Levels Of Glycogen Synthase Kinase Research Articles

β-Sitosterol alleviates the malignant phenotype of hepatocellular carcinoma cells via inhibiting GSK3B expression

To explore the effects of β-Sitosterol upon hepatocellular carcinoma cell proliferation, apoptosis, migration, invasion, and epithelial–mesenchymal transition (EMT), and to investigate the underlying mechanism using network pharmacology. Human hepatocellular carcinoma cell lines (Huh-7 and HCCLM3) were expose to gradient concentrations of β-Sitosterol (5 μg/mL, 10 μg/mL, and 20 μg/mL). Cell viability and proliferation were assessed using MTT, CCK-8, colony formation, and EdU assays.Flow cytometry was employed to evaluate cell cycle and apoptosis. Scratch and Transwell assays were performed, respectively, to detect cell migration and invasion. The levels of apoptosis-associated proteins (BAX, BCL2, and cleaved caspase3) as well as EMT-associated proteins (E-cadherin, N-cadherin, Snail, and Vimentin) were detected in Huh-7 and HCCLM3 cell lines using Western blot analysis. The drug target gene for β-Sitosterol was screened via PubChem and subsequently evaluated for expression in the GSE112790 dataset. In addition, the expression level of glycogen synthase kinase 3 beta (GSK3B) within the Cancer Genome Atlas-Liver Hepatocellular Carcinoma (TCGA-LIHC) database was analyzed, along with its correlation to the survival outcomes of patients with hepatocellular carcinoma. The diagnostic efficiency of GSK3B was assessed by analyzing the ROC curve. Subsequently, Huh-7 and HCCLM3 cell lines were transfected with the overexpression vector of GSK3B and then treated with β-Sitosterol to further validate the association between GSK3B and β-Sitosterol. GSK3B demonstrated a significantly elevated expression in patients with hepatocellular carcinoma, which could predict hepatocellular carcinoma patients’ impaired prognosis based on GEO dataset and TCGA database. GSK3B inhibitor (CHIR-98014) notably inhibited cell proliferation and invasion, promoted cell apoptosis and cell cycle arrest at G0/G1 phase in hepatocellular carcinoma cells. β-Sitosterol treatment further promoted the efffects of GSK3B inhibitor on hepatocellular carcinoma cells. GSK3B overexpression has been found to enhance the proliferative and invasive capabilities of hepatocellular carcinoma cells. Furthermore it has been observed that GSK3B overexpression, it has been obsear can partially reverse the inhibitory effect of β-Sitosterol upon hepatocellular. β-Sitosterol suppressed hepatocellular carcinoma cell proliferation and invasion, and enhanced apoptosis via inhibiting GSK3B expression.

Escin Activates Canonical Wnt/β-Catenin Signaling Pathway by Facilitating the Proteasomal Degradation of Glycogen Synthase Kinase-3β in Cultured Human Dermal Papilla Cells.

Abnormal inactivation of the Wnt/β-catenin signaling pathway is involved in skin diseases like androgenetic alopecia, vitiligo and canities, but small-molecule activators are rarely described. In this study, we investigated the stimulatory effects of escin on the canonical Wnt/β-catenin signaling pathway in cultured human dermal papilla cells (hDPCs). Escin stimulated Wnt/β-catenin signaling, resulting in increased β-catenin and lymphoid enhancer-binding factor 1 (LEF1), the accumulation of nuclear β-catenin and the enhanced expression of Wnt target genes in cultured hDPCs. Escin drastically reduced the protein level of glycogen synthase kinase (GSK)-3β, a key regulator of the Wnt/β-catenin signaling pathway, while the presence of the proteasome inhibitor MG-132 fully restored the GSK-3β protein level. The treatment of secreted frizzled-related proteins (sFRPs) 1 and 2 attenuated the activity of escin in Wnt reporter assays. Our data demonstrate that escin is a natural agonist of the canonical Wnt/β-catenin signaling pathway and downregulates GSK-3β protein expression by facilitating the proteasomal degradation of GSK-3β in cultured hDPCs. Our data suggest that escin likely stimulates Wnt signaling through direct interactions with frizzled receptors. This study underscores the therapeutic potential of escin for Wnt-related diseases such as androgenetic alopecia, vitiligo and canities.

Efficacy of active ingredients in Qingdai on ulcerative colitis: a network pharmacology-based evaluation.

To elucidate the protective effect of Qingdai (, QD) on ulcerative colitis (UC) by means of and approaches. A systems pharmacology analysis was per-formed to predict the active components of QD whereas the putative biological targets of QD against UC were obtained through target fishing, network cons-truction and enrichment analyses. Meanwhile, we examined the ameliorative effect of QD in a mouse model of dextran sulfate sodium (DSS)-induced colitis. During the 10-day experiment, the control and diseased mice were given with oral gavages of QD (1.3 g raw herbs·kg·d) or 5-aminosalicylic acid (5-ASA, 100 mg·kg·d) every day. The underlying pharma-cological mechanisms of QD in UC were determined using polymerase chain reaction tests, histological staining, enzyme-linked immunoassays, and Western blotting analysis. Searching from various network pharmacology databases, 29 compounds were identified in QD. According to the screening criteria suggested by TCMSP (i.e. OB ≥ 30% and DL ≥ 0.18), nine of them were considered the active ingredients that contribute to the ameliorative effects of QD on different mouse models of colitis. Most importantly, the protective effect of QD on DSS-induced colitis was significantly associated with modulations of the expression levels of glycogen synthase kinase 3-β (Gsk3-β) and forkhead box p3 (Foxp3), which are widely considered as important regulators of excessive inflammatory responses. The results of this study provide solid scientific evidence for the use of QD or its core active components in the clinical management of UC.

Therapeutic effect of Baimai Ointment on peripheral sensitization of chronic inflammatory pain in mice

Chronic inflammatory pain is mainly manifested by peripheral sensitization. Baimai Ointment(BMO), a classical Tibetan medicine for external use, has good clinical efficacy in the treatment of chronic inflammatory pain, while its pharmacodynamics and mechanism for relieving peripheral sensitization remain unclear. This study established an animal model of chronic inflammatory pain induced by complete Freund's adjuvant to explore the mechanism of BMO in the treatment of chronic inflammatory pain by behavioral test, side effect assessment, network analysis, and experimental verification. The pharmacodynamics experiment showed that BMO increased the thresholds of mechanical pain sensitivity and thermal radiation pain sensitivity of chronic inflammatory pain mice in a dose-dependent manner, and had inhibitory effect on foot swelling, inflammatory mediator, and the expression of transient receptor potential vanilloid-1(TRPV1) and transient receptor potential A1(TRPA1). The results of body weight monitoring, pain sensitivity threshold detection in normal mice, rotarod performance test, and forced swimming test showed that BMO had no obvious toxic or side effect. The network analysis of 51 candidate active molecules selected according to the efficacy of BMO, content of main components, and ADME parameters showed that the inhibitory effect of BMO on chronic inflammatory pain was associated with the core regulatory elements of tumor necrosis factor(TNF) and T cell receptor signaling pathways. BMO down-regulated the protein levels of mitogen-activated protein kinase 14(MAPK14), MAPK1, and prostaglandin-endoperoxide synthase 2(PTGS2), and up-regulated the phosphorylation le-vel of glycogen synthase kinase 3 beta(GSK3 B) in the plantar tissue of mice. In conclusion, BMO can effectively relieve peripheral sensitization of chronic inflammatory pain without inducing tolerance and obvious toxic and side effects. The relevant mechanism may be related to the regulation of BMO on core regulatory elements of TNF and T cell receptor signaling pathways in surrounding tissues.

The Effect of Glucose Administration on Yes‐associated Protein (YAP) Activity in Healthy and Cachectic Mouse Muscles

Yes‐associated protein (YAP) is a downstream effector of Hippo signaling and regulates skeletal muscle size and homeostasis. The change of YAP activity has been reported in aging and dystrophic muscles. Our preliminary data showed that tumor burden alters the YAP activity in skeletal muscle. Recent studies showed that glucose metabolism also regulates YAP activity in cancer cells; however, it is unknown whether glucose administration stimulates YAP activity in skeletal muscle. The purpose of this study was to determine whether the YAP activity would change after acute glucose administration in healthy and cachectic mouse muscles. We hypothesized that glucose could stimulate YAP activity, at least in healthy muscle. Approximately 4‐month‐old wild‐type (WT, n=10) and ApcMin/+ mice (Min, n=10) were used in this study. Each group of mice was assigned to either control (Con, n=5 per group) or glucose administration (Glu n=5 per group) treatment. At first, all the mice fasted for four hours. Glucose dissolved in phosphate‐buffered saline (PBS, 2 mg/kg) was administered intraperitoneally to Glu groups. Con groups received PBS injections. Thirty minutes after glucose administration, the mice were euthanized, and plantaris muscles (PLAN) were removed. The time to sacrifice was based on the peak insulin secretion during a glucose tolerance test. To ensure injection accuracy, the glucose levels were measured using a handheld glucometer before glucose injection and at death. After total protein was extracted, routine Western blotting was performed using 50~100 µg of the total protein. Each phosphorylated protein was normalized to its respective total protein. Unpaired t‐test (WT vs. Min) and a two‐way ANOVA (genotype x glucose treatment) were used for statistical analysis. The levels of significance were set at 0.05. The body weight of Min mice was smaller than that of WT mice (19.8±1.2 mg vs. 17.3±0.5 mg for WT and Min, respectively, p=0.05). Likewise, the PLAN muscle mass of Min mice was smaller than that of WT mice (19.8±1.2 mg vs. 17.3±0.5 mg for WT and Min, respectively, p=0.05). There was no significant difference in fasting glucose levels between WT and Min mice, and glucose administration increased plasma glucose levels regardless of genotype (main effect of glucose administration, p<0.05). The activity levels of glycogen synthase kinase 3 beta (GSK3β) were comparable at the baseline (1.1±0.1 vs. 1.1±0.2 for WT and Min, respectively) and after glucose administration (1.1±0.2 vs. 1.1±0.4 for WT and Min, respectively). Min mice showed a reduction in total ribosomal protein S6 level after fasting (1.0±0.2 vs. 0.4±0.2 for WT and Min, respectively, p<0.05) and in response to glucose administration (2.7±1.5 vs. 0.6±0.4 for WT and Min, respectively, p<0.05). Interestingly, the basal YAP levels were comparable between WT and Min mice (1.0±0.1 vs. 1.1±0.1 for WT and Min, respectively), and the glucose administration did not alter the YAP levels (1.1±0.1 vs. 1.1±0.2 for WT and Min, respectively). These data suggest that YAP activity is independent of acute glucose administration in healthy and cachectic mouse skeletal muscles.

RETRACTED ARTICLE: Propofol inhibits oxidative stress injury through the glycogen synthase kinase 3 beta/nuclear factor erythroid 2-related factor 2/heme oxygenase-1 signaling pathway

Statement of Retraction Ziyin Zhan, BaoFeng Yan, Yuguo Li, Shuo Yang, and Jinfeng Li- 2022, Propofol inhibits oxidative stress injury through the glycogen synthase kinase 3 beta/nuclear factor erythroid 2-related factor 2/heme oxygenase-1 signaling pathway, Bioengineered 13(1), doi: 10.1080/21655979.2021.2021062 Since publication, significant concerns have been raised about the integrity of the data and reported results in the article. When approached for an explanation, the authors did not provide their original data or any necessary supporting information. As verifying the validity of published work is core to the integrity of the scholarly record, we are therefore retracting the article. All authors listed in this publication have been informed. We have been informed in our decision-making by our editorial policies and the COPE guidelines. The retracted article will remain online to maintain the scholarly record, but it will be digitally watermarked on each page as ‘Retracted’.

Lactobacillus plantarum PS128 prevents cognitive dysfunction in Alzheimer’s disease mice by modulating propionic acid levels, glycogen synthase kinase 3 beta activity, and gliosis

BackgroundAccording to recent evidence, psychobiotics exert beneficial effects on central nervous system-related diseases, such as mental disorders. Lactobacillus plantarum PS128 (PS128), a novel psychobiotic strain, improves motor function, depression, and anxiety behaviors. However, the psychobiotic effects and mechanisms of PS128 in Alzheimer’s disease (AD) remain to be explored.ObjectivesThe goal of the current study was to evaluate the beneficial effects of PS128 and to further elucidate its mechanism in AD mice.MethodsPS128 (1010 colony-forming unit (CFU)/ml) was administered via oral gavage (o.g.) to 6-month-old male wild-type B6 and 3 × Tg-AD mice (harboring the PS1M146V, APPswe and TauP30IL transgenes) that received an intracerebroventricular injection of streptozotocin (icv-STZ, 3 mg/kg) or vehicle (saline) for 33 days. After serial behavioral tests, fecal short-chain fatty acid levels and AD-related pathology were assessed in these mice.ResultsOur findings show that intracerebroventricular injection of streptozotocin accelerated cognitive dysfunction associated with increasing levels of glycogen synthase kinase 3 beta (GSK3β) activity, tau protein phosphorylation at the T231 site (pT231), amyloid-β (Aβ) deposition, amyloid-β protein precursor (AβPP), β-site AβPP-cleaving enzyme (BACE1), gliosis, fecal propionic acid (PPA) levels and cognition-related neuronal loss and decreasing postsynaptic density protein 95 (PSD95) levels in 3 × Tg-AD mice. PS128 supplementation effectively prevented the damage induced by intracerebroventricular injection of streptozotocin in 3 × Tg-AD mice.ConclusionsBased on the experimental results, intracerebroventricular injection of streptozotocin accelerates the progression of AD in the 3 × Tg-AD mice, primarily by increasing the levels of gliosis, which were mediated by the propionic acid and glycogen synthase kinase 3 beta pathways. PS128 supplementation prevents damage induced by intracerebroventricular injection of streptozotocin by regulating the propionic acid levels, glycogen synthase kinase 3 beta activity, and gliosis in 3 × Tg-AD mice. Therefore, we suggest that PS128 supplementation is a potential strategy to prevent and/or delay the progression of AD.

Platelet Rich Plasma Regulates Wnt/Beta-Catenin Signal Pathway in Rabbit Chondrocytes by Mediating Interleukin 1 Beta

It was reported that platelet rich plasma can reduce the degeneration of articular cartilage. The purpose of this study was to investigate the effect of platelet rich plasma on Wnt/beta-catenin signal transduction in rabbit chondrocytes. Platelet rich plasma was prepared from the ear vein blood of 3-mo old New Zealand white rabbits. Chondrocytes were isolated from knee cartilage and cultured. The cell identification and the effect of platelet rich plasma on the viability of chondrocytes were measured by the C-terminal telopeptide II and proteoglycan staining. Chondrocytes were divided into 5 groups, control group, interleukin-1 beta group, platelet rich plasma (100 x dilutions) group, Dickkopf WNT signaling pathway inhibitor 1 (100 ng/mL) group and Dickkopf WNT signaling pathway inhibitor 1+platelet rich plasma group. After being treated with interleukin-1 beta (50 μL, 10 μg/mL) for 24 h, the morphology of chondrocytes was observed by an electron microscope. The levels of C-terminal telopeptide II and cartilage oligomeric matrix protein in the culture medium were determined by enzyme linked immunosorbent assay. The messenger RNA and protein expression levels of Wnt-1, beta-catenin and glycogen synthase kinase 3 beta were detected by Real-time polymerase chain reaction and Western blot respectively. Platelet rich plasma promotes the proliferation of chondrocytes. The chondrocytes in the interleukin-1 beta group showed an ultrastructural abnormality, which was not obvious in platelet rich plasma group, Dickkopf WNT signaling pathway inhibitor 1group, and Dickkopf WNT signaling pathway inhibitor 1+platelet rich plasma group. The concentrations of C-terminal telopeptide II and cartilage oligomeric matrix protein in the interleukin-1 beta group were higher than those in the control group, platelet rich plasma group, Dickkopf WNT signaling pathway inhibitor 1group and Dickkopf WNT signaling pathway inhibitor 1+platelet rich plasma group. Compared with the control group, platelet rich plasma group, Dickkopf WNT signaling pathway inhibitor 1group and Dickkopf WNT signaling pathway inhibitor 1+platelet rich plasma group, the expression of Wnt1 and beta-catenin was higher and the level of glycogen synthase kinase 3 beta was lower in the interleukin-1 beta group. Platelet rich plasma may protect interleukin-1 beta activated chondrocytes by inhibiting Wnt/beta-catenin signal.

Tissue-based metabolomics reveals potential biomarkers for cervical carcinoma and HPV infection.

Aberrant metabolic regulation has been observed in human cancers, but the corresponding regulation in human papillomavirus (HPV) infection-associated cervical cancer is not well understood. Here, we explored potential biomarkers for the early prediction of cervical carcinoma based on the metabolic profile of uterine cervical tissue specimens that were positive for HPV16 infection. Fifty-two fresh cervical tissues were collected from women confirmed to have cervical squamous cell carcinoma (SCC; n = 21) or cervical intraepithelial neoplasia (CIN) stages II-III (n = 20). Eleven healthy women constituted the controls (negative controls [NCs]). Real-time polymerase chain reaction (PCR) was performed to detect HPV infection in the tissues. High-resolution magic angle spinning nuclear magnetic resonance was utilized for the analysis of the metabolic profile in the tissues. The expression of rate-limiting enzymes involved in key metabolic pathways was detected by reverse-transcription quantitative PCR. An independent immunohistochemical analysis was performed using 123 cases of paraffin-embedded cervical specimens. A profile of 17 small molecular metabolites that showed differential expression in HPV16-positive cervical SCC or CIN II-III compared with HPV-negative NC group was identified. According to the profile, the levels of α- and β-glucose decreased, those of lactate and low-density lipoproteins increased, and the expression of multiple amino acids was altered. Significantly increased transcript and protein levels of glycogen synthase kinase 3 beta (GSK3β) and glutamate decarboxylase 1 (GAD1) and decreased transcript and protein levels of pyruvate kinase muscle isozyme 2 (PKM2) and carnitine palmitoyltransferase 1A (CPT1A) were observed in the patient group (p < 0.05). HPV infection and cervical carcinogenesis drive metabolic modifications that might be associated with the aberrant regulation of enzymes related to metabolic pathways.

Regulation of AKT phosphorylation by GSK3β and PTEN to control chemoresistance in breast cancer.

Phosphorylated AKT is highly expressed or overexpressed in chemoresistant tumor samples. However, the precise molecular mechanism involved in AKT phosphorylation-related chemoresistance in breast cancer is still elusive. The present research was designed to estimate the effect of AKT phosphorylation on cell viability and chemoresistance in breast cancer. We utilized MCF-7 and MDA-MB468 human breast cancer cell lines and developed multidrug-resistant MCF-7/MDR and cisplatin-resistant MDA-MB-468 cells. Immunofluorescence analysis and Western blotting were employed to test the level of glycogen synthase kinase 3 beta (GSK3β), phosphorylated phosphatase and tension homologue (p-PTEN) and phosphorylated AKT (p-AKT) in MCF-7/MDR and MDA-MB468 cells. Xenograft assays in nude mice were performed with MCF-7/MDR cells to verify chemoresistance and the signaling pathway upstream of phosphatidylinositide 3-kinase (PI3K)/AKT. An increase in GSK3β, p-PTEN and p-AKT expression was strongly induced in MCF-7/MDR and cisplatin-resistant MDA-MB-468 cells, and augmented GSK3β phosphorylation and PTEN inactivation enhanced AKT signaling. The elevation in GSK3β, p-PTEN and p-AKT was associated with cell viability based on a CCK-8 assay. The results of in vivo and in vitro assays indicated that GSK3β knockdown with lentiviral shRNA (shRNA-GSK3β) promoted apoptosis and suppressed the migration of cisplatin-resistant MCF-7/MDR cells, while these effects were reversed by activating p-AKT with the PTEN inhibitor bpV(pic). AKT phosphorylation mediated by GSK3β and PTEN were correlated with cell viability, migration and apoptosis, which may promote chemoresistance in breast cancer. Furthermore, GSK3β can regulate cell viability through the PTEN/PI3K/AKT signaling pathway and induce chemoresistance, serving as a valuable molecular strategy for breast cancer therapy.

Vitamin A status affects weight gain and hepatic glucose metabolism in rats fed a high-fat diet.

Whether vitamin A (VA) has a role in the development of metabolic abnormalities associated with intake of a high-fat diet (HFD) is unclear. Sprague-Dawley rats after weaning were fed an isocaloric VA sufficient HFD (VAS-HFD) or a VA deficient HFD (VAD-HFD) for 8 weeks. Body mass, food intake, liver and adipose tissue mass, and the hepatic expression levels of key proteins for metabolism were determined. VAD-HFD rats had lower body, liver, and epididymal fat mass than VAS-HFD rats. VAD-HFD rats had lower hepatic protein expression levels of cytochrome P450 26A1, glucokinase, and phosphoenolpyruvate carboxykinase than VAS-HFD rats. VAD-HFD rats had higher protein levels of glycogen synthase kinase (GSK)-3α and lower levels of GSK-3β, but not glycogen synthase, than VAS-HFD rats. VAD-HFD rats had higher hepatic levels of insulin receptor substrate-1 (IRS-1), insulin receptor β-subunit, mitogen-activated protein kinase proteins, and peroxisome proliferator-activated receptor-gamma coactivator 1α mRNA, and lower level of IRS-2 protein than VAS-HFD rats. These results indicate that in a HFD setting, VA deficiency attenuated HFD-induced obesity, and VA status altered the expression levels of proteins required for glucose metabolism and insulin signaling. We conclude that VA status contributes to the regulation of hepatic glucose and lipid metabolism in a HFD setting, and may regulate hepatic carbohydrate metabolism.

Transcription factor profiling identifies Sox9 as regulator of proliferation and differentiation in corneal epithelial stem/progenitor cells

Understanding transcription factor (TF) regulation of limbal epithelial stem/progenitor cells (LEPCs) may aid in using non-ocular cells to regenerate the corneal surface. This study aimed to identify and characterize TF genes expressed specifically in LEPCs isolated from human donor eyes by laser capture microdissection. Using a profiling approach, preferential limbal expression was found for SoxE and SoxF genes, particularly for Sox9, which showed predominantly cytoplasmic localization in basal LEPCs and nuclear localization in suprabasal and corneal epithelial cells, indicating nucleocytoplasmic translocation and activation during LEPC proliferation and differentiation. Increased nuclear localization of Sox9 was also observed in activated LEPCs following clonal expansion and corneal epithelial wound healing. Knockdown of SOX9 expression in cultured LEPCs by RNAi led to reduced expression of progenitor cell markers, e.g. keratin 15, and increased expression of differentiation markers, e.g. keratin 3. Furthermore, SOX9 silencing significantly suppressed the proliferative capacity of LEPCs and reduced levels of glycogen synthase kinase 3 beta (GSK-3ß), a negative regulator of Wnt/ß-catenin signaling. Sox9 expression, in turn, was significantly suppressed by treatment of LEPCs with exogenous GSK-3ß inhibitors and enhanced by small molecule inhibitors of Wnt signaling. Our results suggest that Sox9 and Wnt/ß-catenin signaling cooperate in mutually repressive interactions to achieve a balance between quiescence, proliferation and differentiation of LEPCs in the limbal niche. Future molecular dissection of Sox9-Wnt interaction and mechanisms of nucleocytoplasmic shuttling of Sox9 may aid in improving the regenerative potential of LEPCs and the reprogramming of non-ocular cells for corneal surface regeneration.

HnRNP K plays a protective role in TNF-α-induced apoptosis in podocytes.

Apoptosis of podocytes contributes to proteinuria in many chronic kidney diseases. The cytokine, tumor necrosis factor-α (TNF-α) is thought to be involved in podocyte apoptosis, but the underlying mechanism is not understood. In our study, we established a model of TNF-α-induced apoptosis by isolating primary podocytes from mice. After exposing cells to TNF-α, we determined the expression levels of heterogeneous nuclear ribonucleoprotein K (hnRNP K) and cellular FLICE-inhibitory protein (c-FLIP) and the phosphorylation levels of glycogen synthase kinase β (GSK3β) and extracellular signal-regulated kinase (ERK). We then knocked down or overexpressed the levels of hnRNP K and observed its effects on the expressions of c-FLIP, caspase-8, caspase-3, and the phosphorylation of GSK3β and ERK. In addition, we examined the percentage of cells undergoing apoptosis and studied cell cycle distribution. We found that TNF-α induced apoptosis in podocytes and that the expressions of hnRNP K and c-FLIP were significantly decreased, whereas the phosphorylations of GSK3β and ERK were significantly increased. Both gene knockdown and overexpression of hnRPN K resulted in varied expressions/phosphorylations of c-FLIP, GSK3β, and ERK. Moreover, decreased hnRPN K expression contributed to increased levels of caspase-8 and capase-3, as well as an increase in cell apoptosis and G0/G1 arrest. In conclusion, down-regulated expression of hnRNP K by TNF-α resulted in a decrease in the expression of c-FLIP as well as increases in phosphorylated GSK3β, ERK, caspase-8, and caspase-3, and then critically contributed to the podocyte apoptosis.

Lithium-associated transcriptional regulation of CRMP1 in patient-derived olfactory neurons and symptom changes in bipolar disorder

There is growing evidence that lithium used in the treatment of bipolar disorder (BD) affects molecular targets that are involved in neuronal growth, survival, and maturation, but it remains unclear if neuronal alterations in any of these molecules predict specific symptom changes in BD patients undergoing lithium monotherapy. The goals of this study were to (a) determine which molecular changes in the olfactory neurons of symptomatic patients receiving lithium are associated with antimanic or antidepressant response, and (b) uncover novel intraneuronal regulatory mechanisms of lithium therapy. Twenty-two treatment-naïve non-smoking patients, with symptomatic BD underwent nasal biopsies for collection of olfactory tissues, prior to their treatment and following a 6-week course of lithium monotherapy. Sixteen healthy controls were also biopsied. Combining laser capture microdissection with real-time polymerase chain reaction, we investigated baseline and treatment-associated transcriptional changes in candidate molecular targets of lithium action in the olfactory neuroepithelium. Baseline mRNA levels of glycogen synthase kinase 3 beta (GSK3β) and collapsin response mediator protein 1 (CRMP1) genes were significantly associated with BD status and with severity of mood symptoms. Among BD subjects, treatment-associated downregulation of CRMP1 expression was most predictive of decreases in both manic and depressive symptoms. This study provides a novel insight into the relevance of CRMP1, a key molecule in semaphorin-3A signaling during neurodevelopment, in the molecular mechanism of action of lithium, and in the pathophysiology of BD. It supports the use of human-derived olfactory neuronal tissues in the evaluation of treatment response of psychiatric disorders.

Puerarin inhibits β-amyloid peptide 1-42-induced tau hyperphosphorylation via the Wnt/β-catenin signaling pathway

Excessive tau protein phosphorylation is important in the pathogenesis and early abnormal signal transduction of Alzheimer's disease. Excessive phosphorylation of microtubules is associated with tau accumulation, which induces the formation of neurofibrillary tangles in neurons, leading to synaptic damage and ultimately, neurodegeneration. The present study aimed to investigate the possible mechanism underlying the inhibitory effects of puerarin on β‑amyloid peptide (Aβ)1‑42‑induced tau protein hyperphosphorylation in SH‑SY5Y cells. Following various treatments, the viability of SH‑SY5Y cells was determined using the MTT assay, and cell morphology was observed under an inverted fluorescence microscope. Western blotting was used to detect tau phosphorylation, and the protein expression levels of glycogen synthase kinase (GSK)‑3β, phosphorylated (p)‑GSK‑3β (Ser9), β‑catenin and cyclin D1, which are the key factors mediating the Wnt/β‑catenin signaling pathway in SH‑SY5Y cells. The results demonstrated that puerarin reversed the Aβ1‑42‑induced decrease in SH‑SY5Y cell viability. In addition, puerarin inhibited the degree of Aβ1‑42‑induced tau phosphorylation at Ser396, Ser199 and Thr231 in SH‑SY5Y cells, and reduced the expression of GSK‑3β by increasing the expression of p‑GSK‑3β (Ser9). Furthermore, puerarin increased the protein expression levels of β‑catenin and cyclin D1, which are key factors involved in the Wnt/β‑catenin signaling pathway. The results of the present study demonstrated that puerarin may attenuate Aβ1‑42‑induced tau hyperphosphorylation in SH‑SY5Y cells, by inhibiting the expression of GSK‑3β and activating the Wnt/β‑catenin signaling pathway; therefore, puerarin may exert protective effects against Alzheimer's disease.

Downregulation of β-catenin blocks fibrosis via Wnt2 signaling in human keloid fibroblasts.

Keloid is a disorder of fibroproliferative diseases that occurs in wounds, characterized by an exaggerated response to injury. The key factor responsible for the disease process has not been identified. This study sought to elucidate the role of β-catenin in the regulation of keloid phenotypes and signaling. Expression of β-catenin in keloid and normal non-keloid samples was measured by real-time polymerase chain reaction. Knockdown of β-catenin was achieved by delivering small interfering RNA to target β-catenin. Cell proliferation, cell cycle progression, and apoptosis of keloid cells were measured by functional assays in vitro. The proteins related to keloid fibrosis were measured by Western blotting. β-catenin expression was significantly upregulated in keloid tissue samples compared with the normal non-keloid age-adjusted skin sample counterparts. Functionally, targeting β-catenin with lipofection-delivered small interfering RNA oligonucleotide inhibited the proliferation and cell cycle arrest in G0/G1 phase and increased apoptosis of fibroblast cells, accompanied by downregulation of Wnt2 and cyclin D1 as well as the phosphorylation level of glycogen synthase kinase 3 beta in the keloid fibrosis. Our study supports a crucial role of β-catenin in the regulation of fibroproliferation and extracellular matrix deposition. Targeting β-catenin using small interfering RNA oligonucleotide may be a promising approach for preventing excessive fibroproliferative development after wound healing and may lead to the development of novel strategies for restoring keloid diseases.

Ablation of periostin inhibits post-infarction myocardial regeneration in neonatal mice mediated by the phosphatidylinositol 3 kinase/glycogen synthase kinase 3β/cyclin D1 signalling pathway.

AimsTo resolve the controversy as to whether periostin plays a role in myocardial regeneration after myocardial infarction (MI), we created a neonatal mouse model of MI to investigate the influence of periostin ablation on myocardial regeneration and clarify the underlying mechanisms.Methods and resultsNeonatal periostin-knockout mice and their wildtype littermates were subjected to MI or sham surgery. In the wildtype mice after MI, fibrosis was detectable at 3 days and fibrotic tissue was completely replaced by regenerated myocardium at 21 days. In contrast, in the knockout mice, significant fibrosis in the infarcted area was present at even 3 weeks after MI. Levels of phosphorylated-histone 3 and aurora B in the myocardium, detected by immunofluorescence and western blotting, were significantly lower in knockout than in wildtype mice at 7 days after MI. Similarly, angiogenesis was decreased in the knockout mice after MI. Expression of both the endothelial marker CD-31 and α-smooth muscle actin was markedly lower in the knockout than in wildtype mice at 7 days after MI. The knockout MI group had elevated levels of glycogen synthase kinase (GSK) 3β and decreased phosphatidylinositol 3-kinase (PI3K), phosphorylated serine/threonine protein kinase B (p-Akt), and cyclin D1, compared with the wildtype MI group. Similar effects were observed in experiments using cultured cardiomyocytes from neonatal wildtype or periostin knockout mice. Administration of SB216763, a GSK3β inhibitor, to knockout neonatal mice decreased myocardial fibrosis and increased angiogenesis in the infarcted area after MI.ConclusionAblation of periostin suppresses post-infarction myocardial regeneration by inhibiting the PI3K/GSK3β/cyclin D1 signalling pathway, indicating that periostin is essential for myocardial regeneration.

Chimeric antibody targeting SRPK-1 in the treatment of non-small cell lung cancer by inhibiting growth, migration and invasion.

Non-small cell lung cancer (NSCLC) is one of the most common types of cancer in humans, and is characterized by rapid growth, migration, invasion and reoccurrence. Evidence has indicated that the protein and mRNA levels of serine‑arginine protein kinase‑1 (SRPK‑1) are upregulated in NSCLC tissues. However, the functions of SRPK1 and targeted therapy for SRPK1 in the progression and treatment of NSCLC remain to be fully elucidated. In the present study, the mRNA and protein expression levels of SRPK‑1 in NSCLC cells and tissues were analyzed using reverse transcription‑quantitative polymerase chain reaction analysis and SDS‑PAGE, and the role of SRPK1 in the progression of NSCLC was investigated. In addition, a chimeric antibody target for SRPK‑1 (ChanSRPK‑1) was constructed, and the therapeutic effects of ChanSRPK‑1 were investigated in H358‑bearing mice. The curative effects of ChanSRPK‑1 on the inhibition of growth, migration and invasion of NSCLC were also examined invitro and invivo. The results revealed that the mRNA and protein levels of SRPK‑1 were upregulated in NSCLC cells and tumor tissues. Higher expression of SRPK1 promoted NSCLC cell growth, migration and invasion, whereas lower expression of SRPK‑1 suppressed growth, migration and invasion of the NSCLC cells. Animal experiments demonstrated that ChanSRPK‑1 inhibited the β‑catenin/T‑cell factor complex. ChanSRPK‑1 treatment also downregulated the phosphorylation levels of glycogen synthase kinase 3-β and prolonged the survival of tumor‑bearing mice. Taken together, SRPK‑1 may offer potential as a therapeutic target oncogenic molecular in NSCLC, and ChanSRPK‑1 may be a therapeutic agent with functions as a target and for oncolytic therapy in the treatment of NSCLC.

Etoposide induces pancreatic β-cells cytotoxicity via the JNK/ERK/GSK-3 signaling-mediated mitochondria-dependent apoptosis pathway

Etoposide is widely used in the treatment of the different types of tumors such as pancreatic cancer. However, etoposide also causes several unwanted side-effects in normal viable cells, including pancreatic β-cells, which are vulnerable to chemical-induced injuries, and the molecular mechanisms underlying etoposide-induced apoptosis are still unclear. Here, the results showed that in RIN-m5F cells (a β-cell-derived cell line), the number of viable cells was significantly decreased after 24h of etoposide treatment and underwent mitochondria-dependent apoptotic signals accompanied by mitochondrial dysfunction, and increases in the population of sub-G1 hypodiploid cells and apoptotic cells, caspase-3 activity, and the activation of caspase cascades. Etoposide also increased the phosphorylation levels of glycogen synthase kinase (GSK)-3α/β in treated RIN-m5F cells. Pretreatment with LiCl, a GSK-3 inhibitor, prevented etoposide-induced mitochondria-dependent apoptosis and GSK-3 protein phosphorylation in RIN-m5F cells. Furthermore, exposure of the cells to etoposide induced the phosphorylation of c-Jun N-terminal kinase (JNK) and extracellular signal-related kinase (ERK)1/2 but not p38-MAPK, which was suppressed by the specific JNK inhibitor (SP600125) and ERK1/2 inhibitor (PD98059), respectively. Additionally, pretreatment with both SP600125 and PD98059 effectively suppressed etoposide-induced β-cell cytotoxicity, apoptosis, and GSK-3 protein phosphorylation; however, LiCl did not reverse JNK and ERK1/2 phosphorylation. Taken together, these results suggest that etoposide is capable of causing cytotoxicity on pancreatic β-cells by inducing apoptosis through the JNK/ERK-mediated GSK-3 downstream-triggered mitochondria-dependent signaling pathway.

Donepezil, an acetylcholine esterase inhibitor, and ABT-239, a histamine H3 receptor antagonist/inverse agonist, require the integrity of brain histamine system to exert biochemical and procognitive effects in the mouse

Histaminergic H3 receptors (H3R) antagonists enhance cognition in preclinical models and modulate neurotransmission, in particular acetylcholine (ACh) release in the cortex and hippocampus, two brain areas involved in memory processing. The cognitive deficits seen in aging and Alzheimer’s disease have been associated with brain cholinergic deficits. Donepezil is one of the acetylcholinesterase (AChE) inhibitor approved for use across the full spectrum of these cognitive disorders. We addressed the question if H3R antagonists and donepezil require an intact histamine neuronal system to exert their procognitive effects. The effect of the H3R antagonist ABT-239 and donepezil were evaluated in the object recognition test (ORT), and on the level of glycogen synthase kinase 3 beta (GSK-3β) phosphorylation in normal and histamine-depleted mice. Systemic administration of ABT-239 or donepezil ameliorated the cognitive performance in the ORT. However, these compounds were ineffective in either genetically (histidine decarboxylase knock-out, HDC-KO) or pharmacologically, by means of intracerebroventricular (i.c.v.) injections of the HDC irreversible inhibitor a-fluoromethylhistidine (a-FMHis), histamine-deficient mice. Western blot analysis revealed that ABT-239 or donepezil systemic treatments increased GSK-3β phosphorylation in cortical and hippocampal homogenates of normal, but not of histamine-depleted mice. Furthermore, administration of the PI3K inhibitor LY294002 that blocks GSK-3β phosphorylation, prevented the procognitive effects of both drugs in normal mice. Our results indicate that both donepezil and ABT-239 require the integrity of the brain histaminergic system to exert their procognitive effects and strongly suggest that impairments of PI3K/AKT/GSK-3β intracellular pathway activation is responsible for the inefficacy of both drugs in histamine-deficient animals.