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Background: R-spondins (Rspos) are secreted proteins that modulate Wnt/β-catenin signaling. At the early stages of spinal cord development, Wnts (Wnt1, Wnt3a) and Rspos (Rspo1, Rspo3) are co-expressed in the roof plate, suggesting that Rspos are involved in development of dorsal spinal cord and neural crest cells in cooperation with Wnt ligands., Results: Here, we found that Rspo1 and Rspo3, as well as Wnt1 and Wnt3a, maintained roof-plate-specific expression until late embryonic stages. Rspo1- and Rspo3-double-knock-out (dKO) embryos partially exhibited the phenotype of Wnt1 and Wnt3a dKO embryos. While the number of Ngn2-positive sensory lineage neural crest cells is reduced in Rspo-dKO embryos, development of dorsal spinal cord, including its size and dorso-ventral patterning in early development, elongation of the roof plate, and proliferation of ependymal cells, proceeded normally. Consistent with these slight defects, Wnt/β-catenin signaling was not obviously changed in developing spinal cord of dKO embryos., Conclusions: Our results show that Rspo1 and Rspo3 are dispensable for most developmental processes involving roof plate-derived Wnt ligands, except for specification of a subtype of neural crest cells. Thus, Rspos may modulate Wnt/β-catenin signaling in a context-dependent manner., (© 2023 The Authors. Developmental Dynamics published by Wiley Periodicals LLC on behalf of American Association for Anatomy.)

Retinitis pigmentosa (RP) and macular dystrophy (MD) cause severe retinal dysfunction, affecting 1 in 4000 people worldwide. This disease is currently assumed to be intractable, because effective therapeutic methods have not been established, regardless of genetic or sporadic traits. Here, we examined a RP mouse model in which the Prominin-1 (Prom1) gene was deficient and investigated the molecular events occurring at the outset of retinal dysfunction. We extracted the Prom1-deficient retina subjected to light exposure for a short time, conducted single-cell expression profiling, and compared the gene expression with and without stimuli. We identified the cells and genes whose expression levels change directly in response to light stimuli. Among the genes altered by light stimulation, Igf1 was decreased in rod photoreceptor cells and astrocytes under the light-stimulated condition. Consistently, the insulin-like growth factor (IGF) signal was weakened in light-stimulated photoreceptor cells. The recovery of Igf1 expression with the adeno-associated virus (AAV) prevented photoreceptor cell death, and its treatment in combination with the endothelin receptor antagonist led to the blockade of abnormal glial activation and the promotion of glycolysis, thereby resulting in the improvement of retinal functions, as assayed by electroretinography. We additionally demonstrated that the attenuation of mammalian/mechanistic target of rapamycin (mTOR), which mediates IGF signalling, leads to complications in maintaining retinal homeostasis. Together, we propose that combinatorial manipulation of distinct mechanisms is useful for the maintenance of the retinal condition., (© 2024. The Author(s).)

During development, progenitor cell survival is essential for proper tissue functions, but the underlying mechanisms are not fully understood. Here we show that ERCC6L2, a member of the Snf2 family of helicase-like proteins, plays an essential role in the survival of developing chick neural cells. ERCC6L2 expression is induced by the Sonic Hedgehog (Shh) signaling molecule by a mechanism similar to that of the known Shh target genes Ptch1 and Gli1. ERCC6L2 blocks programmed cell death induced by Shh inhibition and this inhibition is independent of neural tube patterning. ERCC6L2 knockdown by siRNA resulted in the aberrant appearance of apoptotic cells. Furthermore, ERCC6L2 cooperates with the Shh signal and plays an essential role in the induction of the anti-apoptotic factor Bcl-2. Taken together, ERCC6L2 acts as a key factor in ensuring the survival of neural progenitor cells., (© 2023 Japanese Society of Developmental Biologists.)

Neural induction is a process where naive cells are converted into committed cells with neural characteristics, and it occurs at the earliest step during embryogenesis. Although the signaling molecules and chromatin remodeling for neural induction have been identified, the mutual relationships between these molecules are yet to be fully understood. By taking advantage of the neural differentiation system of mouse embryonic stem (ES) cells, we discovered that the BMP signal regulates the expression of several polycomb repressor complex (PRC) component genes. We particularly focused on Polyhomeotic Homolog 1 (Phc1) and established Phc1 -knockout ( Phc1 -KO) ES cells. We found that Phc1 -KO failed to acquire the neural fate, and the cells remained in pluripotent or primitive non-neural states. Chromatin accessibility analysis suggests that Phc1 is essential for chromatin packing. Aberrant upregulation of the BMP signal was confirmed in the Phc1 homozygotic mutant embryos. Taken together, Phc1 is required for neural differentiation through epigenetic modification., Competing Interests: The authors declare no conflict of interest., (© 2023 The Authors.)

Purpose: Spermatogenesis is a complex process orchestrated by several essential genes. Prominin-1 ( Prom1 /PROM1) is a gene that is expressed in the testis but with a poorly understood role in spermatogenesis., Methods: We used Prom1 knockout ( Prom1 KO) mice to assess the role of Prom1 in spermatogenesis. To this end, we performed immunohistochemistry, immunofluorescence, western blotting, β-galactosidase staining, and apoptosis assay. Additionally, we analyzed the morphology of sperm and assessed litter sizes., Results: We observed that PROM1 is localized to the dividing spermatocytes in seminiferous epithelial cells, sperm, and columnar epithelium in the epididymis. In the Prom1 KO testis, an aberrant increase in apoptotic cells and a decrease in proliferating seminiferous epithelial cells were observed. Cellular FLICE-like inhibitory protein (c-FLIP) and extracellular signal-regulated kinase 1/2 (ERK1/2) expression were also significantly decreased in Prom1 KO testis. In addition, a significantly increased number of epididymal spermatozoa with abnormal morphology and less motility was found in Prom1 KO mice., Conclusions: PROM1 maintains spermatogenic cell proliferation and survival via c-FLIP expression in the testis. It is also involved in sperm motility and fertilization potential. The mechanism underlying the effect of Prom1 on sperm morphology and motility remains to be identified., Competing Interests: The authors declare no conflict of interest., (© 2023 The Authors. Reproductive Medicine and Biology published by John Wiley & Sons Australia, Ltd on behalf of Japan Society for Reproductive Medicine.)

During development, the hypothalamus emerges from the ventral diencephalon and is regionalized into several distinct functional domains. Each domain is characterized by a different combination of transcription factors, including Nkx2.1, Nkx2.2, Pax6, and Rx, which are expressed in the presumptive hypothalamus and its surrounding regions, and play critical roles in defining each area. Here, we recapitulated the molecular networks formed by the gradient of Sonic Hedgehog (Shh) and the aforementioned transcription factors. Using combinatorial experimental systems of directed neural differentiation of mouse embryonic stem (ES) cells, as well as a reporter mouse line and gene overexpression in chick embryos, we deciphered the regulation of transcription factors by different Shh signal intensities. We then used CRISPR/Cas9 mutagenesis to demonstrate the mutual repression between Nkx2.1 and Nkx2.2 in a cell-autonomous manner; however, they induce each other in a non-cell-autonomous manner. Moreover, Rx resides upstream of all these transcription factors and determines the location of the hypothalamic region. Our findings suggest that Shh signaling and its downstream transcription network are required for hypothalamic regionalization and establishment., (© The Author(s) 2023. Published by Oxford University Press. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Many young individuals attempt to lose too much weight because of a false body image, which induces low bone mineral density (BMD) resulting from energy restriction. In addition, a decrease in estrogen has been observed along with the decrease in BMD. Estrogen is responsible for maintaining bone mass, and soybeans contain high levels of isoflavones, which have estrogen-like effects. Thus, we hypothesized that soy protein prevents low BMD caused by energy deficiency in young female rats. The purpose of this study was to examine the effect of soy protein intake on bone loss by energy deficiency in young female rats. Female Sprague-Dawley rats (6 weeks old) were randomly divided into the following 4 experimental groups: ad libitum feeding and casein diet (AL-Cas); ad libitum feeding and soy diet (AL-Soy); 40% energy restriction and casein diet (ER-Cas); and 40% energy restriction and soy diet (ER-Soy). The experimental period was 10.5 weeks. The AL-soy group had significantly higher BMD of the femur than the AL-Cas group (AL-Cas = 156 ± 5 mg/cm 2 , AL-Soy = 165 ± 7 mg/cm 2 ; P < .05). Meanwhile, the ER-Soy group had significantly lower BMD of the tibia, femur, and lumbar spine than the ER-Cas group (ER-Cas = 147 ± 7 mg/cm 2 , ER-Soy = 133 ± 10 mg/cm 2 ; P < .01). These results show that compared with ad libitum control groups, soy protein resulted in higher BMD under nonenergy deficiency, but under energy-deficiency conditions, it resulted in lower BMD., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Understanding the molecular mechanisms leading to retinal development is of great interest for both basic scientific and clinical applications. Several signaling molecules and transcription factors involved in retinal development have been isolated and analyzed; however, determining the direct impact of the loss of a specific molecule is problematic, due to difficulties in identifying the corresponding cellular lineages in different individuals. Here, we conducted genome-wide expression analysis with embryonic stem (ES) cells devoid of the Rx gene, which encodes one of several homeobox transcription factors essential for retinal development. We performed three-dimensional differentiation of wild-type and mutant cells and compared their gene-expression profiles. The mutant tissue failed to differentiate into the retinal lineage and exhibited precocious expression of genes characteristic of neuronal cells. Together, these results suggest that Rx expression is an important biomarker of the retinal lineage and that it helps regulates appropriate differentiation stages., (© 2022 Japanese Society of Developmental Biologists.)

Objectives: Mammalian DNA methyltransferases are essential to re-establish global DNA methylation patterns during implantation, which is critical for transmitting epigenetic information to the next generation. In contrast, the significance of methyl-CpG binding proteins (MBPs) that bind methylated CpG remains almost unknown at this stage. We previously demonstrated that Zbtb38 (also known as CIBZ)-a zinc finger type of MBP-is required for mouse embryonic stem (ES) cell proliferation by positively regulating Nanog expression. However, the physiological function of Zbtb38 in vivo remains unclear., Materials and Methods: This study used the Cre-loxP system to generate conditional Zbtb38 knockout mice. Cell proliferation and apoptosis were studied by immunofluorescence staining. Quantitative real-time PCR, immunoblotting and immunofluorescence were performed to investigate the molecular mechanisms., Results: Germline loss of the Zbtb38 single allele resulted in decreased epiblast cell proliferation and increased apoptosis shortly after implantation, leading to early embryonic lethality. Heterozygous loss of Zbtb38 reduced the expression of Nanog, Sox2, and the genes responsible for epiblast proliferation, differentiation, and cell viability. Although this early lethal phenotype, Zbtb38 is dispensable for ES cell establishment and identity., Conclusions: These findings indicate that Zbtb38 is essential for early embryonic development via the suppression of Nanog and Sox2 expression., (© 2022 The Authors. Cell Proliferation published by John Wiley & Sons Ltd.)

Background: The neural tube comprises several different types of progenitors and postmitotic neurons that co-ordinately act with each other to play integrated functions. Its development consists of two phases: proliferation of progenitor cells and differentiation into postmitotic neurons. How progenitor cells differentiate into each corresponding neuron is an important question for understanding the mechanisms of neuronal development., Results: Here we introduce one of the Sox transcription factors, Sox14, which plays an essential role in the promotion of neuronal differentiation. Sox14 belongs to the SoxB2 subclass and its expression starts in the progenitor regions before neuronal differentiation is initiated at the trunk level of the neural tube. After neuronal differentiation is initiated, Sox14 expression gradually becomes confined to the V2a region of the neural tube, where Chx10 is co-expressed. Overexpression of Sox14 restricts progenitor cell proliferation. Conversely, the blockade of Sox14 expression by the RNAi strategy inhibits V2a neuron differentiation and causes expansion of the progenitor domain. We further found that Sox14 acted as a transcriptional activator., Conclusions: Sox14 acts as a modulator of cell proliferation and is essential for initiation of neuronal differentiation in the chick neural tube., (© 2021 American Association for Anatomy.)

Retinitis pigmentosa (RP) and macular dystrophy (MD) are characterized by gradual photoreceptor death in the retina and are often associated with genetic mutations, including those in the prominin-1 (Prom1) gene. Prom1-knockout (KO) mice recapitulate key features of these diseases including light-dependent retinal degeneration and constriction of retinal blood vessels. The mechanisms underlying such degeneration have remained unclear, however. We here analysed early events associated with retinal degeneration in Prom1-KO mice. We found that photoreceptor cell death and glial cell activation occur between 2 and 3 weeks after birth. Whereas gene expression was not affected at 2 weeks, the expression of several genes was altered at 3 weeks in the Prom1-KO retina, with the expression of that for endothelin-2 (Edn2) being markedly upregulated. Expression of Edn2 was also induced by light stimulation in Prom1-KO mice reared in the dark. Treatment with endothelin receptor antagonists attenuated photoreceptor cell death, gliosis and retinal vessel stenosis in Prom1-KO mice. Our findings thus reveal early manifestations of retinal degeneration in a model of RP/MD and suggest potential therapeutic agents for these diseases. This article has an associated First Person interview with the first author of the paper., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2021. Published by The Company of Biologists Ltd.)

Introduction/aims: Lymphatic vessels are responsible for the removal of metabolic waste from body tissues. They also play a crucial role in skeletal muscle functioning thorough their high-energy metabolism. In this study we investigated whether disuse muscle atrophy induced by hindlimb unloading is associated with an alteration in the number of lymphatic vessels and differential expression of lymphangiogenic factors in the soleus muscle., Methods: Male C57BL/6 mice were subjected to tail suspension (TS) for 2 or 4 weeks to induce soleus muscle atrophy. After TS, lymphatic and blood capillaries in the soleus muscle were visualized and counted by double staining with LYVE-1 and CD31. The protein and mRNA levels of vascular endothelial growth factor (VEGF)-C, VEGF-D, and vascular endothelial growth factor receptor-3 were measured by Western blotting and real-time reverse transcript polymerase chain reaction, respectively., Results: TS for 2 weeks resulted in a significant decrease in the number of blood capillaries compared with controls. However, there was no significant change in the number of lymphatic capillaries. By contrast, TS for 4 weeks resulted in a significant decrease in the number of lymphatic and blood capillaries. We observed a significant decrease in the mRNA levels of VEGF-C and VEGF-D in mice subjected to TS for 4 weeks., Discussion: The decrease of intramuscular lymphatic vessels may a crucial role in the process of muscle atrophy., (© 2021 Wiley Periodicals LLC.)

Organisms living in temperate regions are sensitive to seasonal variations in the environment; they are known to accumulate energy as fat in their livers during the winter when days are shorter, temperatures are lower, and food is scarce. However, the effect of variations in photoperiod alone on hepatic lipid metabolism has not been well studied. Therefore, in this study, we analyzed lipid metabolism in the liver of medaka, Oryzias latipes, while varying the length of days at constant temperature. Larger amounts of fatty acids accumulated in the liver after 14 days under short-day conditions than under long-day conditions. Metabolome analysis showed no accumulation of long-chain unsaturated fatty acids, but showed a significant accumulation of long-chain saturated fatty acids. Short-day conditions induced a reduction in the levels of succinate, fumarate, and malate in the tricarboxylic acid cycle, decreased expression of PPARα, and decreased accumulation of acylcarnitine, which suggested inhibition of lipolysis. In addition, transparent medaka fed on a high-fat diet under short-day conditions exhibited greater amounts of fat accumulation and developed fatty liver. The findings of our study will be useful for creating a medaka hepatic steatosis model for future studies of hepatic steatosis-related diseases., (© 2021 The Authors. FEBS Open Bio published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

Embryonic stem (ES) cells are a useful experimental material to recapitulate the differentiation steps of early embryos, which are usually invisible and inaccessible from outside of the body, especially in mammals. ES cells have greatly facilitated the analyses of gene expression profiles and cell characteristics. In addition, understanding the mechanisms during neural differentiation is important for clinical purposes, such as developing new therapeutic methods or regenerative medicine. As neurons have very limited regenerative ability, neurodegenerative diseases are usually intractable, and patients suffer from the disease throughout their lifetimes. The functional cells generated from ES cells in vitro could replace degenerative areas by transplantation. In this review, we will first demonstrate the historical views and widely accepted concepts regarding the molecular mechanisms of neural induction and positional information to produce the specific types of neurons in model animals. Next, we will describe how these concepts have recently been applied to the research in the establishment of the methodology of neural differentiation from mammalian ES cells. Finally, we will focus on examples of the applications of differentiation systems to clinical purposes. Overall, the discussion will focus on how historical developmental studies are applied to state-of-the-art stem cell research., (© 2021 Japanese Society of Developmental Biologists.)

Exercise is known to improve skeletal muscle function. The mechanism involves muscle contraction-induced activation of the mTOR pathway, which plays a central role in protein synthesis. However, mTOR activation blocks autophagy, a recycling mechanism with a critical role in cellular maintenance/homeostasis. These two responses to muscle contraction look contradictory to the functional improvement of exercise. Herein, we investigate these paradoxical muscle responses in a series of active-inactive phases in a cultured myotube model receiving electrical stimulation to induce intermittent muscle contraction. Our model shows that (1) contractile activity induces mTOR activation and muscle hypertrophy but blocks autophagy, resulting in the accumulation of damaged proteins, while (2) cessation of muscle contraction rapidly activates autophagy, removing damaged protein, yet a prolonged inactive state results in muscle atrophy. Our findings provide new insights into muscle biology and suggest that not only muscle contraction, but also the subsequent cessation of contraction plays a substantial role for the improvement of skeletal muscle function., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Osteomodulin (OMD) and proline/arginine-rich end leucine repeat protein (PRELP) are secreted extracellular matrix proteins belonging to the small leucine-rich proteoglycans family. We found that OMD and PRELP were specifically expressed in umbrella cells in bladder epithelia, and their expression levels were dramatically downregulated in all bladder cancers from very early stages and various epithelial cancers. Our in vitro studies including gene expression profiling using bladder cancer cell lines revealed that OMD or PRELP application suppressed the cancer progression by inhibiting TGF-β and EGF pathways, which reversed epithelial-mesenchymal transition (EMT), activated cell-cell adhesion, and inhibited various oncogenic pathways. Furthermore, the overexpression of OMD in bladder cancer cells strongly inhibited the anchorage-independent growth and tumorigenicity in mouse xenograft studies. On the other hand, we found that in the bladder epithelia, the knockout mice of OMD and/or PRELP gene caused partial EMT and a loss of tight junctions of the umbrella cells and resulted in formation of a bladder carcinoma in situ-like structure by spontaneous breakdowns of the umbrella cell layer. Furthermore, the ontological analysis of the expression profiling of an OMD knockout mouse bladder demonstrated very high similarity with those obtained from human bladder cancers. Our data indicate that OMD and PRELP are endogenous inhibitors of cancer initiation and progression by controlling EMT. OMD and/or PRELP may have potential for the treatment of bladder cancer.

The USA300 clone, which produces Panton-Valentine leukocidin (PVL), is a major pathogenic community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) clone that causes intractable skin infections. Recently, PVL-positive CA-MRSA, including USA300 clones, have emerged in both communities and hospitals in Japan. To prevent an outbreak of PVL-positive MRSA, infected patients should be treated with effective antimicrobial agents at community clinics. Herein, we investigate molecular epidemiological characteristics of PVL-positive MRSA isolated from outpatients with skin and soft tissue infections (SSTI), which are common community-onset infectious diseases. The detection rate of MRSA was 24.9% (362 strains) out of 1455 S. aureus strains isolated between 2013 and 2017. Among the MRSA strains, 15.5% (56 strains) were PVL-positive strains and associated with deep-seated skin infections. Molecular epidemiological analyses of PVL-positive MRSA showed that USA300 was the predominant clone (53.6%, 30 strains) and was identified in Kanto (18 strains), Kagawa (nine strains), Tohoku (two strains) and Hokkaido (one strain). Notably, minocycline and fusidic acid were effective against all PVL-positive MRSA strains. Hence, our data reveals the current status of PVL-positive MRSA isolated from patients with SSTI in Japan. Continuous surveillance of CA-MRSA is necessary to monitor latest prevalence rates and identify effective antimicrobial agents for PVL-positive MRSA strains., (© 2020 Japanese Dermatological Association.)

Hepatic stellate cells (HSCs) play a vital role in liver fibrosis, and a greater understanding of their regulation is required. Recent studies have focused on relationships between extracellular matrix (ECM) stiffness and gene expression or cellular metabolism, but none have provided a detailed metabolic analysis of HSC changes in spheroid cultures. Accordingly, in the present study, we created an HSC spheroid culture and analyzed changes in gene expression and metabolism. Expression of α-smooth muscle actin (α-SMA) decreased in the spheroids, suppressing proliferation. Gene expression analysis revealed the cell cycle, sirtuin signaling, mitochondrial dysfunction, and the Hippo pathway to be canonical pathways, believed to result from decreased proliferative ability or mitochondrial suppression. In the Hippo pathway, nuclear translocation of the yes-associated protein (YAP) was decreased in the spheroid, which was associated with the stiffness of the ECM. Metabolome analysis showed glucose metabolism changes in the spheroid, including glutathione pathway upregulation and increased lipid synthesis. Addition of the glycolytic product phosphoenolpyruvate (PEP) led to increased spheroid size, with increased expression of proteins such as α-SMA and S6 ribosomal protein (RPS6) phosphorylation, which was attributed to decreased suppression of translation. The results of our study contribute to the understanding of metabolic changes in HSCs and the progression of hepatic fibrosis., Competing Interests: The authors declare no conflict of interest.

The hedgehog (Hh) family comprises sonic hedgehog (Shh), Indian hedgehog (Ihh), and desert hedgehog (Dhh), which are versatile signaling molecules involved in a wide spectrum of biological events including cell differentiation, proliferation, and survival; establishment of the vertebrate body plan; and aging. These molecules play critical roles from embryogenesis to adult stages; therefore, alterations such as abnormal expression or mutations of the genes involved and their downstream factors cause a variety of genetic disorders at different stages. The Hh family involves many signaling mediators and functions through complex mechanisms, and achieving a comprehensive understanding of the entire signaling system is challenging. This review discusses the signaling mediators of the Hh pathway and their functions at the cellular and organismal levels. We first focus on the roles of Hh signaling mediators in signal transduction at the cellular level and the networks formed by these factors. Then, we analyze the spatiotemporal pattern of expression of Hh pathway molecules in tissues and organs, and describe the phenotypes of mutant mice. Finally, we discuss the genetic disorders caused by malfunction of Hh signaling-related molecules in humans., (Copyright © 2019 Sasai, Toriyama and Kondo.)

Membrane morphology is an important structural determinant as it reflects cellular functions. The pentaspan membrane protein Prominin-1 (Prom1/CD133) is known to be localised to protrusions and plays a pivotal role in migration and the determination of cellular morphology; however, the underlying mechanism of its action have been elusive. Here, we performed molecular characterisation of Prom1, focussing primarily on its effects on cell morphology. Overexpression of Prom1 in RPE-1 cells triggers multiple, long, cholesterol-enriched fibres, independently of actin and microtubule polymerisation. A five amino acid stretch located at the carboxyl cytosolic region is essential for fibre formation. The small GTPase Rho and its downstream Rho-associated coiled-coil-containing protein kinase (ROCK) are also essential for this process, and active Rho colocalises with Prom1 at the site of initialisation of fibre formation. In mouse embryonic fibroblast (MEF) cells we show that Prom1 is required for chloride ion efflux induced by calcium ion uptake, and demonstrate that fibre formation is closely associated with chloride efflux activity. Collectively, these findings suggest that Prom1 affects cell morphology and contributes to chloride conductance.

We examined the effect of jump exercise on bone parameters in young female rats under food restriction. Seven-week-old female rats were divided into four groups: a sedentary and ad libitum feeding group (n = 10), a jump exercise and ad libitum feeding group (n = 9), a sedentary and 30% food restriction group (n = 9), and a jump exercise and 30% food restriction group (n = 10). The jump groups jumped 20 times/day, 5 times/week. The experiment lasted for 13 weeks. There were no interactions of jump exercise and food restriction on bone. Jump exercise under food restriction conditions induced higher bone strength, bone mineral content, bone area, bone mineral density (BMD), and cortical bone volume in young female rats, similar to rats under ad libitum feeding conditions. Bone strength parameters were not significantly different between ad libitum intake and food restriction with jump exercise training; however, BMD, bone size, and bone mass in the food restriction groups did not reach the levels of those in the ad libitum conditions group with jump exercise training. Neither jump exercise nor food restriction had a significant effect on serum estradiol or IGF-1. Our study reveals jump exercise attenuates loss of biomechanical properties and some bone sites with food restriction in young female rats.

The neural tube is composed of a number of neural progenitors and postmitotic neurons distributed in a quantitatively and spatially precise manner. The floor plate, located in the ventral-most region of the neural tube, has a lot of unique characteristics, including a low cell proliferation rate. The mechanisms by which this region-specific proliferation rate is regulated remain elusive. Here we show that the activity of the mTOR signaling pathway, which regulates the proliferation of the neural progenitor cells, is significantly lower in the floor plate than in other domains of the embryonic neural tube. We identified the forkhead-type transcription factor FoxA2 as a negative regulator of mTOR signaling in the floor plate, and showed that FoxA2 transcriptionally induces the expression of the E3 ubiquitin ligase RNF152, which together with its substrate RagA, regulates cell proliferation via the mTOR pathway. Silencing of RNF152 led to the aberrant upregulation of the mTOR signal and aberrant cell division in the floor plate. Taken together, the present findings suggest that floor plate cell number is controlled by the negative regulation of mTOR signaling through the activity of FoxA2 and its downstream effector RNF152., (Copyright © 2019 Kadoya and Sasai.)

Dorsal-ventral pattern formation of the neural tube is regulated by temporal and spatial activities of extracellular signalling molecules. Sonic hedgehog (Shh) assigns ventral neural subtypes via activation of the Gli transcription factors. Shh activity in the neural progenitor cells changes dynamically during differentiation, but the mechanisms regulating this dynamicity are not fully understood. Here, we show that temporal change of intracellular cAMP levels confers the temporal Shh signal, and the purinergic G-protein-coupled receptor GPR17 plays an essential role in this regulation. GPR17 is highly expressed in the ventral progenitor regions of the neural tube and acts as a negative regulator of the Shh signal in chick embryos. Although the activation of the GPR17-related signal inhibits ventral identity, perturbation of Gpr17 expression leads to aberrant expansion of ventral neural domains. Notably, perturbation of Gpr17 expression partially inhibits the negative feedback of Gli activity. Moreover, GPR17 increases cAMP activity, suggesting that it exerts its function by inhibiting the processing of Gli3 protein. GPR17 also negatively regulates Shh signalling in neural cells differentiated from mouse embryonic stem cells, suggesting that GPR17 function is conserved among different organisms. Our results demonstrate that GPR17 is a novel negative regulator of Shh signalling in a wide range of cellular contexts., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2019. Published by The Company of Biologists Ltd.)

Background: In Japan, with increasing age of the population, diabetic patients often become in need of hemodialysis due to diabetic nephropathy, and thus there is a demand for development of diabetic treatments that take into account renal effects in the elderly. No previous studies of alogliptin had focused on Japanese elderly subjects; we therefore assessed the effects of alogliptin in elderly individuals using available data., Methods: Laboratory data were compiled for 1 year at intervals of 3 months following the start of alogliptin treatment. The subjects were divided into three groups by age: < 65 years (n = 110), 65 - 74 years (n = 87), and ≥ 75 years (n = 93). Laboratory values in comparison with baseline were compared within groups at various time points, and changes from baseline were compared among the different groups., Results: Hemoglobin A1c (HbA1c) levels decreased significantly from baseline values in all groups at and after month 3: the change at month 12 was -0.74±1.45% for the age group < 65, -0.47±1.02% for the age group 65 - 74, and -0.42±1.11% for the age group ≥ 75. The 12-month change in estimated glomerular filtration rate (eGFR) was -6.5 ± 12.0 for the age group < 65, -2.0 ± 8.4 for the age group 65 - 74, and -1.5 ± 10.0 for the age group ≥ 75; the reduction in the age group < 65 was significant, whereas the reduction in the age groups ≥ 65 was not., Conclusions: Alogliptin significantly lowers HbA1c levels in the elderly and can be used without posing any safety issues, including renal effects, thus contributing to safe blood glucose control in clinical practice., Competing Interests: This study was designed and planned by members of the Kanagawa Physicians Association and was funded by the Japan Diabetes Foundation. No company was involved in designing the study, patient enrollment, calculation or analysis of the data, interpretation of the findings, or writing the manuscript.

Differences in the culturing conditions of mesenchymal stem cells used in regenerative medicine may affect their differentiation ability, genome instability, and therapeutic effects. In particular, bone marrow-derived mesenchymal stem cells cultured under hypoxia are known to proliferate while maintaining an undifferentiated state and the use of deferoxamine, a hypoxia mimetic reagent, has proven to be a suitable strategy to maintain the cells under hypoxic metabolic state. Here, the deferoxamine effects were investigated in mesenchymal stem cells to gain insights into the mechanisms regulating stem cell survival. A 12-h deferoxamine treatment reduced proliferation, oxygen consumption, mitochondrial activity, and ATP production. Microarray analysis revealed that deferoxamine enhanced the transcription of genes involved in glycolysis and the HIF1α pathway. Among the earliest changes, transcriptional variations were observed in HIF1α, NUPR1, and EGLN, in line with previous reports showing that short deferoxamine treatments induce substantial changes in mesenchymal stem cells glycolysis pathway. NUPR1, which is induced by stress and involved in autophagy-mediated survival, was upregulated by deferoxamine in a concentration-dependent manner. Consistently, NUPR1 knockdown was found to reduce cell proliferation and increase the proapoptotic effect of staurosporine, suggesting that deferoxamine-induced NUPR1 promotes mesenchymal stem cell survival and cytoprotective autophagy. Our findings may substantially contribute to improve the effectiveness of mesenchymal stem cell-based regenerative medicine., Competing Interests: No potential conflicts of interest were disclosed.

Background: Sitagliptin, the first dipeptidyl peptidase-4 inhibitor, has demonstrated efficacy and safety as monotherapy and as add-on therapy to oral antidiabetic agents or insulin. However, there have been few reports about sitagliptin in elderly patients. The ASSIST-K observational study was performed in patients with type 2 diabetes mellitus (T2DM) receiving sitagliptin as add-on therapy to insulin. Changes of hemoglobin A1c (HbA1c), body weight, and the estimated glomerular filtration rate (eGFR), as well as adverse events, were investigated over 12 months in age-stratified groups., Methods: Among outpatients with T2DM treated at member institutions of Kanagawa Physicians Association, those starting sitagliptin as add-on therapy to insulin were followed for 12 months. HbA1c (National Glycohemoglobin Standardization Program), body weight, and eGFR were the efficacy endpoints, while adverse events were investigated to assess safety. Patients were stratified into three age groups (≤ 64 years, 65 - 74 years, and ≥ 75 years) for comparison of the endpoints., Results: Among 937 patients on insulin before starting sitagliptin, 821 patients were analyzed after excluding those without HbA1c data at baseline and 12 months. The two groups of elderly patients (65 - 74 years and ≥75 years) had more complications and their HbA1c was lower at initiation of sitagliptin therapy. The dose of sitagliptin, daily number of insulin injections, and number of concomitant oral antidiabetic agents were all lower in the elderly patients. HbA1c showed a significant decrease after initiation of sitagliptin in all age groups, and there were no significant intergroup differences in the change of HbA1c at 12 months. Body weight did not change significantly in any group. eGFR decreased significantly in all groups, with no significant intergroup differences at 12 months. Regarding adverse events, there were no significant intergroup differences in the incidence of severe hypoglycemia, gastrointestinal symptoms, or constipation., Conclusions: Despite baseline differences in demographic factors and medications, sitagliptin showed good efficacy and safety in all age groups of patients receiving it as add-on therapy to insulin during routine management of T2DM. Adding sitagliptin to insulin achieves similar efficacy and safety outcomes at 12 months in both elderly and non-elderly T2DM patients., Competing Interests: The authors declare no conflicts of interest.

Recently, the USA300 clone, which is a Panton-Valentine leukocidin (PVL)-positive clonal complex 8-staphylococcal cassette chromosome mec type IV (CC8-IV) community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) strain, emerged in community and hospital settings in Japan. Hence, clonal types of CA-MRSA strains are predicted to be changing. Nonetheless, long-term surveillance of CA-MRSA has not been conducted in Japan. Here, we investigated the transition and current status of CA-MRSA strains isolated from outpatients with impetigo; the samples were collected between 2007 and 2016 in Kagawa, Japan. The detection rate (22.8%, 488/2139 strains) of MRSA slightly decreased in these 10 years. Molecular epidemiological analyses showed that the prevalence of the CC89-II clone, which is a typical CA-MRSA genotype of causative agents of impetigo, significantly decreased from 48.0% (48/100 strains) in 2007-2009 to 21.9% (16/73 strains) in 2013-2016. By contrast, a non-USA300 CC8-IV clone, which is a highly pathogenic CA-MRSA/J clone, significantly increased in prevalence from 9.0% (9/100 strains) to 32.9% (24/73 strains). The prevalence of PVL-positive CA-MRSA strains increased annually from 2012 (0%) to 2015 (6.7%), whereas only one of these strains turned out to be the USA300 clone. Antibiotic susceptibility data revealed that the rates of resistance to gentamicin and clindamycin among CA-MRSA strains decreased along with the decreased prevalence of the CC89-II clone and increased prevalence of the CA-MRSA/J clone. Our data strongly suggest that the clonal types and antibiotic susceptibility of CA-MRSA isolated from patients with impetigo dramatically changed during the last 10 years in Japan., (© 2019 Japanese Dermatological Association.)

Recently, the dissemination of community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) into hospitals has frequently been reported worldwide. Hospital-acquired MRSA (HA-MRSA) strains exhibit high-level resistance to multiple antimicrobial agents, whereas CA-MRSA strains are usually susceptible to non-β-lactams. Thus, it is predicted that the antibiogram of the HA-MRSA population would change along with the change in genotype of MRSA. Here, we investigated the changes in the MRSA population along with the MRSA antibiogram in a hospital between 2010 and 2016. Staphylococcal cassette chromosome (SCC) mec typing showed that the predominant HA-MRSA strains in the hospital dramatically changed from SCCmec type II, which is the major type of HA-MRSA, to SCCmec type IV, which is the major type of CA-MRSA. Multilocus sequence typing revealed that the predominant SCCmec type IV strain was a clonal complex (CC) 8 clone, which is mainly found among CA-MRSA. Furthermore, the CC1-SCCmec type IV (CC1-IV) clone significantly increased. Both the CC8-IV and CC1-IV clones exhibited high antimicrobial susceptibility. The antibiogram change of the HA-MRSA population was consistent with the antimicrobial susceptibilities and increased prevalence of the CC8-IV and CC1-IV clones. Our data reveal that the change in the genotypes of MRSA strains could impact the antibiogram of HA-MRSA population., (Copyright © 2018 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Tight junctions are required for the formation of tissue barriers and function as suppressors of signalling mechanisms that control gene expression and cell behaviour; however, little is known about the physiological and developmental importance of such signalling functions. Here, we demonstrate that depletion of MarvelD3, a transmembrane protein of tight junctions, disrupts neural crest formation and, consequently, development of neural crest-derived tissues during Xenopus embryogenesis. Using embryos and explant cultures combined with a small molecule inhibitor or mutant mRNAs, we show that MarvelD3 is required to attenuate JNK signalling during neural crest induction and that inhibition of JNK pathway activation is sufficient to rescue the phenotype induced by MarvelD3 depletion. Direct JNK stimulation disrupts neural crest development, supporting the importance of negative regulation of JNK. Our data identify the junctional protein MarvelD3 as an essential regulator of early vertebrate development and neural crest induction and, thereby, link tight junctions to the control and timing of JNK signalling during early development.

Background: Ipragliflozin is a selective sodium glucose co-transporter 2 inhibitor. The ASSIGN-K study is investigating the efficacy and safety of ipragliflozin for type 2 diabetes mellitus (T2DM) in the real-world clinical setting., Methods: Japanese T2DM patients with inadequate glycemic control despite diet and exercise with/without pharmacotherapy were enrolled in an investigator-driven, multicenter, prospective, observational study examining the efficacy and safety of ipragliflozin treatment (50 mg/day for 52 weeks). We performed interim analysis after 24 weeks., Results: In 367 patients completing 24-week ipragliflozin therapy, hemoglobin A1c (HbA1c) decreased significantly from 8.07% at baseline to 7.26% in week 24 (P < 0.001). The change in HbA1c from treatment initiation to week 24 was -0.88% in patients < 65 years old versus -0.55% in those ≥ 65 years and -0.92% in men versus -0.70% in women (all P < 0.001). When baseline HbA1c was < 7%, 7% to < 8%, and ≥ 8%, the change was -0.18%, -0.45%, and -1.48%, respectively (P = 0.5352, P < 0.001, and P < 0.001, respectively). When baseline body mass index (BMI) was < 25, 25 to < 30, and ≥ 30, the change was -1.05%, -0.65%, and -0.87%, respectively (all P < 0.001). Multiple regression analysis showed that HbA1c decreased more in patients with a higher baseline HbA1c or shorter duration of diabetes. An HbA1c < 7% was achieved in 33.3% of the patients, and their baseline HbA1c was significantly lower than that of patients failing to achieve it (P < 0.001). Adverse events (AEs) occurred in 106/451 patients (23.5%), including 29.1% of patients aged 65 or older. Common AEs were vulvovaginal candidiasis (3.1%) and genital pruritus (1.8%). Serious AEs included urinary tract infection, unstable angina, and ketosis, which occurred in patients who did not suspend medication during acute illness., Conclusions: Ipragliflozin significantly improved HbA1c in T2DM patients with inadequate glycemic control. Improvement in HbA1c was significant irrespective of age, sex, baseline HbA1c, or BMI, but efficacy was greater with a higher baseline HbA1c and shorter duration of diabetes. For safe continuation of treatment, patients should be advised to suspend medication during acute illness.

Background: Ipragliflozin, a sodium-glucose transporter 2 inhibitor, was administered to patients with type 2 diabetes mellitus for 24 weeks to evaluate its effect on glycemic control and body composition., Methods: This was an investigator-initiated multicenter prospective intervention study in which ipragliflozin (50 mg) was administered once daily and glycemic control, blood pressure, body weight (BW), body composition (measured by a biological impedance method), the lipid profile, and adverse events were evaluated after 4, 12, and 24 weeks of treatment., Results: Efficacy and safety up to 24 weeks of ipragliflozin therapy were analyzed in 367 patients and 451 patients, respectively. Hemoglobin A1c decreased significantly from 8.07% at the start of ipragliflozin therapy to 7.26% in week 24 (P < 0.001). Fasting and postprandial blood glucose levels were significantly reduced by ipragliflozin. In week 24, there were significant decreases from baseline in BW (-2.6 kg), waist circumference (-2.9 cm), and body fat mass (-1.9 kg) (P < 0.001). The body water mass and mineral mass were decreased significantly by 0.5 and by 0.1 kg, respectively (P < 0.001), whereas the protein mass did not change significantly. Intracellular water mass did not change significantly, whereas extracellular water mass showed a significant decrease of 0.5 kg (P < 0.001). Muscle mass did not change in the upper and lower limbs, but that of the trunk decreased significantly (P < 0.001). There was a significant decrease in the fasting triglyceride level and a significant increase in fasting high-density lipoprotein cholesterol level, while low-density lipoprotein cholesterol was unchanged. Adverse events occurred in 23.5% of the patients, with a high frequency of genital infections, such as vulvovaginal candidiasis (3.1%) and genital pruritus (1.8%). Adverse drug reactions were noted in 13.7% of the patients., Conclusions: Administration of ipragliflozin for 24 weeks improved glycemic control and decreased BW. Reduction of body fat accounted for more than 70% of the total weight loss and reduction of extracellular water accounted for about 20%.

Ocular morphogenesis requires several signalling pathways controlling the expression of transcription factors and cell-cycle regulators. However, despite a well-known mechanism, the dialogue between those signals and factors remains to be unveiled. Here, we identify a requirement for MarvelD3, a tight junction transmembrane protein, in eye morphogenesis in Xenopus MarvelD3 depletion led to an abnormally pigmented eye or even an eye-less phenotype, which was rescued by ectopic MarvelD3 expression. Altering MarvelD3 expression led to deregulated expression of cell-cycle regulators and transcription factors required for eye development. The eye phenotype was rescued by increased c-Jun terminal Kinase activation. Thus, MarvelD3 links tight junctions and modulation of the JNK pathway to eye morphogenesis., Competing Interests: The authors declare no competing or financial interests., (© 2016. Published by The Company of Biologists Ltd.)

Background: Ipragliflozin is a selective sodium glucose co-transporter 2 (SGLT2) inhibitor that blocks glucose reabsorption in the proximal tubules. SGLT2 inhibitors are expected to be effective in patients with insulin resistance and obesity, but it is important to select treatment according to patient background factors that minimizes the risk of adverse events. There have been a limited number of investigations into the relationship between the clinical efficacy (reducing hemoglobin A1c (HbA1c) and body weight (BW)) or safety of SGLT2 inhibitors and patient characteristics., Methods: ASSIGN-K is an investigator-initiated, multicenter, prospective observational study examining the efficacy and safety of ipragliflozin (50 - 100 mg/day for 52 weeks) in Japanese patients with type 2 diabetes mellitus (T2DM) who had inadequate glycemic control with HbA1c ≥ 6.0% (National Glycohemoglobin Standardization Program) despite diet and exercise therapy or diet and exercise plus antidiabetic drug therapy. We conducted an interim analysis of the relationship between changes in HbA1c or BW and characteristics in patients who had been on treatment for more than 12 weeks., Results: In 257 patients completing 12 weeks of treatment, HbA1c decreased significantly from 8.23% to 7.55% (-0.68%, P < 0.01). The change in HbA1c after 12 weeks was -0.17%, -0.33%, and -1.16% when baseline HbA1c was < 7%, 7% to < 8%, and ≥ 8%, respectively (P < 0.05, P < 0.01, and P < 0.01, respectively), and -1.30%, -0.62%, and -0.62% when baseline body mass index (BMI) was < 25, 25 to < 30, and ≥ 30, respectively (all P < 0.01). Stratified analysis showed that age, gender, or BMI did not have a significant influence on the improvement in HbA1c. Multiple regression analysis showed that reduction in HbA1c was greater as baseline HbA1c increased and the duration of diabetes decreased. A higher baseline HbA1c was associated with less weight loss., Conclusions: Ipragliflozin significantly improved HbA1c in patients with T2DM. HbA1c improved more when baseline HbA1c was higher and the duration of diabetes was shorter, suggesting that current treatment policies for diabetes could be re-examined.

In the vertebrate neural tube, a morphogen-induced transcriptional network produces multiple molecularly distinct progenitor domains, each generating different neuronal subtypes. Using an in vitro differentiation system, we defined gene expression signatures of distinct progenitor populations and identified direct gene-regulatory inputs corresponding to locations of specific transcription factor binding. Combined with targeted perturbations of the network, this revealed a mechanism in which a progenitor identity is installed by active repression of the entire transcriptional programs of other neural progenitor fates. In the ventral neural tube, sonic hedgehog (Shh) signaling, together with broadly expressed transcriptional activators, concurrently activates the gene expression programs of several domains. The specific outcome is selected by repressive input provided by Shh-induced transcription factors that act as the key nodes in the network, enabling progenitors to adopt a single definitive identity from several initially permitted options. Together, the data suggest design principles relevant to many developing tissues., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Ipragliflozin is a sodium-glucose co-transporter 2 inhibitor that can improve glycemic control and reduce body weight and blood pressure in patients with type 2 diabetes mellitus (T2DM). We evaluated the efficacy and safety of ipragliflozin in the real-world clinical setting, with a focus on the changes of body composition up to 3 months of treatment., Methods: This was a prospective multicenter interventional trial. We investigated changes of the blood pressure, body composition, blood glucose, hemoglobin A1c (HbA1c), ketone bodies, lipids, and insulin after treatment with ipragliflozin (50 - 100 mg/day) for 12 weeks in Japanese patients with T2DM who showed poor glycemic control despite receiving diet and exercise therapy with or without oral antidiabetic drugs for more than 12 weeks., Results: Two hundred and fifty-seven subjects were included in the efficacy analysis up to 12 weeks of treatment and 301 subjects were included in the safety analysis. From baseline to 12 weeks, HbA1c showed a change of -0.68% (95% confidence interval (CI): -0.83, -0.53) and fasting blood glucose showed a change of -23.9 mg/dL (95% CI: -30.5, -17.2), with both parameters displaying a significant reduction (P < 0.001). The difference of body weight from baseline was -1.82 kg (95% CI: -2.14, -1.50), and it also showed significant reduction (P < 0.001). Analysis of body composition revealed that body fat changed by -1.46 kg (95% CI: -1.79, -1.14, P < 0.001) and body water changed by -0.37 kg (95% CI: -0.60, -0.14, P < 0.01). Laboratory tests demonstrated improvement of liver function and the lipid profile. Adverse events (AEs) occurred in 22.6% of the subjects, with frequent events being vulvovaginal candidiasis in 2.7% and cystitis in 2.0%. Serious AEs occurred in three subjects., Conclusions: In patients with T2DM, ipragliflozin improved glycemic control after 1 month of treatment and caused weight loss by reducing body fat more than body water.

Background: Dipeptidyl peptidase-4 (DPP-4) inhibitors have been shown to reduce hemoglobin A1c (HbA1c) in patients with type 2 diabetes, but the reduction varies between patients and adequate glycemic control may not be achieved. We evaluated the efficacy and safety of the DPP-4 inhibitor alogliptin in the real clinical setting, and analyzed factors associated with the improvement of HbA1c by alogliptin treatment., Methods: A retrospective observational study was performed in patients with type 2 diabetes attending hospitals or clinics belonging to the Kanagawa Physicians Association who received treatment with alogliptin for 1 year or longer. Patients using insulin were excluded from the study. The efficacy endpoints were HbA1c (National Glycohemoglobin Standardization Program value), blood glucose (fasting/postprandial), body weight, blood pressure (systolic/diastolic), liver function (glutamate oxaloacetate transaminase, glutamate pyruvate transaminase, and γ-glutamyl transpeptidase), kidney function (serum creatinine and estimated glomerular filtration rate), serum lipids (total cholesterol, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, and triglycerides), and serum amylase. Adverse events were compiled to assess safety., Results: Of 330 patients whose case records were collected, 27 patients were excluded for protocol violations, leaving 303 patients to form the full analysis set. Compared with baseline, HbA1c showed a decrease by 0.54±1.22% (mean ± standard deviation) after 12 months of alogliptin treatment. Factor analysis demonstrated that the change of HbA1c after 12 months was significantly influenced by the baseline HbA1c level, duration of diabetes, concomitant use of sulfonylureas, and compliance with diet therapy. In addition, there was a significant reduction of total cholesterol, low-density lipoprotein cholesterol, and the estimated glomerular filtration rate after 12 months of alogliptin treatment, as well as a significant increase in serum creatinine. No significant changes of body weight, blood pressure, or liver function were observed. Symptoms of hypoglycemia occurred in two patients (0.6%)., Conclusions: Alogliptin displayed a significant hypoglycemic effect and excellent safety in routine clinical use. Factors influencing the change of HbA1c with alogliptin therapy may include the HbA1c at the start of treatment, the duration of diabetes, use of sulfonylureas, and compliance with diet therapy.

Congenital hereditary endothelial dystrophy 1 (CHED1) and posterior polymorphous corneal dystrophy 1 (PPCD1) are autosomal-dominant corneal endothelial dystrophies that have been genetically mapped to overlapping loci on the short arm of chromosome 20. We combined genetic and genomic approaches to identify the cause of disease in extensive pedigrees comprising over 100 affected individuals. After exclusion of pathogenic coding, splice-site, and copy-number variations, a parallel approach using targeted and whole-genome sequencing facilitated the identification of pathogenic variants in a conserved region of the OVOL2 proximal promoter sequence in the index families (c.-339&#95;361dup for CHED1 and c.-370T>C for PPCD1). Direct sequencing of the OVOL2 promoter in other unrelated affected individuals identified two additional mutations within the conserved proximal promoter sequence (c.-274T>G and c.-307T>C). OVOL2 encodes ovo-like zinc finger 2, a C2H2 zinc-finger transcription factor that regulates mesenchymal-to-epithelial transition and acts as a direct transcriptional repressor of the established PPCD-associated gene ZEB1. Interestingly, we did not detect OVOL2 expression in the normal corneal endothelium. Our in vitro data demonstrate that all four mutated OVOL2 promoters exhibited more transcriptional activity than the corresponding wild-type promoter, and we postulate that the mutations identified create cryptic cis-acting regulatory sequence binding sites that drive aberrant OVOL2 expression during endothelial cell development. Our data establish CHED1 and PPCD1 as allelic conditions and show that CHED1 represents the extreme of what can be considered a disease spectrum. They also implicate transcriptional dysregulation of OVOL2 as a common cause of dominantly inherited corneal endothelial dystrophies., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Small leucine-rich repeat proteoglycan (SLRP) family proteins play important roles in a number of biological events. Here, we demonstrate that the SLRP family member Asporin (ASPN) plays a crucial role in the early stages of eye development in Xenopus embryos. During embryogenesis, ASPN is broadly expressed in the neuroectoderm of the embryo. Overexpression of ASPN causes the induction of ectopic eyes. By contrast, blocking ASPN function with a morpholino oligonucleotide (ASPN-MO) inhibits eye formation, indicating that ASPN is an essential factor for eye development. Detailed molecular analyses revealed that ASPN interacts with insulin growth factor receptor (IGFR) and is essential for activating the IGF receptor-mediated intracellular signalling pathway. Moreover, ASPN perturbed the Wnt, BMP and Activin signalling pathways, suggesting that ASPN thereby creates a favourable environment in which the IGF signal can dominate. ASPN is thus a novel secreted molecule essential for eye induction through the coordination of multiple signalling pathways., (© 2015. Published by The Company of Biologists Ltd.)

Background: It is unclear whether dipeptidyl peptidase-4 inhibitors decrease hemoglobin A1c (HbA1c) in a glucose-dependent manner in patients on insulin therapy who have impaired insulin secretion. This study investigated factors influencing the efficacy of sitagliptin when used concomitantly with insulin to treat type 2 diabetes mellitus (T2DM) in the real-world setting., Methods: A retrospective study was conducted of 1,004 T2DM patients at 36 Japanese clinics associated with the Diabetes Task Force of the Kanagawa Physicians Association. Eligible patients had been on insulin for at least 6 months, with a baseline HbA1c of 7.0% (53 mmol/mol) or higher. Baseline characteristics and laboratory data from 495 patients were subjected to multiple regression analysis to identify factors influencing the change of HbA1c., Results: Most patients (n = 809) received sitagliptin at a dose of 50 mg. In the 1,004 patients, HbA1c decreased by 0.74% (6 mmol/mol) and body weight increased by 0.1 kg after 6 months of combination therapy. Multiple regression analysis showed that a higher baseline HbA1c, older age, and lower body mass index influenced the change of HbA1c after 6 months. Hypoglycemic symptoms occurred in 7.4%, but none were severe., Conclusions: These results emphasize the importance of a higher HbA1c at the commencement of sitagliptin therapy in patients on insulin. Glucose-dependent suppression of glucagon secretion by sitagliptin may be useful in patients with impaired insulin secretion. Sitagliptin can be used concomitantly with insulin irrespective of the insulin regimen, duration of insulin treatment, and concomitant medications.

Background: There have only been a few reports about use of dipeptidyl peptidase 4 (DPP-4) inhibitors in elderly patients with type 2 diabetes mellitus (T2DM), suggesting that the safety of these agents has not been sufficiently demonstrated. We performed a comparative review of the efficacy and safety of sitagliptin for Japanese patients with T2DM managed in the real-world clinical setting., Methods: An age-stratified analysis was performed of 831 patients who were treated with sitagliptin for 2 years. Parameters assessed included the hemoglobin A1c (HbA1c), body weight, serum creatinine, and adverse events. HbA1c and the incidence of hypoglycemia were also evaluated in patients treated with sitagliptin and a sulfonylurea (SU), who were divided into three age groups (<65 years, 65-74 years, and ≥75 years)., Results: Comparison of glycemic control parameters, laboratory values, and adverse events revealed significant improvement of HbA1c, casual postprandial plasma glucose, and fasting plasma glucose in each age group with no change in body weight. Serum creatinine increased significantly in all age groups. Hypoglycemia only occurred in patients who received combined treatment with an SU and sitagliptin, and there was no age-related difference in its incidence., Conclusions: HbA1c was improved by 2 years of sitagliptin therapy in all three age groups, and age did not seem to influence the incidence of hypoglycemic events. These results confirm the efficacy and safety of sitagliptin in patients ≥ 75 years old, suggesting that it is also useful for treating elderly patients with T2DM.