Identification of ABCG2 as an Exporter of Uremic Toxin Indoxyl Sulfate in Mice and as a Crucial Factor Influencing CKD Progression.

Chronic kidney disease (CKD) patients accumulate uremic toxins in the body, potentially require dialysis, and can eventually develop cardiovascular disease. CKD incidence has increased worldwide, and preventing CKD progression is one of the most important goals in clinical treatment. In this study, we conducted a series of in vitro and in vivo experiments and employed a metabolomics approach to investigate CKD. Our results demonstrated that ATP-binding cassette transporter subfamily G member 2 (ABCG2) is a major transporter of the uremic toxin indoxyl sulfate. ABCG2 regulates the pathophysiological excretion of indoxyl sulfate and strongly affects CKD survival rates. Our study is the first to report ABCG2 as a physiological exporter of indoxyl sulfate and identify ABCG2 as a crucial factor influencing CKD progression, consistent with the observed association between ABCG2 function and age of dialysis onset in humans. The above findings provided valuable knowledge on the complex regulatory mechanisms that regulate the transport of uremic toxins in our body and serve as a basis for preventive and individualized treatment of CKD.

TBI and melphalan followed by allogeneic hematopoietic SCT in children with advanced hematological malignancies.

We evaluated the efficacy and safety of the conditioning regimen that consisted of TBI and melphalan (L-PAM), followed by hematopoietic SCT (HSCT) in 23 children with advanced hematological malignancies. The median age at HSCT was 9 (range, 2-15) years. The underlying diseases were ALL in 16 patients (5 in CR2, 3 in CR3, 6 in relapse (RP) and 2 in induction failure (IF)), AML in 4 patients (3 in RP and 1 in IF) and non-Hodgkin's lymphoma in 3 patients (1 in CR3, 1 in CR4 and 1 in RP). The stem cell sources were BM for 19 patients and cord blood for 4 patients. All patients received the conditioning regimen that consisted of TBI 12 or 13.2 Gy and L-PAM 210 mg/m(2). In all, 22 patients engrafted on the median of day 16 (range, 10-23). The regimen was well tolerated and common regimen-related toxicities (RRTs) included grade II stomatitis and grade I hepatic toxicity. The cumulative incidences of RP and TRM were 47.6 and 21.5%, respectively. At a median follow-up of 24.4 months, the probability of disease-free survival was 41.0%. The regimen may provide sufficient anti-leukemic effect without increased RRT for advanced pediatric hematological malignancies.

Unilateral expansion of a narrow mandibular dental arch combined with bimaxillary osteotomies in a patient with hypoglossia.

A 23-year-old female with hypoglossia, who had a narrow mandibular dental arch, was treated using the gradual expansion technique. Three lower incisors were missing and the right molar occlusion showed a scissor bite. Her speech was acceptable. Gradual unilateral expansion of the mandibular alveolar bone was performed. Orthodontic tooth alignment was performed prior to surgical treatment. A tooth-borne expander was devised using a hyrax-type screw to move the inclined right alveolar bone into an upright position. Alveolar bone osteotomies were performed under general anesthesia and the expander was placed in the mandibular dental arch. After a 5-day latency period, the screw was activated for 21 days. After expansion, the width of the mandibular dental arch increased by 10mm at the first molar region and the right molars were moved to an upright position. After a consolidation period of 7 days, simultaneous two-jaw surgery that combined Le Fort I osteotomy and intraoral vertical ramus osteotomies was performed to obtain a stable occlusion. After post-surgical orthodontic and prosthodontic treatment, her occlusion improved without deterioration of her speech. The results indicate that this technique is useful for unilateral expansion of distorted mandibular alveolar process.

Low-dose MTX for the treatment of acute and chronic graft-versus-host disease in children.

We report the results of a retrospective analysis in 27 pediatric patients who received low-dose MTX as the second-line treatment for steroid-refractory or -dependent acute and chronic GVHD. Between July 2000 and May 2006, 10 patients with aGVHD and 17 with cGVHD were treated with MTX at a dose of 3-10 mg/m(2) weekly. Seven of ten patients (70%) with aGVHD responded well to MTX, thus resulting in the achievement of either a complete response (CR) or a partial response (PR). The dose of prednisone could be reduced to equal to or lower than 1 mg/kg in the responding patients at the end of MTX therapy. The median number of MTX administrations was five (range, 1-7). Ten (58.8%) of seventeen patients with cGVHD achieved CR or PR. The dose of prednisone could be reduced to lower than 0.4 mg/kg in 16 of 17 patients and seven patients could discontinue prednisone. The median duration of MTX administration was 18 months (range, 1-68). The toxicities of grade III to IV occurred in only six patients presenting cytopenias or elevated levels of serum transaminases. Low-dose MTX was tolerable and effective for the steroid-refractory or -dependent GVHD in reducing the dose of steroid without increasing the risk of opportunistic infection.

Regimen-related toxicity following reduced-intensity stem-cell transplantation (RIST): comparison between Seattle criteria and National Cancer Center Common Toxicity Criteria (NCI-CTC) version 2.0.

Acute regimen-related toxicity (RRT) is minimal in reduced-intensity stem-cell transplantation (RIST). However, the Seattle RRT grading (Bearman et al), developed in the context of conventional-intensity transplantation, is frequently applied to RIST. We compared the National Cancer Institute Common Toxicity Criteria (NCI-CTC) version 2.0 with the Seattle criteria after RIST in 86 patients. RRT within 30 days of transplant graded by both sets of criteria were significantly associated with the outcome confirming the predictive value of both the systems. A total of 15 patients died of disease progression, and 12 of transplant-related mortality: RRT (n = 2), graft-versus-host disease (GVHD) (n = 7), infection (n = 1), and others (n = 2). GVHD-related deaths primarily resulted from infections after steroid treatment (n = 6) and bronchiolitis obliterans (n = 1). This study shows that NCI-CTC is appropriate in toxicity evaluation of RIST, and that its application to RIST enables a toxicity comparison between RIST and other types of cancer treatments. Since GVHD is a significant problem in RIST, modifications are required to evaluate immunological complications following RIST.

Reduced-intensity stem cell transplantation from an HLA-identical sibling donor in patients with myeloid malignancies.

The purpose of this study was to evaluate the feasibility and efficacy of allogeneic hematopoietic stem cell transplantation with a reduced-intensity regimen (RIST) in patients with acute myeloid leukemia (AML) or myelodysplastic syndrome (MDS). In all, 36 patients (median age 55 years) underwent RIST from an HLA-matched related donor between September 1999 and December 2002. The diagnoses included AML (n=14), leukemia evolving from MDS (n=10), and MDS (refractory anemia with excess blasts n=6, refractory anemia n=6). The RIST regimen consisted of purine analog (cladribine or fludarabine)/busulfan, with or without antithymocyte globulin. The regimen was well tolerated, and 34 patients achieved durable engraftment and most achieved remission after RIST. A total of 17 patients developed grade II-IV acute GVHD, and 27 developed chronic GVHD. Eight patients relapsed, and five of them received antithymocyte globulin (ATG) as part of the preparative regimen. A total of 12 patients died (four disease progression, six transplantation-related complications, and two others). Estimated 1-year disease-free survival (DFS) in low- and high-risk groups was 85 and 64%, respectively. We conclude that RIST can be performed safely in elderly patients with myeloid malignancies, and has therapeutic potential for those who fail conventional chemotherapy. In view of the significant association between GVHD or ATG and DFS, defined management of GVHD following RIST should become a major target of clinical research.

Induction of graft-versus-autoimmune (GVA) disease effect against refractory psoriasis by complete donor-type chimerism and graft-versus-host disease after allogeneic hematopoietic stem cell transplantation.

A 67-year-old man with AML, who had a 21-year history of psoriasis without remission, received a reduced-intensity transplantation from an HLA-identical sibling. The preparative regimen consisted of busulfan and fludarabine. Graft-versus-host-disease (GVHD) prophylaxis was cyclosporine and methotrexate. Psoriasis was completely resolved on day 18. The subsequent clinical course was uneventful until day 42, when psoriasis recurred at the same sites as before RIST. Peripheral blood examined on day 63 showed mixed chimerism with 54% recipient type. Cyclosporine was rapidly tapered off over the next 2 weeks. On day 90, 100% donor-type chimerism was confirmed. Subsequently, psoriasis improved simultaneously with the occurrence of mucositis and rash as a manifestation of GVHD. Scattered erythematous patches of psoriasis disappeared again by day 105. We initiated 0.5 mg/kg prednisolone on day 119, and resumed cyclosporine on day 133. At 7 months after RIST, he still suffers from chronic GVHD, but his psoriasis remains in remission for the first time in 21 years. The anti-psoriasis effect of the conditioning is mild and transient, while the graft-versus-autoimmunity effect, related to the induction of complete donor-type chimerism and GVHD, is more profound and persisting. A graft-versus-autoimmunity effect lies in the delicate balance between alloimmunity and immunosuppressant used for GVHD prophylaxis/treatment.

Menaquinol oxidase activity and primary structure of cytochrome bd from the amino-acid fermenting bacterium Corynebacterium glutamicum.

Cytochrome d was spectroscopically detected in membrane fractions of the amino-acid-fermenting, high-G+C gram-positive bacterium Corynebacterium glutamicum. Inhibition of NADH oxidase activity in the membranes by cyanide suggested that the main terminal respiratory oxidase during the stationary phase was a type of cytochrome bd. Cytochrome bd-type quinol oxidase, purified from the membranes, was composed of two subunits. Its reduced form showed absorption peaks at 627, 595, and 560 nm, which were due to haem d, high-spin protohaem, and low-spin protohaem, respectively. The air-oxidised form showed a peak at 645 nm, which might be due to oxygenated ferrous haem d. The spectral features and the size of subunit I are more similar to the properties of cytochromes bd from Proteobacteria, such as Escherichia coli, than to those of cytochrome bd from low-G+C gram-positive bacteria, such as Bacillus stearothermophilus. The menaquinol oxidase activity of the purified cytochrome bd was low, but was enhanced about fivefold by pre-incubating the enzyme with menaquinones. The order of effectiveness of quinols as oxidase substrates was clearly different from that of quinones as the activators of enzyme activity. Furthermore, activation was destroyed by ultraviolet irradiation of the pre-incubated enzyme and then restored by a second incubation with menaquinone. These results indicate that the enzymatic properties of this new oxidase are more similar to the properties of cytochromes bd from low-G+C gram-positive bacterial than to those of proteobacterial counterparts. They also suggest that the enzyme has a second quinone-binding site essential for full activity, in addition to the active centre for substrate oxidation. By using probes based on partial peptide sequences of the subunits, the genes for the two subunits of C. glutamicum cytochrome bd were cloned. The deduced amino acid sequence demonstrated that subunit I lacks the C-terminal half of the Q loop and that the primary structure of C. glutamicum cytochrome bd is more similar to that of other gram-positive bacteria than to proteobacterial cytochromes bd.

Calorimetric study of the effect of intrachain cross-linking on lysozyme unfolding.

Thermodynamics of unfolding of lysozyme cross-linked between Glu 35 and Trp 108 were studied in solutions of various concentrations of 1-propanol (1-PrOH) at pH 3.7 by means of scanning microcalorimetry. The transition temperature for the cross-linked lysozyme increases by 17-19 degrees C due to cross-linking at every concentration of 1-PrOH. This corresponds to the increase in the unfolding Gibbs free energy of about 28 kJ.mol-1, which is independent of the concentration of 1-PrOH. It was found that the unfolding enthalpy of cross-linked lysozyme is only slightly larger than that of intact one, and the unfolding entropy of the cross-linked one is nearly equal to that of the intact one, if both are compared at the same temperature. The stabilization mechanism for the cross-linked lysozyme is discussed on the basis of these calorimetric data.