Gemigliptin attenuates the expression of sodium phosphate co-transporter PiT-1.

<p>The expression of PiT-1 was evaluated by qRT-PCR and western blot after treatment with 3 mM phosphate and/or 50 μM gemigliptin for 2 days (A) or 4 days (B). GAPDH was used as the loading control. The results are expressed as percentage of control. Results are presented as mean ± SEM (n = 5–6 in each group). Con, control; Gemi50, 50 μM gemigliptin; Pi, 3mM phosphate; PG50, 3mM phosphate and 50 μM gemigliptin. *P<0.5, **P<0.01, ***P<0.001 compared with control and #P<0.5, ##P<0.01, ###P<0.001 compared with phosphate group.</p

Free Thyroxine Level as an Independent Predictor of Infection-Related Mortality in Patients on Peritoneal Dialysis: A Prospective Multicenter Cohort Study

<div><p>Background</p><p>Previous studies have reported the relationship between thyroid hormone levels and mortality in dialysis patients. However, little is known about the association of free thyroxine (fT4) and mortality in patients on peritoneal dialysis (PD). This study investigated the association between basal and annual variation in fT4 level and mortality in PD patients.</p><p>Methods</p><p>Patients on maintenance PD were enrolled from a prospective multicenter cohort study in Korea; their serum triiodothyronine, fT4, and thyroid-stimulating hormone levels were measured 12 months apart. Patients with overt thyroid disease and those receiving thyroid hormone replacement therapy were excluded from the analysis. Patients were divided into two groups based on the median levels of fT4. The differences of all-cause, infection-related, and cardiovascular mortalities were analyzed between the two groups. The association of basal levels and annual variation with mortality was investigated with Kaplan–Meier curves and Cox proportional hazard models.</p><p>Results</p><p>Among 235 PD patients, 31 (13.2%) deaths occurred during the mean follow-up period of 24 months. Infection (38.7%) was the most common cause of death. Lower basal fT4 levels were an independent predictor of all-cause and infection-related death (hazard ratio [HR] = 2.74, 95% confidence interval [CI] 1.27–5.90, <i>P</i> = 0.01, and HR = 6.33, 95% CI 1.16–34.64, <i>P</i> = 0.03, respectively). Longitudinally, patients with persistently lower fT4 levels during the 12-month period had significantly higher all-cause mortality than those with persistently higher levels (HR = 3.30, 95% CI 1.15–9.41, <i>P</i> = 0.03). The area under the receiver operating characteristic curve of fT4 for predicting all-cause and infection-related mortality was 0.60 and 0.68, respectively.</p><p>Conclusions</p><p>fT4 level is an independent predictor of mortality and is especially attributable to infection in PD patients. This predictor was consistent when considering both baseline measurements and annual variation patterns. Close attention to infection in PD patients with relatively lower fT4 levels should be considered.</p></div

Gemigliptin modulates ROS generation and the expression of NADPH oxidase <i>NOX4</i> and NADPH oxidase subunit <i>p22</i>^<i>phox</i>.

<p>(A) Intracellular ROS generation was visualized by fluorescence microscopy, and (B) quantified in the cellular lysate after staining with 10 μM H₂DCF-DA. (C) The level of H₂O₂ generation was measured using the Amplex red hydrogen peroxide assay kit. The mRNA expression of <i>NOX4</i> and <i>p22</i>^<i>phox</i> was analyzed by qRT-PCR after treating VSMCs with phosphate and/or gemigliptin for (D) 7 days and (E) 14 days. The results are expressed as percentage of control. Results are presented as mean ± SEM (n = 6–7 in each group). Con, control; Pi, 3 mM phosphate; PG50 –PG500, 3 mM phosphate and 50 μM– 500 μM gemigliptin. *P<0.5, **P<0.01, ***P<0.001 compared with control and #P<0.5, ##P<0.01, ###P<0.001 compared with phosphate group.</p

Gemigliptin attenuates vascular calcification in the abdominal aorta.

<p>(A) Representative photomicrographs of the abdominal aorta stained by Von Kossa staining (original magnification, × 17 and × 100). (B) The percentage of calcified area was calculated as the ratio of the Von Kossa positive area versus the total tissue area. (C) Representative photomicrographs of immunohistochemistry for RUNX2 (original magnification, × 200) are shown. (D) Representative immunoblot images and the quantification of RUNX2 expression are shown. GAPDH was used as the loading control. The results are expressed as percentage of control. Results are presented as mean ± SEM (n = 5 for LP, low protein group; n = 5 for adenine group; n = 6 for AG10, adenine-gemigliptin [10mg/kg] group; n = 6 for AG20; adenine-gemigliptin [20mg/kg] group). RUNX2, runt-related transcription factor-2. *P<0.5, **P<0.01, and ***P<0.001 compared with LP group, and #P<0.5, ###P<0.001 compared with adenine group.</p

A flow diagram of the effect of gemigliptin against high-phosphate-induced vascular calcification.

<p>Gemigliptin attenuated vascular calcification and osteogenic trans-differentiation in VSMCs via multiple steps including downregulation of PiT-l expression, and suppression of ROS generation, phosphor-PI3K/AKT, and Wnt signaling pathway.</p

Gemigliptin attenuates high phosphate-induced Wnt signaling.

<p>The expression of <i>FDZ3</i> and <i>DKK-</i>1 was evaluated by qRT-PCR (A and B) and western blot (C and D) in VSMCs after incubating with 3 mM phosphate and/or 50 μM gemigliptin for 7 days (A and C) and 14 days (B and D). The results are expressed as percentage of control. Results are presented as mean ± SEM (n = 5–6 in each group). FDZ3, frizzled-3; DKK-1, dickkopf-1; Con, control; Gemi50, 50 μM gemigliptin; Pi, 3mM phosphate; PG50, 3mM phosphate and 50 μM gemigliptin. *P<0.5, **P<0.01, ***P<0.001 compared with control and #P<0.5, ##P<0.01, ###P<0.001 compared with phosphate group.</p

Gemigliptin attenuates phosphate-induced phospho-PI3K-AKT signaling pathway.

<p>(A) VSMCs were treated with 3 mM phosphate and/or 50 μM gemigliptin for 24 h. Representative immunoblot images are shown. Graph represents quantitative data as (B) ratio of phospho-AKT/total-AKT or (C) phospho-PI3K/total PI3K. The results are expressed as percentage of control. Results are presented as mean ± SEM (n = 4–5 in each group). Con, control; Gemi50, 50 μM gemigliptin; Pi, 3 mM phosphate; PG50, 3 mM phosphate and 50 μM gemigliptin. *P<0.5, **P<0.01, ***P<0.001 compared with control, #P<0.5, ##P<0.01, ###P<0.001 compared with phosphate group.</p

Gemigliptin attenuates high phosphate-induced vascular calcification in VSMCs.

<p>(A) Visualization of calcium deposition on VSMCs was assessed by alizarin red staining. Representative pictures were shown. (B) After HCl decalcification, calcium content of the cells was assayed. The value was normalized by the total protein and expressed as percentage of control. (C) Cell viability was determined using MTT assay. The results are expressed as percentage of control. Results are presented as mean ± SEM (n = 6–7 in each group). Con, control; Gemi50, 50 μM gemigliptin; Pi, 3mM phosphate; PG50, 3 mM phosphate and 50 μM gemigliptin. ***P<0.001 compared with control and ###P<0.001 compared with phosphate group.</p

Hazard ratios of mortality based on time-dependent hemoglobin (Hb) levels in all the patients.

<p>Multivariate analysis sequentially adjusted models for covariates as follows. Model 1: age, sex, dialysis vintage, diabetes, cardiovascular disease, albumin, and high-sensitivity C-reactive protein; Model 2: Model 1 + erythropoiesis-stimulating agent (ESA) dose, ferritin, and transferrin saturation (TSAT). ESA dose was categorized as untreated (reference), 1 to <5000, 5000 to <10000, 10000 to <15000, and ≥15000 IU/week. Ferritin was categorized as <100, 100 to <500, and ≥500 ng/mL. TSAT was categorized as <20, 20 to <50, and ≥50%.</p

Baseline demographic, clinical, and biochemical data according to fT4 dichotomization in 235 peritoneal dialysis patients.

<p>Values are shown as mean ± standard deviation.</p><p>APD, automated peritoneal dialysis; CAPD, continuous ambulatory peritoneal dialysis; CRP, C-reactive protein; eGFR, estimated glomerular filtration rate; LDL, low-density lipoprotein; T3, triiodothyronine; TSH, thyroid-stimulating hormone.</p><p>Baseline demographic, clinical, and biochemical data according to fT4 dichotomization in 235 peritoneal dialysis patients.</p