A novel role of PRL in regulating epithelial cell density by inducing apoptosis at confluence

Maintaining proper epithelial cell density is essential for the survival of multicellular organisms. While regulation of cell density through apoptosis is well known, its mechanistic details remain elusive. Here, we report the involvement of membrane-anchored phosphatase of regenerating liver (PRL), originally known for its role in cancer malignancy, in this process. In epithelial MDCK cells, upon confluence, doxycycline-induced expression of PRL upregulated apoptosis, reducing the cell density. This could be circumvented by artificially reducing the cell density via stretching the cell-seeded silicon chamber. Moreover, siRNA-mediated knockdown of endogenous PRL blocked apoptosis, leading to greater cell density. Mechanistically, PRL promoted apoptosis by upregulating the translation of E-cadherin and activating TGF-β pathway. Morpholino-mediated inhibition of PRL expression in zebrafish embryos caused developmental defect with reduced apoptosis and increased epithelial cell density during convergent extension. This study revealed a novel role of PRL in regulating density-dependent apoptosis in vertebrate epithelium.

Increased Phosphatase of Regenerating Liver-1 by Placental Stem Cells Promotes Hepatic Regeneration in a Bile-Duct-Ligated Rat Model

Phosphatase of regenerating liver-1 (PRL-1) controls various cellular processes and liver regeneration. However, the roles of PRL-1 in liver regeneration induced by chorionic-plate-derived mesenchymal stem cells (CP-MSCs) transplantation remain unknown. Here, we found that increased PRL-1 expression by CP-MSC transplantation enhanced liver regeneration in a bile duct ligation (BDL) rat model by promoting the migration and proliferation of hepatocytes. Engrafted CP-MSCs promoted liver function via enhanced hepatocyte proliferation through increased PRL-1 expression in vivo and in vitro. Moreover, higher increased expression of PRL-1 regulated CP-MSC migration into BDL-injured rat liver through enhancement of migration-related signals by increasing Rho family proteins. The dual effects of PRL-1 on proliferation of hepatocytes and migration of CP-MSCs were substantially reduced when PRL-1 was silenced with siRNA-PRL-1 treatment. These findings suggest that PRL-1 may serve as a multifunctional enhancer for therapeutic applications of CP-MSC transplantation.

The mechanisms of colorectal cancer cell mesenchymal-epithelial transition induced by hepatocyte exosome-derived miR-203a-3p

Background Liver metastasis is the most common cause of death in patients with colorectal cancer (CRC). Phosphatase of regenerating liver-3 induces CRC metastasis by epithelial-to-mesenchymal transition, which promotes CRC cell liver metastasis. Mesenchymal-to-epithelial transition (MET), the opposite of epithelial-to-mesenchymal transition, has been proposed as a mechanism for the establishment of metastatic neoplasms. However, the molecular mechanism of MET remains unclear. Methods Using Immunohistochemistry, western blotting, invasion assays, real-time quantitative PCR, chromatin immunoprecipitation, luciferase reporter assays, human miRNA arrays, and xenograft mouse model, we determined the role of hepatocyte exosome-derived miR-203a-3p in CRC MET. Results In our study, we found that miR-203a-3p derived from hepatocyte exosomes increased colorectal cancer cells E-cadherin expression, inhibited Src expression, and reduced activity. In this way miR-203a-3p induced the decreased invasion rate of CRC cells. Coclusion MiR-203a-3p derived from hepatocyte exosomes plays an important role of CRC cells to colonize in liver.

The phosphatase PRL-3 affects intestinal homeostasis by altering the crypt cell composition

Expression of the phosphatase of regenerating liver-3 (PRL-3) is known to promote tumor growth in gastrointestinal adenocarcinomas, and the incidence of tumor formation upon inflammatory events correlates with PRL-3 levels in mouse models. These carcinomas and their onset are associated with the impairment of intestinal cell homeostasis, which is regulated by a balanced number of Paneth cells and Lgr5 expressing intestinal stem cells (Lgr5+ ISCs). Nevertheless, the consequences of PRL-3 overexpression on cellular homeostasis and ISC fitness in vivo are unexplored. Here, we employ a doxycycline-inducible PRL-3 mouse strain to show that aberrant PRL-3 expression within a non-cancerous background leads to the death of Lgr5+ ISCs and to Paneth cell expansion. A higher dose of PRL-3, resulting from homozygous expression, led to mice dying early. A primary 3D intestinal culture model obtained from these mice confirmed the loss of Lgr5+ ISCs upon PRL-3 expression. The impaired intestinal organoid formation was rescued by a PRL inhibitor, providing a functional link to the observed phenotypes. These results demonstrate that elevated PRL-3 phosphatase activity in healthy intestinal epithelium impairs intestinal cell homeostasis, which correlates this cellular mechanism of tumor onset with PRL-3-mediated higher susceptibility to tumor formation upon inflammatory or mutational events.Key messages• Transgenic mice homozygous for PRL-3 overexpression die early.• PRL-3 heterozygous mice display disrupted intestinal self-renewal capacity.• PRL-3 overexpression alone does not induce tumorigenesis in the mouse intestine.• PRL-3 activity leads to the death of Lgr5+ ISCs and Paneth cell expansion.• Impairment of cell homeostasis correlates PRL-3 action with tumor onset mechanisms.

Combination of PRL-3 Inhibitor with Sorafenib Synergistically Promotes AML Apoptosis

Phosphatase of regenerating liver-3 (PRL-3) is recognized as a novel independent crucial driver for AML progression. Thus, the specific inhibitor of PRL-3 would be a potential therapeutic agent to AML in clinic, but there is no such preclinical application reported yet. Here we evaluated the cytotoxicity of PRL-3 inhibitor, BR-1, against AML cells ML-1 and MOLM-13. Meanwhile, the effect of BR-1 on the biological characteristics of AML cells and the underlying mechanism were investigated along with the combination of BR-1 and sorafenib on AML cell viability. Our results show that BR-1 promotes apoptosis by inactivation of JAK/STAT5 and PI3K/AKT pathways, while inhibits cell proliferation through arresting cell cycle in the S phase. In addition, combination of BR-1 with a FLT3 inhibitor, sorafenib can further improve the therapeutic effect on AML. Thus, our results demonstrated that BR-1 would be a novel and potent therapeutic agent to AML, and its combination with other anti-AML drugs would be a promising strategy to AML therapy.

The Mechanisms of Colorectal Cancer Cell mesenchymal-epithelial Transition Induced by Hepatocyte Exosome-derived miR-203a-3p

Background: Liver metastasis is the most common cause of death in patients with colorectal cancer (CRC). Phosphatase of regenerating liver-3 induces CRC metastasis by epithelial-to-mesenchymal transition, which promotes CRC cell liver metastasis. Mesenchymal-to-epithelial transition (MET), the opposite of epithelial-to-mesenchymal transition, has been proposed as a mechanism for the establishment of metastatic neoplasms. However, the molecular mechanism of MET remains unclear. Methods: Using Immunohistochemistry, western blotting，invasion assays, real-time quantitative PCR, chromatin immunoprecipitation, luciferase reporter assays, human miRNA arrays, and xenograft mouse model, we determined the role of hepatocyte exosome-derived miR-203a-3p in CRC MET.Results: In our study, we found that miR-203a-3p derived from hepatocyte exosomes increased colorectal cancer cells E-cadherin expression, inhibited Src expression, and reduced activity. In this way miR-203a-3p induced the decreased invasion rate of CRC cells.Coclusion: MiR-203a-3p derived from hepatocyte exosomes plays an important role of CRC cells to colonize in liver.

Fast Diffusion Sustains Plasma Membrane Accumulation of Phosphatase of Regenerating Liver-1

It has been proposed that the accumulation of farnesylated phosphatase of regenerating liver-1 (PRL-1) at the plasma membrane is mediated by static electrostatic interactions of a polybasic region with acidic membrane lipids and assisted by oligomerization. Nonetheless, localization at early and recycling endosomes suggests that the recycling compartment might also contribute to its plasma membrane accumulation. Here, we investigated in live cells the dynamics of PRL-1 fused to the green fluorescent protein (GFP-PRL-1). Blocking the secretory pathway and photobleaching techniques suggested that plasma membrane accumulation of PRL-1 was not sustained by recycling endosomes but by a dynamic exchange of diffusible protein pools. Consistent with this idea, fluorescence correlation spectroscopy in cells overexpressing wild type or monomeric mutants of GFP-PRL-1 measured cytosolic and membrane-diffusing pools of protein that were not dependent on oligomerization. Endogenous expression of GFP-PRL-1 by CRISPR/Cas9 genome edition confirmed the existence of fast diffusing cytosolic and membrane pools of protein. We propose that plasma membrane PRL-1 replenishment is independent of the recycling compartment and the oligomerization state and mainly driven by fast diffusion of the cytosolic pool.

Functionally enhanced placenta-derived mesenchymal stem cells inhibit adipogenesis in orbital fibroblasts with Graves’ ophthalmopathy

Background Placenta-derived mesenchymal stem cells (PD-MSCs) have unique immunomodulatory properties. Phosphatase of regenerating liver-1 (PRL-1) regulates the self-renewal ability of stem cells and promotes proliferation. Graves’ ophthalmopathy (GO) is an autoimmune inflammatory disease of the orbit and is characterized by increased orbital levels of adipose tissue. Here, we evaluated the therapeutic mechanism for regulation of adipogenesis by PRL-1-overexpressing PD-MSCs (PD-MSCsPRL-1, PRL-1+) in orbital fibroblast (OF) with GO patients. Methods PD-MSCs isolated from human placenta were transfected with the PRL-1 gene using nonviral transfection method. Primary OFs were isolated from orbital adipose tissue specimens from GO patients. After maturation as adipogenic differentiation, normal and GO-derived OFs were cocultured with naïve and PD-MSCsPRL-1. We analyzed the protein levels of adipogenesis markers and their signaling pathways in OFs from GO patients. Results The characteristics of PD-MSCsPRL-1 were similar to those of naïve cells. OFs from GO patients induced adipocyte differentiation and had significantly decreased a lipid accumulation after coculture with PD-MSCsPRL-1 compared to naïve cells. The mRNA and protein expression of adipogenic markers was decreased in PD-MSCsPRL-1. Insulin-like growth factor-binding proteins (IGFBPs) secreting PD-MSCsPRL-1 downregulated the phosphorylated PI3K/AKT/mTOR expression in OFs from GO patients. Interestingly, IGFBP2, − 4, − 6, and − 7 expression in PD-MSCsPRL-1, which was mediated by integrin alpha 4 (ITGA4) and beta 7 (ITGB7), was higher than that in naïve cells and upregulated phosphorylated FAK downstream factor. Conclusion In summary, IGFBPs secreting PD-MSCPRL-1 inhibit adipogenesis in OFs from GO patients by upregulating phosphorylated FAK and downregulating PI3K/AKT/mTOR signaling pathway. The functional enhancement of PD-MSCs by nonviral gene modification provides a novel therapeutic strategy for the treatment of degenerative diseases.