Dynamic landscape of immune cell-specific gene regulation in immune-mediated diseases.

Genetic studies have revealed many variant loci that are associated with immune-mediated diseases. To elucidate the disease pathogenesis, it is essential to understand the function of these variants, especially under disease-associated conditions. Here, we performed a large-scale immune cell gene-expression analysis, together with whole-genome sequence analysis. Our dataset consists of 28 distinct immune cell subsets from 337 patients diagnosed with 10 categories of immune-mediated diseases and 79 healthy volunteers. Our dataset captured distinctive gene-expression profiles across immune cell types and diseases. Expression quantitative trait loci (eQTL) analysis revealed dynamic variations of eQTL effects in the context of immunological conditions, as well as cell types. These cell-type-specific and context-dependent eQTLs showed significant enrichment in immune disease-associated genetic variants, and they implicated the disease-relevant cell types, genes, and environment. This atlas deepens our understanding of the immunogenetic functions of disease-associated variants under in vivo disease conditions.

Jacob-induced transcriptional inactivation of CREB promotes Aß-induced synapse loss in Alzheimer's disease.

Synaptic dysfunction caused by soluble ß-amyloid peptide (Aß) is a hallmark of early-stage Alzheimer's disease (AD), and is tightly linked to cognitive decline. By yet unknown mechanisms, Aß suppresses the transcriptional activity of cAMP-responsive element-binding protein (CREB), a master regulator of cell survival and plasticity-related gene expression. Here, we report that Aß elicits nucleocytoplasmic trafficking of Jacob, a protein that connects a NMDA-receptor-derived signalosome to CREB, in AD patient brains and mouse hippocampal neurons. Aß-regulated trafficking of Jacob induces transcriptional inactivation of CREB leading to impairment and loss of synapses in mouse models of AD. The small chemical compound Nitarsone selectively hinders the assembly of a Jacob/LIM-only 4 (LMO4)/ Protein phosphatase 1 (PP1) signalosome and thereby restores CREB transcriptional activity. Nitarsone prevents impairment of synaptic plasticity as well as cognitive decline in mouse models of AD. Collectively, the data suggest targeting Jacob protein-induced CREB shutoff as a therapeutic avenue against early synaptic dysfunction in AD.

4-META/MMA-TBB resin containing nano hydroxyapatite induces the healing of periodontal tissue repair in perforations at the pulp chamber floor.

We aimed to evaluate the materials based on 4-methacryloxyethyl trimellitate anhydride/methyl methacrylate tri-n-butylborane (Super-bond [SB]) and nano hydroxyapatite (naHAp) for the repair of perforation at pulp chamber floor (PPF) in vitro and in vivo models. SB and naHAp were mixed in the mass ratio of 10% or 30% to produce naHAp/SB. Human periodontal ligament stem cells (HPDLSCs) were cultured on resin discs of SB or naHAp/SB to analyze the effects of naHAp/SB on cell adhesion, proliferation, and cementoblastic differentiation. A rat PPF model was treated with SB or naHAp/SB to examine the effects of naHAp/SB on the healing of defected cementum and periodontal ligament (PDL) at the site of PPF. HPDLSCs were spindle-shaped and adhered to all resin discs. Changing the resin from SB to naHAp/SB did not significantly alter cell proliferation. Both 10% and 30% naHAp/SB were more effective than SB in promoting cementoblastic differentiation of HPDLSCs. In the rat PPF model, 30% naHAp/SB was more effective than SB in promoting the formation Sharpey's fiber-like structures with expression of the PDL-related marker and cementum-like structures with expression of cementum-related markers. In conclusion, 30% naHAp/SB can be the new restorative material for PPF because it exhibited the abilities of adhering to dentin and healing of defected periodontal tissue.

The emerging threat of methicillin-resistant Staphylococcus aureus (MRSA) clone ST22-PT, carrying both Panton-Valentine leucocidin and toxic shock syndrome toxin 1 genes.

OBJECTIVES: Some MRSA strains produce Panton-Valentine leucocidin (PVL) and/or toxic shock syndrome toxin 1 (TSST-1), which are associated with severe infectious diseases. Although PVL- or TSST-1-positive strains have been isolated worldwide, strains carrying both PVL and TSST-1 genes are rare and sporadic. The objective of this study was to characterize these strains from Japan. METHODS: A total of 6433 MRSA strains isolated in Japan between 2015 and 2021 were analysed. Molecular epidemiological and comparative genomic analyses were conducted on PVL- and TSST-1-positive MRSA strains. RESULTS: A total of 26 strains from 12 healthcare facilities were PVL positive and TSST-1 positive, and all were classified as clonal complex (CC) 22. These strains exhibited similar genetic features to each other and were named as ST22-PT according to a previous report. Twelve and one of the ST22-PT strains were identified in patients with deep-seated skin infections and toxic shock syndrome-like symptoms, which are typical clinical features of PVL-positive and TSST-1-positive Staphylococcus aureus, respectively. Whole-genome comparative analysis revealed that the ST22-PT strains were highly similar to PVL- and TSST-1-positive CC22 strains isolated in several countries. Evaluation of the genome structure showed that ST22-PT possessed ΦSa2 harbouring PVL genes and a unique S. aureus pathogenicity island harbouring the TSST-1 gene. CONCLUSIONS: ST22-PT strains have recently emerged from several healthcare facilities in Japan, and ST22-PT-like strains have been identified in several countries. Our report highlights that the risk of international spread of PVL- and TSST-1-positive MRSA clone ST22-PT needs to be further investigated.

Extracellular aaRSs drive autoimmune and inflammatory responses in rheumatoid arthritis via the release of cytokines and PAD4.

OBJECTIVES: Recent studies demonstrate that extracellular-released aminoacyl-tRNA synthetases (aaRSs) play unique roles in immune responses and diseases. This study aimed to understand the role of extracellular aaRSs in the pathogenesis of rheumatoid arthritis (RA). METHODS: Primary macrophages and fibroblast-like synoviocytes were cultured with aaRSs. aaRS-induced cytokine production including IL-6 and TNF-α was detected by ELISA. Transcriptomic features of aaRS-stimulated macrophages were examined using RNA-sequencing. Serum and synovial fluid (SF) aaRS levels in patients with RA were assessed using ELISA. Peptidyl arginine deiminase (PAD) 4 release from macrophages stimulated with aaRSs was detected by ELISA. Citrullination of aaRSs by themselves was examined by immunoprecipitation and western blotting. Furthermore, aaRS inhibitory peptides were used for inhibition of arthritis in two mouse RA models, collagen-induced arthritis and collagen antibody-induced arthritis. RESULTS: All 20 aaRSs functioned as alarmin; they induced pro-inflammatory cytokines through the CD14-MD2-TLR4 axis. Stimulation of macrophages with aaRSs displayed persistent innate inflammatory responses. Serum and SF levels of many aaRSs increased in patients with RA compared with control subjects. Furthermore, aaRSs released PAD4 from living macrophages, leading to their citrullination. We demonstrate that aaRS inhibitory peptides suppress cytokine production and PAD4 release by aaRSs and alleviate arthritic symptoms in a mouse RA model. CONCLUSIONS: Our findings uncovered the significant role of aaRSs as a novel alarmin in RA pathogenesis, indicating that their blocking agents are potent antirheumatic drugs.

Depressive symptoms predict antibody titers after a second dose of the SARS-CoV-2 BNT162b2 vaccine among hospital workers in Japan.

BACKGROUND: Although factors associated with the antibody response to the BNT162b2 mRNA COVID-19 vaccine have been reported, psychological factors have not been examined. Depression or anxiety may affect vaccine reactions because these factors influence immune responses. This study aimed to determine whether psychological status at the time of vaccination predicts antibody responses. METHODS: A prospective observational study of the BNT162b2 mRNA COVID-19 vaccine response was carried out among individuals attending for an annual health check-up. Participants included 78 volunteers out of 80 hospital workers in Nagoya, Japan. No participants had been infected with COVID-19 and all gave written informed consent to participate in the study. Blood samples were obtained approximately 28 days after the second dose of the vaccine, and antibody titers of the SARS-CoV-2 spike protein were determined using the SARS-CoV-2 IgG II Quant assay. Participants completed the Japanese version of the hospital anxiety and depression scale (HADS) questionnaire, one day before both vaccinations. Participants also recorded any adverse reactions, such as body temperature and other side effects, every day for two weeks after each dose. The relationships between antibody titers and the predictive factors were analyzed using multiple linear regression analysis, with the antibody titers as the dependent variables, followed by univariate analysis. RESULTS: Multiple linear regression analysis revealed that no or excessive alcohol intake (p = 0.039), poor results from a health check-up (p = 0.011), a longer duration between the second dose and blood collection (p = 0.039), and increased degree of depressive symptoms (p = 0.041) were significant negative predictors of antibody titers, while body temperature one day after the second dose as a significant positive predictor of antibody titers (p < 0.0005). CONCLUSION: We identified that depressive symptoms just before the second dose of the BNT162b2 mRNA COVID-19 were an independent negative predictor of antibody responses, in addition to other factors. Our results highlight the importance of mental health at the time of vaccination to achieve the higher antibody responses necessary to acquire humoral immunity.

Appropriate Surgical Timing of Salter Innominate Osteotomy for Residual Acetabular Dysplasia in Children.

BACKGROUND: Salter innominate osteotomy (SIO) provides favorable results for treating residual acetabular dysplasia in young children. In this study, we examined the midterm results of SIO according to the age at surgery to determine the optimal timing of this procedure. METHODS: We retrospectively examined 50 hips of 42 patients (8 boys and 34 girls) with acetabular dysplasia who underwent SIO and were followed up until skeletal maturity. The center-edge angle (CEA) was measured based on the anteroposterior radiographs of the hip obtained before surgery, 5 weeks after surgery, and at the latest follow-up. Severin classification was evaluated at the latest follow-up. Patients were categorized into 3 groups according to age at surgery: younger than 7 years of age (group A), 7 to 8 years of age (group B), and 9 years of age or older (group C). RESULTS: The mean preoperative CEA level of 0.9 degrees improved to 17.1 degrees postoperatively, which was increased to 28.1 degrees at the latest examination. Overall, 45 hips (90%) were classified as Severin I or II, with 96% in group A, 94% in group B, and 57% in group C. In group C, postoperative acetabular coverage was similar to that in the other groups (16.6 degrees in group A, 14.8 degrees in group B, and 18.1 degrees in group C), although the final outcome was unsatisfactory. The average improvement in CEA from postoperative to skeletal maturity was significantly smaller in group C than in the other groups (12.7 degrees in group A, 11.3 degrees in group B, and 3.0 degrees in group C). CONCLUSIONS: SIO showed favorable outcomes with satisfactory acetabular coverage at skeletal maturity. However, satisfactory acetabular coverage could not be obtained in some older patients because of limited postoperative remodeling capacity and smaller secondary improvement of CEA. We recommend that SIO should be performed in patients aged 8 years or younger. LEVEL OF EVIDENCE: Level III-retrospective comparative study.

Decorin Promotes Osteoblastic Differentiation of Human Periodontal Ligament Stem Cells.

The aim of this study is to clarify the biological functions of decorin (DCN) in the healing and regeneration of wounded periodontal tissue. We investigated the expression pattern of DCN during the healing of wounded periodontal tissue in rats by immunohistochemistry and the effects of DCN on the osteoblastic differentiation of human periodontal ligament (PDL) stem cells (HPDLSCs) and preosteoblasts by Alizarin red S staining, quantitative reverse transcription-polymerase chain reactions, and western blotting. The expression of DCN was increased around the wounded PDL tissue on day 5 after surgery compared with the nonwounded PDL tissue, whereas its expression was not changed in the osteoblastic layer around the wounded alveolar bone. Furthermore, DCN promoted the osteoblastic differentiation of HPDLSCs, but it did not affect the osteoblastic differentiation of preosteoblasts. ERK1/2 phosphorylation was upregulated during the DCN-induced osteoblastic differentiation of HPDLSCs. DCN did not affect proliferation, migration, or the PDL-related gene expression of HPDLSCs. In conclusion, this study demonstrates that DCN has a role in the healing of wounded periodontal tissue. Furthermore, DCN secreted from PDL cells may contribute to bone healing by upregulating osteoblastic differentiation through ERK1/2 signaling in HPDLSCs, indicating a therapeutic effect of DCN in periodontal tissue regeneration.

Acute exacerbation of rheumatoid arthritis-associated interstitial lung disease: clinical features and prognosis.

OBJECTIVES: RA-associated interstitial lung disease (RA-ILD) is commonly associated with acute exacerbations (ILD-AE). This study examined the clinical characteristics and risk factors of ILD-AE and mortality of RA-ILD. METHODS: We retrospectively collected data on 165 RA-ILD patients who visited or were admitted to our hospital between January 2007 and December 2019. We compared the clinical characteristics of patients who did and did not develop ILD-AE and identified variables significantly associated with ILD-AE. We also compared the admission characteristics of those who survived and those who died after admission for ILD-AE. ILD-AE was defined using previously proposed criteria, modified slightly for application to RA-ILD. RESULTS: The mean patient age was 73.6 years (s.d. 9.7) and 97 (71.9%) patients were female. Thirty (22.2%) patients developed ILD-AE, 13 (43.3%) of whom died. In univariate analyses, neither the usual interstitial pneumonia (UIP) pattern nor MTX was associated with ILD-AE. In multivariate analyses, the UIP pattern was significantly associated with ILD-AE [odds ratio (OR) 2.55 (95% CI 1.05, 6.20), P = 0.038]. In the Cox proportional hazards model, the UIP pattern [hazard ratio (HR) 4.67 (95% CI 1.02, 21.45), P = 0.048] was significantly associated with death, while MTX use [HR 0.16 (95% CI 0.04, 0.72), P = 0.016] was significantly associated with survival. CONCLUSION: Our data suggest that the UIP pattern is related to ILD-AE. Furthermore, both the UIP pattern and non-use of MTX might be related to death from ILD-AE in RA-ILD.

Adverse drug reactions to trimethoprim-sulfamethoxazole in systemic lupus erythematosus.

OBJECTIVES: Trimethoprim-sulfamethoxazole (TMP-SMX), a prophylactic agent against pneumocystis pneumonia (PCP), can cause adverse drug reactions (ADRs), particularly in patients with systemic lupus erythematosus (SLE). However, the risk factors for ADRs remain unclear. Thus, we sought to examine the prevalence of TMP-SMX-related ADRs in patients with SLE and identify specific risk factors for ADR development in these patients. METHODS: We retrospectively reviewed data from patients with connective tissue disease (CTD) who were administered TMP-SMX as a PCP prophylactic. The prevalence of ADRs was compared between patients with SLE and those with other CTDs. Univariate and multivariate analyses were conducted to identify risk factors for ADRs in patients with SLE. RESULTS: Of the 424 patients with CTD included in our study (SLE, n = 162; other CTDs, n = 262), 22 with SLE (13.6%) developed ADRs, and this rate was significantly higher than that observed in patients with non-SLE CTDs (n = 18 [6.9%], p = 0.033). In patients with SLE, univariate analyses revealed direct associations of ADRs with anti-Sm (p < 0.001), anti-RNP (p = 0.02), and anti-Ro/SS-A antibodies (p = 0.042). Multivariate analysis identified a significant association between anti-Sm antibody levels and the development of ADRs (adjusted odds ratio 5.27, 95% confidence interval 1.80-15.40, p = 0.002). CONCLUSIONS: Patients with SLE who are prophylactically administered TMP-SMX are at high risk of ADRs. Among these patients, those who display a positive anti-Sm antibody should be carefully monitored for ADRs.

A novel community-acquired MRSA clone, USA300-LV/J, uniquely evolved in Japan.

BACKGROUND: USA300 [ST8-staphylococcal cassette chromosome mec type IVa (ST8-IVa)/arginine catabolic mobile element (ACME) positive] is a major Panton-Valentine leucocidin (PVL)-positive community-acquired MRSA (CA-MRSA) clone. In Japan, we identified USA300-like strains with characteristics (ST8-IVc/ACME negative) similar to those of USA300. OBJECTIVES: To reveal the evolution of the USA300-like strains. METHODS: The whole-genome sequence of a USA300-like strain was determined and genome analysis was performed using Type Strain Genome Server, MUSCLE and progressiveMauve. RESULTS: Genome-based phylogenetic analysis showed that the USA300-like strain is more similar to the USA300-Latin American variant (USA300-LV), which is a PVL-positive CA-MRSA clone identified in South America, than to USA300. Instead of the ACME, copper and mercury resistance mobile elements were located on the genome of the USA300-like strain. In addition, the USA300-like strain possessed a unique mobile genetic element, ICE6013. Therefore, we named this novel USA300-LV variant identified in Japan as USA300-LV/J. CONCLUSIONS: Our findings strongly suggest that a PVL-positive CA-MRSA USA300-LV/J clone originating from abroad has uniquely evolved and disseminated in Japan.

Fluorodeoxyglucose PET/CT of Arthritis in Rheumatic Diseases: A Pictorial Review.

Rheumatic diseases are various painful conditions that affect joints, bones, cartilage, tendons, ligaments, and muscles. Arthritis is a typical condition of rheumatic disease. Although rheumatoid arthritis is a representative rheumatic disease, various diseases other than rheumatoid arthritis can also affect joints, and differential diagnosis of rheumatic diseases is often difficult owing to the similar clinical manifestations. However, accurate diagnosis is crucial for an appropriate treatment strategy. The utility of fluorine 18 fluorodeoxyglucose (FDG) PET/CT has been established, and it is widely used for assessing malignancies. In addition to accumulating in tumor cells, FDG also accumulates in inflammatory tissue, allowing FDG PET/CT to demonstrate arthritis. PET/CT allows evaluation of whole-body articular and extra-articular lesions in one examination, representing a key advantage over US and MRI, which allow assessment of only a few regions because of their limited field of view. Although FDG PET/CT is sensitive for detecting inflammatory lesions, the uptake itself is nonspecific; therefore, knowledge of characteristic uptake patterns is necessary to narrow the differential diagnosis in rheumatic disease. Furthermore, pathognomonic extra-articular findings such as vasculitis, skin lesions, lymphadenopathy, and chondritis play an important role in achieving accurate diagnosis. The authors present the FDG PET/CT appearances of (a) rheumatoid arthritis and allied disorders (polymyalgia rheumatica, remitting seronegative symmetrical synovitis with pitting edema, adult-onset Still disease), (b) spondyloarthritis (ankylosing spondylitis, psoriatic arthritis, reactive arthritis, inflammatory bowel disease arthritis, SAPHO [synovitis, acne, pustulosis, hyperostosis, and osteitis] syndrome, chronic recurrent multifocal osteomyelitis), and (c) miscellaneous systemic disorders with arthropathy (relapsing polychondritis, multicentric reticulohistiocytosis, amyloidosis, sarcoidosis, hemophilia). ©RSNA, 2020.

Sagittal plane deformity after temporary epiphysiodesis of the distal femur for correcting limb length discrepancy.

BACKGROUND: Outcome of the temporary epiphysiodesis procedure for limb length discrepancy (LLD) is commonly evaluated in the coronal plane. The purpose of this study was to investigate implant position and complications of the distal femur and patella in the sagittal plane after the temporary epiphysiodesis. METHODS: We retrospectively reviewed 27 patients with LLD who underwent temporary epiphysiodesis of the distal femur using staples (11 patients) or eight-plates (16 patients) between 2007 and 2015. The mean age was 9.7 years (range, 6.3-13.8) at the time of epiphysiodesis. The implants were removed after a mean period of 2.6 years (range, 0.8-4.8) from the epiphysiodesis. Correction amount of LLD was measured on anteroposterior long leg standing radiographs. Implant position, extension deformity of the distal femur (>5° from epiphysiodesis to removal of implant) and patella baja (the epiphyseal line midpoint method < 1.0) were evaluated using lateral knee radiographs. RESULTS: The average correction amount of LLD was 17.4 mm (range, 2-34). The average implant position was 43.1% (range, 35-55) from the anterior edge of the distal femoral epiphysis. At removal surgery, 16 patients (59%) had extension deformity of the distal femur and 14 patients (52%) showed patella baja. There were significant correlations between implant position and extension deformity (r = -0.51, p < 0.01) and as well as between correction amount of LLD and patella baja (r = -0.64, p < 0.01). CONCLUSION: After temporary epiphysiodesis for the treatment of LLD, extension deformity of the distal femur and patella baja occurred frequently. Anterior placement of the implants is associated with extension deformity of the distal femur. The implant should be placed in the center of distal femoral physis, not the center of femoral shaft. Excessive correction of LLD should be avoided due to a risk of patella baja.

m6A modification-mediated CBX8 induction regulates stemness and chemosensitivity of colon cancer via upregulation of LGR5.

BACKGROUND: Colon cancer (CC) cells can exhibit stemness and expansion capabilities, which contribute to resistance to conventional chemotherapies. Aberrant expression of CBX8 has been identified in many types of cancer, but the cause of this aberrant CBX8 expression and whether CBX8 is associated with stemness properties in CC remain unknown. METHODS: qRT-PCR and IHC were applied to examine CBX8 levels in normal and chemoresistant CC tissues. Cancer cell stemness and chemosensitivity were evaluated by spheroid formation, colony formation, Western blot and flow cytometry assays. RNA-seq combined with ChIP-seq was used to identify target genes, and ChIP, IP and dual luciferase reporter assays were applied to explore the underlying mechanisms. RESULTS: CBX8 was significantly overexpressed in chemoresistant CC tissues. In addition, CBX8 could promote stemness and suppress chemosensitivity through LGR5. Mechanistic studies revealed that CBX8 activate the transcription of LGR5 in a noncanonical manner with assistance of Pol II. CBX8 recruited KMT2b to the LGR5 promoter, which maintained H3K4me3 status to promote LGR5 expression. Moreover, m6A methylation participated in the upregulation of CBX8 by maintaining CBX8 mRNA stability. CONCLUSIONS: Upon m6A methylation-induced upregulation, CBX8 interacts with KMT2b and Pol II to promote LGR5 expression in a noncanonical manner, which contributes to increased cancer stemness and decreased chemosensitivity in CC. This study provides potential new therapeutic targets and valuable prognostic markers for CC.

Classifications of scleroderma renal crisis and reconsideration of its pathophysiology.

Categorization of scleroderma renal crisis (SRC) as hypertensive or normotensive can potentially overlook the underlying pathophysiology that might be unique in each patient, as they often exhibit a mixture of distinct pathological characteristics of narrowly defined SRC (nd-SRC) and systemic sclerosis associated thrombocytic micro-angiopathy (SSc-TMA). In this review, we provide evidence suggesting that better categorization of patients presenting with certain clinical features of both nd-SRC and TMA will improve treatment approaches. Based on our clinical experience and literature review, distinguishing between nd-SRC and SSc-TMA is important because the association of SSc-TMA with prior steroid administration and poor prognosis was stronger than that of nd-SRC. Although the two pathological entities cannot be easily distinguished based on blood pressure, we suggest that the detailed clinical course is helpful. Typically, nd-SRC exhibits prominently elevated blood pressure and worsening of renal function initially, followed by mild thrombocytopenia. Conversely, SSc-TMA presents first with severe thrombocytopenia, followed by elevated blood pressure and renal function deterioration. The degree of involvement in each pathological condition should be considered for determination of appropriate therapeutic interventions and prognostic prediction.

Clinical characteristics and prognosis of polymyositis and dermatomyositis associated with malignancy: a 25-year retrospective study.

The objective of this study was to identify factors predictive of malignancy in patients with polymyositis (PM) and dermatomyositis (DM) in Japan. We conducted a retrospective study of PM and DM patients who were admitted to our hospital between January 1992 and September 2017. Among 134 patients, 29 (21.6%) were diagnosed with cancer in the 3 years prior to and 3 years after the initial diagnosis of PM or DM. According to multivariate analyses, male sex [odds ratio (OR) = 3.65, p = 0.03], old age (OR = 1.05, p = 0.02), and a past history of diabetes mellitus (OR = 10.4, p = 0.005) were associated with an increased risk of malignancy. The absence of interstitial lung disease (ILD) (OR = 0.25, p = 0.03) was also associated with an increased risk of malignancy. Diabetes mellitus was observed in 28.6% of PM and DM patients with malignancy, but in only 7.3% of those with malignancy. Survival was significantly lower in patients with malignancy than in those without malignancy (p < 0.001). Independent factors associated with malignancies in patients with PM or DM were male sex, old age, the absence of ILD, and, especially, a past history of diabetes mellitus.

Induction of erythropoietin gene expression in epithelial cells by chemicals identified in GATA inhibitor screenings.

Erythropoietin (EPO) is a hormone that promotes proliferation, differentiation and survival of erythroid progenitors. EPO gene expression is regulated in a tissue-specific and hypoxia-inducible manner and is mainly restricted to renal EPO-producing cells after birth. Chronic kidney disease (CKD) confers high risk for renal anemia due to lower EPO production from injured kidneys. In transgenic reporter lines of mice, disruption of a GATA-binding motif within the Epo gene promoter-proximal region restores constitutive reporter expression in epithelial cells. Here, mitoxantrone and its analogues, identified as GATA factor inhibitors through high-throughput chemical library screenings, markedly induce EPO/Epo gene expression in epithelium-derived cell lines and mice regardless of oxygen levels. In contrast, mitoxantrone interferes with hypoxia-induced EPO gene expression in Hep3B cells. Cryptic promoters are created for the EPO/Epo gene expression in epithelial cells upon mitoxantrone treatment, and consequently, unique 5'-untranslated regions are generated. The mitoxantrone-induced aberrant transcripts contribute to the reporter protein production in epithelial cells that carry the reporter gene in the proper reading frame of mouse Epo gene. Thus, EPO production in uninjured adult epithelial cells may be a therapeutic approach for renal anemia in patients with CKD.