Ochratoxin A induces hepatic and renal toxicity in mice through increased oxidative stress, mitochondrial damage, and multiple cell death mechanisms.

Ochratoxin A (OTA) is a widespread food toxin produced by Aspergillus ochraceus and other molds. In this study, we developed and established acute OTA toxicity conditions in mice, which received daily oral doses of OTA between 0.5 up to 8 mg/kg body weight up to 7 days and were subjected to histological and biochemical analysis to characterize renal and hepatic damage. Oral administration of OTA for 7 days resulted in loss of body weight in a dose-dependent manner and increased the levels of serum biomarkers of hepatic and renal damage. The kidney was more sensitive to OTA-induced damage than the liver. In addition to necrosis, OTA induced hepatic and renal apoptosis in dose- and time-dependent manners. Especially, a high dose of OTA (8 mg/kg body weight) administered for 7 days led to necroptosis in both liver and kidney tissues. OTA dose-dependently increased the oxidative stress levels, including lipid peroxidation, in the liver and kidneys. OTA disrupted mitochondrial dynamics and structure in hepatic and renal cells, leading to the dysregulation of mitochondrial homeostasis. OTA increased transferrin receptor 1 and decreased glutathione peroxidase 4 levels in a dose- and time-dependent manner. These results suggest the induction of ferroptosis. Collectively, this study highlighted the characteristics of acute OTA-induced hepatic and renal toxicity in mice in terms of oxidative stress, mitochondrial damage, and multiple cell death mechanisms, including necroptosis and ferroptosis.

p90RSK Inhibition Ameliorates TGF-ß1 Signaling and Pulmonary Fibrosis by Inhibiting Smad3 Transcriptional Activity.

BACKGROUND/AIMS: Idiopathic pulmonary fibrosis (IPF) is a specific form of progressive and chronic interstitial lung disease of unknown cause. IPF is characterized by excessive deposition of extracellular matrix (ECM) and destructive pathological remodeling due to epithelial-to-mesenchymal transition (EMT). Eventually, lung interstitium thickens and stiffens and breathing becomes difficult. It has been well established that the transforming growth factor-ß1 (TGF-ß1)/Smad signaling pathway plays a critical role in the pathogenesis of pulmonary fibrosis. TGF-ß1-mediated activation of mitogen activated protein kinase (MAPK) family affects Smad signaling. p90RSK is a serine/threonine kinase and is activated by the extracellular signal-regulated kinase (ERK) signaling pathway. However, the roles played by p90RSK in TGF-ß1 signaling and the pathogenesis of pulmonary fibrosis remain unknown. METHODS: We investigated whether p90RSK regulates the pathogenesis of pulmonary fibrosis using in vitro and in vivo systems and Western blotting, real-time quantitative PCR, transcriptional activity assays and immunofluorescence studies. RESULTS: Pharmacological inhibition of p90RSK by FMK or inhibition of p90RSK with adenoviral vector encoding a dominant negative form of p90RSK suppressed TGF-ß1-induced ECM accumulation and EMT in lung epithelial cells and fibroblasts. Interestingly, FMK significantly inhibited TGF-ß1-induced Smad3 nuclear translocation and smad binding element-dependent transcriptional activity, but not Smad3 phosphorylation. Furthermore, in a mouse model of bleomycin-induced lung fibrosis, FMK ameliorated pulmonary fibrosis. CONCLUSION: These findings indicate that p90RSK plays critical roles in pulmonary fibrosis, which suggests it be viewed as a novel therapeutic target for the treatment of lung fibrosis.

A common signaling pathway leading to degranulation in mast cells and its regulation by CCR1-ligand.

BACKGROUND: Signal transduction pathways mediated by various receptors expressed on mast cells are thought to be complex, and inhibitory signals that turn off activating signals are not known. METHODS: Upstream signaling cascades mediated by several known receptors in bone marrow-derived mast cells that lead to degranulation and mediator release were studied by immunoblotting and immunoprecipitation. Small interfering RNAs and knockout mice were used to confirm findings. RESULTS: All ligands tested including IgE/Ag, SCF, HSP70, CCL3, and its valiant eMIP induced phosphorylation of linker for activation of T cells (LAT), which triggered their receptor-mediated downstream signaling cascades that controlled degranulation and mediator release. Phosphorylation of lymphocyte-specific protein kinase (Lck) was induced by each ligand, which commonly played an indispensable role in LAT phosphorylation. In contrast, phosphorylation of spleen tyrosine kinase was additionally induced in cells stimulated only with IgE/Ag and SCF, which is also associated with LAT phosphorylation in part. Degranulation and mediator release induced by IgE/Ag, SCF, or HSP70 were enhanced by nanomolar doses of CCR1 ligands CCL3 and eMIP via enhanced LAT phosphorylation. On the other hand, micromolar doses of CCR1 ligand inhibited degranulation and mediator release from mast cells stimulated with IgE/Ag, SCF, or HSP70 by de-phosphorylation of phosphorylated Lck with Src homology region 2 domain-containing phosphatase-1. CONCLUSIONS: Linker for activation of T cells plays a central role in signal transduction pathways in mast cells stimulated with any ligand tested. Dose-dependent alternate costimulation and inhibition of CCR1 ligands in IgE/Ag-, SCF-, or HSP70-stimulated mast cells occur at the level of Lck-LAT phosphorylation.

Quercetin-induced apoptosis ameliorates vascular smooth muscle cell senescence through AMP-activated protein kinase signaling pathway.

Aging is one of the risk factors for the development of cardiovascular diseases. During the progression of cellular senescence, cells enter a state of irreversible growth arrest and display resistance to apoptosis. As a flavonoid, quercetin induces apoptosis in various cells. Accordingly, we investigated the relationship between quercetin-induced apoptosis and the inhibition of cellular senescence, and determined the mechanism of oxidative stress-induced vascular smooth muscle cell (VSMC) senescence. In cultured VSMCs, hydrogen peroxide (H2O2) dose-dependently induced senescence, which was associated with increased numbers of senescence-associated ß-galactosidase-positive cells, decreased expression of SMP30, and activation of p53-p21 and p16 pathways. Along with senescence, expression of the anti-apoptotic protein Bcl-2 was observed to increase and the levels of proteins related to the apoptosis pathway were observed to decrease. Quercetin induced apoptosis through the activation of AMP-activated protein kinase. This action led to the alleviation of oxidative stress-induced VSMC senescence. Furthermore, the inhibition of AMPK activation with compound C and siRNA inhibited apoptosis and aggravated VSMC senescence by reversing p53-p21 and p16 pathways. These results suggest that senescent VSMCs are resistant to apoptosis and quercetin-induced apoptosis attenuated the oxidative stress-induced senescence through activation of AMPK. Therefore, induction of apoptosis by polyphenols such as quercetin may be worthy of attention for its anti-aging effects.

Correlation among lipid parameters, pulse wave velocity and central blood pressure in young Korean population.

BACKGROUND: Central blood pressure is closely related to the important cardiovascular intermediate end points, such as vascular hypertrophy and extent of carotid atherosclerosis. Therefore it is prudent to study correlation among central aortic blood pressure, body composition, lipid profiles, and pulse wave velocity in population-based study. Consequently, we investigate the correlation between central aortic blood pressure and other risk parameters of hypertension such as body composition and lipid profile. METHODS: We recruited 20 young participants diagnosed with hypertension as well as 30 without hypertension. The study used an X-SCAN PLUS 950 machine for measurement of overall body composition. Measurements of central blood pressure and carotid-femoral pulse wave velocity were carried out using SphygmoCor XCEL. RESULTS: The hypertensive participants had significantly higher total weight without fat, body moisture mass, muscle mass, body mass index, basal metabolic rate, intracellular and extracellular water contents, protein and mineral contents along with brachial and central aortic blood pressures. In both hypertensive and non-hypertensive participants, central aortic diastolic blood pressure were significantly related to the lipid parameters. CONCLUSION: Overall, the correlations between central blood pressure, pulse wave velocity, and lipid profile in hypertensive and non-hypertensive participants were substantial.

Correction: Cho, K.H., et al. Blood pressure Lowering Effect of Cuban Policosanol is Accompanied by Improvement of Hepatic Inflammation, Lipoprotein profile, and HDL Quality in Spontaneously Hypertensive Rats. Molecules 2018, 23, 1080.

The authors wish to make the following correction to their paper [...].

Engineered islet cell clusters transplanted into subcutaneous space are superior to pancreatic islets in diabetes.

An alternative route for pancreatic islet transplantation is the subcutaneous space; however, inadequate vascularization in the subcutaneous space limits the availability of oxygen and nutrients to the subcutaneously transplanted islets, which leads to the development of a necrotic core in the islets, thereby causing islet dysfunction. Thus, we aimed to prevent the early apoptosis of pancreatic islets after transplantation into subcutaneous space by preparing islet clusters of appropriate size. We prepared fully functional islet cell clusters (ICCs) by using the hanging-drop technique. We optimized the size of ICCs on the basis of viability and functionality after culture in an hypoxic environment. We transplanted ICCs into the subcutaneous space of diabetic mice and evaluated the viability of the islets at the transplantation site. In an hypoxic environment, ICCs exhibited improved viability and functionality compared with control islets. ICCs, upon transplantation into the hypoxic subcutaneous space of diabetic mice, showed better glycemic control compared with control islets. Live/dead imaging of the islets after retrieval from the transplanted area revealed significantly reduced apoptosis in ICCs. Transplantation of ICCs may be an attractive strategy to prevent islet cell apoptosis that results from nonimmune-mediated physiologic stress at the transplantation site.-Pathak, S., Regmi, S., Gupta, B., Pham, T. T., Yong, C. S., Kim, J. O., Yook, S., Kim, J.-R., Park, M. H., Bae, Y. K., Jeong, J.-H. Engineered islet cell clusters transplanted into subcutaneous space are superior to pancreatic islets in diabetes.

Blood Pressure Lowering Effect of Cuban Policosanol is Accompanied by Improvement of Hepatic Inflammation, Lipoprotein Profile, and HDL Quality in Spontaneously Hypertensive Rats.

We investigated the antihypertensive effect of policosanol on spontaneously hypertensive rats (SHR). For this, we analyzed blood pressure, blood lipid, and lipoprotein properties in male SHR after consumption of Cuban policosanol (PCO). The experimental groups were as follows: normotensive Wistar Kyoto (WKY) control, SHR group fed normal diet (ND), SHR group fed 20 mg of PCO, SHR group fed 100 mg of PCO, and SHR group fed 200 mg of PCO per kg of body weight. After eight weeks, the SHR control group showed gradual increases up to 21% in systolic blood pressure (SBP) and diastolic blood pressure (DBP) compared with values at week 0. However, policosanol consumption had a dose-dependent reduction effect on SBP and also reduced DBP up to 17% in a dose-dependent manner. Heart rate (HR) bpm increased by six percent in the SHR control, whereas the 20 mg, 100 mg, and 200 mg of policosanol groups showed a reduction of 36%, 28%, and 34% respectively. Although serum total cholesterol (TC) level of SHR was not affected by policosanol consumption (70⁻80 mg/dL), serum triglyceride (TG) level significantly decreased in the SHR + 200 mg of PCO group. Serum high-density lipoprotein cholesterol (HDL-C) level was also significantly elevated by policosanol consumption. The % HDL-C/TC ratio was elevated in the policosanol group up to 67⁻70%, whereas the SHR control group showed a ratio of 58%. Serum cholesteryl ester transfer protein (CETP) activity was reduced by policosanol in a dose-dependent manner. Although the serum glutamate oxaloacetate transaminase (GOT)/ glutamate pyruvate transaminase (GPT) were similar across all groups, policosanol consumption caused reduction of reactive oxygen species (ROS) levels in hepatic tissue. The SHR control group showed a 2.1-fold higher serum C-reactive protein (CRP) level than the WKY group, whereas the CRP level decreased in the SHR + 200 mg of PCO group (up to 45%) than SHR control group. Aldosterone level was reduced in the policosanol group (up to 34%) in a dose-dependent manner compared to the control. In conclusion, eight weeks of policosanol consumption in SHR resulted in remarkable reduction of blood pressure, serum aldosterone, and serum TG levels along with the elevation of HDL-C and improvement of hepatic inflammation.

Elevated Interleukin-13 Receptor Alpha 1 Expression in Tumor Cells Is Associated with Poor Prognosis in Patients with Invasive Breast Cancer.

BACKGROUND: Interleukin (IL)-13 is an immunoregulatory, anti-inflammatory cytokine that is produced by numerous immune cells, and plasma membrane receptor for IL-13 (IL-13R) is known to be expressed in various human malignancies and in immune cells. METHODS: The authors evaluated the expression of IL-13R alpha 1 (IL-13Rα1, an IL-13R subtype) by immunohistochemistry in tissue microarrays of 1213 invasive breast cancer (IBC) samples to determine the prognostic value of IL-13Rα1 expression. RESULTS: High IL-13Rα1 expression was observed in 619 (51%) cases and was found to be associated with an older (≥50 years) age (p = 0.022), lymph node metastasis (p = 0.015), ductal and micropapillary histologic subtypes (p < 0.001), lymphovascular invasion (p = 0.012), HER2 positivity (p < 0.001), and a high (>20%) Ki-67 index (p = 0.039). No significant correlation was found between IL-13Rα1 expression and clinicopathological variables, including tumor size, histological grade, hormone receptor expressions, and tumor-infiltrating lymphocyte levels. Patients with high IL-13Rα1 expression showed poorer overall survival (p = 0.044) and disease-free survival (DFS, p = 0.001) than those with low/negative expression. Subgroup analysis revealed an association between IL-13Rα1 expression and survival for HER2-negative, but not for HER2-positive tumors. Multivariate analysis showed high IL-13Rα1 expression was an independent negative prognostic factor of DFS (p = 0.019). CONCLUSIONS: The results of this study suggest the IL-13 and IL-13Rα1 interaction promotes cancer cell growth and metastasis, and IL-13Rα1 expression is a potential prognostic marker in IBC.

Antisenescence effect of mouse embryonic stem cell conditioned medium through a PDGF/FGF pathway.

Cellular senescence, an irreversible state of growth arrest, underlies organismal aging and age-related diseases. Recent evidence suggests that aging intervention based on inhibition of cellular senescence might be a promising strategy for treatment of aging and age-related diseases. Embryonic stem cells (ESCs) and ESC conditioned medium (CM) have been suggested as a desirable source for regenerative medicine. However, effects of ESC-CM on cellular senescence remain to be determined. We found that treatment of senescent human dermal fibroblasts with CM from mouse ESCs (mESCs) decreases senescence phenotypes. We found that platelet-derived growth factor BB in mESC-CM plays a critical role in antisenescence effect of mESC-CM through up-regulation of fibroblast growth factor 2. We confirmed that mESC-CM treatment accelerates the wound-healing process by down-regulating senescence-associated p53 expression in in vivo models. Taken together, our results suggest that mESC-CM has the ability to suppress cellular senescence and maintain proliferative capacity. Therefore, this strategy might emerge as a novel therapeutic strategy for aging and age-related diseases.

CD9 monoclonal antibody-conjugated PEGylated liposomes for targeted delivery of rapamycin in the treatment of cellular senescence.

Premature cellular senescence refers to the state of irreversible cell cycle arrest due to DNA damage or other stresses. In this study, CD9 monoclonal antibody (CD9mAb) was successfully conjugated to the surface of PEGylated liposomes for targeted delivery of rapamycin (LR-CD9mAb) to overcome senescence of CD9 receptor-overexpressing cells. LR-CD9mAb has a small particle size (143.3 ± 2.4 nm), narrow size distribution (polydispersity index: 0.220 ± 0.036), and negative zeta potential (-14.6 ± 1.2 mV). The uptake of CD9-targeted liposomes by premature senescent human dermal fibroblasts (HDFs) was higher than that by young HDFs, as displayed by confocal microscopic images. The senescence might not be reversed by treatment with rapamycin; however, the drug promoted cell proliferation and reduced the number of cells that expressed the senescence-associated-ß-galactosidase (SA-ß-gal). These effects were further confirmed by cell viability, cell cycle, and Western blotting analyses. Moreover, CD9-targeted liposomes showed better anti-senescence activity, in comparison with free rapamycin or the conventional liposomal formulation, suggesting the potential application of this system in further in vivo studies.

Fructated apolipoprotein A-I exacerbates cellular senescence in human umbilical vein endothelial cells accompanied by impaired insulin secretion activity and embryo toxicity.

Glycation of apolipoproteins is a major feature of the production of dysfunctional high-density lipoprotein (HDL), which is associated with the incidence of several metabolic diseases such as coronary artery disease and diabetes. In this report, fructated apoA-I (fA-I) induced by fructose treatment showed a covalently multimerized band without cross-linking, and lysine residues were irreversibly modified to prevent crosslinking. Using pancreatic ß-cells, insulin secretion was impaired by fA-I in the lipid-free and reconstituted HDL (rHDL) states, by up to 35%, and 40%, respectively, under hyperglycemic conditions (25 mmol/L glucose). Treatment of human umbilical vein endothelial cells (HUVECs) with fA-I and HDL from elderly patients caused a 1.8-fold and 1.5-fold increased cellular senescence, respectively, along with increased lysosomal enlargement. In the lipid-free and rHDL states, fA-I increased embryo death by 1.5-fold and 2.5-fold, respectively, along with the production of oxidized species. Furthermore, rHDL containing fA-I (fA-I-rHDL) showed a higher isoelectric point (pI, approximately 8.5), whereas rHDL containing nA-I (nA-I-rHDL) showed a narrow band range with lower pI (around 8.0) as well as a much smaller particle size than that of nA-I-rHDL. In conclusion, fructose-mediated apoA-I fructation resulted in the severe loss of several beneficial functions of apoA-I and HDL, including anti-senescence and insulin secretion activities, accompanied with increased susceptibility to protein degradation and structural modification.

Estrogenic compound attenuates angiotensin II-induced vascular smooth muscle cell proliferation through interaction between LKB1 and estrogen receptor α.

The prevalence rate of cardiovascular disease is higher for males than females, and estradiol (E2) induces AMP-activated protein kinase (AMPK) activation, which is known to regulate proliferation of VSMC. We identified the estrogenic properties of nordihydroguaiaretic acid (NDGA, a lignan phytoestrogen) that inhibit VSMC proliferation and explored the underlying mechanisms. Both the phosphorylation and expression of LKB1 were increased by NDGA. In addition, NDGA significantly attenuated angiotensin II (Ang II)-induced VSMC proliferation. To elucidate the estrogenic effects, we confirmed that NDGA increased estrogen receptor α (ERα) expression, similar to treatment with E2 and estriol (E3). Furthermore, tamoxifen and ERα siRNA obstructed the effects of NDGA including ERα expression, AMPK phosphorylation and both LKB1 phosphorylation and expression. VSMC proliferation was restored by tamoxifen and ERα siRNA. LKB1 siRNA also reversed the NDGA-mediated inhibition of VSMC proliferation. The estrogenic activity of NDGA induced LKB1 translocation from nucleus to cytosol, and tamoxifen obstructed LKB1 translocation. The absence of LKB1 completely abolished the increase of ERα expression induced by NDGA. Taken together, the beneficial effects of estrogenic compound (E2 and NDGA) on inhibition of VSMC proliferation are mediated by interaction between LKB1 and ERα, suggesting a potential mechanism for females having less cardiovascular disease.

Delphinidin prevents high glucose-induced cell proliferation and collagen synthesis by inhibition of NOX-1 and mitochondrial superoxide in mesangial cells.

This study examined the effect of delphinidin on high glucose-induced cell proliferation and collagen synthesis in mesangial cells. Glucose dose-dependently (5.6-25 mM) increased cell proliferation and collagen I and IV mRNA levels, whereas pretreatment with delphinidin (50 µM) prevented cell proliferation and the increased collagen mRNA levels induced by high glucose (25 mM). High glucose increased reactive oxygen species (ROS) generation, and this was suppressed by pretreating delphinidin or the antioxidant N-acetyl cysteine. NADPH oxidase (NOX) 1 was upregulated by high glucose, but pretreatment with delphinidin abrogated this upregulation. Increased mitochondrial superoxide by 25 mM glucose was also suppressed by delphinidin. The NOX inhibitor apocynin and mitochondria-targeted antioxidant Mito TEMPO inhibited ROS generation and cell proliferation induced by high glucose. Phosphorylation of extracellular signal regulated kinase (ERK)1/2 was increased by high glucose, which was suppressed by delphinidin, apocynin or Mito TEMPO. Furthermore, PD98059 (an ERK1/2 inhibitor) prevented the high glucose-induced cell proliferation and increased collagen mRNA levels. Transforming growth factor (TGF)-ß protein levels were elevated by high glucose, and pretreatment with delphinidin or PD98059 prevented this augmentation. These results suggest that delphinidin prevents high glucose-induced cell proliferation and collagen synthesis by inhibition of NOX-1 and mitochondrial superoxide in mesangial cells.

Delayed-onset hearing loss in pediatric candidates for cochlear implantation.

The objective of this study was to evaluate the clinical significance of delayed-onset hearing loss in children. Seventy-three children who underwent cochlear implantation (CI) were included. They were divided into a congenital hearing loss group (n = 50) and a delayed-onset hearing loss group (n = 23). The age at diagnosis of hearing loss, age at the beginning of auditory habilitation, the age at CI, and the postimplant speech perception abilities were compared between the two groups. Children in the congenital hearing loss group were confirmed to have hearing loss at a mean age of 0.3 years, and those in the delayed-onset hearing loss group were diagnosed with hearing loss at a mean age of 2.0 years. Auditory habilitation began at a mean age of 0.4 and 2.0 years, and CI was performed at a mean age of 1.4 and 2.6 years, respectively. Children in the congenital hearing loss group had better scores on speech perception tests than those in the delayed-onset hearing loss group, but the differences were not significant. About half of the children with delayed-onset hearing loss (57 %) had risk factors associated with delayed-onset hearing loss. A high prevalence of delayed-onset hearing loss was noted in the group of children who underwent CI. Risk factors for hearing loss were not found in 43 % of children with delayed-onset hearing loss. Universal screening for delayed-onset hearing loss needs to be performed during early childhood.

Chemical constituents of Hericium erinaceum associated with the inhibitory activity against cellular senescence in human umbilical vascular endothelial cells.

Hericium erinaceum is an edible and medicinal mushroom widely used in Korea, Japan, and China. On the search for biologically active compounds supporting the medicinal usage, the MeOH extract of the fruiting bodies of H. erinaceum was investigated for its chemical constituents. Six compounds were isolated and identified as hericenone D (1), (22E,24R)-5α,8α-epidioxyergosta-6,22-dien-3ß-ol (2), erinacerin B (3), hericenone E (4), hericenone F (5) and isohericerin (6) by comparing their spectroscopic data with previously reported values. The inhibitory effects on adriamycin-induced cellular senescence in human dermal fibroblasts (HDFs) and human umbilical vein endothelial cells (HUVECs) of the isolates (1-6) were studied. Among the isolated compounds, ergosterol peroxide (2) reduced senescence associated ß-galactosidase (SA-ß-gal) activity increased in HUVECs treated with adriamycin. According to experimental data obtained, the active compound may inspire the development of a new pharmacologically useful substance to be used in the treatment and prevention of age-related diseases.

Intermittent fasting promotes rejuvenation of immunosenescent phenotypes in aged adipose tissue.

The aging of white adipose tissue (WAT) involves senescence of adipose stem and progenitor cells (ASPCs) and dysregulation of immune cell populations, serving as a major driver of age-associated adipose dysfunction and metabolic diseases. Conversely, the elimination of senescent ASPCs is associated with improvements in overall health. Intermittent fasting (IF), a dietary intervention that incorporates periodic cycles of fasting and refeeding, has been reported to promote weight loss and fat mass reduction and improve glucose and insulin homeostasis in both murine and human studies. While previous studies have assessed the effects of IF on obesity-associated metabolic dysfunction, few studies have examined the aging-specific changes to ASPCs and immune cell populations in WAT. Here, we show that IF in 18-20-month-old mice reduced senescent phenotypes of ASPCs and restored their adipogenic potential. Intriguingly, IF-treated mice exhibited an increase in adipose eosinophils, which has been reported to be associated with improved WAT homeostasis and immunological fitness in aged mice. The observed cellular and metabolic changes suggest that IF may be a feasible lifestyle regimen to reduce cellular senescence which could result in attenuation of downstream aging-induced WAT dysfunction and metabolic diseases.

FLRT2 prevents endothelial cell senescence and vascular aging by regulating the ITGB4/mTORC2/p53 signaling pathway.

The roles of fibronectin leucine-rich transmembrane protein 2 (FLRT2) in physiological and pathological processes are not well known. Here, we identify a potentially novel function of FLRT2 in preventing endothelial cell senescence and vascular aging. We found that FLRT2 expression was lower in cultured senescent endothelial cells as well as in aged rat and human vascular tissues. FLRT2 mediated endothelial cell senescence via the mTOR complex 2, AKT, and p53 signaling pathway in human endothelial cells. We uncovered that FLRT2 directly associated with integrin subunit beta 4 (ITGB4) and thereby promoted ITGB4 phosphorylation, while inhibition of ITGB4 substantially mitigated the induction of senescence triggered by FLRT2 depletion. Importantly, FLRT2 silencing in mice promoted vascular aging, and overexpression of FLRT2 rescued a premature vascular aging phenotype. Therefore, we propose that FLRT2 could be targeted therapeutically to prevent senescence-associated vascular aging.

Hepatocyte growth factor-mediated gastrin-releasing peptide induces IL-8 expression through Ets-1 in gastric cancer cells.

Gastric cancer cells secrete a variety of proangiogenic molecules, including IL-8 and VEGF. However, factors regulating the expression of proangiogenic genes for gastric cancer remain largely undefined. We investigated the role of HGF-induced activation of GRP and Ets-1 transcription factor in expression of the proangiogenic factor IL-8. The genes associated with angiogenesis induced by HGF were screened using cDNA micro-array technology in two gastric cancer cell lines (NUGC-3 and MKN-28). First, GRP RNA and protein were confirmed to be upregulated. Then, expression of GRP, Ets-1, and IL-8 were further estimated by Western blot analysis. A role for Ets-1 in HGF-induced upregulation of IL-8 was determined by knockdown of Ets-1 with Ets-1 sh-RNA and a chromatin immune precipitation assay. The levels of GRP, Ets-1, and IL-8 were upregulated in cells treated with HGF in a dose-dependent manner. HGF-induced expression of Ets-1 and IL-8 was increased more by GRP treatment and inhibited by pretreatment with an ERK 1/2 inhibitor (PD098059). HGF-induced upregulation of IL-8 was repressed by Ets-1 knockdown. HGF enhanced the binding activity of Ets-1 to the IL-8 promoter in control cells, but not in the Ets-1 shRNA cells. We confirmed the functional role of HGF-induced Ets-1 in activation of the IL-8 promoter by the reporter gene assay. Downregulation of IL-8 also decreased in vitro cell invasion. In conclusion, HGF mediated the GRP induction of IL-8 expression through Ets-1, which thus might serve as a promising target for gastric cancer therapy.

α-Lipoic acid attenuates vascular calcification via reversal of mitochondrial function and restoration of Gas6/Axl/Akt survival pathway.

Vascular calcification is prevalent in patients with chronic kidney disease and leads to increased cardiovascular morbidity and mortality. Although several reports have implicated mitochondrial dysfunction in cardiovascular disease and chronic kidney disease, little is known about the potential role of mitochondrial dysfunction in the process of vascular calcification. This study investigated the effect of α-lipoic acid (ALA), a naturally occurring antioxidant that improves mitochondrial function, on vascular calcification in vitro and in vivo. Calcifying vascular smooth muscle cells (VSMCs) treated with inorganic phosphate (Pi) exhibited mitochondrial dysfunction, as demonstrated by decreased mitochondrial membrane potential and ATP production, the disruption of mitochondrial structural integrity and concurrently increased production of reactive oxygen species. These Pi-induced functional and structural mitochondrial defects were accompanied by mitochondria-dependent apoptotic events, including release of cytochrome c from the mitochondria into the cytosol, subsequent activation of caspase-9 and -3, and chromosomal DNA fragmentation. Intriguingly, ALA blocked the Pi-induced VSMC apoptosis and calcification by recovery of mitochondrial function and intracellular redox status. Moreover, ALA inhibited Pi-induced down-regulation of cell survival signals through the binding of growth arrest-specific gene 6 (Gas6) to its cognate receptor Axl and subsequent Akt activation, resulting in increased survival and decreased apoptosis. Finally, ALA significantly ameliorated vitamin D(3) -induced aortic calcification and mitochondrial damage in mice. Collectively, the findings suggest ALA attenuates vascular calcification by inhibiting VSMC apoptosis through two distinct mechanisms; preservation of mitochondrial function via its antioxidant potential and restoration of the Gas6/Axl/Akt survival pathway.