Crosstalk between oxygen signaling and iron metabolism in renal interstitial fibroblasts.

To maintain the oxygen supply, the production of red blood cells (erythrocytes) is promoted under low-oxygen conditions (hypoxia). Oxygen is carried by hemoglobin in erythrocytes, in which the majority of the essential element iron in the body is contained. Because iron metabolism is strictly controlled in a semi-closed recycling system to protect cells from oxidative stress caused by iron, hypoxia-inducible erythropoiesis is closely coordinated by regulatory systems that mobilize stored iron for hemoglobin synthesis. The erythroid growth factor erythropoietin (EPO) is mainly secreted by interstitial fibroblasts in the renal cortex, which are known as renal EPO-producing (REP) cells, and promotes erythropoiesis and iron mobilization. Intriguingly, EPO production is strongly induced by hypoxia through iron-dependent pathways in REP cells. Here, we summarize recent studies on the network mechanisms linking hypoxia-inducible EPO production, erythropoiesis and iron metabolism. Additionally, we introduce disease mechanisms related to disorders in the network mediated by REP cell functions. Furthermore, we propose future studies regarding the application of renal cells derived from the urine of kidney disease patients to investigate the molecular pathology of chronic kidney disease and develop precise and personalized medicine for kidney disease.

Ruxolitinib treatment of a patient with steroid-dependent severe autoimmunity due to STAT1 gain-of-function mutation.

Signal transducer and activator of transcription 1 gain-of-function (STAT1 GOF) mutations are the most common cause of chronic mucocutaneous candidiasis (CMC). We report the effect of oral ruxolitinib, an inhibitor of Janus kinase (JAK) family tyrosine kinases, on the clinical and immune status of a 3-year-old male with steroid-dependent severe autoimmunity due to a STAT1 GOF T385M mutation. The patient's susceptibility to infection improved with antimicrobial prophylaxis and immunoglobulin replacement therapy, but he continued to exhibit severely disabling symptoms of autoimmunity. More than one-third of patients with STAT1 GOF mutations present with autoimmune manifestations, and this patient's mutation was reported to cause CMC with autoimmunity. We analyzed the interleukin (IL)-17A and IFN-γ levels and immunophenotype by flow cytometry before and during treatment with ruxolitinib. The peripheral IL-17A level did not increase, but the IFN-γ level decreased after 4 months of therapy. The STAT1 phosphorylation level decreased significantly upon stimulation of patient cells with IFN-γ. Clinically, cytomegalovirus reactivation occurred, but was controlled. No other adverse effect was noted. We report the potential of JAK1/2 inhibition with ruxolitinib for both CMC and steroid-dependent autoimmunity. However, long-term administration is necessary, as the effect is not sustained after treatment is discontinued.

Application of Muse Cell Therapy for Kidney Diseases.

The kidney plays an essential role in the maintenance of homeostasis in healthy individuals, e.g., by regulating the amount of water and concentration of electrolyte in the body. Owing to the structural complexity, renal dysfunction is caused by a myriad of diseases and conditions, and in severe cases, it progresses to end-stage renal disease in which patients require renal replacement therapy, i.e., maintenance dialysis or kidney transplantation. The currently available therapeutic modalities, with the exception of renal transplantation, cannot recover severely deteriorated renal function. Thus, regenerative medicine holds considerable promise as a potential means for developing next-generation renal therapeutics. Mesenchymal stem cell (MSC) transplantation has been investigated in acute kidney injury and chronic kidney disease models, and clinical studies have already been started for some kinds of kidney diseases. However, most of these studies concluded that the main underlying mechanism of therapeutic effect of MSC transplantation was paracrine. Recently, we reported that Muse cell therapy in a murine model of chronic kidney disease resulted in differentiation of intravenously injected Muse cells into glomerular cells after preferential homing to damaged glomerulus and improvement in renal function. The result suggested the potentiality of Muse cell therapy for glomerular regeneration. Muse cells are a promising cell source for regenerative therapy for kidney diseases.

Beneficial Effects of Systemically Administered Human Muse Cells in Adriamycin Nephropathy.

Multilineage-differentiating stress-enduring (Muse) cells are nontumorigenic endogenous pluripotent-like stem cells that can be collected from various organs. Intravenously administered Muse cells have been shown to spontaneously migrate to damaged tissue and replenish lost cells, but the effect in FSGS is unknown. We systemically administered human bone marrow-derived Muse cells without concurrent administration of immunosuppressants to severe combined immune-deficient (SCID) and BALB/c mouse models with adriamycin-induced FSGS (FSGS-SCID and FSGS-BALB/c, respectively). In FSGS-SCID mice, human Muse cells preferentially integrated into the damaged glomeruli and spontaneously differentiated into cells expressing markers of podocytes (podocin; 31%), mesangial cells (megsin; 13%), and endothelial cells (CD31; 41%) without fusing to the host cells; attenuated glomerular sclerosis and interstitial fibrosis; and induced the recovery of creatinine clearance at 7 weeks. Human Muse cells induced similar effects in FSGS-BALB/c mice at 5 weeks, despite xenotransplant without concurrent immunosuppressant administration, and led to improvement in urine protein, creatinine clearance, and plasma creatinine levels more impressive than that in the FSGS-SCID mice at 5 weeks. However, functional recovery in FSGS-BALB/c mice was impaired at 7 weeks due to immunorejection, suggesting the importance of Muse cell survival as glomerular cells in the FSGS kidney for tissue repair and functional recovery. In conclusion, Muse cells are unique reparative stem cells that preferentially home to damaged glomeruli and spontaneously differentiate into glomerular cells after systemic administration. Introduction of genes to induce differentiation is not required before Muse cell administration; thus, Muse cells may be a feasible therapeutic strategy in FSGS.

Early RAAS Blockade Exerts Renoprotective Effects in Autosomal Recessive Alport Syndrome.

Alport syndrome is a progressive renal disease caused by mutations in COL4A3, COL4A4, and COL4A5 genes that encode collagen type IV alpha 3, alpha 4, and alpha 5 chains, respectively. Because of abnormal collagen chain, glomerular basement membrane becomes fragile and most of the patients progress to end-stage renal disease in early adulthood. COL4A5 mutation causes X-linked form of Alport syndrome, and two mutations in either COL4A3 or COL4A4 causes an autosomal recessive Alport syndrome. Recently, renin-angiotensin-aldosterone system (RAAS) blockade has been shown to attenuate effectively disease progression in Alport syndrome. Here we present three Japanese siblings and their father all diagnosed with autosomal recessive Alport syndrome and with different clinical courses, suggesting the importance of the early initiation of RAAS blockade. The father was diagnosed with Alport syndrome. His consanguineous parents and his wife were healthy. All three siblings showed hematuria since infancy. Genetic analysis revealed that they shared the same gene mutations in COL4A3 in a compound heterozygous state: c.2330G>A (p.Gly777Ala) from the mother and c.4354A>T (p.Ser1452Cys) from the father. Although RAAS blockade was initiated for the older sister and brother when their renal function was already impaired, it did not attenuate disease progression. In the youngest brother, RAAS blockade was initiated during normal renal function stage. After the initiation, his renal function has been normal with the very mild proteinuria to date at the age of 17 years. We propose that in Alport syndrome, RAAS blockade should be initiated earlier than renal function is impaired.

Two novel mutations in the lactase gene in a Japanese infant with congenital lactase deficiency.

Intestinal lactase is required for the hydrolysis of lactose that is the most essential carbohydrate in milk and the primary diet source of newborn. Congenital lactase deficiency [CLD (MIM 223000)] is a severe gastrointestinal disorder and is characterized by watery diarrhea due to an extremely low or the lack of lactase activity in the intestinal wall from birth. CLD is a rare disease and occurs more frequently in Finland. Recent studies have shown that mutations in the coding region of the lactase (LCT) gene underlie CLD in patients from Finland and other European countries. Here, we report two novel mutations in the LCT gene in a Japanese female infant with clinical features consistent with those of CLD. She suffered from severe watery diarrhea from the age of 2 days on breast milk/lactose containing cow's milk formula. With the lactose-free hydrolyzed cow's milk formula, diarrhea was stopped, and she has now developed well on a lactose-free diet. She shows a lactose-intolerance pattern on the lactose challenge test. Sequence analysis revealed the two mutations in her LCT gene: c.4419C>G (p.Y1473X) in exon 10 transmitted from her mother and c.5387delA (p.D1796fs) in exon 16 transmitted from her father. Both mutations cause premature truncation of lactase polypeptide and are supposed to be responsible for CLD. To our knowledge, this is the first report on mutations in the LCT gene in Japan. We suggest that an increased awareness is required regarding CLD.

Expression of a biologically active recombinant follicle stimulating hormone of Japanese Eel Anguilla japonica using methylotropic yeast, Pichia pastoris.

In the Japanese eel Anguilla japonica, the administration of exogenous GTH is necessary for the artificial induction and completion of gonadal maturation due to its GTH deficiency under captive conditions. The isolation of native eel GTH has not been accomplished, which has made it difficult to fully elucidate the biological functioning of the two GTHs (FSH and LH) in eel. In this study, we attempted to produce a recombinant Japanese eel GTH (rjeFSH) having biological activity using methylotrophic yeast, Pichia pastoris in order to gain more understanding of the functioning of GTH in this species. An expression vector in which jeFSHbeta and GTHalpha subunit cDNAs were tandemly connected was constructed. P. pastoris was transformed with the vector, and rjeFSH was expressed. The rjeFSH thus expressed was detected by Western blot analysis. The glycoprotein fraction of the yeast culture supernatant was separated by native PAGE, and a band showed positive reaction with anti-GTHalpha and FSHbeta antisera similarly, suggesting that both subunits are associated. After deglycosylation, both subunits were decreased in molecular mass, indicating that rjeFSH was glycosylated. In in vitro assay, rjeFSH stimulated the release of testosterone and 11-ketotestosterone from immature eel testis, whereas release was not stimulated in maturing eel testis. This is the first report investigating the biological activity of eel GTH using the recombinant eel FSH.