Hatta K.,Queens University |
Chen Z.,Queens University |
Carter A.L.,Queens University |
Leno-Duran E.,Queens University |
And 7 more authors.
Placenta | Year: 2010
Objective: Wingless-type mouse mammary tumor virus integration site family, member 5A (WNT5A), is expressed in mouse decidua and is thought to play an important role in decidualization. We examined expression of the receptor for WNT5A, receptor tyrosine kinase-like orphan receptor 2 (ROR2), in the uteri of cycling and pregnant mice. Study design: Reverse transcription (RT)-PCR and immunohistochemistry were performed. Results: RT-PCR revealed that transcripts for Ror2, Wnt3a, Wnt5a and inhibitor of WNT signaling, Dickkopf homolog 1 (Dkk1), were present in the pregnant uterus. Immunohistochemistry revealed that in the virgin uterus, ROR2 is expressed in stromal cells and on the basal side of uterine gland and endometrial epithelial cells. During pregnancy, both the luminal and basal side of uterine gland epithelial cells expressed ROR2, stromal cell expression of ROR2 became more frequent and ROR2 expressing uterine Natural Killer (NK) cells and cells lining the maternal vascular space emerged. Immunofluorescence imaging and flow cytometry revealed that although uterine NK cells expressed ROR2, NK cells of the spleen were ROR2 negative. Conclusion: The expression of ROR2 by endometrial epithelial cells may suggest WNT signaling has roles in uterine epithelial cell polarity or implantation. Expression of ROR2 by uterine NK cells may suggest WNT signaling regulates uterine NK cell functions such angiogenesis and regulation of trophoblast migration. In summary, our results show that ROR2 expression by maternal uterine cells is influenced by pregnancy. © 2010 Elsevier Ltd. All rights reserved.
Bautista-Cruz F.,Gastrointestinal Disease Research Unit |
Nair D.G.,Gastrointestinal Disease Research Unit |
Nair D.G.,Queens University |
Lourenssen S.,Gastrointestinal Disease Research Unit |
And 5 more authors.
Canadian Journal of Physiology and Pharmacology | Year: 2014
We have previously demonstrated that lower esophageal sphincter (LES) circular smooth muscle (CSM) is functionally impaired in W/Wv mutant mice that lack interstitial cells of Cajal, and speculated that this could be due to altered smooth muscle differentiation. Platelet-derived growth factor (PDGF) is involved in the maturation and differentiation of smooth muscle. To determine whether PDGF expression and (or) function is altered in W/Wv mutant mice, PDGF-Rβ expression was measured using RT-PCR, qPCR, and immunocytochemistry, and Ca2+ imaging and perforated patch clamp recordings performed in isolated LES CSM cells. RT-PCR and immunocytochemistry showed significantly reduced PDGF-Rβ expression in the LES from mutant as opposed to wild-type mice. Quantitative comparison of CSM cell numbers in histological specimens revealed a significantly increased average cell size in the mutant tissue. The specific PDGF-Rβ ligand, PDGF-BB, caused a significant increase in intracellular Ca2+ in cells from the wild-type mice compared with the mutants. Using a ramp protocol, PDGF-BB caused a 2-fold increase in outward K+ currents in cells from the wild-type mice, whereas no significant increase was measured in the cells from the mutants. We conclude that the expression and function of PDGF-Rβ in LES CSM from W/Wv mice is impaired, providing further evidence that LES CSM is abnormal in W/Wv mutants.
Cheung R.,Queens University |
Cheung R.,Gastrointestinal Disease Research Unit |
Kelly J.,Queens University |
Kelly J.,Gastrointestinal Disease Research Unit |
And 2 more authors.
Frontiers in Physiology | Year: 2011
Regulation of expression of the intestinal epithelial actin-binding protein, villin, is poorly understood. The aim of this study was to determine whether Wnt5a stimulates Ror2 in intestinal epithelia caused transient increases in phospho-ERK1/2 (pERK1/2) and subsequently increased expression of villin transcript and protein. To demonstrate Wnt5a-Ror2 regulation of villin expression, we overexpressed wild-type, truncated, or mutant Ror2 constructs in HT29 adenocarcinoma cells and non-transformed fetally derived human intestinal epithelial cells, added conditioned media containing Wnt5a and measured changes in ERK1/2 phosphorylation, villin amplicons, and protein expression by RT-PCR and Western blot techniques. Wnt5a addition caused a transient increase in pERK1/2, which was maximal at 10 min but extinguished by 30 min.Transient transfection with a siRNA duplex against Ror2 diminished Ror2 amplicons and protein and reduced the extent of pERK1/2 activation. Structure-function analysis revealed that the deletion of the cysteine-rich, kringle, or tyrosine kinase domain or substitution mutations of tyrosine residues in the intracellular Ser/Thr-1 region of Ror2 prevented the Wnt5a stimulation of pERK1/2. Deletion of the intracellular proline and serine/threonine-rich regions of Ror2 had no effect onWnt5a stimulation of pERK1/2. The increase in villin expression was blocked by pharmacological inhibition of MEK-1 and casein kinase 1, but not by PKC and p38 inhibitors. Neither Wnt3a nor epidermal growth factor addition caused increases in villin protein. Our findings suggest that Wnt5a/Ror2 signaling can regulate villin expression in the intestine. © 2011 Cheung, Kelly and Macleod.