Browsing by Subject "KNOCKOUT MICE"

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  • Saarimaki-Vire, Jonna; Balboa, Diego; Russell, Mark A.; Saarikettu, Juha; Kinnunen, Matias; Keskitalo, Salla; Malhi, Amrinder; Valensisi, Cristina; Andrus, Colin; Eurola, Solja; Grym, Heli; Ustinov, Jarkko; Wartiovaara, Kirmo; Hawkins, R. David; Silvennoinen, Olli; Varjosalo, Markku; Morgan, Noel G.; Otonkoski, Timo (2017)
    Activating germline mutations in STAT3 were recently identified as a cause of neonatal diabetes mellitus associated with beta-cell autoimmunity. We have investigated the effect of an activating mutation, STAT3(K392R,) on pancreatic development using induced pluripotent stem cells (iPSCs) derived from a patient with neonatal diabetes and pancreatic hypoplasia. Early pancreatic endoderm differentiated similarly from STAT3(K392R) and healthy-control cells, but in later stages, NEUROG3 expressionwas upregulated prematurely in STAT3(K392R) cells together with insulin (INS) and glucagon (GCG). RNA sequencing (RNA-seq) showed robust NEUROG3 downstream targets upregulation. STAT3 mutation correction with CRISPR/Cas9 reversed completely the disease phenotype. STAT3(K392R) -activating properties were not explained fully by altered DNA-binding affinity or increased phosphorylation. Instead, reporter assays demonstrated NEUROG3 promoter activation by STAT3 in pancreatic cells. Furthermore, proteomic and immunocytochemical analyses revealed increased nuclear translocation of STAT3(K392R). Collectively, our results demonstrate that the STAT3(K392R) mutation causes premature endocrine differentiation through direct induction of NEUROG3 expression.
  • Hlushchenko, Iryna; Khanal, Pushpa; Abouelezz, Amr; Paavilainen, Ville O.; Hotulainen, Pirta (2018)
    Many actin cytoskeleton-regulating proteins control dendritic spine morphology and density, which are cellular features often altered in autism spectrum disorder (ASD). Recent studies using animal models show that autism-related behavior can be rescued by either manipulating actin regulators or by reversing dendritic spine density or morphology. Based on these studies, the actin cytoskeleton is a potential target pathway for developing new ASD treatments. Thus, it is important to understand how different ASD-associated actin regulators contribute to the regulation of dendritic spines and how ASD-associated mutations modulate this regulation. For this study, we selected five genes encoding different actin-regulating proteins and induced ASD-associated de novo missense mutations in these proteins. We assessed the functionality of the wild-type and mutated proteins by analyzing their subcellular localization, and by analyzing the dendritic spine phenotypes induced by the expression of these proteins. As the imbalance between excitation and inhibition has been suggested to have a central role in ASD, we additionally evaluated the density, size and subcellular localization of inhibitory synapses. Common for all the proteins studied was the enrichment in dendritic spines. ASD-associated mutations induced changes in the localization of alpha-actinin-4, which localized less to dendritic spines, and for SWAP-70 and SrGAP3, which localized more to dendritic spines. Among the wild-type proteins studied, only alpha-actinin-4 expression caused a significant change in dendritic spine morphology by increasing the mushroom spine density and decreasing thin spine density. We hypothesized that mutations associated with ASD shift dendritic spine morphology from mushroom to thin spines. An M554V mutation in alpha-actinin-4 (ACTN4) resulted in the expected shift in dendritic spine morphology by increasing the density of thin spines. In addition, we observed a trend toward higher thin spine density withmutations inmyosin IXb and SWAP-70. Myosin IIb and myosin IXb expression increased the proportion of inhibitory synapses in spines. The expression of mutated myosin IIb (Y265C), SrGAP3 (E469K), and SWAP-70 (L544F) induced variable changes in inhibitory synapses.
  • Puskarjov, Martin; Ahmad, Faraz; Khirug, Stanislav; Sivakumaran, Sudhir; Kaila, Kai; Blaesse, Peter (2015)
  • Aitta-aho, Teemu; Möykkynen, Tommi Petteri; Panhelainen, Anne E.; Vekovischeva, Olga Yu; Bäckström, Pia; Korpi, Esa R. (2012)
  • Sidorova, Yulia A.; Perepechaeva, Maria L.; Pivovarova, Elena N.; Markel, Arkady L.; Lyakhovich, Vyacheslav V.; Grishanova, Alevtina Y. (2016)
    Oxidative reactions that are catalyzed by cytochromes P450 1A (CYP1A) lead to formation of carcinogenic derivatives of arylamines and polycyclic aromatic hydrocarbons (PAHs), such as the widespread environmental pollutant benzo(a) pyrene (BP). These compounds upregulate CYP1A at the transcriptional level via an arylhydrocarbon receptor (AhR)-dependent signaling pathway. Because of the involvement of AhR-dependent genes in chemically induced carcinogenesis, suppression of this signaling pathway could prevent tumor formation and/or progression. Here we show that menadione (a water-soluble analog of vitamin K-3) inhibits BP-induced expression and enzymatic activity of both CYP1A1 and CYP1A2 in vivo (in the rat liver) and BP-induced activity of CYP1A1 in vitro. Coadministration of BP and menadione reduced DNA-binding activity of AhR and increased DNA-binding activity of transcription factors Oct-1 and CCAAT/enhancer binding protein (C/EBP), which are known to be involved in negative regulation of AhR-dependent genes, in vivo. Expression of another factor involved in downregulation of CYP1A-pAhR repressor (AhRR)-was lower in the liver of the rats treated with BP and menadione, indicating that the inhibitory effect of menadione on CYP1A is not mediated by this protein. Furthermore, menadione was well tolerated by the animals: no signs of acute toxicity were detected by visual examination or by assessment of weight gain dynamics or liver function. Taken together, our results suggest that menadione can be used in further studies on animal models of chemically induced carcinogenesis because menadione may suppress tumor formation and possibly progression.
  • Fickert, Peter; Hirschfield, Gideon M.; Denk, Gerald; Marschall, Hanns-Ulrich; Altorjay, Istvan; Farkkila, Martti; Schramm, Christoph; Spengler, Ulrich; Chapman, Roger; Bergquist, Annika; Schrumpf, Erik; Nevens, Frederik; Trivedi, Palak; Reiter, Florian P.; Tornai, Istvan; Halilbasic, Emina; Greinwald, Roland; Pröls, Markus; Manns, Michael P.; Trauner, Michael; European PSC norUDCA Study Grp (2017)
    Background & Aim: Primary sclerosing cholangitis (PSC) represents a devastating bile duct disease, currently lacking effective medical therapy. 24-norursodeoxycholic acid (norUDCA) is a side chain-shortened C-23 homologue of UDCA and has shown potent anti-cholestatic, anti-inflammatory and anti-fibrotic properties in a preclinical PSC mouse model. A randomized controlled trial, including 38 centers from 12 European countries, evaluated the safety and efficacy of three doses of oral norUDCA (500 mg/d, 1,000 mg/d or 1,500 mg/d) compared with placebo in patients with PSC. Methods: One hundred sixty-one PSC patients without concomitant UDCA therapy and with elevated serum alkaline phosphatase (ALP) levels were randomized for a 12-week treatment followed by a 4-week follow-up. The primary efficacy endpoint was the mean relative change in ALP levels between baseline and end of treatment visit. Results: norUDCA reduced ALP levels by -12.3%, -17.3%, and -26.0% in the 500, 1,000, and 1,500 mg/d groups (p = 0.029, tively, while a +1.2% increase was observed in the placebo group. Similar dose-dependent results were found for secondary end-points, such as ALT, AST, gamma-GT, or the rate of patients achieving ALP levels <1.5 x ULN. Serious adverse events occurred in seven patients in the 500 mg/d, five patients in the 1,000 mg/d, two patients in the 1500 mg/d group, and three in the placebo group. There was no difference in reported pruritus between treatment and placebo groups. Conclusions: norUDCA significantly reduced ALP values dose-dependently in all treatment arms. The safety profile of norUDCA was excellent and comparable to placebo. Consequently, these results justify a phase III trial of norUDCA in PSC patients. Lay summary: Effective medical therapy for primary sclerosing cholangitis (PSC) is urgently needed. In this phase II clinical study in PSC patients, a side chain-shortened derivative of ursodeoxycholic acid, norursodeoxycholic acid (norUDCA), significantly reduced serum alkaline phosphatase levels in a dose-dependent manner during a 12-week treatment. Importantly, norUDCA showed a favorable safety profile, which was similar to placebo. The use of norUDCA in PSC patients is promising and will be further evaluated in a phase III clinical study. (C) 2017 European Association for the Study of the Liver. Published by Elsevier B.V.
  • Ribeiro, Deidiane Elisa; Casarotto, Plinio C.; Staquini, Laura; Pinto e Silva, Maria Augusta; Biojone, Caroline; Wegener, Gregers; Joca, Samia (2019)
    Purinergic receptors, especially P2RX, are associated to the severity of symptoms in patients suffering from depressive and bipolar disorders, and genetic deletion or pharmacological blockade of P2RX7 induces antidepressant-like effect in preclinical models. However, there is scarce evidence about the alterations in P2RX7 or P2RX4 levels and in behavioral consequences induced by previous exposure to stress, a major risk factor for depression in humans. In the present study, we evaluated the effect of imipramine (IMI) on P2RX7 and P2RX4 levels in dorsal and ventral hippocampus as well as in the frontal cortex of rats submitted to the pretest session of learned helplessness (LH) paradigm. Repeated, but not acute administration of IMI (15 mg/kg ip) reduced the levels of both P2RX7 and P2RX4 in the ventral, but not in dorsal hippocampus or frontal cortex. In addition, we tested the effect of P2RX7/ P2RX4 antagonist brilliant blue G (BBG: 25 or 50 mg/kg ip) on the LH paradigm. We observed that repeated (7 days) but not acute (1 day) treatment with BBG (50 mg) reduced the number of failures to escape the shocks in the test session, a parameter mimicked by the same regimen of IMI treatment. Taken together, our data indicates that pharmacological blockade or decrease in the expression of P2RX7 is associated to the antidepressant-like behavior observed in the LH paradigm after repeated drug administration.
  • Leppä, Elli; Linden, Anni-Maija; Vekovischeva, Olga Y.; Swinny, Jerome D.; Rantanen, Ville; Toppila, Esko; Hoeger, Harald; Sieghart, Werner; Wulff, Peer; Wisden, William; Korpi, Esa R. (2011)
  • Kainulainen, Veera; Tang, Yurui; Spillmann, Thomas; Kilpinen, Susanne; Reunanen, Justus; Saris, Per E. J.; Satokari, Reetta (2015)
  • Tachmazidou, Ioanna; Suveges, Daniel; Min, Josine L.; Ritchie, Graham R. S.; Steinberg, Julia; Walter, Klaudia; Iotchkova, Valentina; Schwartzentruber, Jeremy; Huang, Jie; Memari, Yasin; McCarthy, Shane; Crawford, Andrew A.; Bombieri, Cristina; Cocca, Massimiliano; Farmaki, Aliki-Eleni; Gaunt, Tom R.; Jousilahti, Pekka; Kooijman, Marjolein N.; Lehne, Benjamin; Malerba, Giovanni; Mannisto, Satu; Matchan, Angela; Medina-Gomez, Carolina; Metrustry, Sarah J.; Nag, Abhishek; Ntalla, Ioanna; Paternoster, Lavinia; Rayner, Nigel W.; Sala, Cinzia; Scott, William R.; Shihab, Hashem A.; Southam, Lorraine; St Pourcain, Beate; Traglia, Michela; Trajanoska, Katerina; Zaza, Gialuigi; Zhang, Weihua; Artigas, Maria S.; Bansal, Narinder; Benn, Marianne; Chen, Zhongsheng; Danecek, Petr; Lin, Wei-Yu; Locke, Adam; Luan, Jian'an; Manning, Alisa K.; Mulas, Antonella; Sidore, Carlo; Tybjaerg-Hansen, Anne; Perola, Markus; SpiroMeta Consortium; GoT2D Consortium; ArcOGEN Consortium; Understanding Soc Sci Grp; UK10KConsortium (2017)
    Deep sequence-based imputation can enhance the discovery power of genome-wide association studies by assessing previously unexplored variation across the common-and low-frequency spectra. We applied a hybrid whole-genome sequencing (WGS) and deep imputation approach to examine the broader allelic architecture of 12 anthropometric traits associated with height, body mass, and fat distribution in up to 267,616 individuals. We report 106 genome-wide significant signals that have not been previously identified, including 9 low-frequency variants pointing to functional candidates. Of the 106 signals, 6 are in genomic regions that have not been implicated with related traits before, 28 are independent signals at previously reported regions, and 72 represent previously reported signals for a different anthropometric trait. 71% of signals reside within genes and fine mapping resolves 23 signals to one or two likely causal variants. We confirm genetic overlap between human monogenic and polygenic anthropometric traits and find signal enrichment in cis expression QTLs in relevant tissues. Our results highlight the potential of WGS strategies to enhance biologically relevant discoveries across the frequency spectrum.