Classical research has suggested that early palate formation develops via epithelial-mesenchymal interactions, and in this study we reveal which signals control this process. Using Fgf10–/–, FGF receptor 2b–/– (Fgfr2b–/–), and Sonic hedgehog (Shh) mutant mice, which all exhibit cleft palate, we show that Shh is a downstream target of Fgf10/Fgfr2b signaling. Our results demonstrate that mesenchymal Fgf10 regulates the epithelial expression of Shh, which in turn signals back to the mesenchyme. This was confirmed by demonstrating that cell proliferation is decreased not only in the palatal epithelium but also in the mesenchyme of Fgfr2b–/– mice. These results reveal a new role for Fgf signaling in mammalian palate development. We show that coordinated epithelial-mesenchymal interactions are essential during the initial stages of palate development and require an Fgf-Shh signaling network.
Ritva Rice, Bradley Spencer-Dene, Elaine C. Connor, Amel Gritli-Linde, Andrew P. McMahon, Clive Dickson, Irma Thesleff, David P.C. Rice
Congenital obstructive nephropathy is the principal cause of renal failure in infants and children. The underlying molecular and cellular mechanisms of this disease, however, remain largely undetermined. We generated a mouse model of congenital obstructive nephropathy that resembles ureteropelvic junction obstruction in humans. In these mice, calcineurin function is removed by the selective deletion of Cnb1 in the mesenchyme of the developing urinary tract using the Cre/lox system. This deletion results in reduced proliferation in the smooth muscle cells and other mesenchymal cells in the developing urinary tract. Compromised cell proliferation causes abnormal development of the renal pelvis and ureter, leading to defective pyeloureteral peristalsis, progressive renal obstruction, and, eventually, fatal renal failure. Our study demonstrates that calcineurin is an essential signaling molecule in urinary tract development and is required for normal proliferation of the urinary tract mesenchymal cells in a cell-autonomous manner. These studies also emphasize the importance of functional obstruction, resulting from developmental abnormality, in causing congenital obstructive nephropathy.
Ching-Pin Chang, Bradley W. McDill, Joel R. Neilson, Heidi E. Joist, Jonathan A. Epstein, Gerald R. Crabtree, Feng Chen
Embryo liver morphogenesis takes place after gastrulation and starts with a ventral foregut evagination that reacts to factor signaling from both cardiac mesoderm and septum transversum mesenchyme. Current knowledge of the progenitor stem cell populations involved in this early embryo liver development is scarce. We describe here a population of 11-day postcoitus c-Kitlow(CD45/TER119)– liver progenitors that selectively expressed hepatospecific genes and proteins in vivo, was self-maintained in vitro by long-term proliferation, and simultaneously differentiated into functional hepatocytes and bile duct cells. Purified c-Kitlow(CD45/TER119)– liver cells cocultured with cell-depleted fetal liver fragments engrafted and repopulated the hepatic cell compartments of the latter organoids, suggesting that they may include the embryonic stem cells responsible for liver development.
Susana Minguet, Isabel Cortegano, Pilar Gonzalo, José-Alberto Martínez-Marin, Belén de Andrés, Clara Salas, David Melero, Maria-Luisa Gaspar, Miguel A.R. Marcos
Kidney disease affects over 20 million people in the United States alone. Although the causes of renal failure are diverse, the glomerular filtration barrier is often the target of injury. Dysregulation of VEGF expression within the glomerulus has been demonstrated in a wide range of primary and acquired renal diseases, although the significance of these changes is unknown. In the glomerulus, VEGF-A is highly expressed in podocytes that make up a major portion of the barrier between the blood and urinary spaces. In this paper, we show that glomerular-selective deletion or overexpression of VEGF-A leads to glomerular disease in mice. Podocyte-specific heterozygosity for VEGF-A resulted in renal disease by 2.5 weeks of age, characterized by proteinuria and endotheliosis, the renal lesion seen in preeclampsia. Homozygous deletion of VEGF-A in glomeruli resulted in perinatal lethality. Mutant kidneys failed to develop a filtration barrier due to defects in endothelial cell migration, differentiation, and survival. In contrast, podocyte-specific overexpression of the VEGF-164 isoform led to a striking collapsing glomerulopathy, the lesion seen in HIV-associated nephropathy. Our data demonstrate that tight regulation of VEGF-A signaling is critical for establishment and maintenance of the glomerular filtration barrier and strongly supports a pivotal role for VEGF-A in renal disease.
Vera Eremina, Manish Sood, Jody Haigh, András Nagy, Ginette Lajoie, Napoleone Ferrara, Hans-Peter Gerber, Yamato Kikkawa, Jeffrey H. Miner, Susan E. Quaggin
Preadipocyte factor-1 (Pref-1) is a transmembrane protein highly expressed in preadipocytes. Pref-1 expression is, however, completely abolished in adipocytes. The extracellular domain of Pref-1 undergoes two proteolytic cleavage events that generate 50 and 25 kDa soluble products. To understand the function of Pref-1, we generated transgenic mice that express the full ectodomain corresponding to the large cleavage product of Pref-1 fused to human immunoglobulin-γ constant region. Mice expressing the Pref-1/hFc transgene in adipose tissue, driven by the adipocyte fatty acid–binding protein (aP2, also known as aFABP) promoter, showed a substantial decrease in total fat pad weight. Moreover, adipose tissue from transgenic mice showed reduced expression of adipocyte markers and adipocyte-secreted factors, including leptin and adiponectin, whereas the preadipocyte marker Pref-1 was increased. Pref-1 transgenic mice with a substantial, but not complete, loss of adipose tissue exhibited hypertriglyceridemia, impaired glucose tolerance, and decreased insulin sensitivity. Mice expressing the Pref-1/hFc transgene exclusively in liver under the control of the albumin promoter also showed a decrease in adipose mass and adipocyte marker expression, suggesting an endocrine mode of action of Pref-1. These findings demonstrate the inhibition of adipogenesis by Pref-1 in vivo and the resulting impairment of adipocyte function that leads to the development of metabolic abnormalities.
Kichoon Lee, Josep A. Villena, Yang Soo Moon, Kee-Hong Kim, Sunjoo Lee, Chulho Kang, Hei Sook Sul
Christine Fritsch, Elzbieta A. Swietlicki, Olivier Lefebvre, Michele Kedinger, Hristo Iordanov, Marc S. Levin, Deborah C. Rubin
Akiyoshi Uemura, Minetaro Ogawa, Masanori Hirashima, Takashi Fujiwara, Shinji Koyama, Hitoshi Takagi, Yoshihito Honda, Stanley J. Wiegand, George D. Yancopoulos, Shin-Ichi Nishikawa
Wolfgang E. Gallwitz, Theresa A. Guise, Gregory R. Mundy
Lydie Cassard, Joël F.G. Cohen-Solal, Annie Galinha, Xavier Sastre-Garau, Claire Mathiot, Jérôme Galon, Thierry Dorval, Alain Bernheim, Wolf H. Fridman, Catherine Sautès-Fridman
Dipak Panigrahy, Samuel Singer, Lucy Q. Shen, Catherine E. Butterfield, Deborah A. Freedman, Emy J. Chen, Marsha A. Moses, Susan Kilroy, Stefan Duensing, Christopher Fletcher, Jonathan A. Fletcher, Lynn Hlatky, Philip Hahnfeldt, Judah Folkman, Arja Kaipainen
Sumayah Jamal, Robert J. Schneider
Flavio Curnis, Angelina Sacchi, Angelo Corti
Dinesh S. Rao, Teresa S. Hyun, Priti D. Kumar, Ikuko F. Mizukami, Mark A. Rubin, Peter C. Lucas, Martin G. Sanda, Theodora S. Ross
April L. Blajeski, Vy A. Phan, Timothy J. Kottke, Scott H. Kaufmann
TGF-βs are potent inhibitors of epithelial cell proliferation. However, in established carcinomas, autocrine/paracrine TGF-β interactions can enhance tumor cell viability and progression. Thus, we studied the effect of a soluble Fc:TGF-β type II receptor fusion protein (Fc:TβRII) on transgenic and transplantable models of breast cancer metastases. Systemic administration of Fc:TβRII did not alter primary mammary tumor latency in MMTV-Polyomavirus middle T antigen transgenic mice. However, Fc:TβRII increased apoptosis in primary tumors, while reducing tumor cell motility, intravasation, and lung metastases. These effects correlated with inhibition of Akt activity and FKHRL1 phosphorylation. Fc:TβRII also inhibited metastases from transplanted 4T1 and EMT-6 mammary tumors in syngeneic BALB/c mice. Tumor microvessel density in a mouse dorsal skin window chamber was unaffected by Fc:TβRII. Therefore, blockade of TGF-β signaling may reduce tumor cell viability and migratory potential and represents a testable therapeutic approach against metastatic carcinomas.
Rebecca S. Muraoka, Nancy Dumont, Christoph A. Ritter, Teresa C. Dugger, Dana M. Brantley, Jin Chen, Evangeline Easterly, L. Renee Roebuck, Sarah Ryan, Philip J. Gotwals, Victor Koteliansky, Carlos L. Arteaga
Whole-body irradiation at the minimal lethal dose causes bone marrow failure and death within 12–18 days. To identify the principal components of the hematopoietic system that are radioprotective, we transplanted lethally irradiated mice with purified progenitors: common myeloid progenitors (CMPs), megakaryocyte/erythrocyte-restricted progenitors (MEPs), or granulocyte/monocyte-restricted progenitors (GMPs). Transplanted CMPs gave rise to cells both of the granulocyte/monocyte (GM) series and the megakaryocyte/erythrocyte series, whereas GMPs or MEPs showed reconstitution of only GM or ME cells, respectively. CMPs and MEPs but not GMPs protected mice in a dose-dependent manner, suggesting that erythrocytes, platelets, or both are the critical effectors of radioprotection. Accordingly, CMPs and MEPs formed robust colonies in recipient bone marrow and spleen, whereas GMPs formed small colonies that rapidly disappeared. Direct comparisons of spleen CFU (CFU-S) potentials among each progenitor subset showed that MEPs contain the vast majority of day 8 CFU-S activity, suggesting that day 8 CFU-S are the precursors of radioprotective cell subsets. All animals radioprotected for 30 days subsequently survived for at least 6 months post-transplant, and showed only host-derived hematopoiesis after 30 days. These findings suggest that rare hematopoietic stem cells survive myeloablation that can eventually repopulate irradiated hosts if myeloerythroid-restricted progenitors transiently rescue ablated animals through the critical window of bone marrow failure.
Thanyaphong Na Nakorn, David Traver, Irving L. Weissman, Koichi Akashi
TGF-βs play diverse and complex roles in many biological processes. In tumorigenesis, they can function either as tumor suppressors or as pro-oncogenic factors, depending on the stage of the disease. We have developed transgenic mice expressing a TGF-β antagonist of the soluble type II TGF-β receptor:Fc fusion protein class, under the regulation of the mammary-selective MMTV-LTR promoter/enhancer. Biologically significant levels of antagonist were detectable in the serum and most tissues of this mouse line. The mice were resistant to the development of metastases at multiple organ sites when compared with wild-type controls, both in a tail vein metastasis assay using isogenic melanoma cells and in crosses with the MMTV-neu transgenic mouse model of metastatic breast cancer. Importantly, metastasis from endogenous mammary tumors was suppressed without any enhancement of primary tumorigenesis. Furthermore, aged transgenic mice did not exhibit the severe pathology characteristic of TGF-β null mice, despite lifetime exposure to the antagonist. The data suggest that in vivo the antagonist may selectively neutralize the undesirable TGF-β associated with metastasis, while sparing the regulatory roles of TGF-βs in normal tissues. Thus this soluble TGF-β antagonist has potential for long-term clinical use in the prevention of metastasis.
Yu-an Yang, Oksana Dukhanina, Binwu Tang, Mizuko Mamura, John J. Letterio, Jennifer MacGregor, Sejal C. Patel, Shahram Khozin, Zi-yao Liu, Jeffrey Green, Miriam R. Anver, Glenn Merlino, Lalage M. Wakefield
In gastrointestinal epithelium, metaplastic conversion between predominant cell types is associated with an increased risk of neoplasia. However, the mechanisms regulating metaplastic transitions in adult epithelia are largely undefined. Here we show that matrix metalloproteinase-7 (MMP-7) is expressed not only in the majority of human pancreatic ductal adenocarcinoma specimens, but also in human pancreatic intraepithelial neoplasia and metaplastic duct lesions in human and mouse. In a mouse model of pancreatic acinar-to-ductal metaplasia, MMP-7 progressively accumulates during the metaplastic transition, resulting in a concomitant increase in solubilization of Fas ligand (FasL). Under identical conditions, mice either deficient in MMP-7 or carrying an inactive FasL gene are severely inhibited in development of progressive metaplasia and acinar cell apoptosis. Thus, MMP-7 and FasL influence the initiation and maintenance of metaplastic events in pancreatic epithelium, explaining the observed link between metaplasia and apoptosis in pancreas and other gastrointestinal tissues.
Howard C. Crawford, Charles R. Scoggins, M. Kay Washington, Lynn M. Matrisian, Steven D. Leach
Terminal epithelial cell differentiation is a crucial step in development. In the kidney, failure of terminal differentiation causes dysplasia, cystogenesis, and cancer. The present study provides multiple lines of evidence implicating the tumor suppressor protein p53 in terminal differentiation of the renal epithelium. In the developing kidney, p53 is highly enriched in epithelial cells expressing renal function genes (RFGs), such as receptors for vasoactive hormones, the sodium pump, and epithelial sodium and water channels. In comparison, proliferating renal progenitors express little if any p53 or RFGs. p53 binds to and transactivates the promoters of RFGs. In contrast, expression of a dominant negative mutant form of p53 inhibits endogenous RFG expression. Moreover, binding of endogenous p53 to the promoters of RFGs coincides with the differentiation process and is attenuated once renal epithelial cells are fully differentiated. Finally, p53-null pups exhibit a previously unrecognized aberrant renal phenotype and spatial disorganization of RFGs. Interestingly, the p53-related protein p73 is unable to functionally compensate for the loss of p53 and fails to efficiently activate RFG transcription. We conclude that p53 promotes the biochemical and morphological differentiation of the renal epithelium. Aberrations in p53-mediated terminal differentiation may therefore play a role in the pathogenesis of nephron dysgenesis and dysfunction.
Zubaida Saifudeen, Susana Dipp, Samir S. El-Dahr
Myb34.5 is a herpes simplex virus 1 (HSV-1) mutant deleted in the gene for ribonucleotide reductase (ICP6). It also carries a version of γ134.5 (a viral gene product that promotes the dephosphorylation of eIF-2α) that is under control of the E2F-responsive cellular B-myb promoter, rather than of its endogenous promoter. Myb34.5 replication in tumor cells results in their destruction (oncolysis). γ134.5 expression by HSV-1 subverts an important cell defense mechanism against viral replication by preventing shutoff of protein synthesis after viral infection. Infection of colon carcinoma cells with Myb34.5 results in greater eIF-2α dephosphorylation and viral replication compared with infection with HSV-1 mutants completely defective in γ134.5 expression. In contrast, infection of normal hepatocytes with Myb34.5 results in low levels of eIF-2α dephosphorylation and viral replication that are similar to those observed with HSV-1 mutants completely defective in γ134.5 and ICP6. When administered intravascularly into mice with diffuse liver metastases, Myb34.5 has greater antineoplastic activity than HSV-1 mutants with completely defective γ134.5 expression and more restricted biodistribution compared with HSV-1 mutants with wild-type γ134.5 expression. Myb34.5 displays reduced virulence and toxicity compared to HSV-1 mutants with wild-type γ134.5 expression. Portal venous administration of Myb34.5 significantly reduces liver tumor burden in and prolongs the life of mice with diffuse liver metastases. Preexisting Ab’s to HSV-1 do not reduce the antitumor efficacy of Myb34.5 in vivo.
Hideo Nakamura, Hideki Kasuya, John T. Mullen, Sam S. Yoon, Timothy M. Pawlik, Soundararajalu Chandrasekhar, James M. Donahue, E. Antonio Chiocca, Richard Y. Chung, Kenneth K. Tanabe
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