A, 1) and 82 non-CPE strains (extended spectrum ß-lactamase, 61; AmpC, 21). The sCIM was conducted by loading bacteria onto imipenem and meropenem disks. When imipenem disks with a 1+ bacterial load were used, the sensitivity and specificity of the sCIM were 97.0% and 100%, and those of the mCIM were 97.0% and 96.3%, respectively. The specificity of the sCIM decreased to 57.3% when the bacterial load on imipenem disks was increased to 2+. In contrast, when meropenem disks with a 1+ bacterial load were used, the sCIM had a lower sensitivity (78.2%) and an equivalent specificity (100%). When meropenem disks with a bacterial load of 2+ were used, the sensitivity and specificity of the sCIM increased to 96.2% and 93.9%, respectively. The diameter of the inhibition zone on meropenem disks was larger than that on imipenem disks, and the sCIM was less sensitive when meropenem disks were used. In addition, sCIM detection rates when using meropenem disks were particularly low for OXA-48-like producers (bacterial load 1+, 0/12; bacterial load 2+, 10/12). Our results indicate that the sensitivity and specificity of the sCIM was dependent on the bacterial load and that large bacterial loads led to false positives for AmpC and extended spectrum ß-lactamase producers. Thus, the sCIM has high sensitivity and specificity for appropriate bacterial loads when imipenem disks are used.

Combination of a SARS-CoV-2 IgG Assay and RT-PCR for Improved COVID-19 Diagnosis.

BACKGROUND: Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is generally diagnosed by reverse transcription (RT)-PCR or serological assays. The SARS-CoV-2 viral load decreases a few days after symptom onset. Thus, the RT-PCR sensitivity peaks at three days after symptom onset (approximately 80%). We evaluated the performance of the ARCHITECT® SARS-CoV-2 IgG assay (henceforth termed IgG assay; Abbott Laboratories, Lake County, IL, USA), and the combination of RT-PCR and the IgG assay for COVID-19 diagnosis. METHODS: In this retrospective study, 206 samples from 70 COVID-19 cases at two hospitals in Tokyo that were positive using RT-PCR were used to analyze the diagnostic sensitivity. RT-PCR-negative (N=166), COVID-19-unrelated (N=418), and Japanese Red Cross Society (N=100) samples were used to evaluate specificity. RESULTS: Sensitivity increased daily after symptom onset and exceeded 84.4% after 10 days. Specificity ranged from 98.2% to 100% for samples from the three case groups. Seroconversion was confirmed from 9 to 20 days after symptom onset in 18 out of 32 COVID-19 cases with multiple samples and from another case with a positive result in the IgG assay for the first available sample. CONCLUSIONS: The combination of RT-PCR and IgG assay improves the robustness of laboratory diagnostics by compensating for the limitations of each method.

Immunochromatography and chemiluminescent enzyme immunoassay for COVID-19 diagnosis.

INTRODUCTION: The rapid and accurate detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is required to prevent the spread of COVID-19. This study evaluated the utility of two SARS-CoV-2 antigen detection methods. METHODS: We evaluated two types of antigen detection methods using immunochromatography (Espline) and quantitative chemiluminescent enzyme immunoassay (Lumipulse). RT-PCR was performed as a standard procedure for COVID-19 diagnosis. Lumipulse and RT-PCR were performed for all 486 nasopharyngeal swabs and 136 saliva samples, and the Espline test was performed for 271 nasopharyngeal swabs and 93 saliva samples. RESULTS: The sensitivity and specificity of the Espline test were 10/11 and 260/260 (100%), respectively for the nasopharyngeal swabs and 3/9 and 84/84 (100%), respectively for the saliva samples. High sensitivities for both saliva (8/9) and nasopharyngeal swabs (22/24) were observed in the Lumipulse test. The specificities of the Lumipulse test for nasopharyngeal swabs and saliva samples were 460/462 (99.6%) and 123/127 (96.9%), respectively. CONCLUSION: The Espline test is not effective for saliva samples but is useful for simple and rapid COVID-19 tests using nasopharyngeal swabs because it does not require special devices. The Lumipulse test is a powerful high-throughput tool for COVID-19 diagnosis because it has high detection performance for nasopharyngeal swabs and saliva samples.

Complete whole-genome sequence of the novel Pseudomonas species strain TUM18999, isolated from a patient with a burn wound in Japan.

OBJECTIVES: Pseudomonas is a Gram-negative bacterial genus with numerous member species. In this study, using whole-genome sequencing, we characterized a novel Pseudomonas sp. strain TUM18999, isolated as a pathogen from a human patient. METHODS: The TUM18999 strain was isolated from a patient's burn wound. Minimum inhibitory concentrations (MICs) were determined using the broth microdilution method. The whole-genome sequence was obtained using Miseq and MinION, and we conducted phylogenetic analysis based on single nucleotide polymorphisms of the core genome. RESULTS: Antimicrobial susceptibility testing revealed a high ceftazidime MIC (32 mg/L). Moreover, carbapenemase production was confirmed using the modified carbapenem inactivation method. We found that the complete genome of TUM18999 was 6,826,062 bp long, with 6175 coding sequences (CDS) and a DNA G+C content (non-plasmid) of 66.4 mol%. Consistent with the high similarities with the 16S rRNA sequences of P. otitidis MCC10330 (98.6%) and P. alcaligenes NBRC 14159 (99.2%), similarities (<90%) were also observed with the gyrB genes of both strains. The average nucleotide identities for P. alcaligenes NBRC 14159 and P. otitidis MCC10330 were also <90%. The core-genome single nucleotide polymorphism phylogenetic tree indicated that the TUM18999 strain was most closely related to P. otitidis MCC10330. In addition, the TUM18999 strain carried the novel gene, species-specific subclass B3 metallo-ß-lactamase (MBL), and its similarities with P. alcaligenes metallo-ß-lactamase-1 (PAM-1) and P. otitidis metallo-ß-lactamase-1 (POM-1) were 90.24% and 73.14%, respectively. CONCLUSION: We characterized the complete whole genome sequence of the novel Pseudomonas sp. TUM18999 carrying the novel gene species-specific subclass B3 MBL.

Plasma indoxyl sulfate levels predict cardiovascular events in patients with mild chronic heart failure.

Indoxyl sulfate (IS) is associated with either chronic kidney disease or renal failure, which may predict cardiovascular events via cardiorenal syndrome. The present study aimed to elucidate whether the plasma levels of IS can predict the occurrence of cardiovascular events in patients with chronic heart failure (CHF) and investigate which causes of CHF leading to cardiovascular events are highly influenced by plasma IS levels. We measured the plasma IS levels in 165 patients with CHF [valvular disease: 78, dilated cardiomyopathy: 29, hypertrophic cardiomyopathy (HCM): 25 and others: 33] admitted to our hospital in 2012, and we followed up these patients for more than 5 years (the median follow-up period: 5.3 years). We measured the plasma IS level in 165 patients with CHF, and Kaplan-Meier analyses showed that high plasma IS levels (≥ 0.79 µg/mL, the median value) could predict the occurrence of cardiovascular events, i.e., cardiovascular death or rehospitalization due to the worsening of CHF. The sub-analyses showed that the high IS level could predict cardiovascular events in patients with CHF due to HCM and that the plasma IS levels were closely associated with left ventricular (LV) dimension, LV systolic dysfunction, and plasma B-type natriuretic peptide levels, rather than LV diastolic dysfunction. Plasma IS level predicts cardiovascular events in patients with CHF, especially those with HCM along with cardiac dysfunction. Besides, IS may become a proper biomarker to predict cardiovascular events in patients with CHF.

Plasma matrix metalloproteinase 7, CC-chemokine ligand 18, and periostin as markers for pulmonary sarcoidosis.

BACKGROUND: Some patients with sarcoidosis experience worsening of pulmonary lesions. However, no biomarker has been identified that reflects pulmonary disease status in sarcoidosis. We investigated the usefulness of potential markers of pulmonary fibrosis in patients with sarcoidosis. METHODS: Plasma matrix metalloproteinase 7 (MMP-7), CC-chemokine ligand 18 (CCL-18), and periostin levels were evaluated in 60 patients with sarcoidosis and 30 healthy controls; bronchoalveolar lavage fluid levels were analyzed in 22 patients with sarcoidosis. To determine the usefulness of these markers, we explored potential correlations between these markers and sarcoidosis clinical characteristics. RESULTS: Plasma MMP-7, CCL-18, and periostin concentrations were significantly higher in patients with sarcoidosis than those in healthy controls. MMP-7 concentrations in plasma and bronchoalveolar lavage fluid were higher in patients with sarcoidosis with parenchymal infiltration than in those without lung lesions. Moreover, MMP-7 concentration was negatively correlated with pulmonary function. CONCLUSION: Among these novel biomarkers, MMP-7 most precisely reflected pulmonary sarcoidosis disease status and thus, might be useful for diagnosing and evaluating sarcoidosis, particularly in patients with pulmonary parenchymal lesions.

Role of growth hormone signaling pathways in the development of atherosclerosis.

OBJECTIVE: The direct actions of growth hormone (GH) in the development of atherosclerosis are unclear. The goal of this study was to characterize GH-induced changes in expression of signaling pathway elements and other proteins that may be related to atherosclerosis. METHODS: Human umbilical vein endothelial cells (HUVEC) and THP-1, a human acute monocytic leukemia cell line, were stimulated by exposure to 10-9 M or 10-8 M human GH with or without pretreatment with a mitogen-activated protein kinase kinase (MEK) 1 inhibitor. Levels of transcripts encoding vascular cell adhesion molecule (VCAM) -1, E-selectin, monocyte chemotactic protein (MCP-1), interleukin (IL) -6, and IL-8 were investigated by reverse transcription (RT) -PCR. For the quantitative adhesion assay, THP-1 cells or human primary monocytes were fluorescently labeled with 3'-O-acetyl-2',7'-bis(carboxyethyl) -4 diacetoxymethyl ester (BCECF/AM). HUVEC treated with human GH were co-incubated with BCECF-labeled THP-1 cells. One hour later, the number of BCECF-labeled THP-1 cells was assessed. An equivalent experiment was performed using BCECF-labeled primary monocytes, and the number of monocytes adhering to HUVEC was counted. RESULTS: Treatment with hGH increased the levels of E-selectin- and VCAM-1-encoding mRNAs in HUVEC. This effect was attenuated by pretreatment with a MEK1 inhibitor. Furthermore, hGH treatment increased adhesion of BCECF-labeled THP-1 cells or primary monocytes to HUVEC, and this effect was attenuated by pretreatment with a MEK1 inhibitor. CONCLUSIONS: VCAM-1 and E-selectin expression was stimulated by GH via the mitogen-activated protein kinase pathway, resulting in augmented adhesion of THP-1 cells and monocytes to HUVEC. These data suggested that GH directly stimulates the development of atherosclerosis.

Carbapenem inactivation method using bacterial lysate and MOPS (LCIM): a very sensitive method for detecting carbapenemase-producing Acinetobacter species.

OBJECTIVES: Detection of carbapenem-hydrolysing class D ß-lactamase (CHDL)-producing Acinetobacter spp. is critical for understanding antibiotic resistance. In this study, we compared the available detection techniques derived from the carbapenem inactivation method (CIM), using CHDL-producing Acinetobacter spp., and developed a modified method that uses bacterial lysate (lysate CIM; LCIM). METHODS: A total of 159 Acinetobacter spp. (102 carbapenemase producers and 57 non-producers) and 14 Pseudomonas spp. (7 carbapenemase producers and 7 non-producers) were tested. Modified CIM, simplified CIM, CIMTris, Triton-CIM and LCIM were compared using these strains. Distinct from the CIM, LCIM includes a longer incubation period (4 h) with 2.0% Triton X-100 (v/v) in 20 mM MOPS buffer instead of water. RESULTS: The sensitivity/specificity of the modified CIM, simplified CIM, CIMTris, Triton-CIM and LCIM were 71.6%/100%, 66.1%/89.1%, 88.1%/95.3%, 80.7%/100% and 97.2%/100%, respectively. LCIM was the most sensitive and specific. CONCLUSIONS: Use of bacterial lysate and MOPS increased the sensitivity of the CIM in detecting CHDL-producing Acinetobacter spp.

The Difference in the Changes of Indoxyl Sulfate after Catheter Ablation among Atrial Fibrillation Patients with and without Kidney Dysfunction.

Indoxyl sulfate (IS), a protein-bound uremic toxin, induces chronic kidney disease (CKD) and atrial fibrillation (AF). Catheter ablation (CA) of AF improves the renal function. However, the transition of uremic toxins is unclear. This study aimed to investigate the transition of the serum IS level in AF patients with and without CKD after CA. A total of 138 consecutive AF patients who underwent CA and maintained sinus rhythm were prospectively enrolled (paroxysmal AF 67.4%). The patients were divided into 4 groups (non-CKD/low-IS:68, non-CKD/high-IS:28, CKD/low-IS:13, and CKD/high-IS:29). The plasma IS levels and estimated glomerular filtration rate (eGFR) were determined before and 1-year after CA. CKD was defined as CKD stage III and a high-IS according to the mean IS (IS ≥ 1.1 µg/ml). CA significantly improved the eGFR in CKD patients (p < 0.001). The serum IS level in the non-CKD/high-IS group was significantly decreased (from 1.7 ± 0.7 to 1.1 ± 0.6 µg/ml, p < 0.001). However, the serum IS level in the CKD/high-IS group did not improve (from 1.9 ± 0.9 to 1.7 ± 0.7 µg/ml, p = 0.22). The change in the IS in the CKD patients significantly differed from that in those without CKD. In the CKD patients, CA did not significantly decrease the IS, a risk factor of CKD, regardless of an improved eGFR.

Evaluation of inhibitor-combination mCIM for detecting MBL-producing Enterobacterales using three MBL inhibitors.

The increase in carbapenemase-producing Enterobacterales (CPE), including metallo-ß-lactamase (MBL) producers, is a severe global health concern. Thus, highly sensitive and specific methods for detecting MBL producers are needed. In this study, we tested the detectability of MBL-producing Enterobacterales against three types of MBL inhibitors (sodium mercaptoacetate, SMA; ethylenediaminetetraacetic acid, EDTA; and dipicolinic acid, DPA) used in combination with a modified carbapenem inactivation method (mCIM). These inhibitor-combination mCIMs were tested against 129 CPE (IMP, 93; NDM, 11; KPC, 13; NMC, 1; OXA-48, 11) and 75 non-CPE. For evaluation of MBL inhibitors, we used two concentrations for each of the three inhibitors: DPA (200 and 300 mg l- 1), EDTA (5 and 10 mM), and SMA (1500 and 3000 mg l- 1). The overall sensitivities of SMA, EDTA and DPA were 97.1-99.0 %, 81.7-99.0 % and 88.5-96.2 %, respectively. Moreover, each method showed high specificity (99.0-100 %). Although inhibitor-combination mCIMs were highly sensitive and specific for the detection of MBL producers, we found that sensitivity was dependent on the concentration of inhibitors.

Chemerin-9, a potent agonist of chemerin receptor (ChemR23), prevents atherogenesis.

Plasma levels of chemerin, an adipocytokine produced from the adipose tissues and liver, are associated with metabolic syndrome and coronary artery disease (CAD). Chemerin and its analog, chemerin-9, are known to bind to their receptor, ChemR23. However, whether chemerin and chemerin-9 affect atherogenesis remains to be elucidated. We investigated the expression of chemerin and ChemR23 in human coronary arteries and cultured human vascular cells. The effects of chemerin and chemerin-9 on atheroprone phenomena were assessed in human THP1 monocytes, human umbilical vein endothelial cells (HUVECs), and human aortic smooth muscle cells (HASMCs) and aortic lesions in Apoe-/- mice. In patients with CAD, a small amount of ChemR23, but not chemerin, was expressed within atheromatous plaques in coronary arteries. Chemerin and ChemR23 were expressed at high levels in THP1 monocytes, THP1-derived macrophages, and HUVECs; however, their expression in HASMCs was weak. Chemerin and chemerin-9 significantly suppressed the tumor necrosis factor-α (TNF-α)-induced mRNA expression of adhesion and pro-inflammatory molecules in HUVECs. Chemerin and chemerin-9 significantly attenuated the TNF-α-induced adhesion of THP1 monocytes to HUVECs and macrophage inflammatory phenotype. Chemerin and chemerin-9 suppressed oxidized low-density lipoprotein (oxLDL)-induced macrophage foam cell formation associated with down-regulation of CD36 and up-regulation of ATP-binding cassette transporter A1 (ABCA1). In HASMCs, chemerin and chemerin-9 significantly suppressed migration and proliferation without inducing apoptosis. In the Apoe-/- mice, a 4-week infusion of chemerin-9 significantly decreased the areas of aortic atherosclerotic lesions by reducing intraplaque macrophage and SMC contents. Our results indicate that chemerin-9 prevents atherosclerosis. Therefore, the development of chemerin analogs/ChemR23 agonists may serve as a novel therapeutic target for atherosclerotic diseases.

Performance evaluation of an Indoxyl Sulfate Assay Kit "NIPRO".

Background The relationship between renal disease and cardiovascular disease (CVD) is currently known as cardiorenal syndrome. Indoxyl sulfate (IS) is one of the uremic toxins that accelerates the progression of cardiorenal syndrome. This report presents a new method for measuring IS in a simpler way. Methods We evaluated the analytical performance of an IS Assay Kit "NIPRO" loaded on LABOSPECT 008. The evaluated analytical performances included accuracy, precision, dilution linearity, limit of detection (LOD), limit of quantitation (LOQ), recovery test, interference test and comparison against assays performed by high-performance liquid chromatography (HPLC). Results Total precision showed a <5.3% coefficient of variation at IS concentrations of 3.57-277.73 µmol/L, and satisfactory results were observed in the dilution linearity, LOD, LOQ, recovery and interference tests. The IS Assay Kit "NIPRO" showed a high correlation with the HPLC conventional method (r = 0.993). Conclusions The IS Assay Kit "NIPRO" demonstrated satisfactory analytical performance, and this suggests it could shortly become another common method to measure circulating IS.

AST-120, an Adsorbent of Uremic Toxins, Improves the Pathophysiology of Heart Failure in Conscious Dogs.

PURPOSE: Several lines of evidence suggest that renal dysfunction is associated with cardiovascular toxicity through the action of uremic toxins. The levels of those uremic toxins can be reportedly reduced by the spherical carbon adsorbent AST-120. Because heart failure (HF) causes renal dysfunction by low cardiac output and renal edema, the removal of uremic toxins could be cardioprotective. METHOD: To determine whether blood levels of the uremic toxin indoxyl sulfate (IS) increase in HF and whether AST-120 can reduce those levels and improve HF. We induced HF in 12 beagle dogs by 6 weeks of rapid right ventricular pacing at 230 beats per min. We treated six dogs with a 1-g/kg/day oral dosage of AST-120 for 14 days from week 4 after the start of rapid ventricular pacing. The other six dogs did not receive any treatment (control group). RESULTS: In the untreated dogs, IS levels increased as cardiac function deteriorated. In contrast, plasma IS levels in the treated dogs decreased to baseline levels, with both left ventricular fractional shortening and pulmonary capillary wedge pressure also improving when compared with untreated dogs. Finally, AST-120 treatment was shown to reduce both myocardial apoptosis and fibrosis along with decreases in extracellular signal-regulated kinase phosphorylation, the Bax/Bcl-2 ratio, and TGF-ß1 expression and increases in AKT phosphorylation. CONCLUSIONS: IS levels are increased in HF. AST-120 treatment reduces the levels of IS and improves the pathophysiology of HF in a canine model. AST-120 could be a novel candidate for the treatment of HF.

Prognostic Utility of Indoxyl Sulfate for Patients with Acute Coronary Syndrome.

AIM: We investigated whether indoxyl sulfate (IS), a protein-bound uremic toxin, predicts prognosis after acute coronary syndrome (ACS). METHODS: Serum IS level was determined prospectively in 98 patients who underwent successful primary percutaneous coronary intervention for ACS. Patients on hemodialysis were excluded. The endpoint of this study was six-month composite events including death, nonfatal myocardial infarction, heart failure requiring hospitalization, and adverse bleeding events. RESULTS: During the mean follow-up period of 168 days, composite events occurred in 13.3% of cases. Serum IS level was significantly higher in subjects who developed composite events than in those without events (0.14±0.11 mg/dl vs. 0.06±0.04 mg/dl; p＜0.001). After adjusting for confounding factors, a Cox proportional hazard analysis revealed that the IS level (hazard ratio (HR): 10.6; 95% confidence interval (CI): 1.63-69.3, p=0.01), hemoglobin level (HR: 0.61; 95% CI: 0.43-0.87; p＜0.01), and left ventricular ejection fraction (LVEF) (HR: 0.95; 95% CI: 0.91-0.99; p=0.03) were independent predictive factors of composite events. Furthermore, IS level significantly conferred additional value to the combined established risks of LVEF and hemoglobin level for predicting the incidence of composite events (area under the curve: 0.82 vs. 0.88, p=0.01; net reclassification improvement: 0.67, p=0.01; and integrated discrimination improvement: 0.15, p＜0.01). CONCLUSIONS: The assessment of serum IS level has prognostic utility for the management of ACS.

The relationship between serum indoxyl sulfate and the renal function after catheter ablation of atrial fibrillation in patients with mild renal dysfunction.

Indoxyl sulfate (IS), a protein-bound uremic toxin, induces renal disorders and atrial fibrillation (AF). It is well known that renal dysfunction is a risk factor for AF and radiofrequency catheter ablation (RFCA) improves the renal function. However, the improvement in the renal function after RFCA in patients with early stage chronic kidney disease (CKD) and the serial changes in the IS level have not been fully elucidated. This study aimed to investigate whether IS affects the improvement in the renal function. A total of 91 consecutive patients with mild kidney dysfunction (CKD stage I-II) who underwent RFCA and maintained sinus rhythm were prospectively enrolled. The plasma IS level and estimated glomerular filtration rate (eGFR) were determined before, 3 months, and 1 year after RFCA. The patients were divided according to the IS quartiles (Q1-4; < 0.4, 0.4-0.7, 0.7-1.2, and > 1.2 µg/ml). There was no significant difference in the eGFR among the IS quartiles. A significantly higher eGFR improvement rate was obtained for IS-Q4 than the other quartiles (p = 0.039). The IS-Q4 IS level significantly decreased at 1 year after RFCA (1.8 ± 0.8 to 1.2 ± 0.7 µg/ml, p < 0.01). The multivariable logistic model revealed that a high-IS level (IS-Q4) was an independent predictor of an eGFR improvement (OR 3.33; 95% CI 1.16-9.59; p = 0.026). A high-IS level reduction after RFCA improved the renal function in AF patients with mild kidney dysfunction.