A common variant of MAF/c-MAF, transcriptional factor gene in the kidney, is associated with gout susceptibility.

Gout is a multifactorial disease characterized by acute inflammatory arthritis, and it is caused as a consequence of hyperuricemia. A recent meta-analysis of genome-wide association studies has newly identified the relationship between serum uric acid (SUA) levels and rs889472, a single nucleotide polymorphism of musculoaponeurotic fibrosarcoma oncogene (MAF/c-MAF). However, it remained unclear whether rs889472 is associated with gout susceptibility. In the present study, we investigate the association between c-MAF rs889472 and gout in Japanese male population. We genotyped 625 male patients who were clinically diagnosed as gout and 1221 male control subjects without hyperuricemia or a history of gout by TaqMan method. As a result, the major allele (C), which reportedly increases SUA levels, had a higher frequency in the gout cases (58.8%) than in the controls (55.0%). A logistic regression analysis showed a significant association between rs889472 and gout (p = 0.029, odds ratio = 1.17; 95% confidence interval 1.02-1.34). C-MAF is reported as a pivotal transcriptional factor in the development and differentiation of renal proximal tubular cells. Because urate is mainly regulated in renal proximal tubular cells, c-MAF may have an important role in urate regulation in the kidney and influence not only SUA but also gout susceptibility. Our finding shows that rs889472 of c-MAF is associated with gout susceptibility.

Activation and increase of radio-sensitive CD11b+ recruited Kupffer cells/macrophages in diet-induced steatohepatitis in FGF5 deficient mice.

We have recently reported that Kupffer cells consist of two subsets, radio-resistant resident CD68+ Kupffer cells and radio-sensitive recruited CD11b+ Kupffer cells/macrophages (Mφs). Non-alcoholic steatohepatitis (NASH) is characterized not only by hepatic steatosis but also chronic inflammation and fibrosis. In the present study, we investigated the immunological mechanism of diet-induced steatohepatitis in fibroblast growth factor 5 (FGF5) deficient mice. After consumption of a high fat diet (HFD) for 8 weeks, FGF5 null mice developed severe steatohepatitis and fibrosis resembling human NASH. F4/80+ Mφs which were both CD11b and CD68 positive accumulated in the liver. The production of TNF and FasL indicated that they are the pivotal effectors in this hepatitis. The weak phagocytic activity and lack of CRIg mRNA suggested that they were recruited Mφs. Intermittent exposure to 1 Gy irradiation markedly decreased these Mφs and dramatically inhibited liver inflammation without attenuating steatosis. However, depletion of the resident subset by clodronate liposome (c-lipo) treatment increased the Mφs and tended to exacerbate disease progression. Recruited CD11b+ CD68+ Kupffer cells/Mφs may play an essential role in steatohepatitis and fibrosis in FGF5 null mice fed with a HFD. Recruitment and activation of bone marrow derived Mφs is the key factor to develop steatohepatitis from simple steatosis.

Histology and immunohistochemistry of the developing juxta-oral organ in mice.

The juxta-oral organ is a persistent bilateral organ in the mammalian bucca. It consists of epithelial parenchyma and surrounding mesenchymal sheath with connective tissue rich in nerve fibers and sensory receptors between them. The organ develops from the embryonic oral epithelium and is separated from the oral mucosa. The morphology of this organ is very different in mice compared to humans. Although several reports have described the histology of the juxta-oral organ in rodents, its developmental changes have rarely been reported. In the present study, histological development and ultrastructure of the juxta-oral organ, with special attention to its epithelial parenchyma and mesenchymal sheath, were investigated in mice during several developmental stages. Immunohistochemical staining for pan-cytokeratin (CK) on serial paraffin sections of the head allowed easier detection of this organ. The juxta-oral organ was innervated by the buccal nerve from an early stage of organogenesis at E13.5. The organ was located adjacent to the fascia of the masticatory muscles during all developmental stages examined. The parenchymal cells were immunohistochemically positive for p63 and CK14 in both newborn and adult mice, suggesting that these epithelial cells possess characteristics of the basal cells of the stratified epithelium. Furthermore, the parenchyma was ensheathed by layers of mesenchymal cells with perineurial characteristics as shown by immunohistochemical staining and electron microscopy. Unique sensory endings and bundles of elastic fibers were observed between the epithelial parenchyma and mesenchymal sheath. The present findings concerning the structure of mouse juxta-oral organ, especially that of epithelial parenchyma and mesenchymal sheath provide baseline knowledge for the further understanding of the unique morphological features, developmental changes and functional details of this organ.

Fibroblast growth factor-5 participates in the progression of hepatic fibrosis.

Non-alcoholic steatohepatitis (NASH) is characterized by the presence of steatosis, inflammation, and fibrosis and is believed to develop via a "two-hit process"; however, its pathophysiology remains unclear. Fibroblast growth factors (FGFs) are heparin-binding polypeptides with diverse biological activities in many developmental and metabolic processes. In particular, FGF5 is associated with high blood pressure. We investigated the function of FGF5 in vivo using spontaneously Fgf5 null mice and explored the role of diet in the development of NASH. Mice fed a high-fat diet gained little weight and had higher serum alanine transaminase, aspartate amino transferase, and non-high-density lipoprotein-cholesterol levels. Liver histology indicated marked inflammation, focal necrosis, fat deposition, and fibrosis, similar to the characteristics of NASH. FGF5 and a high-fat diet play significant roles in the pathophysiology of hepatic fibrosis and Fgf5 null mice may provide a suitable model for liver fibrosis or NASH.

Systemic endotoxin induces gene expression of inducible nitric oxide synthase in fetal rat brain.

BACKGROUND: Few studies have examined the response of the fetus under stress, such as with maternal infection. Recent work has indicated that nitric oxide (NO) modulates corticotropin-releasing hormone (CRH) secretion by the hypothalamus, but details of the action of NO on the fetus remain unclear. Therefore, we investigated the expression of inducible nitric oxide synthase (iNOS) mRNA and the response pattern following lipopolysaccharide (LPS) loading using a rat model of fetal infection. METHODS: Fetuses were delivered by cesarean section on day 20 of gestation and immediately placed in a chamber maintained at 37 degrees C and 100% relative humidity. The LPS group (n=12) was given 400 microg of LPS/100 g body weight, and the physiologic saline group (n=12) was given physiologic saline. Fetuses were then incubated for a further 3 hours. Fetuses were decapitated, the trunk blood was collected immediately after cesarean section or after 3 hours of incubation, and the fetal brains were fixed in formaldehyde and cryopreserved. Coronal cryosections of the brains were prepared, and a (35)S-uridine triphosphate-labeled antisense RNA probe for iNOS was then prepared. In situ hybridization was performed, and iNOS expression was evaluated semiquantitatively on the basis of optical density. In both groups, plasma corticosterone levels were determined with radioimmunoassay. RESULTS: Expression of iNOS mRNA was not noted in the physiologic saline group (3 hours postpartum). In the LPS group, iNOS mRNA expression was observed in the subfornical organ, but not in the paraventricular nucleus. Plasma corticosterone levels were significantly elevated in the LPS group. CONCLUSIONS: In 20-day-old rat fetuses, the hypothalamic-pituitary-adrenal axis was already mobilized in response to LPS-induced stress. These results suggest that iNOS is not involved in the acute response of the hypothalamic-pituitary-adrenal axis to LPS challenge in 20-day-old rat fetuses.

Organogenesis of the juxta-oral organ in mice.

The juxta-oral organ is a bilateral organ in the mammalian bucca. It consists of epithelial cords with surrounding mesenchyme. It develops from embryonic oral epithelium, but its macroscopic morphology in mice is less studied and seems to be very different from that of humans. The juxta-oral organ in mice extends more widely from the subcutaneous tissue of the mandible near the lateral fascia of the masseter to the submucosa of the soft palate. In this paper, we report that the mutant mouse allele Bmp7(lacZ) presented intense lacZ expression in the epithelial component of the juxta-oral organ in its homo- and heterozygous states. The main aims of this study were to show that this mutant mouse allele is suitable for observing macroscopic structure of the juxta-oral organ and to describe the development of this organ during embryonic and postnatal stages. Whole-mount beta-gal staining of this strain of mouse showed that the juxta-oral organ in mice appeared at E12.0 from oral epithelium and lost connection with it before E12.5. Then, the juxta-oral organ extended anteriorly to the lateral fascia of the masseter and posteriorly to the submucosal layer of the soft palate via the orbit. The mature juxta-oral organ had no connection to other epithelia such as those of the bucca and parotid duct. It persisted until adulthood and there seemed to be no tendency to regress. Transmission electron microscopy showed that each part of the juxta-oral organ was an epithelial cord surrounded by a basement membrane and mesenchymal tissue.

Neonatal exposure to sevoflurane induces abnormal social behaviors and deficits in fear conditioning in mice.

BACKGROUND: Neonatal exposure to anesthetics that block N-methyl-D-aspartate receptors and/or hyperactivate gamma-aminobutyric acid type A receptor has been shown to cause neuronal degeneration in the developing brain, leading to functional deficits later in adulthood. The authors investigated whether exposure of neonatal mice to inhaled sevoflurane causes deficits in social behavior as well as learning disabilities. METHODS: Six-day-old C57BL/6 mice were exposed to 3% sevoflurane for 6 h. Activated cleaved caspase-3 immunohistochemical staining was used for detection of apoptosis. Cognitive functions were tested by pavlovian conditioned fear test. Social behavior was tested by social recognition and interaction tests. RESULTS: Neonatal exposure to sevoflurane significantly increased the number of apoptotic cells in the brain immediately after anesthesia. It caused persistent learning deficits later in adulthood as evidenced by decreased freezing response in both contextual and cued fear conditioning. The social recognition test demonstrated that mice with neonatal exposure to sevoflurane did not develop social memory. Furthermore, these mice showed decreased interactions with a social target compared with controls in the social interaction test, indicating a social interaction deficit. The authors did not attribute these abnormalities in social behavior to impairments of general interest in novelty or olfactory sensation, because they did not detect significant differences in the test for novel inanimate object interaction or for olfaction. CONCLUSIONS: This study shows that exposure of neonatal mice to inhaled sevoflurane could cause not only learning deficits but also abnormal social behaviors resembling autism spectrum disorder.

Purification and characterization of mouse lacrimal gland epithelial cells and reconstruction of an acinarlike structure in three-dimensional culture.

PURPOSE: To clarify which factors are involved in postnatal lacrimal gland development, the expressions of several growth factors and their receptors were analyzed in purified lacrimal gland epithelial and mesenchymal cells. Reconstruction of acinarlike structures in three-dimensional culture conditions from this epithelial cell population was attempted. METHODS: Lacrimal gland epithelial and mesenchymal cells were isolated from newborn mice and purified using nylon mesh and collagenase. By real-time reverse transcription-polymerase chain reaction, immunocytochemistry, and Western blotting, the expressions of several growth factors and their receptors were analyzed. Responses of epithelial cells to the growth factors were analyzed by the addition of these factors to the culture medium. RESULTS: Fibroblast growth factor (FGF)10 and hepatocyte growth factor (HGF) were more intensely expressed in mesenchymal cells than in epithelial cells, and their receptors (FGFR2IIIb and c-Met, respectively) were less intensely expressed, whereas the expression of epidermal growth factor (EGF) and its receptor showed no significant difference between cell types. In the monolayer culture, cell viability was activated by the addition of EGF or HGF to epithelial cells, but no response was observed when FGF10 was added. The epithelial cells formed clusters with lumina when cultured on basement membrane matrix. These clusters contained secretion granules and showed positive immunostaining for aquaporin-5. CONCLUSIONS: EGF and HGF were considered to act in an autocrine/paracrine manner in and around the postnatal lacrimal gland, whereas epithelial cells did not respond to FGF10. It was suggested that certain extracellular matrix conditions accommodate these epithelial cells to reconstruct functional acinarlike structures.

Identification of ERK2-binding domain of EBITEIN1, a novel ERK2-binding protein.

We recently cloned a cDNA encoding a novel extracellular signal-regulated kinase 2 (ERK2) binding protein, EBITEIN1, by yeast two-hybrid screening. In this study, we further characterized EBITEIN1. Binding experiments using various deletion mutants identified a 40-amino acid minimal sequence for binding ERK2. Binding experiments using substitution mutants indicated the crucial role of arginine residues in this sequence. Based on empirical and bioinformatic analyses, we propose two domains in EBITEIN1. One is the minimal sequence for binding ERK2 (EB domain) and the other is the EBITEIN1 C-terminal domain (ECT domain). These results might pave the way for further empirical and bioinformatic analyses of EBITEIN1- and ERK2-mediated events.

Molecular cloning of Ebitein1: a novel extracellular signal-regulated kinase 2-binding protein in testis.

We cloned a cDNA encoding a novel extracellular signal-regulated kinase (ERK) 2-binding protein, EBITEIN1, by yeast two-hybrid screening. Northern and Western blotting experiments showed that the transcript and protein were expressed in the testes. Furthermore, immunohistochemical experiments showed that EBITEIN1 existed at high levels in round spermatids, but at very low levels or not at all in other testicular cells. During spermatogenesis, EBITEIN1 was first translated after meiosis when cells became haploid, then the amount of EBITEIN1 protein gradually increased, reaching a maximum at Oakberg's stage 9. Subsequently, the level of EBITEIN1 decreased such that it was undetectable when the flagellum of the spermatozoon was generated. On a subcellular level, EBITEIN1 localized in the cytoplasm. Based on these results, we propose that EBITEIN1 is an interactor of ERK2 in the intracellular signal transduction pathway that occurs during the morphogenetic development of round spermatids to spermatozoa. The existence of this novel ERK2-interactor indicates that there could be a novel intracellular signaling pathway and/or regulatory mechanism by which ERK2 regulates intracellular events.

Increased pentosidine, an advanced glycation end-product, in urine and tissue reflects disease activity in inflammatory bowel diseases.

BACKGROUND: Under inflammatory conditions with strong oxidative stresses, advanced glycation end-products (AGE), carbonyl compounds, are produced. The concentration of pentosidine, an AGE, reportedly correlates with complications of diabetes mellitus and worsening of rheumatoid arthritis, but its role in the pathogenesis of inflammatory bowel diseases (IBD) is unclear. METHODS: Immunohistochemistry was performed with antibodies against pentosidine, and 8-OH-2-deoxyguanosine. The urinary concentration of pentosidine was also quantified by enzyme-linked immunosorbent assay method. RESULTS: Pentosidine expression was up-regulated in the inflamed tissue of IBD. The expression of both pentosidine and 8-OH-2-deoxyguanosine was similar and increased in the inflamed epithelium and infiltrating cells (neutrophils and lymphocytes). The urinary concentration of pentosidine in active ulcerative colitis was significantly greater than that in inactive ulcerative colitis (0.12+/-0.15 vs 0.021+/-0.011 microg/mg of Cr, P<0.05), and was greater in active Crohn's disease than in inactive Crohn's disease (0.071+/-0.086 vs 0.039+/-0.023 microg/mg of Cr). CONCLUSIONS: The urinary pentosidine level correlated with the activity of ulcerative colitis and may be a marker for disease activity in ulcerative colitis.

Increased expression of long pentraxin PTX3 in inflammatory bowel diseases.

The aims of this study were to investigate the expression of pentraxin-3 in inflamed gastrointestinal tissue in patients with inflammatory bowel diseases and to elucidate the usefulness of plasma pentraxin-3 level as an inflammation marker in patients with inflammatory bowel diseases. Pentraxin-3 immunoreactivity was found in infiltrating neutrophils and vessels in the inflamed gut. Plasma pentraxin-3 concentration in patients with active inflammatory bowel diseases was significantly higher than that of normal subjects and patients with inactive inflammatory bowel diseases. Significant positive correlations of clinical disease activity with plasma pentraxin-3 concentration and serum CRP concentration were found in patients with inflammatory bowel diseases. Pentraxin-3 is directly produced from the inflamed gut in inflammatory bowel diseases. In conclusion, plasma pentraxin-3 concentration is a useful marker for understanding the disease activity in patients with inflammatory bowel diseases.

An optimized immunohistochemical method for detection of phosphorylated mitogen-activated protein kinases.

Study of the in vivo spatio-temporal localization of modified proteins is likely to become a major focus of proteomics research in the near future. In this study we optimized and tested an immunohistochemical procedure for detecting unstable phosphorylated mitogen-activated protein (MAP) kinases. Using our method, phosphorylated MAP kinases can be sensitively and reproducibly localized in the developing white matter of murine spinal cord on embryonic day 15. Our method is simple and effective, and so may be useful in future proteomics research.

A matrix metalloproteinase inhibitor, ONO-4817, suppresses the development of aortic intimal hyperplasia in experimental hyperlipidemic rabbit.

Inhibition of matrix metalloproteinases (MMPs) would be expected to suppress atherosclerotic neointimal proliferation and thus limit atheromatous plaque progression, but this has not yet been demonstrated morphologically in atherosclerotic intimal hyperplasia induced by cholesterol loading in experimental animals. We therefore investigated whether a broad-spectrum MMP inhibitor (MMPi), ONO-4817, could inhibit the development of intimal hyperplasia in male hyperlipidemic rabbits (n = 6) fed laboratory chow supplemented with 1% cholesterol for 2 months followed by a 1% cholesterol diet plus 100 mg/kg ONO-4817 for another month (Chol + ONO group). Control animals (n = 6) received no ONO-4817. When the aortas were studied both histologically and immunohistochemically, intimal hyperplasia was inhibited in Chol + ONO rabbits. The distribution of macrophages and MMP-12 in the hyperplastic tissue of the Chol + ONO rabbits was limited to the luminal side of the lesions. No such limitation in the distribution of macrophages and MMP-12 was observed in the control group. The distribution of smooth muscle cells in the hyperplastic tissue was not different between the Chol + ONO and control groups. However, the distribution of MMP-2 and MMP-12 was limited to the luminal side of lesions in the Chol + ONO group. This is the first reported evidence that an MMPi can suppress the development of intimal hyperplasia in hyperlipidemic rabbits.

Histogenesis of the intravitreal membrane and secondary vitreous in the mouse.

PURPOSE: The intravitreal membrane (IVM) is a membranous structure between the primary and secondary vitreous bodies in developing mammalian eyes. In this study, for the first time the histogenesis of the IVM and the relationship between the hyaloid vasculature and the IVM was characterized in newborn mice. METHODS: Eyes of mice less than 12 days old were fixed and embedded. From these, serial paraffin-embedded sections were made for lectin histochemistry, immunohistochemistry, and picrosirius red (PSR) staining, and ultrathin sections were made for transmission electron microscopy (TEM). Eight biotinylated lectins and antibodies for laminin and type IV collagen were used. RESULTS: Among the eight lectins tested, concanavalin A (Con A) agglutinin, Ricinus communis agglutinin I, and wheat germ agglutinin demonstrated strong positive staining in the IVM and vitreous fibrils of the primary and secondary vitreous bodies. They also bound to the internal limiting membrane (ILM) of the retina. At postgestational day 4, the secondary vitreous first appeared between the ILM and the vasa hyaloidea propria (VHP). Immunohistochemical staining revealed that the IVM consists of extracellular matrix components including laminin and type IV collagen, whereas PSR staining and TEM showed that collagen fibrils in the IVM are bundled and continuous with the basement membrane of hyaloid capillaries or the VHP. CONCLUSIONS: Lectin histochemistry and immunohistochemistry provided good methods for visualizing the structures of the IVM and vitreous fibrils. These results suggest that the IVM is separated from the basement membrane of the retinal ILM along with the vascular network of the VHP when the secondary vitreous begins to form.

Phosphorylated extracellular signal-regulated kinase 1/2 is localized to the XY body of meiotic prophase spermatocytes.

We found that phosphorylated extracellular signal-regulated kinase 1/2 (phospho-ERK1/2) is localized to the XY body of meiotic prophase spermatocytes. A more detailed surface spread analysis showed that phospho-ERK1/2 is localized to the synaptonemal complex of the XY pair of pachytene spermatocytes or the entire XY body of zygotene spermatocytes. In the XY body of meiotic prophase spermatocytes, both transcription and homologous recombination are inactivated. These results suggest a novel function of ERK1/2 in meiotic sex chromosome inactivation.

Activation of connective tissue growth factor gene by the c-Maf and Lc-Maf transcription factors.

The Maf family of transcription factors is expressed during development of various organs and tissues, and is involved in a variety of developmental and cellular differentiation processes. We previously found that c-maf and mafB are strongly expressed in hypertrophic chondrocytes during cartilage development. Connective tissue growth factor (CTGF) is also expressed in hypertrophic chondrocytes. Adenovirus mediated introduction of c-maf gene into the mouse fibroblast cell line C3H10T1/2 strongly induced CTGF expression. CTGF can be induced by TGF-beta via the SMAD pathway; however, the c-Maf could not induce TGF-beta, nor could TGF-beta induce the c-Maf, suggesting that activation of CTGF by Maf is TGF-beta independent. Reporter transfection analysis using C3H10T1/2 cells shows that c-Maf stimulates a CTGF reporter gene. Lc-Maf, a splice variant of c-Maf containing an extra 10 amino acids in the carboxyl terminus, was a stronger inducer of the CTGF reporter gene than c-Maf. Chromatin immunoprecipitation analysis showed that c-Maf binds to the promoter region of the CTGF gene, indicating that Maf directly activates the CTGF gene. Taken together, these data indicate that the CTGF gene is a target of c-Maf and Lc-Maf in cartilage development.

Disappearance of p27(KIP1) and increase in proliferation of the lens cells after extraction of most of the fiber cells of the lens.

PURPOSE: Proliferation of the lens epithelial cells is involved in the fibrotic changes of lens capsules after cataract extraction. However, the mechanisms of the proliferation of the lens epithelial cells are largely unknown. The purpose of this study was to examine the correlation between the expression of p27(KIP1) and cell proliferation in the lens cells after the extraction of lens fiber cells. METHODS: At embryonic days (E) 14 and 18, the C57Bl6 mice were anesthetized and the embryos were surgically removed. The eyes were dissected from these embryos and also from mice 12 weeks after birth. The 12-week-old mice were anesthetized, and then the lens fiber cells were extracted. Normal eyes at E14 and 18 and eyes fixed at 15 min and 48 hr after the extraction of the lens fiber cells were analyzed using immunohistochemistry with anti-p27(KIP1), p57(KIP2), and phosphorylated extracellular signal-regulated kinase (phospho-ERK) 1/2 antibodies, and cells in the S phase of the cell cycle were also examined using anti-bromodeoxyuridine (BrdU) antibody. RESULTS: p27(KIP1) and p57(KIP2)-positive cells were present in the equatorial region of E14 mice. At E18, many lens fiber cells showed nuclear immunoreactivity for p27(KIP1), whereas a small number of cells were positive for p57(KIP2) in the equatorial region. At 12 weeks of age, all nuclei of the lens epithelial cells as well as lens fiber cells showed nuclear immunoreactivity for p27(KIP1). In contrast, p57(KIP2) and phospho-ERK1/2 were not expressed in the lens cells. At 15 min after the extraction of lens fiber cells, phospho-ERK1/2 as well as p27(KIP1) were detected in the lens cells. At 48 hr after the extraction of the lens fiber cells, a few p27(KIP1)-positive nuclei were observed in the equatorial region of the lens capsule. In contrast, many lens cells showed nuclear immunoreactivity for BrdU. CONCLUSIONS: These findings suggest that degradation of p27(KIP1) mediated by phosphorylation of ERK 1/2 is correlated with proliferation of the epithelial cells after the extraction of the lens fiber cells.

Expression of maf-B mRNA in the epithelium around the eyelid closure of the mouse eye at embryonic day 18.

Maf encodes a transcription factor protein containing a typical basic leucine zipper domain structure, a motif for protein dimerization and DNA binding. We examined the expression of maf-B mRNA in the epithelium around the eyelid closure. Expression of maf-B mRNA was examined in C57Bl6 mice at the embryonic stages in 12.5 days of gestation (E12.5) and E18 using in situ hybridization with 35S-labeled antisense riboprobes. In embryos studied 12.5 days postconception, a message specific for maf-B was not detected around the developing eyelid. In contrast, maf-B was strongly expressed in the epithelium of the eyelid closure at E18. Expression of maf-B was strongly noted in the suprabasal differentiating cells derived from the basal layer of the conjunctiva and epidermis. In contrast, basal cells in the eyelid closure and in the epidermis, as well as keratinizing cells, did not express maf-B. These data indicate that maf-B mRNA is expressed during development of the eyelid closure.

Developmental contribution of c-maf in the kidney: distribution and developmental study of c-maf mRNA in normal mice kidney and histological study of c-maf knockout mice kidney and liver.

Maf is a family of oncogenes which encodes a nuclear bZip transcription factor protein and has been originally identified from the avian oncogenic retrovirus, AS42. Maf genes have been reported to have critical roles in embryological development and cellular differentiation. In this study, in situ hybridization with (35)S-labeled antisense riboprobes was used to investigate the distribution of c-maf mRNA in balb/c mouse kidneys from 12 (E12) through 17 days (E17) of gestation and then 1 and 4 weeks after birth. Immunocytochemistry of 4-week-old mouse kidney using anti-c-maf antisera was also performed. Kidney and liver sections from c-maf knockout mice at 4 weeks were stained with hematoxylin-eosin, and their histological features were examined. Expression of c-maf mRNA was first detected on E16 in the renal proximal tubules, and it was expressed through 4 weeks after birth. In the c-maf knockout mice at 4 weeks the cytoplasmic volume of the proximal tubule and liver cell was smaller. These findings suggest that expression of the c-maf gene may be involved in the embryological development and/or cell differentiation of kidney and liver cells.