Association of impaired neuronal migration with cognitive deficits in extremely preterm infants

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Abstract

Many extremely preterm infants (born before 28 gestational weeks [GWs]) develop cognitive impairment in later life, although the underlying pathogenesis is not yet completely understood. Our examinations of the developing human neocortex confirmed that neuronal migration continues beyond 23 GWs, the gestational week at which extremely preterm infants have live births. We observed larger numbers of ectopic neurons in the white matter of the neocortex in human extremely preterm infants with brain injury and hypothesized that altered neuronal migration may be associated with cognitive impairment in later life. To confirm whether preterm brain injury affects neuronal migration, we produced brain damage in mouse embryos by occluding the maternal uterine arteries. The mice showed delayed neuronal migration, ectopic neurons in the white matter, altered neuronal alignment, and abnormal corticocortical axonal wiring. Similar to human extremely preterm infants with brain injury, the surviving mice exhibited cognitive deficits. Activation of the affected medial prefrontal cortices of the surviving mice improved working memory deficits, indicating that decreased neuronal activity caused the cognitive deficits. These findings suggest that altered neuronal migration altered by brain injury might contribute to the subsequent development of cognitive impairment in extremely preterm infants.

Authors

Ken-ichiro Kubo, Kimiko Deguchi, Taku Nagai, Yukiko Ito, Keitaro Yoshida, Toshihiro Endo, Seico Benner, Wei Shan, Ayako Kitazawa, Michihiko Aramaki, Kazuhiro Ishii, Minkyung Shin, Yuki Matsunaga, Kanehiro Hayashi, Masaki Kakeyama, Chiharu Tohyama, Kenji F. Tanaka, Kohichi Tanaka, Sachio Takashima, Masahiro Nakayama, Masayuki Itoh, Yukio Hirata, Barbara Antalffy, Dawna D. Armstrong, Kiyofumi Yamada, Ken Inoue, Kazunori Nakajima

p63⁺ ureteric bud tip cells are progenitors of intercalated cells

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Abstract

During renal branching morphogenesis, ureteric bud tip cells (UBTC) serve as the progenitor epithelium for all cell types of the collecting duct. While the transcriptional circuitry of ureteric bud (UB) branching has been intensively studied, the transcriptional control of UBTC differentiation has been difficult to ascertain. This is partly due to limited knowledge of UBTC-specific transcription factors that mark the progenitor state. Here, we identify the transcription factor p63 (also known as TP63), a master regulator of basal stem cells in stratified epithelia, as a specific marker of mouse and human UBTC. Nuclear p63 marks Ret⁺ UBTC transiently and is silenced by the end of nephrogenesis. Lineage tracing revealed that a subset of UBTC expressing the ΔNp63 isoform (N-terminus truncated p63) is dedicated to generating cortical intercalated cells. Germline targeting of ΔNp63 in mice caused a marked reduction in intercalated cells near the time of birth, indicating that p63 not only marks UBTC, but also is essential for their differentiation. We conclude that the choice of UBTC progenitors to differentiate is determined earlier than previously recognized and that UBTC progenitors are prepatterned and fate restricted. These findings prompt the rethinking of current paradigms of collecting duct differentiation and may have implications for regenerative renal medicine.

Authors

Samir S. El-Dahr, Yuwen Li, Jiao Liu, Elleny Gutierrez, Kathleen S. Hering-Smith, Sabina Signoretti, Jean-Christophe Pignon, Satrajit Sinha, Zubaida Saifudeen

Point mutations in murine Nkx2-5 phenocopy human congenital heart disease and induce pathogenic Wnt signaling

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Abstract

Mutations in the Nkx2-5 gene are a main cause of congenital heart disease. Several studies have addressed the phenotypic consequences of disrupting the Nkx2-5 gene locus, although animal models to date failed to recapitulate the full spectrum of the human disease. Here, we describe a new Nkx2-5 point mutation murine model, akin to its human counterpart disease–generating mutation. Our model fully reproduces the morphological and physiological clinical presentations of the disease and reveals an understudied aspect of Nkx2-5–driven pathology, a primary right ventricular dysfunction. We further describe the molecular consequences of disrupting the transcriptional network regulated by Nkx2-5 in the heart and show that Nkx2-5–dependent perturbation of the Wnt signaling pathway promotes heart dysfunction through alteration of cardiomyocyte metabolism. Our data provide mechanistic insights on how Nkx2-5 regulates heart function and metabolism, a link in the study of congenital heart disease, and confirms that our models are the first murine genetic models to our knowledge to present all spectra of clinically relevant adult congenital heart disease phenotypes generated by NKX2-5 mutations in patients.

Authors

Milena B. Furtado, Julia C. Wilmanns, Anjana Chandran, Joelle Perera, Olivia Hon, Christine Biben, Taylor J. Willow, Hieu T. Nim, Gurpreet Kaur, Stephanie Simonds, Qizhu Wu, David Willians, Ekaterina Salimova, Nicolas Plachta, James M. Denegre, Stephen A. Murray, Diane Fatkin, Michael Cowley, James T. Pearson, David Kaye, Mirana Ramialison, Richard P. Harvey, Nadia A. Rosenthal, Mauro W. Costa

Eomesodermin and T-bet mark developmentally distinct human natural killer cells

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Abstract

Immaturity of the immune system of human fetuses and neonates is often invoked to explain their increased susceptibility to infection; however, the development of the fetal innate immune system in early life remains incompletely explored. We now show that the most mature NK cells found in adult (or postnatal) human circulation (CD94^–CD16⁺) are absent during ontogeny. Human fetal NK cells were found to express the 2 signature T-box transcription factors essential for the development of all murine NK and NK-like cells, eomesodermin (Eomes) and T-bet. The single-cell pattern of Eomes and T-bet expression during ontogeny, however, revealed a stereotyped pattern of reciprocal dominance, with immature NK cells expressing higher amounts of Eomes and more mature NK cells marked by greater abundance of T-bet. We also observed a stereotyped pattern of tissue-specific NK cell maturation during human ontogeny, with fetal liver being more restrictive to NK cell maturity than fetal bone barrow, spleen, or lung. These results support the hypothesis that maturation of human NK cells has a discrete restriction until postnatal life, and provide a framework to better understand the increased susceptibility of fetuses and newborns to infection.

Authors

Amélie Collins, Nyanza Rothman, Kang Liu, Steven L. Reiner

Alterations in sarcomere function modify the hyperplastic to hypertrophic transition phase of mammalian cardiomyocyte development

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Abstract

It remains unclear how perturbations in cardiomyocyte sarcomere function alter postnatal heart development. We utilized murine models that allowed manipulation of cardiac myosin-binding protein C (MYBPC3) expression at critical stages of cardiac ontogeny to study the response of the postnatal heart to disrupted sarcomere function. We discovered that the hyperplastic to hypertrophic transition phase of mammalian heart development was altered in mice lacking MYBPC3 and this was the critical period for subsequent development of cardiomyopathy. Specifically, MYBPC3-null hearts developed evidence of increased cardiomyocyte endoreplication, which was accompanied by enhanced expression of cell cycle stimulatory cyclins and increased phosphorylation of retinoblastoma protein. Interestingly, this response was self-limited at later developmental time points by an upregulation of the cyclin-dependent kinase inhibitor p21. These results provide valuable insights into how alterations in sarcomere protein function modify postnatal heart development and highlight the potential for targeting cell cycle regulatory pathways to counteract cardiomyopathic stimuli.

Authors

Benjamin R. Nixon, Alexandra F. Williams, Michael S. Glennon, Alejandro E. de Feria, Sara C. Sebag, H. Scott Baldwin, Jason R. Becker

Repair after nephron ablation reveals limitations of neonatal neonephrogenesis

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Abstract

The neonatal mouse kidney retains nephron progenitor cells in a nephrogenic zone for 3 days after birth. We evaluated whether de novo nephrogenesis can be induced postnatally beyond 3 days. Given the long-term implications of nephron number for kidney health, it would be useful to enhance nephrogenesis in the neonate. We induced nephron reduction by cryoinjury with or without contralateral nephrectomy during the neonatal period or after 1 week of age. There was no detectable compensatory de novo nephrogenesis, as determined by glomerular counting and lineage tracing. Contralateral nephrectomy resulted in additional adaptive healing, with little or no fibrosis, but did not also stimulate de novo nephrogenesis. In contrast, injury initiated at 1 week of age led to healing with fibrosis. Thus, despite the presence of progenitor cells and ongoing nephron maturation in the newborn mouse kidney, de novo nephrogenesis is not inducible by acute nephron reduction. This indicates that additional nephron progenitors cannot be recruited after birth despite partial renal ablation providing a reparative stimulus and suggests that nephron number in the mouse is predetermined at birth.

Authors

Florian Tögel, M. Todd Valerius, Benjamin S. Freedman, Rossella Latrino, Mor Grinstein, Joseph V. Bonventre

A xenogeneic-free system generating functional human gut organoids from pluripotent stem cells

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Abstract

Functional intestines are composed of cell types from all 3 primary germ layers and are generated through a highly orchestrated and serial developmental process. Directed differentiation of human pluripotent stem cells (hPSCs) has been shown to yield gut-specific cell types; however, these structures do not reproduce critical functional interactions between cell types of different germ layers. Here, we developed a simple protocol for the generation of mature functional intestinal organoids from hPSCs under xenogeneic-free conditions. The stem cell–derived gut organoids produced here were found to contain distinct types of intestinal cells, including enterocytes, goblet cells, Paneth cells, and enteroendocrine cells, that were derived from all 3 germ layers; moreover, they demonstrated intestinal functions, including peptide absorption, and showed innervated bowel movements in response to stimulation with histamine and anticholinergic drugs. Importantly, the gut organoids obtained using this xenogeneic-free system could be stably maintained in culture for prolonged periods and were successfully engrafted in vivo. Our xenogeneic-free approach for generating gut organoids from hPSCs provides a platform for studying human intestinal diseases and for pharmacological testing.

Authors

Hajime Uchida, Masakazu Machida, Takumi Miura, Tomoyuki Kawasaki, Takuya Okazaki, Kengo Sasaki, Seisuke Sakamoto, Noriaki Ohuchi, Mureo Kasahara, Akihiro Umezawa, Hidenori Akutsu

Gene pathway development in human epicardial adipose tissue during early life

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Abstract

Studies in rodents and newborn humans demonstrate the influence of brown adipose tissue (BAT) in temperature control and energy balance and a critical role in the regulation of body weight. Here, we obtained samples of epicardial adipose tissue (EAT) from neonates, infants, and children in order to evaluate changes in their transcriptional landscape by applying a systems biology approach. Surprisingly, these analyses revealed that the transition to infancy is a critical stage for changes in the morphology of EAT and is reflected in unique gene expression patterns of a substantial proportion of thermogenic gene transcripts (~10%). Our results also indicated that the pattern of gene expression represents a distinct developmental stage, even after the rebound in abundance of thermogenic genes in later childhood. Using weighted gene coexpression network analyses, we found precise anthropometric-specific correlations with changes in gene expression and the decline of thermogenic capacity within EAT. In addition, these results indicate a sequential order of transcriptional events affecting cellular pathways, which could potentially explain the variation in the amount, or activity, of BAT in adulthood. Together, these results provide a resource to elucidate gene regulatory mechanisms underlying the progressive development of BAT during early life.

Authors

Shalini Ojha, Hernan P. Fainberg, Victoria Wilson, Giuseppe Pelella, Marcos Castellanos, Sean T. May, Attilio A. Lotto, Harold Sacks, Michael E. Symonds, Helen Budge

Epidermal CYLD inactivation sensitizes mice to the development of sebaceous and basaloid skin tumors

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Abstract

The deubiquitinase-encoding gene Cyld displays a dominant genetic linkage to a wide spectrum of skin-appendage tumors, which could be collectively designated as CYLD mutant–syndrome (CYLD^m-syndrome). Despite recent advances, little is understood about the molecular mechanisms responsible for this painful and difficult-to-treat skin disease. Here, we generated a conditional mouse model with epidermis-targeted expression of a catalytically deficient CYLD^m through K14-Cre–mediated deletion of exon 9 (hereafter refer to Cyld^EΔ9/Δ9). Cyld^EΔ9/Δ9 mice were born alive but developed hair and sebaceous gland abnormalities and dental defects at 100% and 60% penetrance, respectively. Upon topical challenge with DMBA/TPA, these animals primarily developed sebaceous and basaloid tumors resembling human CYLD^m-syndrome as opposed to papilloma, which is most commonly induced in WT mice by this treatment. Molecular analysis revealed that TRAF6-K63-Ubiquitination (K63-Ub), c-Myc-K63-Ub, and phospho-c–Myc (S62) were markedly elevated in Cyld^EΔ9/Δ9 skin. Topical treatment with a pharmacological c-Myc inhibitor induced sebaceous and basal cell apoptosis in Cyld^EΔ9/Δ9 skin. Consistently, c-Myc activation was readily detected in human cylindroma and sebaceous adenoma. Taken together, our findings demonstrate that Cyld^EΔ9/Δ9 mice represent a disease-relevant animal model and identify TRAF6 and c-Myc as potential therapeutic targets for CYLD^m-syndrome.

Authors

Yingai Jane Jin, Sally Wang, Joshua Cho, M. Angelica Selim, Tim Wright, George Mosialos, Jennifer Y. Zhang

Restoration of lymphatic function rescues obesity in Prox1-haploinsufficient mice

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Abstract

Prox1 heterozygous mice have a defective lymphatic vasculature and develop late-onset obesity. Chyle abnormally leaks from those vessels, accumulates in the surrounding tissues, and causes an increase in adipose tissue. We characterized the lymphatics of Prox1^+/– mice to determine whether the extent of obesity correlated with the severity of lymphatic defects. The lymphatic vasculature in Prox1^+/– mice exhibited reduced tracer clearance from the ear skin, dysfunctional perfusion of the lower legs, and reduced tracer uptake into the deep lymphatic collectors during mechanostimulation prior to the onset of obesity. Ear lymphatic vessels and leg collectors in Prox1^+/– mice were disorganized and irregular, further confirming that defective lymphatic vessels are associated with obesity in Prox1^+/– mice. We now provide conclusive in vivo evidence that demonstrates that leaky lymphatics mediate obesity in Prox1^+/– mice, as restoration of lymphatic vasculature function was sufficient to rescue the obesity features in Prox1^+/– mice. Finally, depth-lipomic profiling of lymph contents showed that free fatty acids induce adipogenesis in vitro.

Authors

Noelia Escobedo, Steven T. Proulx, Sinem Karaman, Miriam E. Dillard, Nicole Johnson, Michael Detmar, Guillermo Oliver