Developmental expression of Smoc1 and Smoc2 suggests potential roles in fetal gonad and reproductive tract differentiation

SMOC1 and SMOC2 are matricellular proteins thought to influence growth factor signaling, migration, proliferation, and angiogenesis. We examined the expression and regulation of Smoc1 and Smoc2 in fetal gonad/mesonephros complexes to discover possible roles for these genes in gonad and mesonephros development. Smoc1 was upregulated at ～E10.75 in a center‐to‐poles wave in pre‐Sertoli and pre‐granulosa cells and its expression was greatly reduced in Wt1, Sf1, and Fog2 mutants. After E13.5, Smoc1 was downregulated in an anterior‐to‐posterior wave in granulosa cells but persisted in Sertoli cells, suggesting a sexually dimorphic requirement in supporting cell lineage differentiation. Smoc2 was expressed in Leydig cells, mesonephroi, and Wnt4 mutant ovaries, but not wildtype ovaries. Using organ culture, we determined that Smoc2 expression was dependent on Hedgehog signaling in testes, mesonephroi, and kidneys. Overall, these results demonstrate that SMOC1 and SMOC2 may mediate intercellular signaling and cell type–specific differentiation during gonad and reproductive tract development. Developmental Dynamics 238:2877–2890, 2009. © 2009 Wiley‐Liss, Inc.     

Dmrt1 mutation causes a male-to-female sex reversal after the sex determination by Dmy in the medaka

DMRT1, which is found in many vertebrates, exhibits testis-specific expression during the sexual differentiation period, suggesting a conserved function of DMRT1 in the testicular development of vertebrate gonads. However, functional analyses have been reported only in mammals. The current study focused on the Dmrt1 function in the teleost medaka, Oryzias latipes, which has an XX–XY sex determination system. Although medaka sex is determined by the presence or absence of the Y chromosome-specific gene Dmy, we demonstrated that in one Dmrt1 mutant line, which was found by screening a gene-driven mutagenesis library, XY mutants developed into normal females and laid eggs. Histological analyses of this mutant revealed that the XY mutant gonads first developed into the normal testis type. However, the gonads transdifferentiated into the ovary type. The mutant phenotype could be rescued by transgenesis of the Dmrt1 genomic region. These results show that Dmrt1 is essential to maintain testis differentiation after Dmy-triggered male differentiation pathway.     

Two distinct types of theca cells in the medaka gonad: Germ cell‐dependent maintenance of cyp19a1‐expressing theca cells

Aromatase is a steroidogenic enzyme catalyzing the production of estrogens and is important for the proper development and function of the reproductive system. The lineage of cyp19a1 (ovarian‐type aromatase)‐expressing cells in the developing gonad was analyzed using a transgenic medaka (Oryzias latipes) that recapitulates endogenous cyp19a1 expression with EGFP fluorescence. Our results show that cyp19a1‐expressing cells arise in the ventral stromal cells of the developing female gonad, then expand anteriorly as the gonadal region extends anteriorly. These cells become located close to the developing follicles, and are distinguishable from the P450c17‐I‐expressing theca cells. In the adult ovary, the expression of P450c17‐I and cyp19a1 are mutually exclusive in the outer theca‐cell layer. Cyp19a1 expression in the granulosa cells is found only in the population of large follicles. These observations demonstrate two types of theca cells in the medaka ovary. We also show that the maintenance of cyp19a1‐expressing cells depends on germ cells. Developmental Dynamics 238:2652–2657, 2009. © 2009 Wiley‐Liss, Inc.     

Male Gonadal Differentiation and the Paedomorphic Evolution of the Testis in Teleostei

Testis differentiation from representatives of the Otophysi (Cyprinus carpio), Percomorpha (Amatitlania nigrofasciata), and Atherinomorpha (Poecilia reticulata) was comparatively described. In the undifferentiated gonad of C. carpio, the primordial germ cells (PGCs) are scattered throughout the gonads while in A. nigrofasciata and P. reticulata the PGCs are restricted to the ventral periphery. In the dorsal region of the developing gonads, with the exception of C. carpio, somatic cell rearrangements result in the differentiation of the sperm duct. Pre‐Sertoli cells wrap around single spermatogonia forming cysts that proliferate forming acinar‐clusters. In C. carpio and A. nigrofasciata, the cysts in each acinar‐cluster move away from each other, creating a central lumen. In C. carpio, the acinar‐clusters then fuse to each other forming tubules that become lined by the germinal epithelium. Subsequently, the tubules anastomose dorsally and create the sperm duct. In A. nigrofasciata, the acinar‐clusters elongate, forming lobules that individually connect to the sperm duct. These are lined by the germinal epithelium. In P. reticulata, the spermatogonial cysts remain in the acinar‐cluster organization. Subsequently, developing ducts connect each cluster to the sperm duct and lobules subsequently develop. In the differentiated testis of C. carpio and A. nigrofasciata, spermatogonia are distributed along the lengths of the anastomosing tubules or lobules, respectively. However, in P. reticulata, the spermatogonia remain restricted to the terminal end of the lobules. Considering testis ontogeny, the spermatogonial acinar‐cluster is the adult characteristic of more derived taxa that approximate the early gonad developmental stages of the basal taxa. Anat Rec, 297:1137–1162, 2014. © 2014 Wiley Periodicals, Inc.     

Activation of beta-catenin signaling by Rspo1 controls differentiation of the mammalian ovary

The sex of an individual is determined by the fate of the gonad.While the expression of Sry and Sox9 is sufficient to inducemale development, we here show that female differentiation requiresactivation of the canonical β-catenin signaling pathway.β-catenin activation is controlled by Rspo1 in XX gonadsand Rspo1 knockout mice show masculinized gonads. Molecularanalyses demonstrate an absence of female-specific activationof Wnt4 and as a consequence XY-like vascularization and steroidogenesis.Moreover, germ cells of XX knockout embryos show changes incellular adhesions and a failure to enter XX specific meiosis.Sex cords develop around birth, when Sox9 becomes strongly activated.Thus, a balance between Sox9 and β-catenin activation determinesthe fate of the gonad, with Rspo1 acting as a crucial regulatorof canonical β-catenin signaling required for female development.     

MicroRNAs regulate T‐cell production of interleukin‐9 and identify hypoxia‐inducible factor‐2α as an important regulator of T helper 9 and regulatory T‐cell differentiation

MicroRNAs (miRNAs) regulate many aspects of helper T cell (Th) development and function. Here we found that they are required for the suppression of interleukin‐9 (IL‐9) expression in Th9 cells and other Th subsets. Two highly related miRNAs (miR‐15b and miR‐16) that we previously found to play an important role in regulatory T (Treg) cell differentiation were capable of suppressing IL‐9 expression when they were over‐expressed in Th9 cells. We used these miRNAs as tools to identify novel regulators of IL‐9 expression and found that they could regulate the expression of Epas1, which encodes hypoxia‐inducible factor (HIF)‐2α. HIF proteins regulate metabolic pathway usage that is important in determining appropriate Th differentiation. The related protein, HIF‐1α enhances Th17 differentiation and inhibits Treg cell differentiation. Here we found that HIF‐2α was required for IL‐9 expression in Th9 cells, but its expression was not sufficient in other Th subsets. Furthermore, HIF‐2α suppressed Treg cell differentiation like HIF‐1α, demonstrating both similar and distinct roles of the HIF proteins in Th differentiation and adding a further dimension to their function. Ironically, even though miR‐15b and miR‐16 suppressed HIF‐2α expression in Treg cells, inhibiting their function in Treg cells did not lead to an increase in IL‐9 expression. Therefore, the physiologically relevant miRNAs that regulate IL‐9 expression in Treg cells and other subsets remain unknown. Nevertheless, the analysis of miR‐15b and miR‐16 function led to the discovery of the importance of HIF‐2α so this work demonstrated the utility of studying miRNA function to identify novel regulatory pathways in helper T‐cell development.     

Desert Hedgehog/Patched 1 signaling specifies fetal Leydig cell fate in testis organogenesis

Establishment of the steroid-producing Leydig cell lineage is an event downstream of Sry that is critical for masculinization of mammalian embryos. Neither the origin of fetal Leydig cell precursors nor the signaling pathway that specifies the Leydig cell lineage is known. Based on the sex-specific expression patterns of Desert Hedgehog (Dhh) and its receptor Patched 1 (Ptch1) in XY gonads, we investigated the potential role of DHH/PTCH1 signaling in the origin and specification of fetal Leydig cells. Analysis of Dhh(-/-) XY gonads revealed that differentiation of fetal Leydig cells was severely defective. Defects in Leydig cell differentiation in Dhh(-/-) XY gonads did not result from failure of cell migration from the mesonephros, thought to be a possible source of Leydig cell precursors. Nor did DHH/PTCH1 signaling appear to be involved in the proliferation or survival of fetal Leydig precursors in the interstitium of the XY gonad. Instead, our results suggest that DHH/PTCH1 signaling triggers Leydig cell differentiation by up-regulating Steroidogenic Factor 1 and P450 Side Chain Cleavage enzyme expression in Ptch1-expressing precursor cells located outside testis cords.     

A five‐gene signature as a potential predictor of metastasis and survival in colorectal cancer

To understand the molecular mechanisms of metastasis and prognosis of colorectal cancer (CRC), we isolated single cell‐derived progenies (SCPs) from SW480 cells in vitro and compared their metastatic potential in an orthotopic CRC tumour model in vivo. Two groups of SCPs with the capability of high and low metastasis, respectively, were obtained. By analysing the gene expression profiles of high (SCP51), low (SCP58) metastatic SCPs, and their parental cell line (SW480/EGFP), we demonstrated that 143 genes were differentially expressed either between SCP51 and SCP58 or between SCP58 and SW480/EGFP. Gene‐annotation enrichment analysis of DAVID revealed 80 genes in the top ten clusters of the analysis (gene enrichment score > 1). Of the 80‐gene set, 32 genes are potentially involved in metastasis, as revealed by Geneclip. Five putative metastatic genes (LYN, SDCBP, MAP4K4, DKK1, and MID1) were selected for further validations. Immunohistochemical analysis in a cohort of 181 CRC clinical samples showed that the individual expression of LYN, MAP4K4, and MID1, as well as the five‐gene signature, was closely correlated with lymph node metastasis in CRC patients. More importantly, the individual expression of LYN, MAP4K4, SDCBP, and MID1, as well as the five‐gene signature, was significantly correlated with overall survival in CRC patients. Thus, our five‐gene signature may be able to predict metastasis and survival of CRC in the clinic, and opens new perspectives on the biology of CRC. Copyright © 2009 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Immunisation route‐dependent expression of IL‐4/IL‐13 can modulate HIV‐specific CD8+ CTL avidity

All HIV‐1 ‘systemic vaccine trials’ in humans have yielded poor outcomes. Thus, it is important to understand whether the route of delivery influences the quality of protective CTL immunity. Using heterologous poxvirus immunisation we have shown that systemically (i.m./i.m.) immunised CD8⁺ T cells generated higher levels of IL‐4/IL‐13 compared to mucosal delivery and expression also correlated with i.m./i.m. immunised mice eliciting CTL of lower avidity. Studies using IL‐4^?/? and IL‐13^?/? KO mice have shown that the capacity to express IFN‐γ, IL‐4 and/or IL‐13 by K^dGag_197–205‐specific CTL differed between these groups and was inversely correlated with CTL avidity (IL‐13^?/?>IL‐4^?/?>BALB/c), although no significant differences in the magnitude of CTL responses were observed between IL‐13^?/? and wild type mice. When IL‐13 was reconstituted in IL‐13^?/? splenocytes in vitro, their ability to bind tetramers also decreased significantly. Our data reveal that total absence of IL‐13 can greatly enhance CTL avidity. In contrast, extracellular IL‐4 appears to be important in maintaining long‐term Th1/Th2 balance in CTL, even though expression of IL‐4 by CTL markedly reduced avidity. STAT6^?/? mice also showed memory CTL of higher avidity. Furthermore, CCL5 expression in K^dGag_197–205‐specific CTL was also regulated by IL‐4/IL‐13.     

Ph‐like acute lymphoblastic leukemia with a novel PAX5‐KIDINS220 fusion transcript

Although “paired box 5” (PAX5)‐related fusion genes are well documented in childhood B‐cell precursor acute lymphoblastic leukemia (ALL), these types of fusion with the exception of PAX5‐JAK2 are rarely seen in patients with gene expression profiles similar to those of BCR‐ABL1 (Philadelphia)‐positive ALL (Ph‐like ALL). We report a novel fusion of the genes PAX5 and “kinase D‐interacting substrate of 220 kDa” (KIDINS220, also known as ARMS) in a Ph‐like ALL. As PAX5 is a master regulator of B‐lymphocyte differentiation, PAX5 rearrangements induce a differentiation block in B lymphocytes. KIDINS220 is a mediator of multiple receptor signaling pathways, interacts with both T‐ and B‐cell receptors, and is necessary for sustained extracellular signal‐regulated kinase (ERK) signaling. Although functional studies are needed, the PAX5‐KIDINS220 fusion protein might not only inhibit wild‐type PAX5 function, but also promote sustained activation of the ERK signaling pathway through upregulation of KIDINS220. © 2016 Wiley Periodicals, Inc.