Citations

Abstract We have previously shown that a frequent loss of endosulfatase HSulf-1 in ovarian cancer enhances the heparin binding growth factor signaling which subsequently induces and potentiates tumorigenesis. Our recent data indicate that knockdown of HSulf-1 in ovarian cancer cells significantly alter levels of several metabolites which is comparable to the deranged metabolite profiles in primary ovarian tumors. Here we report that the metabolites associated with lipid metabolism are altered in such a way that the cells start manifesting a “lipogenic phenotype” characterized by an increased fatty acid synthesis, storage and mitochondrial beta oxidation. The lipogenic phenotype is associated with an accumulation of cytoplasmic lipid droplets (LDs) in a stably HSulf-1 knockdown cells [OV202 (Sh1/2) and OV2223 cells]. Presence of LDs is evidenced by BODIPY staining and transmission electron microscopy (TEM) analysis in Sh1/2 and a non-transduced control (NTC) cells. Conversely, enhanced expression of HSulf-1 in TOV21G, SKOV3, and C13 cells also resulted in reducing the number of LDs. More importantly, MEFs (mouse embryonic fibroblasts) from HSulf-1 knockout mice also showed an increased LD biogenesis compared to the wild type MEFs. Microarray analysis identified cytoplasmic phospholipase A2 (cPLA2; an enzyme involved in LD biogenesis) as one of the genes that was significantly upregulated in OV202 Sh1 and Sh2 cells. We found that Sh1 and Sh2 cells show an increased phosphorylation (ser505) of cytoplasmic phospholipase A2 (p-cPLA2) compared to NTC cells. Phosphorylation at ser505 is vital for the enzyme activity of cPLA2 and also necessary for LD biogenesis. We inhibited the LD biogenesis in Sh1 cells by a pharmacological inhibitor of cPLA2 (AACOCF3 and MAFP) as well by shRNA mediated knockdown of endogenous cPLA2. Further, we stimulated NTC and Sh1 cells with heparin binding growth factor (50ng/ml) for different time points and found an elevated phosphorylation of cPLA2 and extracellular signal related kinase (ERK), which is also inhibited by a pre-treatment of cells with cPLA2 inhibitor. Interestingly, inhibition of phospho-ERK by U0126 (MEK inhibitor) also inhibits the phosphorylation of cPLA2. These findings strongly suggest that loss of HSulf-1 promotes lipid metabolism in the ovarian cancer cells via activation of cPLA2 via an ERK dependent pathway. The present work highlights a novel role HSulf-1 in regulating lipid metabolism which could lead to a better understanding of tumorigenesis and identification of new targets for tumor therapy. Citation Format: Debarshi Roy, Xiaoping He, Ashwani Khurana, Susmita Mondal, Deok-Beom Jung, Sung Kim, Thomas Dierks, Clifford Folmes, Andre Terzic, Viji Shridhar. HSulf-1 loss in ovarian cancer cells induces lipid droplet biogenesis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2439. doi:10.1158/1538-7445.AM2014-2439

We report generation and characterization of pain‐related behavior in a minimally invasive facet joint degeneration (FJD) animal model in rats. FJD was produced by a non‐open percutaneous puncture‐induced injury on the right lumbar FJs at three consecutive levels. Pressure hyperalgesia in the lower back was assessed by measuring the vocalization response to pressure from a force transducer. After hyperalgesia was established, pathological changes in lumbar FJs and alterations of intervertebral foramen size were assessed by histological and imaging analyses. To investigate treatment options for lumber FJ osteoarthritis‐induced pain, animals with established hyperalgesia were administered with analgesic drugs, such as morphine, a selective COX‐2 inhibitor, a non‐steroidal anti‐inflammatory drug (NSAID) (ketorolac), or pregabalin. Effects were assessed by behavioral pain responses. One week after percutaneous puncture‐induced injury of the lumbar FJs, ipsilateral primary pressure hyperalgesia developed and was maintained for at least 12 weeks without foraminal stenosis. Animals showed decreased spontaneous activity, but no secondary hyperalgesia in the hind paws. Histopathological and microfocus X‐ray computed tomography analyses demonstrated that the percutaneous puncture injury resulted in osteoarthritis‐like structural changes in the FJs cartilage and subchondral bone. Pressure hyperalgesia was completely reversed by morphine. The administration of celecoxib produced moderate pain reduction with no statistical significance while the administration of ketorolac and pregabalin produced no analgesic effect on FJ osteoarthritis‐induced back pain. Our animal model of non‐open percutanous puncture‐induced injury of the lumbar FJs in rats shows similar characteristics of low back pain produced by human facet arthropathy. J. Cell. Physiol. 9999: 2837–2847, 2015. © 2015 Wiley Periodicals, Inc.

Circadian rhythm dysfunction is linked to many diseases, yet pathophysiological roles in articular cartilage homeostasis and degenerative joint disease including osteoarthritis (OA) remains to be investigated in vivo. Here, we tested whether environmental or genetic disruption of circadian homeostasis predisposes to OA‐like pathological changes. Male mice were examined for circadian locomotor activity upon changes in the light:dark (LD) cycle or genetic disruption of circadian rhythms. Wild‐type (WT) mice were maintained on a constant 12 h:12 h LD cycle (12:12 LD) or exposed to weekly 12 h phase shifts. Alternatively, male circadian mutant mice ( Clock Δ19 or Csnk1e tau mutants) were compared with age‐matched WT littermates that were maintained on a constant 12:12 LD cycle. Disruption of circadian rhythms promoted osteoarthritic changes by suppressing proteoglycan accumulation, upregulating matrix‐degrading enzymes and downregulating anabolic mediators in the mouse knee joint. Mechanistically, these effects involved activation of the PKCδ‐ERK‐RUNX2/NFκB and β‐catenin signaling pathways, stimulation of MMP‐13 and ADAMTS‐5, as well as suppression of the anabolic mediators SOX9 and TIMP‐3 in articular chondrocytes of phase‐shifted mice. Genetic disruption of circadian homeostasis does not predispose to OA‐like pathological changes in joints. Our results, for the first time, provide compelling in vivo evidence that environmental disruption of circadian rhythms is a risk factor for the development of OA‐like pathological changes in the mouse knee joint. J. Cell. Physiol. 230: 2174–2183, 2015. © 2015 Wiley Periodicals, Inc.

AbstractObjectivesDiffuse large B‐cell lymphoma (DLBCL) is an aggressive subtype of non‐Hodgkin lymphoma (NHL) and is the most common NHL subtype diagnosed worldwide. The first large‐scale genome‐wide association study (GWAS) of DLBCL with over 4000 cases conducted among individuals of European ancestry recently identified five independent SNPs that achieved genome‐wide significance, and two SNPs that showed a suggestive association with DLBCL risk.MethodsTo evaluate whether Eastern Asians and individuals of European ancestry share similar genetic risk factors for this disease, we attempted to replicate these GWAS findings in a pooled series of 1124 DLBCL cases and 3596 controls from Hong Kong, South Korea, and Thailand.ResultsThree of the five genome‐wide significant SNPs from the DLBCL GWAS were significantly associated with DLBCL in our study population, including the top finding from the GWAS, EXOC2 rs116446171, which achieved genome‐wide significance in our data (per allele OR = 2.04, 95% CI = 1.63–2.56; ptrend = 3.9 × 10−10). Additionally, we observed a significant association with PVT1 rs13255292 (per allele OR = 1.34, 95% CI = 1.19–1.52; ptrend = 2.1 × 10−6), which was the second strongest finding in the GWAS, and with HLA‐B rs2523607 (per allele OR = 3.05, 95% CI = 1.32–7.05; ptrend = 0.009).ConclusionsOur study, which provides the first evaluation in Eastern Asians of SNPs definitively associated with DLBCL risk in individuals of European ancestry, indicates that at least some of the genetic factors associated with risk of DLBCL are similar between these populations.

AbstractBackgroundHigh‐resolution manometry using the Chicago classification, which utilizes parameters including integrated relaxation pressure (IRP), distal contractile integral (DCI), and contractile front velocity (CFV), shows better diagnostic ability than previous conventional criteria. However, the current normal cut‐off values for the Chicago classification are based on individuals aged 19–48 years and do not include older people. Here, we aimed to assess the normal values for the Chicago classification in individuals aged 20–67 years and compare the parameters across age groups.MethodsFifty‐four asymptomatic healthy individuals (27 male and 27 female; age range. 20–67 years) were prospectively enrolled. To evaluate the effect of age and sex on manometric profiles, we attempted to enroll equal numbers of male and female subjects for each decade. Manometry was performed in both the supine and sitting positions.Key ResultsThe distal latency (DL) was significantly shorter with increasing age in both measurement positions. Furthermore, IRP was significantly higher with increasing age in both positions. Spearman's ranked correlation coefficient analysis indicated that DCI and IRP in both positions were positively correlated with age.Conclusions & InferencesAge affects the key parameters currently used in the Chicago classification, including IRP, DCI, and DL. Larger prospective studies with older subjects are needed to determine the age‐related normal values for the Chicago classification system.

Chronic passive hepatic congestion (congestive hepatopathy) leads to hepatic fibrosis; however, the mechanisms involved in this process are not well understood. We developed a murine experimental model of congestive hepatopathy through partial ligation of the inferior vena cava (pIVCL). C57BL/6 and transgenic mice overexpressing tissue factor pathway inhibitor (SM22α‐TFPI) were subjected to pIVCL or sham. Liver and blood samples were collected and analyzed in immunohistochemical, morphometric, real‐time polymerase chain reaction, and western blot assays. Hepatic fibrosis and portal pressure were significantly increased after pIVCL concurrent with hepatic stellate cell (HSC) activation. Liver stiffness, as assessed by magnetic resonance elastography, correlated with portal pressure and preceded fibrosis in our model. Hepatic sinusoidal thrombosis as evidenced by fibrin deposition was demonstrated both in mice after pIVCL as well as in humans with congestive hepatopathy. Warfarin treatment and TFPI overexpression both had a protective effect on fibrosis development and HSC activation after pIVCL. In vitro studies show that congestion stimulates HSC fibronectin (FN) fibril assembly through direct effects of thrombi as well as by virtue of mechanical strain. Pretreatment with either Mab13 or Cytochalasin‐D, to inhibit β‐integrin or actin polymerization, respectively, significantly reduced fibrin and stretch‐induced FN fibril assembly. Conclusion : Chronic hepatic congestion leads to sinusoidal thrombosis and strain, which in turn promote hepatic fibrosis. These studies mechanistically link congestive hepatopathy to hepatic fibrosis. (H epatology 2015;61:648‐659)

AbstractImproving the effectiveness of adipose‐tissue derived human mesenchymal stromal/stem cells (AMSCs) for skeletal therapies requires a detailed characterization of mechanisms supporting cell proliferation and multi‐potency. We investigated the molecular phenotype of AMSCs that were either actively proliferating in platelet lysate or in a basal non‐proliferative state. Flow cytometry combined with high‐throughput RNA sequencing (RNASeq) and RT‐qPCR analyses validate that AMSCs express classic mesenchymal cell surface markers (e.g., CD44, CD73/NT5E, CD90/THY1, and CD105/ENG). Expression of CD90 is selectively elevated at confluence. Self‐renewing AMSCs express a standard cell cycle program that successively mediates DNA replication, chromatin packaging, cyto‐architectural enlargement, and mitotic division. Confluent AMSCs preferentially express genes involved in extracellular matrix (ECM) formation and cellular communication. For example, cell cycle‐related biomarkers (e.g., cyclins E2 and B2, transcription factor E2F1) and histone‐related genes (e.g., H4, HINFP, NPAT) are elevated in proliferating AMSCs, while ECM genes are strongly upregulated (>10‐fold) in quiescent AMSCs. AMSCs also express pluripotency genes (e.g., POU5F1, NANOG, KLF4) and early mesenchymal markers (e.g., NES, ACTA2) consistent with their multipotent phenotype. Strikingly, AMSCs modulate expression of WNT signaling components and switch production of WNT ligands (from WNT5A/WNT5B/WNT7B to WNT2/WNT2B), while upregulating WNT‐related genes (WISP2, SFRP2, and SFRP4). Furthermore, post‐proliferative AMSCs spontaneously express fibroblastic, osteogenic, chondrogenic, and adipogenic biomarkers when maintained in confluent cultures. Our findings validate the biological properties of self‐renewing and multi‐potent AMSCs by providing high‐resolution quality control data that support their clinical versatility. J. Cell. Biochem. 115: 1816–1828, 2014. © 2014 Wiley Periodicals, Inc.

AbstractBackground & AimsSurvival of patients with hepatocellular carcinoma (HCC) is determined by hepatic function and tumour extent. Recently, a new Model to Estimate Survival in Ambulatory HCC patients (MESIAH) was proposed to predict overall survival in ambulatory HCC patients. This study aimed to evaluate the prognostic performance of the MESIAH score in an independent cohort of HCC patients.MethodsA cohort of 1969 patients newly diagnosed with HCC at the National Cancer Center, Korea between January 2004 and December 2009 was used for validation of the MESIAH score. The model's performance was assessed using C‐statistics, the likelihood ratio (LR) χ2 value and Akaike information criterion (AIC).ResultsPatients in the cohort had a median age of 56 years and 83.2% were men. Hepatitis B virus infection was present in 74.6 and 81.6% had a Child–Pugh class A. The median overall survival was 21.4 months. The MESIAH score had a higher degree of discrimination, with a C‐statistic of 0.792 [95% confidence interval (CI), 0.782–0.803], when compared with the Barcelona Clinic Liver Cancer (BCLC) staging system [0.665 (95% CI, 0.653–0.678), P < 0.001]. The LR χ2 value and the AIC of MESIAH were also better than those of BCLC, Cancer of the Liver Italian Program, Japan Integrated Scoring and Tokyo score. The observed survival in the cohort closely matched that predicted by the MESIAH score.ConclusionsThe new prognostication model MESIAH accurately estimated the overall survival of Korean HCC patients and may be useful in future research as well as individual patient care.