Ethanol-induced ceramide production causes neuronal apoptosis by increasing MCL-1S-mediated ER-mitochondria contacts.

Heavy alcohol consumption causes neuronal cell death and cognitive impairment. Neuronal cell death induced by ethanol may result from increased production of the sphingolipid metabolite ceramide. However, the molecular mechanisms of neuronal cell death caused by ethanol-induced ceramide production have not been elucidated. Therefore, we investigated the mechanism through which ethanol-induced ceramide production causes neuronal cell apoptosis using human induced-pluripotent stem cell-derived neurons and SH-SY5Y cells and identified the effects of ceramide on memory deficits in C57BL/6 mice. First, we found that ethanol-induced ceramide production was decreased by inhibition of the de novo synthesis pathway, mediated by serine palmitoyltransferase (SPT). The associated alterations of the molecules related to the ceramide pathway suggest that the elevated level of ceramide activated protein phosphatase 1 (PP1), which inhibited the nuclear translocation of serine/arginine-rich splicing factor 1 (SRSF1). This led to aberrant splicing of myeloid cell leukemia 1 (MCL-1) pre-mRNA, which upregulated MCL-1S expression. Our results demonstrated that the interaction of MCL-1S with the inositol 1, 4, 5-trisphosphate receptor (IP3R) increases calcium release from the endoplasmic reticulum (ER) and then activated ER-bound inverted formin 2 (INF2). In addition, we discovered that F-actin polymerization through INF2 activation promoted ER-mitochondria contacts, which induced mitochondrial calcium influx and mitochondrial reactive oxygen species (mtROS) production. Markedly, MCL-1S silencing decreased mitochondria-associated ER membrane (MAM) formation and prevented mitochondrial calcium influx and mtROS accumulation, by inhibiting INF2-dependent actin polymerization interacting with mitochondria. Furthermore, the inhibition of ceramide production in ethanol-fed mice reduced MCL-1S expression, neuronal cell death, and cognitive impairment. In conclusion, we suggest that ethanol-induced ceramide production may lead to mitochondrial calcium overload through MCL-1S-mediated INF2 activation-dependent MAM formation, which promotes neuronal apoptosis.

Prenatal glucocorticoid exposure selectively impairs neuroligin 1-dependent neurogenesis by suppressing astrocytic FGF2-neuronal FGFR1 axis.

Exposure to maternal stress irreversibly impairs neurogenesis of offspring by inducing life-long effects on interaction between neurons and glia under raging differentiation process, culminating in cognitive and neuropsychiatric abnormalities in adulthood. We identified that prenatal exposure to stress-responsive hormone glucocorticoid impaired neurogenesis and induced abnormal behaviors in ICR mice. Then, we used human induced pluripotent stem cell (iPSC)-derived neural stem cell (NSC) to investigate how neurogenesis deficits occur. Following glucocorticoid treatment, NSC-derived astrocytes were found to be A1-like neurotoxic astrocytes. Moreover, cortisol-treated astrocytic conditioned media (ACM) then specifically downregulated AMPA receptor-mediated glutamatergic synaptic formation and transmission in differentiating neurons, by inhibiting localization of ionotropic glutamate receptor (GluR)1/2 into synapses. We then revealed that downregulated astrocytic fibroblast growth factor 2 (FGF2) and nuclear fibroblast growth factor receptor 1 (FGFR1) of neurons are key pathogenic factors for reducing glutamatergic synaptogenesis. We further confirmed that cortisol-treated ACM specifically decreased the binding of neuronal FGFR1 to the synaptogenic NLGN1 promoter, but this was reversed by FGFR1 restoration. Upregulation of neuroligin 1, which is important in scaffolding GluR1/2 into the postsynaptic compartment, eventually normalized glutamatergic synaptogenesis and subsequent neurogenesis. Moreover, pretreatment of FGF2 elevated neuroligin 1 expression and trafficking of GluR1/2 into the postsynaptic compartment of mice exposed to prenatal corticosterone, improving spatial memory and depression/anxiety-like behaviors. In conclusion, we identified neuroligin 1 restoration by astrocytic FGF2 and its downstream neuronal nuclear FGFR1 as a critical target for preventing prenatal stress-induced dysfunction in glutamatergic synaptogenesis, which recovered both neurogenesis and hippocampal-related behaviors.

Cyanidin 3-O-arabinoside suppresses DHT-induced dermal papilla cell senescence by modulating p38-dependent ER-mitochondria contacts.

BACKGROUND: Androgenetic alopecia (AGA) is a genetic disorder caused by dihydrotestosterone (DHT), accompanied by the senescence of androgen-sensitive dermal papilla cells (DPCs) located in the base of hair follicles. DHT causes DPC senescence in AGA through mitochondrial dysfunction. However, the mechanism of this pathogenesis remains unknown. In this study, we investigated the protective role of cyanidins on DHT-induced mitochondrial dysfunction and DPC senescence and the regulatory mechanism involved. METHODS: DPCs were used to investigate the effect of DHT on mitochondrial dysfunction with MitoSOX and Rhod-2 staining. Senescence-associated ß-galactosidase activity assay was performed to examine the involvement of membrane AR-mediated signaling in DHT-induced DPC senescence. AGA mice model was used to study the cyanidins on DHT-induced hair growth deceleration. RESULTS: Cyanidin 3-O-arabinoside (C3A) effectively decreased DHT-induced mtROS accumulation in DPCs, and C3A reversed the DHT-induced DPC senescence. Excessive mitochondrial calcium accumulation was blocked by C3A. C3A inhibited p38-mediated voltage-dependent anion channel 1 (VDAC1) expression that contributes to mitochondria-associated ER membrane (MAM) formation and transfer of calcium via VDAC1-IP3R1 interactions. DHT-induced MAM formation resulted in increase of DPC senescence. In AGA mice models, C3A restored DHT-induced hair growth deceleration, which activated hair follicle stem cell proliferation. CONCLUSIONS: C3A is a promising natural compound for AGA treatments against DHT-induced DPC senescence through reduction of MAM formation and mitochondrial dysfunction.

Low Bone Mineral Density in Young Patients Newly Diagnosed with Inflammatory Bowel Disease.

BACKGROUND: The prevalence and risk factors of low bone mineral density (BMD) in Asian patients newly diagnosed with inflammatory bowel disease (IBD) have not been fully suggested. AIMS: We aimed to examine the prevalence and risk factors of low BMD in young Korean patients newly diagnosed with IBD. METHODS: We prospectively enrolled 132 patients aged less than 50 years and newly diagnosed with IBD from six tertiary referral centers in Korea between November 2014 and April 2017. BMD was measured by dual-energy X-ray absorptiometry, and then the Z-score was determined. We defined low BMD as a Z-score ≤ - 1.0. RESULTS: Of 68 patients with ulcerative colitis (UC), 22 (32.4%) had low BMD. Also, of 64 patients with Crohn's disease (CD), 24 (37.5%) showed low BMD. Results from multivariate regression analysis identified the risk factors for low BMD as a high level of alkaline phosphatase (ALP) (≥ 140 U/L) (P = 0.010) in UC patients, and being underweight (body mass index ≤ 18.5 kg/m2) (P = 0.017) in CD patients. CONCLUSIONS: Our study showed that about one-third of newly diagnosed IBD Asian patients had low BMD. The clinical factors associated with low BMD were a high level of ALP in UC patients, and being underweight, in CD patients. Therefore, measurements of BMD in young patients should be considered at the diagnosis of IBD.

Melatonin restores Muc2 depletion induced by V. vulnificus VvpM via melatonin receptor 2 coupling with Gαq.

BACKGROUND: Melatonin (5-methoxy-N-acetyltryptamine), a hormone produced in the pineal gland, has a variety of biological functions as an antioxidant, but a functional role of melatonin in the regulation of intestinal mucin (Muc) production during bacterial infection has yet to be described in detail. In this study, we investigate the effects of melatonin during Muc2 repression elicited by the Gram-negative bacterium V. vulnificus. METHODS: Mucus-secreting human HT29-MTX cells were used to study the functional role of melatonin during Muc2 depletion induced by the recombinant protein (r) VvpM produced by V. vulnificus. The regulatory effects of melatonin coupling with melatonin receptor 2 (MT2) on the production of reactive oxygen species (ROS), the activation of PKCδ and ERK, and the hypermethylation of the Muc2 promoter as induced by rVvpM were examined. Experimental mouse models of V. vulnificus infection were used to study the role of melatonin and how it neutralizes the bacterial toxin activity related to Muc2 repression. RESULTS: Recombinant protein (r) VvpM significantly reduced the level of Muc2 in HT29-MTX cells. The repression of Muc2 induced by rVvpM was significantly restored upon a treatment with melatonin (1 µM), which had been inhibited by the knockdown of MT2 coupling with Gαq and the NADPH oxidase subunit p47 phox. Melatonin inhibited the ROS-mediated phosphorylation of PKCδ and ERK responsible for region-specific hypermethylation in the Muc2 promoter in rVvpM-treated HT29-MTX cells. In the mouse models of V. vulnificus infection, treatment with melatonin maintained the level of Muc2 expression in the intestine. In addition, the mutation of the VvpM gene from V. vulnificus exhibited an effect similar to that of melatonin. CONCLUSIONS: These results demonstrate that melatonin acting on MT2 inhibits the hypermethylation of the Muc2 promoter to restore the level of Muc2 production in intestinal epithelial cells infected with V. vulnificus.

Ethanol-activated CaMKII signaling induces neuronal apoptosis through Drp1-mediated excessive mitochondrial fission and JNK1-dependent NLRP3 inflammasome activation.

BACKGROUND: Neurodegeneration is a representative phenotype of patients with chronic alcoholism. Ethanol-induced calcium overload causes NOD-like receptor protein 3 (NLRP3) inflammasome formation and an imbalance in mitochondrial dynamics, closely associated with the pathogenesis of neurodegeneration. However, how calcium regulates this process in neuronal cells is poorly understood. Therefore, the present study investigated the detailed mechanism of calcium-regulated mitochondrial dynamics and NLRP3 inflammasome formation in neuronal cells by ethanol. METHODS: In this study, we used the SK-N-MC human neuroblastoma cell line. To confirm the expression level of the mRNA and protein, real time quantitative PCR and western blot were performed. Co-immunoprecipitation and Immunofluorescence staining were conducted to confirm the complex formation or interaction of the proteins. Flow cytometry was used to analyze intracellular calcium, mitochondrial dysfunction and neuronal apoptosis. RESULTS: Ethanol increased cleaved caspase-3 levels and mitochondrial reactive oxygen species (ROS) generation associated with neuronal apoptosis. In addition, ethanol increased protein kinase A (PKA) activation and cAMP-response-element-binding protein (CREB) phosphorylation, which increased N-methyl-D-aspartate receptor (NMDAR) expression. Ethanol-increased NMDAR induced intracellular calcium overload and calmodulin-dependent protein kinase II (CaMKII) activation leading to phosphorylation of dynamin-related protein 1 (Drp1) and c-Jun N-terminal protein kinase 1 (JNK1). Drp1 phosphorylation promoted Drp1 translocation to the mitochondria, resulting in excessive mitochondrial fission, mitochondrial ROS accumulation, and loss of mitochondrial membrane potential, which was recovered by Drp1 inhibitor pretreatment. Ethanol-induced JNK1 phosphorylation activated the NLRP3 inflammasome that induced caspase-1 dependent mitophagy inhibition, thereby exacerbating ROS accumulation and causing cell death. Suppressing caspase-1 induced mitophagy and reversed the ethanol-induced apoptosis in neuronal cells. CONCLUSIONS: Our results demonstrated that ethanol upregulated NMDAR-dependent CaMKII phosphorylation which is essential for Drp1-mediated excessive mitochondrial fission and the JNK1-induced NLRP3 inflammasome activation resulting in neuronal apoptosis. Video abstract.

Role of Microtubule-Associated Factors in HIF1α Nuclear Translocation.

Adaptation to hypoxia is essential for regulating the survival and functions of hypoxic cells; it is mainly mediated by the hypoxia-inducible factor 1 (HIF1). The alpha subunit of HIF1 (HIF1α) is a well-known regulatory component of HIF1, which is tightly controlled by various types of HIF1α-regulating processes. Previous research has shown that microtubule-regulated HIF1α nuclear translocation is a key factor for HIF1 activation under hypoxia. In this review, we summarize experimental reports on the role of microtubule-associated factors, such as microtubule, dynein, and dynein adaptor protein, in nuclear translocation of HIF1α. Based upon scientific evidence, we propose a bicaudal D homolog (BICD) as a novel HIF1α translocation regulating factor. A deeper understanding of the mechanism of the action of regulatory factors in controlling HIF1α nuclear translocation will provide novel insights into cell biology under hypoxia.

Membrane-Associated Effects of Glucocorticoid on BACE1 Upregulation and Aß Generation: Involvement of Lipid Raft-Mediated CREB Activation.

Glucocorticoid has been widely accepted to induce Alzheimer's disease, but the nongenomic effect of glucocorticoid on amyloid ß (Aß) generation has yet to be studied. Here, we investigated the effect of the nongenomic pathway induced by glucocorticoid on amyloid precursor protein processing enzymes as well as Aß production using male ICR mice and human neuroblastoma SK-N-MC cells. Mice groups exposed to restraint stress or intracerebroventricular injection of Aß showed impaired cognition, decreased intracellular glucocorticoid receptor (GR) level, but elevated level of membrane GR (mGR). In this respect, we identified the mGR-dependent pathway evoked by glucocorticoid using impermeable cortisol conjugated to BSA (cortisol-BSA) on SK-N-MC cells. Cortisol-BSA augmented the expression of ß-site amyloid precursor protein cleaving enzyme 1 (BACE1), the level of C-terminal fragment ß of amyloid precursor protein (C99) and Aß production, which were maintained even after blocking intracellular GR. We also found that cortisol-BSA enhanced the interaction between mGR and Gαs, which colocalized in the lipid raft. The subsequently activated CREB by cortisol-BSA bound to the CRE site of the BACE1 promoter increasing its expression, which was downregulated by inhibiting CBP. Consistently, blocking CBP attenuated cognitive impairment and Aß production induced by corticosterone treatment or intracerebroventricular injection of Aß more efficiently than inhibiting intracellular GR in mice. In conclusion, glucocorticoid couples mGR with Gαs and triggers cAMP-PKA-CREB axis dependent on the lipid raft to stimulate BACE1 upregulation and Aß generation.SIGNIFICANCE STATEMENT Patients with Alzheimer's disease (AD) have been growing sharply and stress is considered as the major environment factor of AD. Glucocorticoid is the primarily responsive factor to stress and is widely known to induce AD. However, most AD patients usually have impaired genomic pathway of glucocorticoid due to intracellular glucocorticoid receptor deficiency. In this respect, the genomic mechanism of glucocorticoid faces difficulties in explaining the consistent amyloid ß (Aß) production. Therefore, it is necessary to investigate the novel pathway of glucocorticoid on Aß generation to find a more selective therapeutic approach to AD patients. In this study, we revealed the importance of nongenomic pathway induced by glucocorticoid where membrane glucocorticoid receptor plays an important role in Aß formation.

High Glucose-Induced Reactive Oxygen Species Stimulates Human Mesenchymal Stem Cell Migration Through Snail and EZH2-Dependent E-Cadherin Repression.

BACKGROUND/AIMS: Glucose plays an important role in stem cell fate determination and behaviors. However, it is still not known how glucose contributes to the precise molecular mechanisms responsible for stem cell migration. Thus, we investigate the effect of glucose on the regulation of the human umbilical cord blood-derived mesenchymal stem cell (hUCB-MSC) migration, and analyze the mechanism accompanied by this effect. METHODS: Western blot analysis, wound healing migration assays, immunoprecipitation, and chromatin immunoprecipitation assay were performed to investigate the effect of high glucose on hUCB-MSC migration. Additionally, hUCB-MSC transplantation was performed in the mouse excisional wound splinting model. RESULTS: High concentration glucose (25 mM) elicits hUCB-MSC migration compared to normal glucose and high glucose-pretreated hUCB-MSC transplantation into the wound sites in mice also accelerates skin wound repair. We therefore elucidated the detailed mechanisms how high glucose induces hUCB-MSC migration. We showed that high glucose regulates E-cadherin repression through increased Snail and EZH2 expressions. And, we found high glucose-induced reactive oxygen species (ROS) promotes two signaling; JNK which regulates γ-secretase leading to the cleavage of Notch proteins and PI3K/Akt signaling which enhances GSK-3ß phosphorylation. High glucose-mediated JNK/Notch pathway regulates the expression of EZH2, and PI3K/Akt/GSK-3ß pathway stimulates Snail stabilization, respectively. High glucose enhances the formation of EZH2/Snail/HDAC1 complex in the nucleus, which in turn causes E-cadherin repression. CONCLUSION: This study reveals that high glucose-induced ROS stimulates the migration of hUCB-MSC through E-cadherin repression via Snail and EZH2 signaling pathways.

Ethanol-induced PGE2 up-regulates Aß production through PKA/CREB signaling pathway.

Ethanol abuse aggravates dementia-associated cognitive defects through the progression of Alzheimer's disease (AD) pathophysiology. Beta-site APP-cleaving enzyme 1 (BACE1) has been considered as a key regulator of AD pathogenesis by controlling amyloid beta peptide (Aß) accumulation. In addition, previous studies reported that endoplasmic reticulum (ER) stress and neuroinflammation have been proposed in ethanol-induced neurodegeneration. Thus, we investigated the role of ER stress and PGE2, a neuroinflammation mediator, in the ethanol-stimulated BACE1 expression and Aß production. Using the human-derived neuroblastoma cell line SK-N-MC, the results show that ethanol up-regulated BACE1 expression in a dose-dependent manner. Ethanol stimulated reactive oxygen species (ROS) production, which induced CHOP expression and eIF2α phosphorylation. PBA (an ER stress inhibitor) attenuated the ethanol-increased cyclooxygenase-2 (COX-2) expression and PGE2 production. By using salubrinal (an eIF2α dephosphorylation inhibitor) or EIF2A siRNA, we found that eIF2α phosphorylation mediated the ethanol-induced COX-2 expression. In addition, COX-2-induced BACE1 up-regulation was abolished by NS-398 (a selective COX-2 inhibitor). And, PF-04418948 (an EP-2 receptor inhibitor) pretreatment reduced ethanol-induced PKA activation and CREB phosphorylation as well as ethanol-stimulated Aß production. Furthermore, 14-22 amide (a PKA inhibitor) pretreatment or CREB1 siRNA transfection suppressed the ethanol-induced BACE1 expression. In conclusion, ethanol-induced eIF2α phosphorylation stimulates COX-2 expression and PGE2 production which induces the BACE1 expression and Aß production via EP-2 receptor-dependent PKA/CREB pathway.

The Significance of the Prognostic Nutritional Index for All Stages of Pancreatic Cancer.

Nutritional status affects the prognosis of various tumors. The prognostic nutritional index (PNI) is the known predictor of postoperative outcome in resectable pancreatic cancer patients. This study aimed to validate the prognostic value of PNI in all stages of pancreatic cancer. We retrospectively reviewed 499 patients with pancreatic cancer who were diagnosed at Severance Hospital between January 2006 and December 2011. The PNI value was calculated as 10 × serum albumin (g/dL) + 0.005 × total lymphocyte count (/mm3) at initial diagnosis. The median patient age was 62 yr, and 289 were men. The study group comprised resectable disease (n = 121), locally advanced disease (n = 118), and metastatic disease (n = 260). Univariate and multivariate analysis revealed that PNI ≤ 49.5 at initial diagnosis, together with performance status, platelet count, and clinical stage, was significantly associated with overall survival (hazard ratio, 1.562; all P < 0.05). Patients with PNI ≤ 49.5 (n = 208) had shorter median overall survival compared to patients with high PNI (9.8 vs. 14.2 mo; log rank, P < 0.001). In clinical stage subgroup analysis, initial PNI ≤49.5 independently predicted shorter overall survival, especially in resectable and metastatic disease (P = 0.041, P = 0.002, respectively).

Enhancement of high glucose-induced PINK1 expression by melatonin stimulates neuronal cell survival: Involvement of MT2 /Akt/NF-κB pathway.

Hyperglycemia is a representative hallmark and risk factor for diabetes mellitus (DM) and is closely linked to DM-associated neuronal cell death. Previous investigators reported on a genome-wide association study and showed relationships between DM and melatonin receptor (MT), highlighting the role of MT signaling by assessing melatonin in DM. However, the role of MT signaling in DM pathogenesis is unclear. Therefore, we investigated the role of mitophagy regulators in high glucose-induced neuronal cell death and the effect of melatonin against high glucose-induced mitophagy regulators in neuronal cells. In our results, high glucose significantly increased PTEN-induced putative kinase 1 (PINK1) and LC-3B expressions; as well it decreased cytochrome c oxidase subunit 4 expression and Mitotracker™ fluorescence intensity. Silencing of PINK1 induced mitochondrial reactive oxygen species (ROS) accumulation and mitochondrial membrane potential impairment, increased expressions of cleaved caspases, and increased the number of annexin V-positive cells. In addition, high glucose-stimulated melatonin receptor 1B (MTNR1B) mRNA and PINK1 expressions were reversed by ROS scavenger N-acetyl cysteine pretreatment. Upregulation of PINK1 expression in neuronal cells is suppressed by pretreatment with MT2 receptor-specific inhibitor 4-P-PDOT. We further showed melatonin stimulated Akt phosphorylation, which was followed by nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) phosphorylation and nuclear translocation. Silencing of PINK1 expression abolished melatonin-regulated mitochondrial ROS production, cleaved caspase-3 and caspase-9 expressions, and the number of annexin V-positive cells. In conclusion, we have demonstrated the melatonin stimulates PINK1 expression via an MT2 /Akt/NF-κB pathway, and such stimulation is important for the prevention of neuronal cell apoptosis under high glucose conditions.

A prospective randomized study for efficacy of an uncovered double bare metal stent compared to a single bare metal stent in malignant biliary obstruction.

BACKGROUND AND AIMS: A biliary self-expandable metal stent (SEMS) is commonly used to relieve malignant biliary obstruction. The aim of this study was to compare the efficacy of a conventional uncovered SEMS with that of a newly developed uncovered double bare metal stent in reducing the risk of stent occlusion caused by tumor ingrowth. PATIENTS AND METHODS: We performed a prospective, open-labeled, randomized trial in 71 patients at Severance Hospital, Yonsei University College of Medicine from June 2013 to June 2014. Patients with inoperable malignant biliary obstruction were included and randomized to receive an uncovered single bare metal stent (SBSs; S&G Biotech Inc.), an uncovered single bare metal stent (SBSt; Taewoong Medical), or an uncovered double bare metal stent (DBS; S&G Biotech Inc.). RESULTS: The mean age was 66.6 years (range, 35-83), and 42 (59.2%) were male. The mean duration of stent patency was 212 days (±152) in the DBS group (n = 24) compared with 124 days (±98) in the SBSs group (n = 23; P = 0.022 for noninferiority) and 116 days (±79) in the SBSt group (n = 24; P = 0.010 for noninferiority). There were no differences in the incidences of early and delayed complications or migration. CONCLUSIONS: The newly developed DBS is noninferior to the conventional uncovered SEMSs on duration of stent patency and tumor ingrowth occurred less frequently in the DBS group. This might decrease the need for reintervention and offer a better quality of life. The trial is registered with Clinicaltrials.gov no: NCT01869894.

Oleic acid enhances the motility of umbilical cord blood derived mesenchymal stem cells through EphB2-dependent F-actin formation.

The role of unsaturated fatty acids (UFAs) is essential for determining stem cell functions. Eph/Ephrin interactions are important for regulation of stem cell fate and localization within their niche, which is significant for a wide range of stem cell behavior. Although oleic acid (OA) and Ephrin receptors (Ephs) have critical roles in the maintenance of stem cell functions, interrelation between Ephs and OA has not been explored. Therefore, the present study investigated the effect of OA-pretreated UCB-MSCs in skin wound-healing and underlying mechanism of Eph expression. OA promoted the motility of UCB-MSCs via EphB2 expression. OA-mediated GPR40 activation leads to Gαq-dependent PKCα phosphorylation. In addition, OA-induced phosphorylation of GSK3ß was followed by ß-catenin nuclear translocation in UCB-MSCs. Activation of ß-catenin was blocked by PKC inhibitors, and OA-induced EphB2 expression was suppressed by ß-cateninsiRNA transfection. Of those Rho-GTPases, Rac1 was activated in an EphB2-dependent manner. Accordingly, knocking down EphB2 suppressed F-actin expression. In vivo skin wound-healing assay revealed that OA-treated UCB-MSCs enhanced skin wound repair compared to UCB-MSCs pretreated with EphB2siRNA and OA. In conclusion, we showed that OA enhances UCB-MSC motility through EphB2-dependent F-actin formation involving PKCα/GSK3ß/ß-catenin and Rac1 signaling pathways.

Novel Pathway for Hypoxia-Induced Proliferation and Migration in Human Mesenchymal Stem Cells: Involvement of HIF-1α, FASN, and mTORC1.

The control of stem cells by oxygen signaling is an important way to improve various stem cell physiological functions and metabolic nutrient alteration. Lipid metabolism alteration via hypoxia is thought to be a key factor in controlling stem cell fate and function. However, the interaction between hypoxia and the metabolic and functional changes to stem cells is incompletely described. This study aimed to identify hypoxia-inducible lipid metabolic enzymes that can regulate umbilical cord blood (UCB)-derived human mesenchymal stem cell (hMSC) proliferation and migration and to demonstrate the signaling pathway that controls functional change in UCB-hMSCs. Our results indicate that hypoxia treatment stimulates UCB-hMSC proliferation, and expression of two lipogenic enzymes: fatty acid synthase (FASN) and stearoyl-CoA desaturase-1 (SCD1). FASN but not SCD1 is a key enzyme for regulation of UCB-hMSC proliferation and migration. Hypoxia-induced FASN expression was controlled by the hypoxia-inducible factor-1 alpha (HIF-1α)/SCAP/SREBP1 pathway. Mammalian target of rapamycin (mTOR) was phosphorylated by hypoxia, whereas inhibition of FASN by cerulenin suppressed hypoxia-induced mTOR phosphorylation as well as UCB-hMSC proliferation and migration. RAPTOR small interfering RNA transfection significantly inhibited hypoxia-induced proliferation and migration. Hypoxia-induced mTOR also regulated CDK2, CDK4, cyclin D1, cyclin E, and F-actin expression as well as that of c-myc, p-cofilin, profilin, and Rho GTPase. Taken together, the results suggest that mTORC1 mainly regulates UCB-hMSC proliferation and migration under hypoxia conditions via control of cell cycle and F-actin organization modulating factors. In conclusion, the HIF-1α/FASN/mTORC1 axis is a key pathway linking hypoxia-induced lipid metabolism with proliferation and migration in UCB-hMSCs. Stem Cells 2015;33:2182-2195.

Novel method for hybrid endo-laparoscopic full-thickness gastric resection using laparoscopic transgastric suture passer device.

BACKGROUND: Current surgical methods for partial gastric full-thickness resections (FTRs) are limited by long operative times and risk of gastric content spillage, especially for lesions located at the posterior wall. We propose a simplified hybrid approach to FTR with reduced risk of spillage. METHODS: Resection margins were marked by endoscopic electrocautery to simulate a gastric lesion in the upper third of the posterior wall in eight pigs. A custom-made laparoscopic "suture passer" was made of a sharpened bendable dissecting forceps. Full-thickness sutures were alternatively passed from the serosa side with the suture passer through the gastric wall and grabbed endoluminally using an endoscopic grasper and vice versa. These transgastric sutures formed either a purse string (PS; n = 4) or a continuous horizontal mattress (HM; n = 4). Sutures were then fastened from the laparoscopic side, resulting in external outpouching of the lesion. The pouch was transected using 45-mm linear staplers. Operative time, resection margins, and number of staplers were evaluated. RESULTS: The combined approach allowed one to precisely place the sutures around the pseudo lesions, despite the inflated stomach, and it included all target markings. PS and HM methods were similar regarding time for transgastric suture (780 s ± 219.1 s vs. 765 s ± 179.2 s, p = .885), resection margins (1.3 ± 1.0 cm vs. 0.8 ± 0.6 cm, p = .248), and number of staplers (3.8 ± 1.0 vs 3.3 ± 0.5, p = .405). Stapling time (600 s ± 189.7 s vs. 330 s ± 24.5 s, p = .028) was significantly shorter in the HM technique. CONCLUSION: FTR with laparo-endoscopic transgastric suture application was feasible in the animal model. This technique allows one to achieve accurate resection margins with minimal risk of spillage.

Clinical predictors associated with proton pump inhibitor-induced hypomagnesemia.

There is increasing evidence and case reports regarding proton pump inhibitor (PPI)-induced hypomagnesemia. Our study aimed to clarify the relationship between PPI use and serum magnesium levels and to specify high-risk patients. We retrospectively studied 112 consecutive patients aged 20 years or older who were treated with PPI for ≥30 days and whose serum magnesium levels were available for the PPI treatment period. We compared the mean level of serum magnesium of the enrolled patients with PPI treatment with matched controls. There were no significant differences between the matched PPI users (n = 105) and nonusers (n = 210) in the magnesium levels (0.85 ± 0.09 vs. 0.86 ± 0.16 mM, P = 0.297). In a subgroup analysis of a PPI user group, hypomagnesemia could be observed in 32 patients but not in 80 patients. In multivariate analyses, PPI use for >1 year, age less than 45 years, and concurrent cisplatin or carboplatin use were significantly associated with PPI-induced hypomagnesemia {P = 0.042, odds ratio [OR; 95% confidence interval (CI)]: 5.388 [1.056-27.493]; P = 0.007, OR [95% CI]: 4.710 [1.523-14.571]; P = 0.007, OR [95% CI]: 13.404 [2.066-86.952], respectively} after adjusting for confounders. This study shows that long-term PPI use is associated with hypomagnesemia in hospitalized adult patients. Therefore, serum magnesium levels should be checked before the initiation of PPI treatment and during the treatment period in patients, particularly those concurrently using platinum-based chemotherapy or who are expected to use PPI for long periods.

Pre-treatment neutrophil to lymphocyte ratio as a prognostic marker to predict chemotherapeutic response and survival outcomes in metastatic advanced gastric cancer.

BACKGROUND: Several studies have shown that the neutrophil to lymphocyte ratio (NLR) in peripheral blood is a prognostic factor of various cancers. However, there is limited information on the clinical and prognostic significance of NLR in patients with metastatic advanced gastric cancer (AGC). Therefore, we examined whether the NLR can be used as a prognostic marker for predicting chemotherapeutic response and survival outcomes in metastatic AGC patients who are receiving palliative chemotherapy. METHOD: A total of 268 patients diagnosed with metastatic AGC were enrolled. NLR was calculated from complete blood cell count taken before the first chemotherapy treatment. Patients were divided into two groups according to the median value of NLR: a high NLR group and a low NLR group. RESULT: The median follow-up period was 340 days (range 72-1796 days) and median NLR was 3.06 (range 0.18-18.16). The high NLR group (NLR >3.0) contained 138 patients and the low NLR group (NLR ≤3.0) contained 130 patients. Low NLR group patients had a significantly higher chemotherapeutic disease control rate (90.0 % vs. 80.4; P = 0.028), and longer progression-free survival (PFS) and overall survival (OS) than the high NLR group patients (186 vs. 146 days; P = 0.001; 414 vs. 280 days; P < 0.001, respectively). In multivariate analysis, NLR showed a significant association with PFS (HR 1.478; 95 % CI 1.154-1.892; P = 0.002) and OS (HR 1.569; 95 % CI 1.227-2.006; P < 0.001). CONCLUSION: Pretreatment NLR is a useful prognostic marker in patients with metastatic AGC who are undergoing palliative chemotherapy.

Carcinomatosis matters: clinical outcomes and prognostic factors for clinical success of stent placement in malignant gastric outlet obstruction.

BACKGROUND: Although carcinomatosis is not a contraindication to stenting in selected patients with malignant gastric outlet obstruction (GOO), associate factors for clinical success rate of self-expandable metallic stent (SEMS) placement in GOO patients with carcinomatosis have not been fully characterized. METHODS: We analyzed a total 228 patients who were scheduled for SEMS placement for malignant GOO in tertiary-care academic medical center. All patients were treated with an uncovered or covered SEMS by using the over-the-wire placement procedure. We retrospectively evaluated clinical outcomes of SEMS placement. RESULTS: Technical success was achieved in all patients. Patients were categorized into two groups according to the presence of carcinomatosis. Clinical success rates of patients without carcinomatosis group and with carcinomatosis group were 93.9 % (92 of 98) and 80.8 % (105 of 130), respectively (P = 0.004). In subgroup analysis of patients with carcinomatosis, the clinical success rate was lower in patients with ascites (64.8 %) than in those without ascites (92.1 %, P < 0.001). Multivariate logistic regression model revealed that carcinomatosis without ascites did not decrease clinical success rate compared with absence of carcinomatosis; meanwhile, carcinomatosis with ascites showed lower clinical success rates compared with absence of carcinomatosis (adjusted odds ratio 0.163, 95 % confidence interval 0.058-0.461). In addition, poor performance status [Eastern Cooperative Oncology Group (ECOG) status ≥ 3, adjusted odds ratio 0.178, 95 % confidence interval 0.078-0.409] was also an independent poor predictive factor for clinical success of SEMS placement. CONCLUSIONS: In palliation for malignant GOO, the status of carcinomatosis with ascites and poor performance status (ECOG status ≥ 3) are significant predictive factors for poor clinical success of SEMS placement.

Safety and patient satisfaction of early diet after endoscopic submucosal dissection for gastric epithelial neoplasia: a prospective, randomized study.

BACKGROUND: Endoscopic submucosal dissection (ESD) is a standard treatment for gastric neoplasia limited to the mucosa without lymph node metastasis. However, there are neither standardized guidelines nor studies on the best time to start oral intake after ESD. The aim of this study was to compare patient satisfaction, safety, length of hospital stay, and economic feasibility between an early post-ESD diet and the conventional immediate fasting protocol. METHODS: A total of 130 patients with 156 gastric epithelial neoplasias who underwent ESD by a single expert endoscopist were consecutively and prospectively enrolled. Enrolled patients were randomized to an early diet group or a control group. The early diet group started meals as a clear liquid diet on day 0, and a soft diet and general diet in sequence on day 1. The fasting group was fasted for 2 days. Patients in both groups underwent second-look endoscopy within 2 days following ESD and follow-up endoscopy after 2 months. RESULTS: In the course of the study, ten patients were excluded. The total number of patients in the early diet group and control group was 63 and 57, respectively. Mean age was 62 years (±9.4). There were no significant differences in clinicopathologic conditions or endoscopic results such as procedure time or size of lesions between the two groups. There were no significant differences in abdominal pain score, rate of post-ESD bleeding or healing rate of ESD-induced ulcer between the two groups. However, the early diet protocol led to significantly higher patient satisfaction (p = 0.001), lower hospital costs (p < 0.001), and shorter hospital stay (p < 0.001) than the conventional fasting protocol. CONCLUSIONS: An early post-ESD diet protocol provides higher patient satisfaction, is more cost effective, decreases hospital stay, and does not influence complication rates such as post-ESD bleeding, abdominal pain, or ulcer healing compared with the conventional fasting protocol.