Vitamin E isoforms differentially regulate intercellular adhesion molecule-1 activation of PKCα in human microvascular endothelial cells

Aims

ICAM-1-dependent leukocyte recruitment in vivo is inhibited by the vitamin E isoform d-α-tocopherol and elevated by d-γ-tocopherol. ICAM-1 is reported to activate endothelial cell signals including protein kinase C (PKC), but the PKC isoform and the mechanism for ICAM-1 activation of PKC are not known. It is also not known whether ICAM-1 signaling in endothelial cells is regulated by tocopherol isoforms. We hypothesized that d-α-tocopherol and d-γ-tocopherol differentially regulate ICAM-1 activation of endothelial cell PKC.

Results

ICAM-1 crosslinking activated the PKC isoform PKCα but not PKCβ in TNFα-pretreated human microvascular endothelial cells. ICAM-1 activation of PKCα was blocked by the PLC inhibitor {"type":"entrez-nucleotide","attrs":{"text":"U73122","term_id":"4098075","term_text":"U73122"}}U73122, ERK1/2 inhibitor PD98059, and xanthine oxidase inhibitor allopurinol. ERK1/2 activation was blocked by inhibition of XO and PLC but not by inhibition of PKCα, indicating that ERK1/2 is downstream of XO and upstream of PKCα during ICAM-1 signaling. During ICAM-1 activation of PKCα, the XO-generated ROS did not oxidize PKCα. Interestingly, d-α-tocopherol inhibited ICAM-1 activation of PKCα but not the upstream signal ERK1/2. The d-α-tocopherol inhibition of PKCα was ablated by the addition of d-γ-tocopherol.

Conclusions

Crosslinking ICAM-1 stimulated XO/ROS which activated ERK1/2 that then activated PKCα. ICAM-1 activation of PKCα was inhibited by d-α-tocopherol and this inhibition was ablated by the addition of d-γ-tocopherol. These tocopherols regulated ICAM-1 activation of PKCα without altering the upstream signal ERK1/2. Thus, we identified a mechanism for ICAM-1 activation of PKC and determined that d-α-tocopherol and d-γ-tocopherol have opposing regulatory functions for ICAM-1-activated PKCα in endothelial cells.     

Protein kinase C delta (PKCδ) affects proliferation of insulin-secreting cells by promoting nuclear extrusion of the cell cycle inhibitor p21Cip1/WAF1

Background

High fat diet-induced hyperglycemia and palmitate-stimulated apoptosis was prevented by specific inhibition of protein kinase C delta (PKCδ) in β-cells. To understand the role of PKCδ in more detail the impact of changes in PKCδ activity on proliferation and survival of insulin-secreting cells was analyzed under stress-free conditions.

Methodology and Principal Findings

Using genetic and pharmacological approaches, the effect of reduced and increased PKCδ activity on proliferation, apoptosis and cell cycle regulation of insulin secreting cells was examined. Proteins were analyzed by Western blotting and by confocal laser scanning microscopy. Increased expression of wild type PKCδ (PKCδWT) significantly stimulated proliferation of INS-1E cells with concomitant reduced expression and cytosolic retraction of the cell cycle inhibitor p21^Cip1/WAF1. This nuclear extrusion was mediated by PKCδ-dependent phosphorylation of p21^Cip1/WAF1 at Ser146. In kinase dead PKCδ (PKCδKN) overexpressing cells and after inhibition of endogenous PKCδ activity by rottlerin or RNA interference phosphorylation of p21^Cip1/WAF1 was reduced, which favored its nuclear accumulation and apoptotic cell death of INS-1E cells. Human and mouse islet cells express p21^Cip1/WAF1 with strong nuclear accumulation, while in islet cells of PKCδWT transgenic mice the inhibitor resides cytosolic.

Conclusions and Significance

These observations disclose PKCδ as negative regulator of p21^Cip1/WAF1, which facilitates proliferation of insulin secreting cells under stress-free conditions and suggest that additional stress-induced changes push PKCδ into its known pro-apoptotic role.     

Non-small cell lung carcinoma cell motility, rac activation and metastatic dissemination are mediated by protein kinase C epsilon

Background

Protein kinase C (PKC) ε, a key signaling transducer implicated in mitogenesis, survival, and cancer progression, is overexpressed in human primary non-small cell lung cancer (NSCLC). The role of PKCε in lung cancer metastasis has not yet been established.

Principal Findings

Here we show that RNAi-mediated knockdown of PKCε in H358, H1299, H322, and A549 NSCLC impairs activation of the small GTPase Rac1 in response to phorbol 12-myristate 13-acetate (PMA), serum, or epidermal growth factor (EGF). PKCε depletion markedly impaired the ability of NSCLC cells to form membrane ruffles and migrate. Similar results were observed by pharmacological inhibition of PKCε with εV1-2, a specific PKCε inhibitor. PKCε was also required for invasiveness of NSCLC cells and modulated the secretion of extracellular matrix proteases and protease inhibitors. Finally, we found that PKCε-depleted NSCLC cells fail to disseminate to lungs in a mouse model of metastasis.

Conclusions

Our results implicate PKCε as a key mediator of Rac signaling and motility of lung cancer cells, highlighting its potential as a therapeutic target.     

Mechanisms for vascular cell adhesion molecule-1 activation of ERK1/2 during leukocyte transendothelial migration

Background

During inflammation, adhesion molecules regulate recruitment of leukocytes to inflamed tissues. It is reported that vascular cell adhesion molecule-1 (VCAM-1) activates extracellular regulated kinases 1 and 2 (ERK1/2), but the mechanism for this activation is not known. Pharmacological inhibitors of ERK1/2 partially inhibit leukocyte transendothelial migration in a multi-receptor system but it is not known whether VCAM-1 activation of ERK1/2 is required for leukocyte transendothelial migration (TEM) on VCAM-1.

Methodology/Principal Findings

In this study, we identified a mechanism for VCAM-1 activation of ERK1/2 in human and mouse endothelial cells. VCAM-1 signaling, which occurs through endothelial cell NADPH oxidase, protein kinase Cα (PKCα), and protein tyrosine phosphatase 1B (PTP1B), activates endothelial cell ERK1/2. Inhibition of these signals blocked VCAM-1 activation of ERK1/2, indicating that ERK1/2 is activated downstream of PTP1B during VCAM-1 signaling. Furthermore, VCAM-1-specific leukocyte migration under physiological laminar flow of 2 dynes/cm² was blocked by pretreatment of endothelial cells with dominant-negative ERK2 K52R or the MEK/ERK inhibitors, PD98059 and U0126, indicating for the first time that ERK regulates VCAM-1-dependent leukocyte transendothelial migration.

Conclusions/Significance

VCAM-1 activation of endothelial cell NADPH oxidase/PKCα/PTP1B induces transient ERK1/2 activation that is necessary for VCAM-1-dependent leukocyte TEM.     

532 nm Low-Power Laser Irradiation Recovers γ-Secretase Inhibitor-Mediated Cell Growth Suppression and Promotes Cell Proliferation via Akt Signaling

Background and Objective

The γ-secretase inhibitor (GSI) has been shown to inhibit expression of amyloid beta (Aβ), but GSI also has a side effect of reducing cell survival. Since low-power laser irradiation (LLI) has been known to promote cell survival, we examined whether 532 nm LLI can rescue the GSI side effect or not.

Study Design/Materials and Methods

The human-derived glioblastoma cells (A-172) were cultured in 35 mm culture dishes or 96-well plate. The center of dish or selected wells was irradiated with 532 nm laser (Nd:YVO4, CW, 60 mW) for 20, 40 and 60 min, respectively. The irradiated cells were photographed at immediately after, 24 and 48 h later and counted. GSI was supplemented in medium 3 h before LLI. The MTT assay was also used to estimate viable cells at 48 h after irradiation. The expression of phosphorylated Akt (p-Akt) or phosphorylated PTEN (p-PTEN) was examined by immunofluorescent staining and measured by fluorescence intensity using the software (BZ-9000, KEYENCE, Japan).

Results

GSI application depressed cell proliferation as well as cell survival compared to control. GSI down-regulated Aβ but up-regulated p-PTEN and suppressed p-Akt. Application of 532 nm LLI in the presence of GSI significantly recovered the GSI-mediated effects, i.e., LLI could decrease elevated p-PTEN, while increased p-Akt expression with keeping Aβ suppression. The LLI effects had a dose-dependency.

Conclusion

We confirmed that GSI potently suppressed intracellular Aβ and decreased cell survival. We conclude that a combination of GSI application and 532 nm LLI can increase cell proliferation via Akt activation while keeping PTEN and Aβ suppressed.     

Magnesium Modifies the Cardiovascular Mortality Risk Associated with Hyperphosphatemia in Patients Undergoing Hemodialysis: A Cohort Study

Background

In vitro studies have shown inhibitory effects of magnesium (Mg) on phosphate-induced calcification of vascular smooth muscle cells, raising the possibility that maintaining a high Mg level may be useful for reducing cardiovascular risks of patients with hyperphosphatemia. We examined how serum Mg levels affect the association between serum phosphate levels and the risk of cardiovascular mortality in patients undergoing hemodialysis.

Methods

A nationwide register-based cohort study was conducted using database of the Renal Data Registry of the Japanese Society for Dialysis Therapy in 2009. We identified 142,069 patients receiving in-center hemodialysis whose baseline serum Mg and phosphate levels were available. Study outcomes were one-year cardiovascular and all-cause mortality. Serum Mg levels were categorized into three groups (lower, <2.7 mg/dL; intermediate, ≥2.7, <3.1 mg/dL; and higher, ≥3.1 mg/dL).

Results

During follow-up, 11,401 deaths occurred, out of which 4,751 (41.7%) were ascribed to cardiovascular disease. In multivariable analyses, an increase in serum phosphate levels elevated the risk of cardiovascular mortality in the lower- and intermediate-Mg groups, whereas no significant risk increment was observed in the higher-Mg group. Moreover, among patients with serum phosphate levels of ≥6.0 mg/dL, the cardiovascular mortality risk significantly decreased with increasing serum Mg levels (adjusted odds ratios [95% confidence intervals] of the lower-, intermediate-, and higher-Mg groups were 1.00 (reference), 0.81 [0.66–0.99], and 0.74 [0.56–0.97], respectively.). An interaction between Mg and phosphate on the risk of cardiovascular mortality was statistically significant (P = 0.03).

Conclusion

Serum Mg levels significantly modified the mortality risk associated with hyperphosphatemia in patients undergoing hemodialysis.     

Palmitate-activated macrophages confer insulin resistance to muscle cells by a mechanism involving protein kinase C θ and ε

Background

Macrophage-derived factors contribute to whole-body insulin resistance, partly by impinging on metabolically active tissues. As proof of principle for this interaction, conditioned medium from macrophages treated with palmitate (CM-PA) reduces insulin action and glucose uptake in muscle cells. However, the mechanism whereby CM-PA confers this negative response onto muscle cells remains unknown.

Methodology/Principal Findings

L6-GLUT4myc myoblasts were exposed for 24 h to palmitate-free conditioned medium from RAW 264.7 macrophages pre-treated with 0.5 mM palmitate for 6 h. This palmitate-free CM-PA, containing selective cytokines and chemokines, inhibited myoblast insulin-stimulated insulin receptor substrate 1 (IRS1) tyrosine phosphorylation, AS160 phosphorylation, GLUT4 translocation and glucose uptake. These effects were accompanied by a rise in c-Jun N-terminal kinase (JNK) activation, degradation of Inhibitor of κBα (IκBα), and elevated expression of proinflammatory cytokines in myoblasts. Notably, CM-PA caused IRS1 phosphorylation on Ser1101, and phosphorylation of novel PKCθ and ε. Co-incubation of myoblasts with CM-PA and the novel and conventional PKC inhibitor Gö6983 (but not with the conventional PKC inhibitor Gö6976) prevented PKCθ and ε activation, JNK phosphorylation, restored IκBα mass and reduced proinflammatory cytokine production. Gö6983 also restored insulin signalling and glucose uptake in myoblasts. Moreover, co-silencing both novel PKC θ and ε isoforms in myoblasts by RNA interference, but not their individual silencing, prevented the inflammatory response and restored insulin sensitivity to CM-PA-treated myoblasts.

Conclusions/Clinical Significance

The results suggest that the block in muscle insulin action caused by CM-PA is mediated by novel PKCθ and PKCε. This study re-establishes the participation of macrophages as a relay in the action of fatty acids on muscle cells, and further identifies PKCθ and PKCε as key elements in the inflammatory and insulin resistance responses of muscle cells to macrophage products. Furthermore, it portrays these PKC isoforms as potential targets for the treatment of fatty acid-induced, inflammation-linked insulin resistance.     

Receptor for Advanced Glycation End Products - Membrane Type1 Matrix Metalloproteinase Axis Regulates Tissue Factor Expression via RhoA and Rac1 Activation in High-Mobility Group Box-1 Stimulated Endothelial Cells

Background

Atherosclerosis is understood to be a blood vessel inflammation. High-mobility group box-1 (HMGB-1) plays a key role in the systemic inflammation. Tissue factor (TF) is known to lead to inflammation which promotes thrombus formation. Membrane type1 matrix metalloprotease (MT1-MMP) associates with advanced glycation endproducts (AGE) triggered-TF protein expression and phosphorylation of NF-κB. However, it is still unclear about the correlation of MT1-MMP and HMBG-1-mediated TF expression. In this study, we investigated the molecular mechanisms of TF expression in response to HMGB-1 stimulation and the involvement of MT1-MMP in endothelial cells.

Methods and Results

Pull-down assays and Western blotting revealed that HMGB-1 induced RhoA/Rac1 activation and NF-kB phosphorylation in cultured human aortic endothelial cells. HMGB-1 increased the activity of MT1-MMP, and inhibition of RAGE or MT1-MMP by siRNA suppressed HMGB-1-induced TF upregulation as well as HMGB-1-triggered RhoA/Rac1 activation and NF-kB phosphorylation.

Conclusions

The present study showed that RAGE/MT1-MMP axis modified HMBG-1-mediated TF expression through RhoA and Rac1 activation and NF-κB phosphorylation in endothelial cells. These results suggested that MT1-MMP was involved in vascular inflammation and might be a good target for treating atherosclerosis.     

Urokinase plasminogen activator inhibits HIV virion release from macrophage-differentiated chronically infected cells via activation of RhoA and PKCε

Background

HIV replication in mononuclear phagocytes is a multi-step process regulated by viral and cellular proteins with the peculiar feature of virion budding and accumulation in intra-cytoplasmic vesicles. Interaction of urokinase-type plasminogen activator (uPA) with its cell surface receptor (uPAR) has been shown to favor virion accumulation in such sub-cellular compartment in primary monocyte-derived macrophages and chronically infected promonocytic U1 cells differentiated into macrophage-like cells by stimulation with phorbol myristate acetate (PMA). By adopting this latter model system, we have here investigated which intracellular signaling pathways were triggered by uPA/uPAR interaction leading the redirection of virion accumulation in intra-cytoplasmic vesicles.

Results

uPA induced activation of RhoA, PKCδ and PKCε in PMA-differentiated U1 cells. In the same conditions, RhoA, PKCδ and PKCε modulated uPA-induced cell adhesion and polarization, whereas only RhoA and PKCε were also responsible for the redirection of virions in intracellular vesicles. Distribution of G and F actin revealed that uPA reorganized the cytoskeleton in both adherent and polarized cells. The role of G and F actin isoforms was unveiled by the use of cytochalasin D, a cell-permeable fungal toxin that prevents F actin polymerization. Receptor-independent cytoskeleton remodeling by Cytochalasin D resulted in cell adhesion, polarization and intracellular accumulation of HIV virions similar to the effects gained with uPA.

Conclusions

These findings illustrate the potential contribution of the uPA/uPAR system in the generation and/or maintenance of intra-cytoplasmic vesicles that actively accumulate virions, thus sustaining the presence of HIV reservoirs of macrophage origin. In addition, our observations also provide evidences that pathways controlling cytoskeleton remodeling and activation of PKCε bear relevance for the design of new antiviral strategies aimed at interfering with the partitioning of virion budding between intra-cytoplasmic vesicles and plasma membrane in infected human macrophages.     

Activation of p38MAPK contributes to expanded polyglutamine-induced cytotoxicity

Background

The signaling pathways that may modulate the pathogenesis of diseases induced by expanded polyglutamine proteins are not well understood.

Methodologies/Principal Findings

Herein we demonstrate that expanded polyglutamine protein cytotoxicity is mediated primarily through activation of p38MAPK and that the atypical PKC iota (PKCι) enzyme antagonizes polyglutamine-induced cell death through induction of the ERK signaling pathway. We show that pharmacological blockade of p38MAPK rescues cells from polyglutamine-induced cell death whereas inhibition of ERK recapitulates the sensitivity observed in cells depleted of PKCι by RNA interference. We provide evidence that two unrelated proteins with expanded polyglutamine repeats induce p38MAPK in cultured cells, and demonstrate induction of p38MAPK in an in vivo model of neurodegeneration (spinocerebellar ataxia 1, or SCA-1).

Conclusions/Significance

Taken together, our data implicate activated p38MAPK in disease progression and suggest that its inhibition may represent a rational strategy for therapeutic intervention in the polyglutamine disorders.     

Anesthetic propofol reduces endotoxic inflammation by inhibiting reactive oxygen species-regulated Akt/IKKβ/NF-κB signaling

Background

Anesthetic propofol has immunomodulatory effects, particularly in the area of anti-inflammation. Bacterial endotoxin lipopolysaccharide (LPS) induces inflammation through toll-like receptor (TLR) 4 signaling. We investigated the molecular actions of propofol against LPS/TLR4-induced inflammatory activation in murine RAW264.7 macrophages.

Methodology/Principal Findings

Non-cytotoxic levels of propofol reduced LPS-induced inducible nitric oxide synthase (iNOS) and NO as determined by western blotting and the Griess reaction, respectively. Propofol also reduced the production of tumor necrosis factor-α (TNF-α), interleukin (IL)-6, and IL-10 as detected by enzyme-linked immunosorbent assays. Western blot analysis showed propofol inhibited LPS-induced activation and phosphorylation of IKKβ (Ser180) and nuclear factor (NF)-κB (Ser536); the subsequent nuclear translocation of NF-κB p65 was also reduced. Additionally, propofol inhibited LPS-induced Akt activation and phosphorylation (Ser473) partly by reducing reactive oxygen species (ROS) generation; inter-regulation that ROS regulated Akt followed by NF-κB activation was found to be crucial for LPS-induced inflammatory responses in macrophages. An in vivo study using C57BL/6 mice also demonstrated the anti-inflammatory properties against LPS in peritoneal macrophages.

Conclusions/Significance

These results suggest that propofol reduces LPS-induced inflammatory responses in macrophages by inhibiting the interconnected ROS/Akt/IKKβ/NF-κB signaling pathways.     

Thrombin induces heme oxygenase-1 expression in human synovial fibroblasts through protease-activated receptor signaling pathways

Introduction

Thrombin is a key factor in the stimulation of fibrin deposition, angiogenesis, and proinflammatory processes. Abnormalities in these processes are primary features of osteoarthritis (OA). Heme oxygenase (HO)-1 is a stress-inducible rate-limiting enzyme in heme degradation that confers cytoprotection against oxidative injury. Here, we investigated the intracellular signaling pathways involved in thrombin-induced HO-1 expression in human synovial fibroblasts (SFs).

Methods

Thrombin-mediated HO-1 expression was assessed with quantitative real-time (q)PCR. The mechanisms of action of thrombin in different signaling pathways were studied by using Western blotting. Knockdown of protease-activated receptor (PAR) proteins was achieved by transfection with siRNA. Chromatin immunoprecipitation assays were used to study in vivo binding of Nrf2 to the HO-1 promoter. Transient transfection was used to examine HO-1 activity.

Results

Osteoarthritis synovial fibroblasts (OASFs) showed significant expression of thrombin, and expression was higher than in normal SFs. OASFs stimulation with thrombin induced concentration- and time-dependent increases in HO-1 expression. Pharmacologic inhibitors or activators and genetic inhibition by siRNA of protease-activated receptors (PARs) revealed that the PAR1 and PAR3 receptors, but not the PAR4 receptor, are involved in thrombin-mediated upregulation of HO-1. Thrombin-mediated HO-1 expression was attenuated by thrombin inhibitor (PPACK), PKCδ inhibitor (rottlerin), or c-Src inhibitor (PP2). Stimulation of cells with thrombin increased PKCδ, c-Src, and Nrf2 activation.

Conclusion

Our results suggest that the interaction between thrombin and PAR1/PAR3 increases HO-1 expression in human synovial fibroblasts through the PKCδ, c-Src, and Nrf2 signaling pathways.     

Prostaglandin E2 reverses aberrant production of an inflammatory chemokine by microglia from Sandhoff disease model mice through the cAMP-PKA pathway

Background

Sandhoff disease (SD) is a neurodegenerative lysosomal β-hexosaminidase (Hex) deficiency involving excessive accumulation of undegraded substrates, including terminal GlcNAc-oligosaccharides and GM2 ganglioside. Microglia-mediated neuroinflammation contributes to the pathogenesis and progression of SD. Our previous study demonstrated that MIP-1α, a putative pathogenic factor for SD, is up-regulated in microglial cells derived from SD model mice (SD-Mg) through activation of Akt and JNK.

Methodology/Principal Findings

In this study, we first demonstrated that prostaglandin E2 (PGE2), which is one of the lipid mediators derived from arachidonic acid and is known to suppress activation of microglia, reduced the aberrant MIP-1α production by SD-Mg to the same level as by WT-Mg. PGE2 also attenuated the activation of Akt and JNK. The inhibition of MIP-1α production and the activation of Akt and JNK occurred through the EP2 and 4/cAMP/PKA signaling pathway in the murine microglia derived from SD model mice.

Conclusions/Significance

We propose that PGE2 plays a role as a negative regulator of MIP-1α production in the pathogenesis of SD, and that PGE2-EP2 and 4/cAMP/PKA signaling could be a target pathway for therapy for SD.     

Ursodeoxycholic acid is conjugated with taurine to promote secretin-stimulated biliary hydrocholeresis in the normal rat

Background & Aims

Secretin induces bicarbonate-rich hydrocholeresis in healthy individuals, but not in untreated patients with primary biliary cirrhosis (PBC). Ursodeoxycholic acid (UDCA) – the first choice treatment for PBC – restores the secretin response. Compared with humans, secretin has poor effect in experimental normal-rat models with biliary drainage, although it may elicit hydrocholeresis when the bile-acid pool is maintained. In view of the benefits of UDCA in PBC, we used normal-rat models to unravel the acute contribution of UDCA (and/or taurine-conjugated TUDCA) for eliciting the biliary secretin response.

Methods

Intravascular and/or intrabiliary administration of agonists and inhibitors was performed in normal rats with biliary monitoring. Secretin/bile-acid interplay was analyzed in 3D cultured rat cholangiocytes that formed expansive cystic structures with intralumenal hydroionic secretion.

Results

In vivo, secretin stimulates hydrocholeresis upon UDCA/TUDCA infusion, but does not modify the intrinsic hypercholeretic effect of dehydrocholic acid (DHCA). The former effect is dependent on microtubule polymerization, and involves PKCα, PI3K and MEK pathways, as shown by colchicine (i.p.) and retrograde biliary inhibitors. In vitro, while secretin alone accelerates the spontaneous expansion of 3D-cystic structures, this effect is enhanced in the presence of TUDCA, but not UDCA or DHCA. Experiments with inhibitors and Ca²⁺-chelator confirmed that the synergistic effect of secretin plus TUDCA involves microtubules, intracellular Ca²⁺, PKCα, PI3K, PKA and MEK pathways. Gene silencing also demonstrated the involvement of the bicarbonate extruder Ae2.

Conclusions

UDCA is conjugated in order to promote secretin-stimulated hydrocholeresis in rats through Ae2, microtubules, intracellular Ca²⁺, PKCα, PI3K, PKA, and MEK.     

Diosgenin, a steroidal saponin, inhibits migration and invasion of human prostate cancer PC-3 cells by reducing matrix metalloproteinases expression

Background

Diosgenin, a steroidal saponin obtained from fenugreek (Trigonella foenum graecum), was found to exert anti-carcinogenic properties, such as inhibiting proliferation and inducing apoptosis in a variety of tumor cells. However, the effect of diosgenin on cancer metastasis remains unclear. The aim of the study is to examine the effect of diosgenin on migration and invasion in human prostate cancer PC-3 cells.

Methods and Principal Findings

Diosgenin inhibited proliferation of PC-3 cells in a dose-dependent manner. When treated with non-toxic doses of diosgenin, cell migration and invasion were markedly suppressed by in vitro wound healing assay and Boyden chamber invasion assay, respectively. Furthermore, diosgenin reduced the activities of matrix metalloproteinase-2 (MMP-2) and MMP-9 by gelatin zymography assay. The mRNA level of MMP-2, -9, -7 and extracellular inducer of matrix metalloproteinase (EMMPRIN) were also suppressed while tissue inhibitor of metalloproteinase-2 (TIMP-2) was increased by diosgenin. In addition, diosgenin abolished the expression of vascular endothelial growth factor (VEGF) in PC-3 cells and tube formation of endothelial cells. Our immunoblotting assays indicated that diosgenin potently suppressed the phosphorylation of phosphatidylinositide-3 kinase (PI3K), Akt, extracellular signal regulating kinase (ERK) and c-Jun N-terminal kinase (JNK). In addition, diosgenin significantly decreased the nuclear level of nuclear factor kappa B (NF-κB), suggesting that diosgenin inhibited NF-κB activity.

Conclusion/Significance

The results suggested that diosgenin inhibited migration and invasion of PC-3 cells by reducing MMPs expression. It also inhibited ERK, JNK and PI3K/Akt signaling pathways as well as NF-κB activity. These findings reveal new therapeutic potential for diosgenin in anti-metastatic therapy.     

Hyperphosphatemia Is an Independent Risk Factor for Mortality in Critically Ill Patients: Results from a Cross-Sectional Study

Background

Phosphate imbalances or disorders have a high risk of morbidity and mortality in patients with chronic kidney disease. It is unknown if this finding extends to mortality in patients presenting at an emergency room with or without normal kidney function.

Methods and Patients

This cross sectional analysis included all emergency room patients between 2010 and 2011 at the Inselspital Bern, Switzerland. A multivariable cox regression model was applied to assess the association between phosphate levels and in-hospital mortality up to 28 days.

Results

22,239 subjects were screened for the study. Plasma phosphate concentrations were measured in 2,390 patients on hospital admission and were included in the analysis. 3.5% of the 480 patients with hypophosphatemia and 10.7% of the 215 patients with hyperphosphatemia died. In univariate analysis, phosphate levels were associated with mortality, age, diuretic therapy and kidney function (all p<0.001). In a multivariate Cox regression model, hyperphosphatemia (OR 3.29, p<0.001) was a strong independent risk factor for mortality. Hypophosphatemia was not associated with mortality (p>0.05).

Conclusion

Hyperphosphatemia is associated with 28-day in-hospital mortality in an unselected cohort of patients presenting in an emergency room.