Cdc42 is crucial for facial and palatal formation during craniofacial development

Abstract

Craniofacial deformities with multifactorial etiologies, such as cleft palate and facial dysmorphism, represent some of the most frequent congenital birth defects seen in humans. Their pathogeneses are often related to cranial neural crest (CNC) cells. During CNC cell migration, changes in cell shape and formation, as well as maintenance of subcellular structures, such as filopodia and lamellipodia, are dependent on the complex functions of Rho family small GTPases, which are regulators of actin cytoskeletal organization. Cdc42, a member of the Rho family of small GTPases, is known to play critical roles in organogenesis of various tissues. To investigate the physiological functions of Cdc42 during craniofacial development, we generated CNC-derived cell-specific inactivated Cdc42 mutant mice (Cdc42fl/fl;P0-cre). Most of the Cdc42fl/fl;P0-cre neonates were viable at birth, though they appeared weaker and no milk was found in their stomachs, and all died within a few days. They had a short face and intracranial bleeding, and abnormal calcification of the cranium. Cdc42fl/fl;P0-cre neonates also demonstrated a cleft palate and there was no fusion of the secondary palate because of failure of palatal shelf elongation for the process of palate closure. Cdc42 is crucial for facial and palatal formation during craniofacial development.

A reliable murine model of bone metastasis by injecting cancer cells through caudal arteries

“…For CA injection, LLC/luc (2 × 10 5 or 1 × 10 6 cells to male C57B/6), MDA-MB-231/luc (5 × 10 5 cells to female NOD-SCID), PC-3/luc (1 × 10 6 cells to male SCID), 786-O/luc (1 × 10 6 cells to male BALB/c-nu), 143B/luc (5 × 10 5 cells to male BALB/c-nu), E0771/mKO-luc2 (2 × 10 5 cells to female C57B/6 albino), 4T1/luc (1 × 10 3 cells to female BALB/c), or MCF7 (1.5 × 10 6 cells to female SCID) suspended in 100 μL PBS was injected into the caudal artery of anesthetized mice using 29 G syringe needle in a short time (<3 s). IC injection was performed as described previously 4 , 5 . LLC/luc (2 × 10 5 cells) suspended in 100 μL PBS and was injected into left cardiac ventricle of 5-week-old-male C57B/6 mice.…”

“…Murine models of bone metastasis using intracardiac (IC) and intratibial injections have been instrumental in revealing molecular mechanisms underlying metastatic processes and translational studies for drug development 3 , 4 . During the past two decades, IC injection has been the gold standard to develop bone metastasis in mice 5 – 9 by injecting cancer cells into the left ventricle to disseminate them to the whole body including bone marrow tissue via the arterial bloodstream, which eventually develop into metastatic colonies in the bone and other organs 10 .…”

“…Unlike intratibial injection that severely damages the tibia, IC injection recapitulates the bone metastasis process, including survival of cancer cells in the bloodstream, extravasation, micro-colony formation, and metastatic progression in the intact bone marrow, and thus provides more relevant information for drug development. IC injection, however, is insufficient for rapid studies in this field, mainly owing to its requirement for high technical proficiency to exactly insert a syringe needle into the left ventricle of a mouse, causing severe cardiac stresses 3 , 4 . This limits the number of cancer cells that can be injected at one time, leading to limited delivery of cancer cells to the bone.…”

Symmetry and fluctuation of cell movements in neural crest-derived facial mesenchyme

“…Mouse gene-targeting experiments also support a specific requirement for Rho GTPase signaling in morphogenesis of the midface. Three tissue-specific knockouts of Cdc42 using P0-cre (a neural crest cell cre driver) ( Oshima-Nakayama et al, 2016 ) or Wnt1Cre (neural crest cells) ( Fuchs et al, 2009 ; Liu et al, 2013 ) or Prrx-Cre (facial mesenchyme) ( Aizawa et al, 2012 ) cause a wide midfacial cleft. Rac1 was also knocked out conditionally with Wnt1-Cre ( Thomas et al, 2010 ) and a transgene expressing a dominant-negative form of Rho kinase ( Phillips et al, 2012 ) also gave rise to the same phenotype.…”

Rho-Family Small GTPases: From Highly Polarized Sensory Neurons to Cancer Cells

“…Functions of Rac1 ( left ) or Cdc42 ( right ) in vivo ( upper ), our recent advances using Rac - [66] or Cdc42 -KO [57,71] mice ( middle ), and congenital/hereditary patients with RAC1 or CDC42 mutants ( lower ). Evidence from animal models helps elucidate the functions of Rac1 [66,67,68,69,72,73,74,75,76,77,78] and Cdc42 [71,74,79,80,81,82,83,84,85,86,87,88,89,90,91,92,93] and the symptoms of congenital/hereditary patients with mutations in RAC1 [20] and CDC42 [29,30,31,32]. Nrg1: neuregulin 1.…”