Lysophosphatidylcholine acyltransferase 3 deficiency impairs 3T3L1 cell adipogenesis through activating Wnt/β-catenin pathway

Levels of polyunsaturated phosphatidylcholine (PC) influence plasma membrane structure and function. Phosphatidylcholine (PC) is synthesized de novo in the Kennedy pathway and then undergoes extensive deacylation/reacylation remodeling via Lands' cycle (non-Kennedy pathway). The reacylation is catalyzed by lysophosphatidylcholine acyltransferase (LPCAT), which adds a polyunsaturated fatty acid at the sn-2 position. Four LPCAT isoforms have been described to date, among which we found LPCAT3 to be the major isoform in adipose tissue, but its exact role in adipogenesis is unclear. In this study, we aimed to investigate whether LPCAT3 activity affects 3T3L1 cell adipogenic differentiation potential and its underline mechanism. Lentivirus-mediated LPCAT3 shRNA expression stably knocked down LPCAT3 in 3T3L1 preadipocytes and LPCAT3 deficiency dramatically reduced the levels of cellular polyunsaturated PCs. Importantly, we found that this deficiency activated the β-catenin dependent Wnt signaling pathway, which suppressed the expression of adipogenesis-related genes, thereby inhibiting 3T3L1 preadipocyte differentiation and lipid accumulation. Moreover, three different Wnt/β-catenin pathway inhibitors reversed the effect of LPCAP3 deficiency, suggesting that Wnt/β-catenin pathway activation is one of the causes for the observed phenotypes. To the best of our knowledge, we show here for the first time that PC remodeling is an important regulator of adipocyte differentiation.     

Caudatin inhibits carcinomic human alveolar basal epithelial cell growth and angiogenesis through modulating GSK3β/β-catenin pathway

In this study, we investigate the anti‐cancer activity of caudatin in carcinomic human alveolar basal epithelial cell line A549 and anti‐angiogenic activity in human umbilical vein endothelial cells (HUVECs). We show that caudatin impairs the cell viability and induces G₀/G₁ phase arrest in A549 cells with a dose dependent manner. A549 cells, not HUVECs, dealing with caudatin exhibited typical characteristics of apoptosis, which were accompanied by activation of caspase‐3, caspase‐9 and Poly(ADP–Ribose) Polymerase (PARP). In addition, caudatin treatment resulted in a decrease of β‐catenin and increase of phosphorylation of β‐catenin, and inhibited phosphorylation levels of GSK3β (Ser 9) in A549 cells. Conditional medium of A549 cells‐induced or growth factors‐induced tube formation of HUVECs was markedly inhibited by caudatin treatment, which was associated with the inhibiting VEGF secretion from A549 cells by caudatin. Our findings suggest that caudatin inhibits carcinomic human alveolar basal epithelial cell growth and angiogenesis by targeting GSK3β/β‐catenin pathway and suppressing VEGF production. J. Cell. Biochem. 113: 3403–3410, 2012. © 2012 Wiley Periodicals, Inc.     

Axed MUC4 (MUC4/X) aggravates pancreatic malignant phenotype by activating integrin-β1/FAK/ERK pathway

Alternative splicing is evolving as an eminent player of oncogenic signaling for tumor development and progression. Mucin 4 (MUC4), a type I membrane-bound mucin, is differentially expressed in pancreatic cancer (PC) and plays a critical role in its progression and metastasis. However, the molecular implications of MUC4 splice variants during disease pathogenesis remain obscure. The present study delineates the pathological and molecular significance of a unique splice variant of MUC4, MUC4/X, which lacks the largest exon 2, along with exon 3. Exon 2 encodes for the highly glycosylated tandem repeat (TR) domain of MUC4 and its absence creates MUC4/X, which is devoid of TR. Expression analysis from PC clinical samples revealed significant upregulation of MUC4/X in PC tissues with most differential expression in poorly differentiated tumors. In vitro studies suggest that overexpression of MUC4/X in wild-type-MUC4 (WT-MUC4) null PC cell lines markedly enhanced PC cell proliferation, invasion, and adhesion to extracellular matrix (ECM) proteins. Furthermore, MUC4/X overexpression leads to an increase in the tumorigenic potential of PC cells in orthotopic transplantation studies. In line with these findings, doxycycline-induced expression of MUC4/X in an endogenous WT-MUC4 expressing PC cell line (Capan-1) also displayed enhanced cell proliferation, invasion, and adhesion to ECM, compared to WT-MUC4 alone, emphasizing its direct involvement in the aggressive behavior of PC cells. Investigation into the molecular mechanism suggested that MUC4/X facilitated PC tumorigenesis via integrin-β1/FAK/ERK signaling pathway. Overall, these findings revealed the novel role of MUC4/X in promoting and sustaining the oncogenic features of PC.     

Prednisolone induces osteocytes apoptosis by promoting Notum expression and inhibiting PI3K/AKT/GSK3β/β-catenin pathway

Journal of Molecular Histology - The apoptosis of mature osteocytes is the main factor causing damage to the microstructure of cortical bone in glucocorticoid-induced osteoporosis (GIOP). Our...     

MicroRNA-182-5p aggravates ulcerative colitis by inactivating the Wnt/β-catenin signaling pathway through DNMT3A-mediated SMARCA5 methylation

This research focused on novel molecular mechanisms underlying microRNA (miR)-182-5p in ulcerative colitis (UC). Colon tissues were obtained from UC patients, and dextrose sodium sulfate (DSS)-induced mouse and interleukin-1β (IL-1β)-induced Caco-2 cell models were generated. Then, miR-182-5p, SMARCA5, and the Wnt/β-catenin signaling pathway were altered in IL-1β-stimulated Caco-2 cells and DSS-treated mice to assess their function. MiR-182-5p and SMARCA5 were upregulated and DNMT3A, β-catenin, and Cyclin D1 were downregulated in UC patients, IL-1β-stimulated Caco-2 cells, and DSS-treated mice. Mechanistically, miR-182-5p targeted DNMT3A to upregulate SMARCA5, thus blocking the Wnt/β-catenin signaling pathway. Moreover, SMARCA5 silencing or Wnt/β-catenin signaling pathway activation repressed apoptosis and augmented proliferation and epithelial barrier function of IL-1β-stimulated Caco-2 cells. SMARCA5 silencing annulled the impacts of miR-182-5p overexpression on IL-1β-stimulated Caco-2 cells. SMARCA5 silencing or miR-182-5p inhibition ameliorated intestinal barrier dysfunction in DSS-treated mice. Collectively, miR-182-5p aggravates UC by inactivating the Wnt/β-catenin signaling pathway through DNMT3A-mediated SMARCA5 methylation.     

The protease Omi regulates mitochondrial biogenesis through the GSK3β/PGC-1α pathway

Loss of the mitochondrial protease activity of Omi causes mitochondrial dysfunction, neurodegeneration with parkinsonian features and premature death in mnd2 (motor neuron degeneration 2) mice. However, the detailed mechanisms underlying this pathology remain largely unknown. Here, we report that Omi participates in the process of mitochondrial biogenesis, which has been linked to several neurodegenerative diseases. The mitochondrial biogenesis is deficit in mnd2 mice, evidenced by severe decreases of mitochondrial components, mitochondrial DNA and mitochondrial density. Omi cleaves glycogen synthase kinase 3β (GSK3β), a kinase promoting PPARγ coactivator-1α (PGC-1α) degradation, to regulate PGC-1α, a factor important for the mitochondrial biogenesis. In mnd2 mice, GSK3β abundance is increased and PGC-1α abundance is decreased significantly. Inhibition of GSK3β by SB216763 or overexpression of PGC-1α can restore mitochondrial biogenesis in mnd2 mice or Omi-knockdown N2a cells. Furthermore, there is a significant improvement of the movement ability of mnd2 mice after SB216763 treatment. Thus, our study identified Omi as a novel regulator of mitochondrial biogenesis, involving in Omi protease-deficient-induced neurodegeneration.Mitochondria have a vital role in neuronal death and survival.¹ As critical cellular organelles, mitochondria have highly dynamic properties, including mitochondrial fission, fusion, transport, biogenesis and degradation. The changes of those properties affect mitochondrial functions, leading to the occurrence of diseases.^{2, 3} Growing lines of evidence suggest that the mitochondrial dysfunction is involved in aging and neurodegenerative diseases, such as Alzheimer''s disease (AD), Huntington''s disease (HD) and Parkinson''s disease (PD).^{4, 5} Similar to other neurodegenerative diseases, PD is a progressive neurological disorder, which is characterized by the development of cytoplasmic aggregates known as Lewy bodies and degeneration of dopaminergic (DA) neurons in the substantia nigra of midbrain and other brain regions.⁶ In PD, dysfunction of mitochondria has been documented to be associated with disease pathogenesis in PD brains and both genetic- and toxin-induced PD animal models. In PD brains, mutations in mitochondrial DNA (mtDNA) occur more frequently than those in age-matched control; and mutations in the nuclear-encoded mtDNA polymerase-γ gene, which impair mtDNA replication and result in multiple mtDNA deletions, cause PD-like symptoms.⁵ Meanwhile, several PD-associated gene products, including α-synuclein, parkin, DJ-1, PINK1 (PTEN-induced putative kinase 1), leucine-rich repeat kinase 2, ubiquitin carboxy-terminal hydrolase L1 and Omi, have been identified to be associated with PD, and lead to mitochondrial dysfunction with changes in mitochondrial morphology, biogenesis and mitophagy in vivo and in vitro.^{5, 7, 8, 9} Besides, mitochondrial toxins, such as MPTP (1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine) and rotenone that inhibit complex I of the mitochondrial respiratory chain, cause clinically parkinsonian phenotype.^{10, 11}The serine protease Omi (also known as HtrA2) belongs to the high-temperature requirement factor A (HtrA) family, and was originally identified as a mammalian homolog of the Escherichia coli heat-shock-induced serine protease HtrA/DegP and DegS.¹² Omi is mainly localized in mitochondria, although a fraction of it is also found in nucleus.¹³ Omi is released from the mitochondria into the cytosol to mediate cell death by caspase-dependent or -independent pathways in response to apoptotic stimuli.^{14, 15} However, the notion that Omi is an apoptosis inducer in the central nervous system was challenged by studies of Omi-overexpressing or -deficient mice. Omi-overexpressing mice show normal development without any sign of apoptotic cell death.¹⁶ On the other hand, mnd2 (motor neuron degeneration 2) mice that harbor protease-deficient Omi S276C mutants, and Omi-knockout mice both suffer from progressive neurodegeneration, especially in striatum, and motor abnormalities similar to PD. Both mice fail to gain weight and die before postnatal day 40 due to neurodegeneration with progressive mitochondrial damage.^{17, 18, 19} Besides, mutations in the Omi gene have also been identified in PD patients.^{20, 21} Previous studies have shown that Omi has a vital role in the mitochondrial integrity, and the loss of protease activity leads to mitochondrial dysfunction, such as abnormal mitochondrial morphology and increased mtDNA mutation and deletions, increased susceptibility of mitochondrial membrane permeabilization, decreased mitochondrial membrane potential, and reduced mitochondrial density in mnd2 mice and Omi-knockout mice.^{17, 18, 22} Omi has been found to act downstream of PINK1, but parallel to parkin, in a mitochondrial stress sensing pathway to sense the different stresses, which may be defective in PD.²³ These findings suggest that the primary function of Omi is involved in neuroprotection, especially in the maintenance of mitochondrial homeostasis.^{23, 24}In this article, we identified that Omi cleaves glycogen synthase kinase 3β (GSK3β) to regulate PPARγ coactivator-1α (PGC-1α) abundance and to ensure mitochondrial biogenesis.     

Fractalkine aggravates LPS-induced macrophage activation and acute kidney injury via Wnt/β-catenin signalling pathway

Fractalkine (CX3CL1, FKN), a CX3C gene sequence inflammatory chemokine, has been found to have pro-inflammatory and pro-adhesion effects. Macrophages are immune cells with a critical role in regulating the inflammatory response. The imbalance of M1/M2 macrophage polarization can lead to aggravated inflammation. This study attempts to investigate the mechanisms through which FKN regulates macrophage activation and the acute kidney injury (AKI) involved in inflammatory response induced by lipopolysaccharide (LPS) by using FKN knockout (FKN-KO) mice and cultured macrophages. It was found that FKN and Wnt/β-catenin signalling have a positive interaction in macrophages. FKN overexpression inhibited LPS-induced macrophage apoptosis. However, it enhanced their cell viability and transformed them into the M2 type. The effects of FKN overexpression were accelerated by activation of Wnt/β-catenin signalling. In the in vivo experiments, FKN deficiency suppressed macrophage activation and reduced AKI induced by LPS. Inhibition of Wnt/β-catenin signalling and FKN deficiency further mitigated the pathologic process of AKI. In summary, we provide a novel mechanism underlying activation of macrophages in LPS-induced AKI. Although LPS-induced murine AKI was unable to completely recapitulate human AKI, the positive interactions between FKN and Wnt/β-catenin signalling pathway may be a therapeutic target in the treatment of kidney injury.     

β-Ionone represses renal cell carcinoma progression through activating LKB1/AMPK-triggered autophagy

β-Ionone, the end ring analog of β-carotenoids, has been proven to have an antitumor effect in a variety of cancers. In this study, we investigated the impact of β-ionone on renal cell carcinoma (RCC) cell lines (786-O and ACHN) using colony formation assays, flow cytometry analysis, and western blot analysis. We found that β-ionone effectively inhibited the proliferation of RCC cells in vitro, which was also confirmed in a xenograft model. Moreover, we found that β-ionone could induce autophagy, as indicated by LC3 puncta in 786-O and ACHN cell lines and the expression of LC3 in β-ionone-treated RCC cells. To further explore the underlying mechanism, we assessed liver kinase B1/AMP-activated protein kinase (LKB1/AMPK) signaling pathway activity, and the results showed that β-ionone inhibited the proliferation of RCC cells by inducing autophagy via the LKB1/AMPK signaling pathway. In summary, our findings provide a new therapeutic strategy of β-ionone-induced autophagy in RCC.     

KIF18B promotes hepatocellular carcinoma progression through activating Wnt/β-catenin-signaling pathway

This study aimed to investigate the functional roles of kinesin family member 18B (KIF18B) in hepatocellular carcinoma (HCC) development, as well as the related molecular mechanisms. Tissue specimens were collected from 105 patients with HCC, and the messenger RNA (mRNA) and protein levels of KIF18B were detected using quantitative real-time polymerase chain reaction and immunohistochemistry assays, respectively. The χ² test was performed to estimate the association of KIF18B with clinical characteristics of patients with HCC. Effects of KIF18B expression on biological behaviors of HCC cells were detected by clone formation, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, and transwell assays. The expression patterns of proteins were investigated using Western blot analysis. HCC tissues and cell lines showed significant upregulation of KIF18B at both mRNA and protein levels (p > .05, for all). Furthermore, the elevated KIF18B expression was positively correlated with the tumor-node-metastasis stage (p = .015) and lymph node metastasis (p = .007). Knockdown of KIF18B might suppress HCC cell clone formation, proliferation, migration, and invasion in vitro. Besides, the activity of Wnt/β-catenin pathway was also significantly inhibited after the KIF18B knockdown. However, the antitumor actions caused by KIF18B knockdown might be reversed by lithium chloride treatment, which was the inducer of Wnt/β-catenin-signaling pathway. KIF18B may serve as an oncogene in HCC through enhancing the activity of Wnt/β-catenin pathway.     

Long non-coding RNA SUMO1P3 promotes hepatocellular carcinoma progression through activating Wnt/β-catenin signalling pathway by targeting miR-320a

In this study, we aimed to investigate expression profile of long non-coding RNA (lncRNA) SUMO1P3, and its role and molecular mechanisms in the progression of hepatocellular carcinoma (HCC). The expression of SUMO1P3 in HCC tissues and cells was detected using quantitative real-time polymerase chain reaction (qRT-PCR). The chi-squared test was used to estimate the relationship between SUMO1P3 levels and clinical characteristics of HCC cases. Cellular biological behaviours were investigated using MTT, transwell assays and wound healing assay. Bioinformatics and dual-luciferase reporter assays were performed to identify potential target of SUMO1P3 in HCC. Additionally, protein analysis was carried out using Western blot. The expression of SUMO1P3 was significantly higher in HCC tissues and cells than in non-cancerous specimens and normal cells (P < .01). Moreover, its up-regulation was closely correlated with lymph node metastasis (P = .027) and TNM stage (P = .019). SUMO1P3 knockdown inhibited the proliferation, migration and invasion of HCC cells. MiR-320a was a potential target of SUMO1P3, and its expression was negatively regulated by SUMO1P3 in HCC SUMO1P3 could activate Wnt/β-catenin pathway, which was mediated by miR-320a. Elevated expression of SUMO1P3 predicts malignant progression among HCC patients. SUMO1P3 enhances Wnt/β-catenin pathway through sponging miR-320a, thus contributing to aggressive progression of HCC.     

Integrinβ3 mediates the protective effects of soluble receptor for advanced glycation end-products during myocardial ischemia/reperfusion through AKT/STAT3 signaling pathway

Soluble receptor for advanced glycation end-product (sRAGE) was reported to protect myocardial ischemia/reperfusion (I/R) injuries via directly interacting with cardiomyocytes besides competing with RAGE for AGEs. However, the specific molecule for the interaction between sRAGE and cardiomyocytes are not clearly defined. Integrins which were reported to interact with RAGE on leukocytes were also expressed on myocardial cells, therefore it was supposed that sRAGE might interact with integrins on cardiomyocytes to protect hearts from ischemia/reperfusion injuries. The results showed that sRAGE increased the expression of integrinβ3 but not integrinβ1, β2, β4 or β5 in cardiomyocytes during I/R injuries. Meanwhile, the suppressive effects of sRAGE on cardiac function, cardiac infraction size and apoptosis in mice were cancelled by inhibition of integrinβ3 with cilengitide (CLG, 75 mg/kg). The results from cultured cardiomyocytes also proved that sRAGE attenuated myocardial apoptosis and autophagy through interacting with integrinβ3 to activate Akt and STAT3 pathway during oxygen and glucose deprivation/reperfusion (OGD/R) treatment. Furthermore, the phosphorylation of STAT3 was significantly downregulated by the inhibition of Akt (LY294002, 10 μM) in OGD/R and sRAGE treated cardiomyocytes, which suggested that STAT3 pathway was induced by Akt in I/R and sRAGE treated cardiomyocytes. The present study contributes to the understanding of myocardial I/R pathogenesis and provided a novel integrinβ3-dependent therapy strategy for sRAGE ameliorating I/R injuries.

     

Anti-IL-23 exerted protective effects on cerebral ischemia–reperfusion injury through JAK2/STAT3 signaling pathway

Molecular Biology Reports - Ischemia–reperfusion frequently occurs in ischemic cerebral vascular disease, during which the inflammatory signaling plays essential roles. The aim of this study...     

Regulation of valproic acid induced EMT by AKT/GSK3β/β-catenin signaling pathway in triple negative breast cancer

Molecular Biology Reports - Valproic acid (VPA) is a selective histone deacetylation (HDAC) inhibitor and exerts anti-cancer properties in different types of cancer. The epithelial-to-mesenchymal...     

Cardiac-specific Traf2 overexpression enhances cardiac hypertrophy through activating AKT/GSK3β signaling

Tumor necrosis factor superfamily ligands provoke a dilated cardiac phenotype signal through a common scaffolding protein termed tumor necrosis factor receptor-associated factor 2 (Traf2); however, Traf2 signaling in the adult mammalian cardiac hypertrophy is not fully understood. This study was aimed to identify the effect of Traf2 on cardiac hypertrophy and the underlying mechanisms. A significant up-regulation of Traf2 expression was observed in mice failing hearts. To further investigate the role of Traf2 in cardiac hypertrophy, we used cultured neonatal rat cardiomyocytes with gain and loss of Traf2 function and cardiac-specific Traf2-overexpressing transgenic (TG) mice. In cultured cardiomyocytes, Traf2 positively regulated angiotensin II (Ang II)-mediated hypertrophic growth, as detected by [³H]-Leucine incorporation, cardiac myocyte area, and hypertrophic marker protein levels. Cardiac hypertrophy in vivo was produced by constriction of transverse aortic (TAC) in TG mice and their wild-type controls. The extent of cardiac hypertrophy was evaluated by echocardiography as well as by pathological and molecular analyses of heart samples. Traf2 overexpression in the heart remarkably enhanced cardiac hypertrophy, left ventricular dysfunction in mice in response to TAC. Further analysis of the signaling pathway in vitro and in vivo suggested that these adverse effects of Traf2 were associated with the activation of AKT/glycogen synthase kinase 3β (GSK3β). The present study demonstrates that Traf2 serves as a novel mediator that enhanced cardiac hypertrophy by activating AKT/GSK3β signaling.