Curie Ahn

Patient-Specific Embryonic Stem Cells Derived from Human SCNT Blastocysts

Patient-specific, immune-matched human embryonic stem cells (hESCs) are anticipated to be of great biomedical importance for studies of disease and development and to advance clinical deliberations regarding stem cell transplantation. Eleven hESC lines were established by somatic cell nuclear transfer (SCNT) of skin cells from patients with disease or injury into donated oocytes. These lines, nuclear transfer (NT)–hESCs, grown on human feeders from the same NT donor or from genetically unrelated individuals, were established at high rates, regardless of NT donor sex or age. NT-hESCs were pluripotent, chromosomally normal, and matched the NT patients DNA. The major histocompatibility complex identity of each NT-hESC when compared to the patients own showed immunological compatibility, which is important for eventual transplantation. With the generation of these NT-hESCs, evaluations of genetic and epigenetic stability can be made. Additional work remains to be done regarding the development of reliable directed differentiation and the elimination of remaining animal components. Before clinical use of these cells can occur, preclinical evidence is required to prove that transplantation of differentiated NT-hESCs can be safe, effective, and tolerated.

Suppression of NF-κB signaling by KEAP1 regulation of IKKβ activity through autophagic degradation and inhibition of phosphorylation

IkappaB kinase beta (IKKbeta) plays a crucial role in biological processes, including immune response, stress response, and tumor development by mediating the activation of various signaling molecules such as NF-kappaB. Extensive studies on the mechanisms underlying IKK activation have led to the identification of new activators and have facilitated an understanding of the cellular responses related to NF-kappaB and other target molecules. However, the molecular processes that modulate IKK activity are still unknown. In this study, we show that KEAP1 is a new IKK binding partner, which is responsible for the down-regulation of TNFalpha-stimulated NF-kappaB activation. The E(T/S)GE motif, which is found only in the IKKbeta subunit of the IKK complex, is essential for interaction with the C-terminal Kelch domain of KEAP1. Reduction of KEAP1 expression by small interfering RNA enhanced NF-kappaB activity, and up-regulated the expression of NF-kappaB target genes. Ectopic expression of KEAP1 decreased the expression of IKKbeta, which was restored by an autophagy inhibitor. IKK phosphorylation stimulated by TNFalpha was blocked by KEAP1. Our data demonstrate that KEAP1 is involved in the negative regulation of NF-kappaB signaling through the inhibition of IKKbeta phosphorylation and the mediation of autophagy-dependent IKKbeta degradation.

Benefits of biocompatible PD fluid for preservation of residual renal function in incident CAPD patients: a 1-year study

BACKGROUND In vitro studies of peritoneal dialysis (PD) solutions demonstrated that a biocompatible fluid with neutral-pH and low glucose degradation products (LF) has better biocompatibility than a conventional acidic lactate-buffered fluid (CF). However, few clinical trials have investigated the effects of the biocompatible solution on residual renal function (RRF). We performed a prospective, randomized trial with patients starting continuous ambulatory peritoneal dialysis (CAPD). METHODS Ninety-one incident patients started CAPD for 12-month treatment with either LF (Balance, Fresenius, n = 48) or CF (CAPD/DPCA, Fresenius, n = 43). RRF, peritoneal solute transport rate and solute clearance were measured every 6 months. RESULTS LF had a significant effect on the change of glomerular filtration rate (GFR) (P = 0.048 by the mixed model). In per-protocol analysis, GFR in the LF group did not decrease over a 12-month period, while GFR in the control group significantly decreased (0.13 +/- 33.4 L/ week/1.73 m(2) for LF versus -13.6 +/- 19.4 L/week/1.73 m(2) for CF, P = 0.049). Subgroup analysis for patients with initial GFR of 2 mL/min/1.73 m(2) or above showed a significantly higher GFR for the LF group over the 12-month period. At Month 13, serum total CO(2) levels were higher and serum albumin levels were lower in the LF group. No differences between the two groups were observed for the C-reactive protein. Over the 12-month period, effluent cancer antigen-125 levels significantly increased in the LF group, compared with those of the CF group, while effluent interleukin-6 levels were not different between the two groups. CONCLUSION Our study suggests that LF may better preserve RRF over the 12-month treatment period in incident CAPD patients.

Cyst Formation in Kidney via B-Raf Signaling in the PKD2 Transgenic Mice

The pathogenic mechanisms of human autosomal dominant polycystic kidney disease (ADPKD) have been well known to include the mutational inactivation of PKD2. Although haploinsufficiency and loss of heterozygosity at the Pkd2 locus can cause cyst formation in mice, polycystin-2 is frequently expressed in the renal cyst of human ADPKD, raising the possibility that deregulated activation of PKD2 may be associated with the cystogenesis of human ADPKD. To determine whether increased PKD2 expression is physiologically pathogenic, we generated PKD2-overexpressing transgenic mice. These mice developed typical renal cysts and an increase of proliferation and apoptosis, which are reflective of the human ADPKD phenotype. These manifestations were first observed at six months, and progressed with age. In addition, we found that ERK activation was induced by PKD2 overexpression via B-Raf signaling, providing a possible molecular mechanism of cystogenesis. In PKD2 transgenic mice, B-Raf/MEK/ERK sequential signaling was up-regulated. Additionally, the transgenic human polycystin-2 partially rescues the lethality of Pkd2 knock-out mice and therefore demonstrates that the transgene generated a functional product. Functional strengthening or deregulated activation of PKD2 may be a direct cause of ADPKD. The present study provides evidence for an in vivo role of overexpressed PKD2 in cyst formation. This transgenic mouse model should provide new insights into the pathogenic mechanism of human ADPKD.

Chronic asymptomatic pyuria precedes overt urinary tract infection and deterioration of renal function in autosomal dominant polycystic kidney disease

BackgroundUrinary tract infection (UTI) occurs in 30%-50% of individuals with autosomal dominant polycystic kidney disease (ADPKD). However, the clinical relevance of asymptomatic pyuria in ADPKD patients remains unknown.MethodsWe retrospectively reviewed medical records of 256 ADPKD patients who registered to the ADPKD clinic at Seoul National University Hospital from Aug 1999 to Aug 2010. We defined the asymptomatic pyuria as more than 5-9 white blood cells in high-power field with no related symptoms or signs of overt UTI. Patients were categorized into 2 groups depending on its duration and frequency: Group A included non-pyuria and transient pyuria patients; Group B included recurrent and persistent pyuria patients. The association between asymptomatic pyuria and both the development of overt UTI and the deterioration of renal function were examined.ResultsWith a mean follow-up duration of 65.3 months, 176 (68.8%) out of 256 patients experienced 681 episodes of asymptomatic pyuria and 50 episodes of UTI. The annual incidence of asymptomatic pyuria was 0.492 episodes/patient/year. The patients in group B showed female predominance (58.5% vs. 42.0%, P=0.01) and experienced an upper UTI more frequently (hazard ratio: 4.612, 95% confidence interval: 1.735-12.258; P=0.002, adjusted for gender and hypertension). The annual change in estimated glomerular filtration rate (ΔeGFR) was significantly larger in magnitude in group B than in group A (-2.7±4.56 vs. -1.17±5.8, respectively; P=0.01). Age and Group B found to be the independent variables for ΔeGFR and developing end-stage renal disease (16.0% vs. 4.3%, respectively; P=0.001).ConclusionsChronic asymptomatic pyuria may increase the risk of developing overt UTI and may contribute to declining renal function in ADPKD.

Hemorrhagic Fever with Renal Syndrome Caused by the Seoul Virus

The Seoul virus is an important etiologic agent in hemorrhagic fever with renal syndrome (HFRS), and infections with the Seoul virus are less severe than those with the Hantaan virus. However, the information on HFRS caused by the Seoul virus is limited in Korea. Retrospective clinical analysis was done on 30 patients with Seoul virus infection who had been diagnosed as having HFRS by clinical features and serologic testing by the plaque reduction neutralization test from 1986 to 1991 at the Seoul National University Hospital. They were compared with 69 patients with Hantaan virus infection. The Seoul virus was the etiologic agent in 25% of Korean HFRS and the major cause of HFRS during the summer season although infections occurred throughout the year. The Seoul virus infection had a milder degree of bleeding and renal derangement but had severer liver dysfunction than the Hantaan virus infection. Renal histopathologic findings revealed a milder degree of hemorrhage and vascular changes than cases involving Hantaan virus infection. The precise mechanisms of vascular dysfunction and organ involvement in Seoul virus infection, however, still remain to be explored.

Cyclosporine-induced Renal Injury Induces Toll-like Receptor and Maturation of Dendritic cells

Background. The toll-like receptor (TLR) is stimulated by not only pathogen-associated molecular patterns but also endogenous TLR ligands provided by injured cells. The influence of cyclosporine A (CsA)-induced renal injury on TLR expression and subsequent signaling pathway was evaluated. Methods. Induction of chronic CsA nephropathy was made by administering CsA (15 mg/kg/day) for 28 days in rats. The TLR2 and TLR4 mRNA and protein expression, TLR-signaling pathway (MYD88, NF-&kgr;B and AP-1), putative TLR ligand (heat shock protein 70 [HSP70]), and maturation of dendritic cells were evaluated in CsA-treated rat kidneys. Results. Long-term CsA treatment upregulated TLR2 and TLR4 mRNA and protein expression on renal tubular cells, and these were accompanied by increased MYD88, NF-&kgr;B and AP-1 expression. Putative TLR ligand (HSP70) was also significantly increased in CsA-treated rat kidney compared with vehicle-treated rat kidney. CsA-treatment increased expression of TNF-α mRNA, the number of dendritic cells, and expression of MHC class II antigen. Double-labeling of markers of dendritic cells and MHC class II antigen revealed that matured dendritic cells increased in CsA-treated rat kidney. Conclusions. CsA-induced renal injury stimulates components of innate immunity, and this finding suggests close association between CsA-induced renal injury and activation of innate immunity.

Influence of strain and age differences on the yields of porcine islet isolation: extremely high islet yields from SPF CMS miniature pigs

Abstract:  Background:  Porcine pancreas is a potential source of material for islet xenotransplantation. However, the difficulty in isolating islets, because of their fragility and the variability of isolation outcome in donor age and breed, represents a major obstacle to porcine islet xenotransplantation. In this study, we compared the islet isolation yield of specific pathogen‐free (SPF) Chicago Medical School (CMS) miniature pigs with that of another miniature pig breed and market pigs from a local slaughterhouse.

IL-2/Anti-IL-2 Complex Attenuates Renal Ischemia-Reperfusion Injury through Expansion of Regulatory T Cells

Regulatory T cells (Tregs) can suppress immunologic damage in renal ischemia-reperfusion injury (IRI), but the isolation and ex vivo expansion of these cells for clinical application remains challenging. Here, we investigated whether the IL-2/anti-IL-2 complex (IL-2C), a mediator of Treg expansion, can attenuate renal IRI in mice. IL-2C administered before bilateral renal IRI induced Treg expansion in both spleen and kidney, improved renal function, and attenuated histologic renal injury and apoptosis after IRI. Furthermore, IL-2C administration reduced the expression of inflammatory cytokines and attenuated the infiltration of neutrophils and macrophages in renal tissue. Depletion of Tregs with anti-CD25 antibodies abrogated the beneficial effects of IL-2C. However, IL-2C-mediated renal protection was not dependent on either IL-10 or TGF-β. Notably, IL-2C administered after IRI also enhanced Treg expansion in spleen and kidney, increased tubular cell proliferation, improved renal function, and reduced renal fibrosis. In conclusion, these results indicate that IL-2C-induced Treg expansion attenuates acute renal damage and improves renal recovery in vivo, suggesting that IL-2C may be a therapeutic strategy for renal IRI.

Intra-peritoneal interleukin-6 system is a potent determinant of the baseline peritoneal solute transport in incident peritoneal dialysis patients

BACKGROUND Interleukin-6 (IL-6) is a key player in modulating inflammation. IL-6 and soluble IL-6 receptor (sIL-6R) complex induces the synthesis and secretion of various chemokines, adhesion molecules and angiogenic molecules. We hypothesized that the baseline peritoneal solute transport rate (PSTR) early after commencing peritoneal dialysis (PD) may depend largely on the IL-6/sIL-6R system. We also hypothesized that the dialysate concentrations of IL-6/sIL-6R could be closely related to local inflammation or angiogenesis in the peritoneal cavity. METHODS Fifty incident patients with a modified peritoneal equilibration test result within 3 months after commencing PD and without a previous history of peritonitis were enrolled. Clinical parameters such as age, sex, comorbid disease, body mass index, residual renal function and C-reactive protein were assessed. Serum and dialysate markers including CA125, IL-6, sIL-6R, monocyte chemoattractant protein-1 (MCP-1), vascular endothelial growth factor (VEGF) and angiopoietin-2 (Ang-2) were measured and correlated with PSTR. RESULTS Dialysate concentrations of IL-6 (r = 0.576, P < 0.001), MCP-1 (r = 0.408, P = 0.003) and Ang-2 (r = 0.408, P = 0.003) correlated with mass transfer area coefficient for creatinine (MTAC(cr)), respectively. Dialysate appearance rate (AR) of albumin correlated with dialysate concentrations of CA125 (r = 0.751, P < 0.001), IL-6 (r = 0.303, P = 0.039), sIL-6R (r = 0.497, P < 0.001), MCP-1 (r = 0.488, P < 0.001), VEGF (r = 0.443, P = 0.004) and Ang-2 (r = 0.488, P < 0.001). Neither MTAC(cr) nor AR of albumin was associated with systemic markers. Multivariate analysis showed that MTAC(cr) is independently associated with dialysate IL-6 and serum albumin. It also showed that AR of albumin is independently predicted by dialysate sIL-6R. Dialysate IL-6 correlated with dialysate concentrations of CA125 MCP-1, VEGF and Ang-2. CONCLUSION Our study from incident PD patients suggested that (i) dialysate the IL-6 system is a potent determinant of baseline PSTR and (ii) elevation of IL-6 in the dialysate is associated with up-regulation of intra-peritoneal inflammatory and angiogenic molecules.