IU Indianapolis ScholarWorks :: Browsing by Author "Kyritsi, Konstantina"

Browsing by Author "Kyritsi, Konstantina"

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Amelioration of Large Bile Duct Damage by Histamine-2 Receptor Vivo-Morpholino Treatment
(Elsevier, 2020-05) Kennedy, Lindsey; Meadows, Vik; Kyritsi, Konstantina; Pham, Linh; Kundu, Debjyoti; Kulkarni, Rewa; Cerritos, Karla; Demieville, Jennifer; Hargrove, Laura; Glaser, Shannon; Zhou, Tianhao; Jaeger, Victoria; Alpini, Gianfranco; Francis, Heather; Medicine, School of Medicine
Histamine binds to one of the four G-protein-coupled receptors expressed by large cholangiocytes and increases large cholangiocyte proliferation via histamine-2 receptor (H2HR), which is increased in patients with primary sclerosing cholangitis (PSC). Ranitidine decreases liver damage in Mdr2-/- (ATP binding cassette subfamily B member 4 null) mice. We targeted hepatic H2HR in Mdr2-/- mice using vivo-morpholino. Wild-type and Mdr2-/- mice were treated with mismatch or H2HR vivo-morpholino by tail vein injection for 1 week. Liver damage, mast cell (MC) activation, biliary H2HR, and histamine serum levels were studied. MC markers were determined by quantitative real-time PCR for chymase and c-kit. Intrahepatic biliary mass was detected by cytokeratin-19 and F4/80 to evaluate inflammation. Biliary senescence was determined by immunofluorescence and senescence-associated β-galactosidase staining. Hepatic fibrosis was evaluated by staining for desmin, Sirius Red/Fast Green, and vimentin. Immunofluorescence for transforming growth factor-β1, vascular endothelial growth factor-A/C, and cAMP/ERK expression was performed. Transforming growth factor-β1 and vascular endothelial growth factor-A secretion was measured in serum and/or cholangiocyte supernatant. Treatment with H2HR vivo-morpholino in Mdr2-/--mice decreased hepatic damage; H2HR protein expression and MC presence or activation; large intrahepatic bile duct mass, inflammation and senescence; and fibrosis, angiogenesis, and cAMP/phospho-ERK expression. Inhibition of H2HR signaling ameliorates large ductal PSC-induced damage. The H2HR axis may be targeted in treating PSC.
Biliary damage and liver fibrosis are ameliorated in a novel mouse model lacking l-histidine decarboxylase/histamine signaling
(Nature Publishing Group, 2020-02-13) Kennedy, Lindsey; Meadows, Vik; Demieville, Jennifer; Hargrove, Laura; Virani, Shohaib; Glaser, Shannon; Zhou, Tianhao; Rinehart, Evan; Jaeger, Victoria; Kyritsi, Konstantina; Pham, Linh; Alpini, Gianfranco; Francis, Heather; Medicine, School of Medicine
Primary sclerosing cholangitis (PSC) is characterized by biliary damage and fibrosis. Multidrug resistance-2 gene knockout (Mdr2−/−) mice and PSC patients have increased histamine (HA) levels (synthesized by l-histidine decarboxylase, HDC) and HA receptor (HR) expression. Cholestatic HDC−/− mice display ameliorated biliary damage and hepatic fibrosis. The current study evaluated the effects of knockout of HDC−/− in Mdr2−/− mice (DKO) on biliary damage and hepatic fibrosis. WT, Mdr2−/− mice and homozygous DKO mice were used. Selected DKO mice were treated with HA. We evaluated liver damage along with HDC expression and HA serum levels. Changes in ductular reaction were evaluated along with liver fibrosis, inflammation and bile acid signaling pathways. The expression of H1HR/PKC-α/TGF-β1 and H2HR/pERK/VEGF-C was determined. In vitro, cholangiocyte lines were treated with HA with/without H1/H2 inhibitors before measuring: H1/H2HR, TGF-β1 and VEGF-C expression. Knockout of HDC ameliorates hepatic damage, ductular reaction, fibrosis, inflammation, bile acid signaling and H1HR/PKC-α/TGF-β1 and H2HR/pERK/VEGF-C signaling. Reactivation of the HDC/HA axis increased these parameters. In vitro, stimulation with HA increased HR expression and PKC-α, TGF-β1 and VEGF-C expression, which was reduced with HR inhibitors. Our data demonstrate the key role for the HDC/HA axis in the management of PSC progression.
Downregulation of hepatic stem cell factor by Vivo-Morpholino treatment inhibits mast cell migration and decreases biliary damage/senescence and liver fibrosis in Mdr2−/− mice
(Elsevier, 2019-12-01) Meadows, Vik; Kennedy, Lindsey; Hargrove, Laura; Demieville, Jennifer; Meng, Fanyin; Virani, Shohaib; Reinhart, Evan; Kyritsi, Konstantina; Invernizzi, Pietro; Yang, Zhihong; Wu, Nan; Liangpunsakul, Suthat; Alpini, Gianfranco; Francis, Heather; Medicine, School of Medicine
Primary sclerosing cholangitis (PSC) is characterized by increased mast cell (MC) infiltration, biliary damage and hepatic fibrosis. Cholangiocytes secrete stem cell factor (SCF), which is a chemoattractant for c-kit expressed on MCs. We aimed to determine if blocking SCF inhibits MC migration, biliary damage and hepatic fibrosis. Methods: FVB/NJ and Mdr2-/- mice were treated with Mismatch or SCF Vivo-Morpholinos. We measured (i) SCF expression and secretion; (ii) hepatic damage; (iii) MC migration/activation and histamine signaling; (iv) ductular reaction and biliary senescence; and (v) hepatic fibrosis. In human PSC patients, SCF expression and secretion were measured. In vitro, cholangiocytes were evaluated for SCF expression and secretion. Biliary proliferation/senescence was measured in cholangiocytes pretreated with 0.1% BSA or the SCF inhibitor, ISK03. Cultured HSCs were stimulated with cholangiocyte supernatant and activation measured. MC migration was determined with cholangiocytes pretreated with BSA or ISK03 loaded into the bottom of Boyden chambers and MCs into top chamber. Results: Biliary SCF expression and SCF serum levels increase in human PSC. Cholangiocytes, but not hepatocytes, from SCF Mismatch Mdr2-/- mice have increased SCF expression and secretion. Inhibition of SCF in Mdr2-/- mice reduced (i) hepatic damage; (ii) MC migration; (iii) histamine and SCF serum levels; and (iv) ductular reaction/biliary senescence/hepatic fibrosis. In vitro, cholangiocytes express and secrete SCF. Blocking biliary SCF decreased MC migration, biliary proliferation/senescence, and HSC activation. Conclusion: Cholangiocytes secrete increased levels of SCF inducing MC migration, contributing to biliary damage/hepatic fibrosis. Targeting MC infiltration may be an option to ameliorate PSC progression.
Downregulation of hepatic stem cell factor by Vivo-Morpholino treatment inhibits mast cell migration and decreases biliary damage/senescence and liver fibrosis in Mdr2−/− mice
(Elsevier, 2019) Meadows, Vik; Kennedy, Lindsey; Hargrove, Laura; Demieville, Jennifer; Meng, Fanyin; Virani, Shohaib; Reinhart, Evan; Kyritsi, Konstantina; Invernizzi, Pietro; Yang, Zhihong; Wu, Nan; Liangpunsakul, Suthat; Alpini, Gianfranco; Francis, Heather; Medicine, School of Medicine
Introduction Primary sclerosing cholangitis (PSC) is characterized by increased mast cell (MC) infiltration, biliary damage and hepatic fibrosis. Cholangiocytes secrete stem cell factor (SCF), which is a chemoattractant for c-kit expressed on MCs. We aimed to determine if blocking SCF inhibits MC migration, biliary damage and hepatic fibrosis. Methods FVB/NJ and Mdr2−/− mice were treated with Mismatch or SCF Vivo-Morpholinos. We measured (i) SCF expression and secretion; (ii) hepatic damage; (iii) MC migration/activation and histamine signaling; (iv) ductular reaction and biliary senescence; and (v) hepatic fibrosis. In human PSC patients, SCF expression and secretion were measured. In vitro, cholangiocytes were evaluated for SCF expression and secretion. Biliary proliferation/senescence was measured in cholangiocytes pretreated with 0.1% BSA or the SCF inhibitor, ISK03. Cultured HSCs were stimulated with cholangiocyte supernatant and activation measured. MC migration was determined with cholangiocytes pretreated with BSA or ISK03 loaded into the bottom of Boyden chambers and MCs into top chamber. Results Biliary SCF expression and SCF serum levels increase in human PSC. Cholangiocytes, but not hepatocytes, from SCF Mismatch Mdr2−/− mice have increased SCF expression and secretion. Inhibition of SCF in Mdr2−/− mice reduced (i) hepatic damage; (ii) MC migration; (iii) histamine and SCF serum levels; and (iv) ductular reaction/biliary senescence/hepatic fibrosis. In vitro, cholangiocytes express and secrete SCF. Blocking biliary SCF decreased MC migration, biliary proliferation/senescence, and HSC activation. Conclusion Cholangiocytes secrete increased levels of SCF inducing MC migration, contributing to biliary damage/hepatic fibrosis. Targeting MC infiltration may be an option to ameliorate PSC progression.
Downregulation of p16 Decreases Biliary Damage and Liver Fibrosis in the Mdr2 / Mouse Model of Primary Sclerosing Cholangitis
(Cognizant Communication Corporation, 2020-11) Kyritsi, Konstantina; Francis, Heather; Zhou, Tianhao; Ceci, Ludovica; Wu, Nan; Yang, Zhihong; Meng, Fanyin; Chen, Lixian; Baiocchi, Leonardo; Kundu, Debjyoti; Kennedy, Lindsey; Liangpunsakul, Suthat; Wu, Chaodong; Glaser, Shannon; Alpini, Gianfranco; Medicine, School of Medicine
Biliary senescence and hepatic fibrosis are hallmarks of cholangiopathies including primary sclerosing cholangitis (PSC). Senescent cholangiocytes display senescence-associated secretory phenotypes [SASPs, e.g., transforming growth factor-1 (TGF-1)] that further increase biliary senescence (by an autocrine loop) and trigger liver fibrosis by paracrine mechanisms. The aim of this study was to determine the effect of p16 inhibition and role of the TGF-1/microRNA (miR)-34a/sirtuin 1 (SIRT1) axis in biliary damage and liver fibrosis in the Mdr2/ mouse model of PSC. We treated (i) in vivo male wild-type (WT) and Mdr2/ mice with p16 Vivo-Morpholino or controls before measuring biliary mass [intrahepatic bile duct mass (IBDM)] and senescence, biliary SASP levels, and liver fibrosis, and (ii) in vitro intrahepatic murine cholangiocyte lines (IMCLs) with small interfering RNA against p16 before measuring the mRNA expression of proliferation, senescence, and fibrosis markers. p16 and miR-34a increased but SIRT1 decreased in Mdr2/ mice and PSC human liver samples compared to controls. p16 immunoreactivity and biliary senescence and SASP levels increased in Mdr2/ mice but decreased in Mdr2/ mice treated with p16 Vivo-Morpholino. The increase in IBDM and hepatic fibrosis (observed in Mdr2/ mice) returned to normal values in Mdr2/ mice treated with p16 Vivo-Morpholino. TGF-1 immunoreactivity and biliary SASPs levels were higher in Mdr2/ compared to those of WT mice but returned to normal values in Mdr2/ mice treated with p16 Vivo-Morpholino. The expression of fibrosis/senescence markers decreased in cholangiocytes from Mdr2/ mice treated with p16 Vivo-Morpholino (compared to Mdr2/ mice) and in IMCLs (after p16 silencing) compared to controls. Modulation of the TGF-1/miR-34a/SIRT1 axis may be important in the management of PSC phenotypes.
Functional Role of the Secretin/Secretin Receptor Signaling During Cholestatic Liver Injury
(AASLD, 2020-12) Wu, Nan; Baiocchi, Leonardo; Zhou, Tianhao; Kennedy, Lindsey; Ceci, Ludovica; Meng, Fanyin; Sato, Keisaku; Wu, Chaodong; Ekser, Burcin; Kyritsi, Konstantina; Kundu, Debjyoti; Chen, Lixian; Meadows, Vik; Franchitto, Antonio; Alvaro, Domenico; Onori, Paolo; Gaudio, Eugenio; Lenci, Ilaria; Francis, Heather; Glaser, Shannon; Alpini, Gianfranco; Medicine, School of Medicine
Liver diseases are a major health concern and affect a large proportion of people worldwide. There are over 100 types of liver disorders, including cirrhosis, cholangiocarcinoma (CCA), hepatocellular carcinoma, and hepatitis. Despite the relevant number of people who are affected by liver diseases, and the increased awareness with regard to these disorders, the number of deaths corresponding to liver injury is expected to increase in the foreseeable future. One of the possible reasons for this is that a complete comprehension of the mechanisms of hepatic damage involving specific liver anatomical districts is lacking, and, as a consequence, current treatments available are suboptimal. A major burden in the clinical setting are chronic cholestatic liver diseases (e.g., primary biliary cholangitis [PBC], primary sclerosing cholangitis [PSC], biliary atresia), which target the biliary epithelium and are characterized by cholestasis.(1, 2) Because the secretin (Sct)/secretin receptor (SR) axis (expressed only by cholangiocytes in the liver)(3, 4) is the major regulator of ductal bile secretion,(5, 6) it is intuitive that this axis plays a key role in the maintenance of biliary homeostasis during the progression of cholangiopathies. For instance, PBC is characterized by reduced bicarbonate secretion, a phenomenon possibly impeding the formation of an HCO3 canalicular film (“bicarbonate umbrella”) on bile ducts, which has protective properties against highly concentrated bile acids (BAs).(1, 7, 8) In this review, we examined the molecular mechanisms by which the Sct/SR axis regulates biliary function and the homeostasis of the biliary epithelium in normal and pathophysiological conditions.
Inhibition of Secretin/Secretin Receptor Axis Ameliorates NAFLD Phenotypes
(Wiley, 2021-10) Chen, Lixian; Wu, Nan; Kennedy, Lindsey; Francis, Heather; Ceci, Ludovica; Zhou, Tianhao; Samala, Niharika; Kyritsi, Konstantina; Wu, Chaodong; Sybenga, Amelia; Ekser, Burcin; Dar, Wasim; Atkins, Constance; Meadows, Vik; Glaser, Shannon; Alpini, Gianfranco; Surgery, School of Medicine
Background & Aims Human non-alcoholic fatty liver disease (NAFLD) is characterized at early stages by hepatic steatosis, which may progress to nonalcoholic steatohepatitis (NASH) when the liver displays microvesicular steatosis, lobular inflammation, and pericellular fibrosis. The secretin (SCT)/secretin receptor (SCTR) axis promotes biliary senescence and liver fibrosis in cholestatic models through downregulation of miR-125b signaling. We aim to evaluate the effect of disrupting biliary SCT/SCTR/miR-125b signaling on hepatic steatosis, biliary senescence and liver fibrosis in NAFLD/NASH. Approach & Results In vivo, 4 wk male WT, Sct-/- and Sctr-/- mice were fed a control diet (CD) or high-fat diet (HFD) for 16 wks. The expression of SCT/SCTR/miR-125b axis was measured in human NAFLD/NASH liver samples and HFD mouse livers by immunohistochemistry (IHC) and qPCR. Biliary/hepatocyte senescence, ductular reaction and liver angiogenesis were evaluated in mouse liver and human NAFLD/NASH liver samples. miR-125b target lipogenesis genes in hepatocytes were screened and validated by custom RT2 Profiler PCR array and luciferase assay. Biliary SCT/SCTR expression was increased in human NAFLD/NASH samples and in livers of HFD mice, whereas the expression of miR-125b was decreased. Biliary/hepatocyte senescence, ductular reaction, and liver angiogenesis were observed in human NAFLD/NASH samples as well as HFD mice, which were decreased in Sct-/- and Sctr-/- HFD mice. Elovl1 is a lipogenesis gene targeted by miR-125b, and its expression was also decreased in HFD mouse hepatocytes following Sct or Sctr knockout. Bile acid profile in fecal samples have the greatest changes between WT mice and Sct-/-/Sctr-/- mice. Conclusion The biliary SCT/SCTR/miR-125b axis promotes liver steatosis by upregulating lipid biosynthesis gene Elovl1. Targeting the biliary SCT/SCTR/miR-125b axis may be key for ameliorating phenotypes of human NAFLD/NASH.
Knockdown of vimentin reduces mesenchymal phenotype of cholangiocytes in the Mdr2-/- mouse model of primary sclerosing cholangitis (PSC)
(Elsevier, 2019-10) Zhou, Tianhao; Kyritsi, Konstantina; Wu, Nan; Francis, Heather; Yang, Zhihong; Chen, Lixian; O'Brien, April; Kennedy, Lindsey; Ceci, Ludovica; Meadows, Vik; Kusumanchi, Praveen; Wu, Chaodong; Baiocchi, Leonardo; Skill, Nicholas J.; Saxena, Romil; Sybenga, Amelia; Xie, Linglin; Liangpunsakul, Suthat; Meng, Fanyin; Alpini, Gianfranco; Glaser, Shannon; Medicine, School of Medicine
BACKGROUND: Cholangiocytes are the target cells of cholangiopathies including primary sclerosing cholangitis (PSC). Vimentin is an intermediate filament protein that has been found in various types of mesenchymal cells. The aim of this study is to evaluate the role of vimentin in the progression of biliary damage/liver fibrosis and whether there is a mesenchymal phenotype of cholangiocytes in the Mdr2-/- model of PSC. METHODS: In vivo studies were performed in 12 wk. Mdr2-/- male mice with or without vimentin Vivo-Morpholino treatment and their corresponding control groups. Liver specimens from human PSC patients, human intrahepatic biliary epithelial cells (HIBEpiC) and human hepatic stellate cell lines (HHSteCs) were used to measure changes in epithelial-to-mesenchymal transition (EMT). FINDINGS: There was increased mesenchymal phenotype of cholangiocytes in Mdr2-/- mice, which was reduced by treatment of vimentin Vivo-Morpholino. Concomitant with reduced vimentin expression, there was decreased liver damage, ductular reaction, biliary senescence, liver fibrosis and TGF-β1 secretion in Mdr2-/- mice treated with vimentin Vivo-Morpholino. Human PSC patients and derived cell lines had increased expression of vimentin and other mesenchymal markers compared to healthy controls and HIBEpiC, respectively. In vitro silencing of vimentin in HIBEpiC suppressed TGF-β1-induced EMT and fibrotic reaction. HHSteCs had decreased fibrotic reaction and increased cellular senescence after stimulation with cholangiocyte supernatant with reduced vimentin levels. INTERPRETATION: Our study demonstrated that knockdown of vimentin reduces mesenchymal phenotype of cholangiocytes, which leads to decreased biliary senescence and liver fibrosis. Inhibition of vimentin may be a key therapeutic target in the treatment of cholangiopathies including PSC. FUND: National Institutes of Health (NIH) awards, VA Merit awards.
Mast Cells Regulate Ductular Reaction and Intestinal Inflammation in Cholestasis Through Farnesoid X Receptor Signaling
(Wiley, 2021-11) Meadows, Vik; Kennedy, Lindsey; Ekser, Burcin; Kyritsi, Konstantina; Kundu, Debjyoti; Zhou, Tianhao; Chen, Lixian; Pham, Linh; Wu, Nan; Demieville, Jennifer; Hargrove, Laura; Glaser, Shannon; Alpini, Gianfranco; Francis, Heather; Medicine, School of Medicine
Background & Aim Cholestasis is characterized by increased total bile acid (TBA) levels, which are regulated by farnesoid X receptor (FXR)/fibroblast growth factor 15 (FGF15). Primary sclerosing cholangitis (PSC) patients typically present with inflammatory bowel disease (IBD). Mast cells (MCs) (i) express FXR and (ii) infiltrate the liver during cholestasis promoting liver fibrosis. In bile duct ligated (BDL) MC-deficient mice (KitW-sh), ductular reaction (DR) and liver fibrosis decrease compared to BDL WT; and MC injection exacerbates liver damage in normal mice. Approach & Results In this study, we demonstrated that MC-FXR regulates biliary FXR/FGF15, DR, hepatic fibrosis and alters intestinal FXR/FGF15. We found increased MC number and biliary FXR expression in patients with liver injury compared to control. Histamine and FGF19 serum levels and small heterodimer partner expression increase in PSC and PSC-IBD patients compared to healthy controls. MC injection increased liver damage, DR, inflammation, biliary senescence/senescence associated secretory phenotype (SASP), fibrosis and histamine in KitW-sh mice. Inhibition of MC-FXR prior to injection reduced these parameters. BDL and KitW-sh mice injected with MCs displayed increased TBA content, biliary FXR/FGF15 and intestinal inflammation, which decreased in BDL KitW-sh and KitW-sh mice injected with MC-FXR. MCs increased ileal FXR/FGF15 expression in KitW-sh mice that was reduced following FXR inhibition. BDL and Mdr2-/- mice, models of PSC, displayed increased intestinal MC infiltration and FXR/FGF15 expression. These were reduced following MC stabilization with cromolyn sodium in Mdr2-/- mice. In vitro, MC-FXR inhibition decreased biliary proliferation/SASP/FGF and hepatic stellate cell activation. Conclusion Our studies demonstrate the novel findings that MC-FXR plays a key role in liver damage and DR, including TBA regulation through alteration of intestinal and biliary FXR/FGF15 signaling.
Melatonin receptor 1A, but not 1B, knockout decreases biliary damage and liver fibrosis during cholestatic liver injury
(Wiley, 2021) Wu, Nan; Carpino, Guido; Ceci, Ludovica; Baiocchi, Leonardo; Francis, Heather; Kennedy, Lindsey; Zhou, Tianhao; Chen, Lixian; Sato, Keisaku; Kyritsi, Konstantina; Meadows, Vik; Ekser, Burcin; Franchitto, Antonio; Mancinelli, Romina; Onori, Onori; Gaudio, Eugenio; Glaser, Shannon; Alpini, Gianfranco; Medicine, School of Medicine
Background and Aims Melatonin reduces biliary damage and liver fibrosis in cholestatic models by interaction with melatonin receptors 1A (MT1) and 1B (MT2). MT1 and MT2 can form heterodimers and homodimers, but MT1 and MT2 can heterodimerize with the orphan receptor G protein–coupled receptor 50 (GPR50). MT1/GPR50 dimerization blocks melatonin binding, but MT2/GPR50 dimerization does not affect melatonin binding. GPR50 can dimerize with TGFβ receptor type I (TGFβRI) to activate this receptor. We aimed to determine the differential roles of MT1 and MT2 during cholestasis. Approach and Results Wild-type (WT), MT1 knockout (KO), MT2KO, and MT1/MT2 double KO (DKO) mice underwent sham or bile duct ligation (BDL); these mice were also treated with melatonin. BDL WT and multidrug resistance 2 KO (Mdr2−/−) mice received mismatch, MT1, or MT2 Vivo-Morpholino. Biliary expression of MT1 and GPR50 increases in cholestatic rodents and human primary sclerosing cholangitis (PSC) samples. Loss of MT1 in BDL and Mdr2−/− mice ameliorated biliary and liver damage, whereas these parameters were enhanced following loss of MT2 and in DKO mice. Interestingly, melatonin treatment alleviated BDL-induced biliary and liver injury in BDL WT and BDL MT2KO mice but not in BDL MT1KO or BDL DKO mice, demonstrating melatonin’s interaction with MT1. Loss of MT2 or DKO mice exhibited enhanced GPR50/TGFβR1 signaling, which was reduced by loss of MT1. Conclusions Melatonin ameliorates liver phenotypes through MT1, whereas down-regulation of MT2 promotes liver damage through GPR50/TGFβR1 activation. Blocking GPR50/TGFβR1 binding through modulation of melatonin signaling may be a therapeutic approach for PSC.

Browsing by Author "Kyritsi, Konstantina"

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