Browsing by Subject "INHIBITION"

Sort by: Order: Results:

Now showing items 1-20 of 143
  • Buettner, Ralf; Le Xuan Truong Nguyen; Kumar, Bijender; Morales, Corey; Liu, Chao; Chen, Lisa S.; Pemovska, Tea; Synold, Timothy W.; Palmer, Joycelynne; Thompson, Ryan; Li, Ling; Dinh Hoa Hoang; Zhang, Bin; Ghoda, Lucy; Kowolik, Claudia; Kontro, Mika; Leitch, Calum; Wennerberg, Krister; Yu, Xiaochun; Chen, Ching-Cheng; Horne, David; Gandhi, Varsha; Pullarkat, Vinod; Marcucci, Guido; Rosen, Steven T. (2019)
    Nucleoside analogs represent the backbone of several distinct chemotherapy regimens for acute myeloid leukemia (AML) and combination with tyrosine kinase inhibitors has improved survival of AML patients, including those harboring the poor-risk FLT3-ITD mutation. Although these compounds are effective in killing proliferating blasts, they lack activity against quiescent leukemia stem cells (LSCs), which contributes to initial treatment refractoriness or subsequent disease relapse. The reagent 8-chloro-adenosine (8-Cl-Ado) is a ribose-containing, RNA-directed nucleoside analog that is incorporated into newly transcribed RNA rather than in DNA, causing inhibition of RNA transcription. In this report, we demonstrate antileukemic activities of 8-Cl-Ado in vitro and in vivo and provide mechanistic insight into the mode of action of 8-Cl-Ado in AML. 8-Cl-Ado markedly induced apoptosis in LSC, with negligible effects on normal stem cells. 8-Cl-Ado was particularly effective against AML cell lines and primary AML blast cells harboring the FLT3-ITD mutation. FLT3-ITD is associated with high expression of miR-155. Furthermore, we demonstrate that 8-Cl-Ado inhibits miR-155 expression levels accompanied by induction of DNA-damage and suppression of cell proliferation, through regulation of miR-155/ErbB3 binding protein 1(Ebp1)/p53/PCNA signaling. Finally, we determined that combined treatment of NSG mice engrafted with FLT3-ITD (+) MV4-11 AML cells with 8-Cl-Ado and the FLT3 inhibitor AC220 (quizartinib) synergistically enhanced survival, compared with that of mice treated with the individual drugs, suggesting a potentially effective approach for FLT3-ITD AML patients.
  • Manoilov, Kyrylo Yu; Verkhusha, Vladislav V.; Shcherbakova, Daria M. (2021)
    Genetically encoded tools for the regulation of endogenous molecules (RNA, DNA elements and protein) are needed to study and control biological processes with minimal interference caused by protein overexpression and overactivation of signaling pathways. Here we focus on light-controlled optogenetic tools (OTs) that allow spatiotemporally precise regulation of gene expression and protein function. To control endogenous molecules, OTs combine light-sensing modules from natural photoreceptors with specific protein or nucleic acid binders. We discuss OT designs and group OTs according to the principles of their regulation. We outline characteristics of OT performance, discuss considerations for their use in vivo and review available OTs and their applications in cells and in vivo. Finally, we provide a brief outlook on the development of OTs. This Review discusses optogenetic tools for manipulating endogenous targets such as genes and signaling pathways in a physiological range.
  • Sathyan, Sabin; Tolmacheva, Aleksandra; Tugin, Sergei; Mäkelä, Jyrki P.; Shulga, Anastasia; Lioumis, Pantelis (2021)
    Paired associative stimulation (PAS) is a stimulation technique combining transcranial magnetic stimulation (TMS) and peripheral nerve stimulation (PNS) that can induce plastic changes in the human motor system. A PAS protocol consisting of a high-intensity single TMS pulse given at 100% of stimulator output (SO) and high-frequency 100-Hz PNS train, or "the high-PAS " was designed to promote corticomotoneuronal synapses. Such PAS, applied as a long-term intervention, has demonstrated therapeutic efficacy in spinal cord injury (SCI) patients. Adding a second TMS pulse, however, rendered this protocol inhibitory. The current study sought for more effective PAS parameters. Here, we added a third TMS pulse, i.e., a 20-Hz rTMS (three pulses at 96% SO) combined with high-frequency PNS (six pulses at 100 Hz). We examined the ability of the proposed stimulation paradigm to induce the potentiation of motor-evoked potentials (MEPs) in five human subjects and described the safety and tolerability of the new protocol in these subjects. In this study, rTMS alone was used as a control. In addition, we compared the efficacy of the new protocol in five subjects with two PAS protocols consisting of PNS trains of six pulses at 100 Hz combined with (a) single 100% SO TMS pulses (high-PAS) and (b) a 20-Hz rTMS at a lower intensity (three pulses at 120% RMT). The MEPs were measured immediately after, and 30 and 60 min after the stimulation. Although at 0 and 30 min there was no significant difference in the induced MEP potentiation between the new PAS protocol and the rTMS control, the MEP potentiation remained significantly higher at 60 min after the new PAS than after rTMS alone. At 60 min, the new protocol was also more effective than the two other PAS protocols. The new protocol caused strong involuntary twitches in three subjects and, therefore, its further characterization is needed before introducing it for clinical research. Additionally, its mechanism plausibly differs from PAS with high-frequency PNS that has been used in SCI patients.
  • Baggio, Francesca; Hetzel, Udo; Nufer, Lisbeth; Kipar, Anja; Hepojoki, Jussi (2021)
    Viruses need cells for their replication and, therefore, ways to hijack cellular functions. Mitochondria play fundamental roles within the cell in metabolism, immunity and regulation of homeostasis due to which some viruses aim to alter mitochondrial functions. Herein we show that the nucleoprotein (NP) of arenaviruses enters the mitochondria of infected cells, affecting the mitochondrial morphology. Reptarenaviruses cause boid inclusion body disease (BIBD) that is characterized, especially in boas, by the formation of cytoplasmic inclusion bodies (IBs) comprising reptarenavirus NP within the infected cells. We initiated this study after observing electron-dense material reminiscent of IBs within the mitochondria of reptarenavirus infected boid cell cultures in an ultrastructural study. We employed immuno-electron microscopy to confirm that the mitochondrial inclusions indeed contain reptarenavirus NP. Mutations to a putative N-terminal mitochondrial targeting signal (MTS), identified via software predictions in both mamm- and reptarenavirus NPs, did not affect the mitochondrial localization of NP, suggesting that it occurs independently of MTS. In support of MTS-independent translocation, we did not detect cleavage of the putative MTSs of arenavirus NPs in reptilian or mammalian cells. Furthermore, in vitro translated NPs could not enter isolated mitochondria, suggesting that the translocation requires cellular factors or conditions. Our findings suggest that MTS-independent mitochondrial translocation of NP is a shared feature among arenaviruses. We speculate that by targeting the mitochondria arenaviruses aim to alter mitochondrial metabolism and homeostasis or affect the cellular defense.
  • Beck, Janina; Fuhr, Olaf; Nieger, Martin; Bräse, Stefan (2020)
    The synthesis of highly substituted hydroanthraquinone derivatives with up to three stereogenic centres via a Diels-Alder reaction, starting from easily accessible 2-substituted naphthoquinones, is described. The [4+2]-cycloaddition is applicable for a broad range of substrates, runs under mild conditions and results in high yields. The highly regioselective outcome of the reactions is enabled by a benzoyl substituent at C2 of the dienophiles. The obtained hydroanthraquinones can be further modified and represent ideal substrates for follow-up intramolecular coupling reactions to create unique bicyclo[3.3.1] or -[3.2.2]nonane ring systems which are important natural product skeletons.
  • Abdurakhmanova, Shamsiiat; Semenova, Svetlana; Piepponen, T. Petteri; Panula, Pertti (2019)
    Hypothalamic histaminergic neurons regulate a variety of homeostatic, metabolic and cognitive functions. Recent data have suggested a modulatory role of histamine and histamine receptors in shaping striatal activity and connected the histaminergic system to neuropsychiatric disorders. We characterized exploratory behavior and striatal neurotransmission in mice lacking the histamine producing enzyme histidine decarboxylase (Hdc). The mutant mice showed a distinct behavioral pattern during exploration of novel environment, specifically, increased frequency of rearing seated against the wall, jumping and head/body shakes. This behavioral phenotype was associated with decreased levels of striatal dopamine and serotonin and increased level of dopamine metabolite DOPAC. Gene expression levels of dynorphin and enkephalin, opioids released by medium spiny neurons of striatal direct and indirect pathways respectively, were lower in Hdc mutant mice than in control animals. A low dose of amphetamine led to similar behavioral and biochemical outcomes in both genotypes. Increased striatal dopamine turnover was observed in Hdc KO mice after treatment with dopamine precursor l-Dopa. Overall, our study suggests a role for striatal dopamine and opioid peptides in formation of distinct behavioral phenotype of Hdc KO mice.
  • Laursen, Jens Christian; Sondergaard-Heinrich, Niels; de Melo, Joana Mendes Lopes; Haddock, Bryan; Rasmussen, Ida Kirstine Bull; Safavimanesh, Farzaneh; Hansen, Christian Stevns; Storling, Joachim; Larsson, Henrik Bo Wiberg; Groop, Per-Henrik; Frimodt-Moller, Marie; Andersen, Ulrik Bjorn; Rossing, Peter (2021)
    Background: Inhibitors of the sodium-glucose cotransporter 2 (SGLT2) slow the progression of diabetic kidney disease, possibly by reducing the proximal tubule transport workload with subsequent improvement of renal oxygenation. We aimed to test this hypothesis in individuals with type 1 diabetes and albuminuria. Methods: A randomised, double-blind, placebo-controlled, crossover trial with a single 50 mg dose of the SGLT2 inhibitor dapagliflozin and placebo in random order, separated by a two-week washout period. Magnetic resonance imaging (MRI) was used to assess renal R-2* (a low value corresponds to a high tissue oxygenation), renal perfusion (arterial spin labelling) and renal artery flow (phase contrast imaging) at baseline, three- and six hours from tablet ingestion. Exploratory outcomes, including baroreflex sensitivity, peripheral blood oxygen saturation, peripheral blood mononuclear cell mitochondrial oxygen consumption rate, and biomarkers of inflammation were evaluated at baseline and 12 h from medication. The study is registered in the EU Clinical Trials Register (EudraCT 2019-004,557-92), on ClinicalTrials.gov (NCT04193566), and is completed. Findings: Between February 3, 2020 and October 23, 2020, 31 individuals were screened, and 19 eligible individuals were randomised. Three dropped out before receiving any of the interventions and one dropped out after receiving only placebo. We included 15 individuals (33% female) in the per-protocol analysis with a mean age of 58 (SD 14) years, median urinary albumin creatinine ratio of 46 [IQR 21-58] mg/g and an eGFR of 73 (32) ml/min/1.73m(2). The mean changes in renal cortical R-2* from baseline to six hours were for dapagliflozin -1.1 (SD 0.7) s(-1) and for placebo +1.3 (0.7) s(-1), resulting in a difference between interventions of -2.3 s(-1) [95% CI -4.0 to -0.6]; p = 0.012. No between-intervention differences were found in any other MRI outcomes, physiological parameters or exploratory outcomes. There were no adverse events. Interpretation: A single dose of 50 mg dapagliflozin acutely improved renal cortical R-2* without changing renal perfusion or blood flow. This suggests improved renal cortical oxygenation due to a reduced tubular transport workload in the proximal tubules. Such improved oxygenation may in part explain the long-term beneficial renal effects seen with SGLT2 inhibitors, but it remains to be determined whether the observed effects can be achieved with lower doses, with chronic treatment and if they occur in type 2 diabetes as well. (C) 2021 The Author(s). Published by Elsevier Ltd.
  • Osipova, Olga; Sharoyko, Vladimir; Zashikhina, Natalia; Zakharova, Natalya; Tennikova, Tatiana; Urtti, Arto; Korzhikova-Vlakh, Evgenia (2020)
    Polyethyleneimine, poly-L-lysine, chitosan and some others cationic polymers have been thoroughly studied as nucleic acid delivery systems in gene therapy. However, the drug release from these systems proceeds at a very low rate due to extremely high binding between a carrier and gene material. To reduce these interactions and to enhance drug release, we developed a set of amphiphilic polypeptides containing positively and negatively charged amino acids as well as a hydrophobic one. The copolymers obtained were characterized by size-exclusion chromatography, static light scattering, HPLC amino acid analysis and (HNMR)-H-1 spectroscopy. All copolymers formed particles due to a self-assembly in aqueous media. Depending on polypeptide composition, the formation of particles with hydrodynamic diameters from 180 to 900 nm was observed. Stability of polymer particles, loading and release efficiency were carefully studied. Cellular uptake of the particles was efficient and their cytotoxicity was negligible. The application of polymer carriers, containing siRNA, to vascular endothelial growth factor (VEGF-A165) silencing of ARPE-19 cells was successful. The gene silencing was confirmed by suppression of both messenger RNA and protein expression.
  • Zhong, Wenbin; Lin, Weize; Yang, Yingjie; Chen, Dan; Cao, Xiuye; Xu, Mengyang; Pan, Guoping; Chen, Huanzhao; Zheng, Jie; Feng, Xiaoqin; Yang, Li hua; Lai, Chaofeng; Olkkonen, Vesa M.; Xu, Jun; Cui, Shuzhong; Yan, Daoguang (2022)
    Lipid remodeling is crucial for malignant cell transformation and tumorigenesis, but the precise molecular processes involved and direct evidences for these in vivo remain elusive. Here, we report that oxysterol-binding protein (OSBP)-related protein 4 L (ORP4L) is expressed in adult T-cell leukemia (ATL) cells but not normal T-cells. In ORP4L knock-in T-cells, ORP4L dimerizes with OSBP to control the shuttling of OSBP between the Golgi apparatus and the plasma membrane (PM) as an exchanger of phosphatidylinositol 4-phosphate [PI(4)P]/cholesterol. The PI(4)P arriving at the PM via this transport machinery replenishes phosphatidylinositol 4,5-bisphosphate [PI(4,5)P-2] and phosphatidylinositol (3,4,5) trisphosphate [PI(3,4,5)P-3] biosynthesis, thus contributing to PI3K/AKT hyperactivation and T-cell deterioration in vitro and in vivo. Disruption of ORP4L and OSBP dimerization disables PI(4)P transport and T-cell leukemogenesis. In summary, we identify a non-vesicular lipid transport machinery between Golgi and PM maintaining the oncogenic signaling competence initiating T-cell deterioration and leukemogenesis. The oxysterol-binding protein-related protein 4 (ORP4L) is expressed in T-cell acute lymphoblastic leukemia and is required for leukemogenesis. Here the authors show that ORP4L orchestrates the transport of the phospholipid PI(4)P from Golgi to the plasma membrane, contributing to PI3K/AKT hyperactivation and T-cell leukemogenesis.
  • Zhu, Ya-Di; Pang, Hui-Lin; Zhou, Qi-Hang; Qin, Zi-Fei; Jin, Qiang; Finel, Moshe; Wang, Yi-Nan; Qin, Wei-Wei; Lu, Yin; Wang, Dan-Dan; Ge, Guang-Bo (2020)
    The human UDP-glucuronosyltransferase 1A1 (UGT1A1), one of the most essential conjugative enzymes, is responsible for the metabolism and detoxification of bilirubin and other endogenous substances, as well as many different xenobiotic compounds. Deciphering UGT1A1 relevance to human diseases and characterizing the effects of small molecules on the activities of UGT1A1 requires reliable tools for probing the function of this key enzyme in complex biological matrices. Herein, an easy-to-use assay for highly-selective and sensitive monitoring of UGT1A1 activities in various biological matrices, using liquid chromatography with fluorescence detection (LC-FD), has been developed and validated. The newly developed LC-FD based assay has been confirmed in terms of sensitivity, specificity, precision, quantitative linear range and stability. One of its main advantages is lowering the limits of detection and quantification by about 100-fold in comparison to the previous assay that used the same probe substrate, enabling reliable quantification of lower amounts of active enzyme than any other method. The precision test demonstrated that both intra- and inter-day variations for this assay were less than 5.5%. Furthermore, the newly developed assay has also been successfully used to screen and characterize the regulatory effects of small molecules on the expression level of UGT1A1 in living cells. Overall, an easy-to-use LC-FD based assay has been developed for ultra-sensitive UGT1A1 activities measurements in various biological systems, providing an inexpensive and practical approach for exploring the role of UGT1A1 in human diseases, interactions with xenobiotics, and characterization modulatory effects of small molecules on this conjugative enzyme. (c) 2020 Xi'an Jiaotong University. Production and hosting by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
  • Puranen, Jooseppi; Koponen, Sanna; Nieminen, Tiina; Kanerva, Iiris; Kokki, Emmi; Toivanen, Pyry; Urtti, Arto; Ylä-Herttuala, Seppo; Ruponen, Marika (2022)
    Pathological angiogenesis related to neovascularization in the eye is mediated through vascular endothelial growth factors (VEGFs) and their receptors. Ocular neovascular-related diseases are mainly treated with anti-VEGF agents. In this study we evaluated the efficacy and safety of novel gene therapy using adeno associated virus 2 vector expressing a truncated form of soluble VEGF receptor-2 fused to the Fc-part of human IgG1 (AAV2-sVEGFR-2-Fc) to inhibit ocular neovascularization in laser induced choroidal neovascularization (CNV) in mice. The biological activity of sVEGFR-2-Fc was determined in vitro. It was shown that sVEGFR-2-Fc secreted from ARPE-19 cells was able to bind to VEGF-A165 and reduce VEGF-A165 induced cell growth and survival. A single intravitreal injection (IVT) of AAV2-sVEGFR-2-Fc (1 mu l, 4.7 x 1012 vg/ml) one-month prior laser photocoagu-lation did not cause any changes in the retinal morphology and significantly suppressed fluorescein leakage at 7, 14, 21 and 28 days post-lasering compared to controls. Macrophage infiltration was observed after the injection of both AAV2-sVEGFR-2-Fc and PBS. Our findings indicate that AAV2 mediated gene delivery of the sVEGFR-2-Fc efficiently reduces formation of CNV and could be developed to a therapeutic tool for the treatment of retinal diseases associated with neovascularization.
  • Windbichler, Katharina; Michalopoulou, Eleni; Palamides, Pia; Pesch, Theresa; Jelinek, Christine; Vapalahti, Olli; Kipar, Anja; Hetzel, Udo; Hepojoki, Jussi (2019)
    Boid Inclusion Body Disease (BIBD) is a potentially fatal disease reported in captive boid snakes worldwide that is caused by reptarenavirus infection. Although the detection of intracytoplasmic inclusion bodies (IB) in blood cells serves as the gold standard for the ante mortem diagnosis of BIBD, the mechanisms underlying IB formation and the pathogenesis of BIBD are unknown. Knowledge on the reptile immune system is sparse compared to the mammalian counterpart, and in particular the response towards reptarenavirus infection is practically unknown. Herein, we investigated a breeding collection of 70 Boa constrictor snakes for BIBD, reptarenavirus viraemia, anti-reptarenavirus IgM and IgY antibodies, and population parameters. Using NGS and RT-PCR on pooled blood samples of snakes with and without BIBD, we could identify three different reptarenavirus S segments in the collection. The examination of individual samples by RT-PCR indicated that the presence of University of Giessen virus (UGV)-like S segment strongly correlates with IB formation. We could also demonstrate a negative correlation between BIBD and the presence of anti-UGV NP IgY antibodies. Further evidence of an association between antibody response and BIBD is the finding that the level of anti-reptarenavirus antibodies measured by ELISA was lower in snakes with BIBD. Furthermore, female snakes had a significantly lower body weight when they had BIBD. Taken together our findings suggest that the detection of the UGV-/S6-like S segment and the presence of anti-reptarenavirus IgY antibodies might serve as a prognostic tool for predicting the development of BIBD.
  • Jantti, Maria H.; Talman, Virpi; Räsänen, Kati; Tarvainen, Ilari; Koistinen, Hannu; Tuominen, Raimo K. (2018)
    Prostate cancer is one of the most common cancers in men. Although it has a relatively high 5-year survival rate, development of resistance to standard androgen-deprivation therapy is a significant clinical problem. Therefore, novel therapeutic strategies are urgently needed. The protein kinase C (PKC) family is a putative prostate cancer drug target, but so far no PKC-targeting drugs are available for clinical use. By contrast to the standard approach of developing PKC inhibitors, we have developed isophthalate derivatives as PKC agonists. In this study, we have characterized the effects of the most potent isophthalate, 5-(hydroxymethyl) isophthalate 1a3 (HMI-1a3), on three prostate cancer cell lines (LNCaP, DU145, and PC3) using both 2D and 3D cell culture models. In 2D cell culture, HMI-1a3 reduced cell viability or proliferation in all cell lines as determined by the metabolic activity of the cells (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay) and thymidine incorporation. However, the mechanism of action in LNCaP cells was different to that in DU145 or PC3 cells. In LNCaP cells, HMI-1a3 induced a PKC-dependent activation of caspase 3/7, indicating an apoptotic response, whereas in DU145 and PC3 cells, it induced senescence, which was independent of PKC. This was observed as typical senescent morphology, increased beta-galactosidase activity, and upregulation of the senescence marker p21 and downregulation of E2F transcription factor 1. Using a multicellular spheroid model, we further showed that HMI-1a3 affects the growth of LNCaP and DU145 cells in a 3D culture, emphasizing its potential as a lead compound for cancer drug development.
  • Onali, Tuulia; Kivimäki, Anne; Mauramo, Matti; Salo, Tuula; Korpela, Riitta (2021)
    Wild berries are part of traditional Nordic diets and are a rich source of phytochemicals, such as polyphenols. Various berry treatments have shown to interfere with cancer progression in vitro and in vivo. Here, we systematically reviewed the anticancer effects of two Nordic wild berries of the Vaccinium genus, lingonberry (Vaccinium vitis-idaea) and bilberry (Vaccinium myrtillus), on digestive tract cancers. The review was conducted according to the PRISMA 2020 guidelines. Searches included four databases: PubMed, Scopus, Web of Science, and CAB abstracts. Publications not written in English, case-reports, reviews, and conference abstracts were excluded. Moreover, studies with only indirect markers of cancer risk or studies with single compounds not derived from lingonberry or bilberry were not included. Meta-analysis was not performed. The majority (21/26) of studies investigated bilberry and colorectal cancer. Experimental studies on colorectal cancer indicated that bilberry inhibited intestinal tumor formation and cancer cell growth. One uncontrolled pilot human study supported the inhibitory potential of bilberry on colorectal cancer cell proliferation. Data from all 10 lingonberry studies suggests potent inhibition of cancer cell growth and tumor formation. In conclusion, in vitro and animal models support the antiproliferative and antitumor effects of various bilberry and lingonberry preparations on digestive tract cancers.
  • Kinnunen, P. T. T.; Murtola, T. J.; Talala, K.; Taari, K.; Tammela, T. L. J.; Auvinen, A. (2019)
    PurposeAnticoagulants may reduce mortality of cancer patients, though the evidence remains controversial. We studied the association between different anticoagulants and cancer death.MethodsAll anticoagulant use during 1995-2015 was analyzed among 75,336 men in the Finnish Randomized Study of Screening for Prostate Cancer. Men with prevalent cancer were excluded. Multivariable Cox regression was performed to compare risk of death from any cancer and disease-specific death from 9 specific cancer types between (1) anticoagulant users overall and (2) warfarin users compared to anticoagulant non-users and (3) warfarin or (4) low-molecular-weight heparins (LMWH) compared to users of other anticoagulants. Medication use was analyzed as time-dependent variable to minimize immortal time bias. 1-, 2- and 3-year lag-time analyses were performed.ResultsDuring a median follow-up of 17.2years, a total of 27,233 men died of whom 8033 with cancer as the primary cause of death. In total, 32,628 men (43%) used anticoagulants. Any anticoagulant use was associated with an increased risk of cancer death (HR=2.50, 95% CI 2.37-2.64) compared to non-users. Risk was similar independent of the amount, duration, or intensity of use. The risk increase was observed both among warfarin and LMWH users, although not as strong in warfarin users. Additionally, cancer-specific risks of death were similar to overall cancer mortality in all anticoagulant categories.ConclusionOur study does not support reduced cancer mortality among anticoagulant users. Future studies on drug use and cancer mortality should be adjusted for anticoagulants as they are associated with significantly higher risk of cancer death.
  • Hanemaaijer, Evelyn S.; Panahi, Mahmod; Swaddiwudhipong, Nol; Tikka, Saara; Winblad, Bengt; Viitanen, Matti; Piras, Antonio; Behbahani, Homira (2018)
    Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is a familial progressive degenerative disorder and is caused by mutations in NOTCH3 gene. Previous study reported that mutant NOTCH3 is more prone to form aggregates than wild-type NOTCH3 and the mutant aggregates are resistant to degradation. We hypothesized that aggregation or accumulation of NOTCH3 could be due to impaired lysosomal-autophagy machinery in VSMC. Here, we investigated the possible cause of accumulation/aggregation of NOTCH3 in CADASIL using cerebral VSMCs derived from control and CADASIL patients carrying NOTCH3(RI33C) mutation. Thioflavin-S-staining confirmed the increased accumulation of aggregated NOTCH3 in VSMCR133C compared to VSMCWT. Increased levels of the lysosomal marker, Lamp2, were detected in VSMCR133C, which also showed co-localization with NOTCH3 using double-immunohistochemistry. Increased level of LC3-II/LC3-I ratio was observed in VSMCR133C suggesting an accumulation of autophagosomes. This was coupled with the decreased co-localization of NOTCH3 with LC3, and Lamp2 and, further, increase of p62/SQSTM1 levels in VSMCR133C compared to the VSMCWT. In addition, Western blot analysis indicated phosphorylation of p-ERK, p-S6RP, and p-P70 S6K. Altogether, these results suggested a dysfunction in the autophagy-lysosomal pathway in VSMCR133C. The present study provides an interesting avenue of the research investigating the molecular mechanism of CADASIL.
  • Landolt, Lea; Furriol, Jessica; Babickova, Janka; Ahmed, Lavina; Eikrem, Oystein; Skogstrand, Trude; Scherer, Andreas; Suliman, Salwa; Leh, Sabine; Lorens, J. B.; Gausdal, Gro; Marti, H.P.; Osman, Tarig (2019)
    The AXL receptor tyrosine kinase (RTK) is involved in partial epithelial-to-mesenchymal transition (EMT) and inflammation - both main promoters of renal fibrosis development. The study aim was to investigate the role of AXL inhibition in kidney fibrosis due to unilateral ureteral obstruction (UUO). Eight weeks old male C57BL/6 mice underwent UUO and were treated with oral AXL inhibitor bemcentinib (n = 22), Angiotensin-converting enzyme inhibitor (ACEI, n = 10), ACEI and bemcentinib (n = 10) or vehicle alone (n = 22). Mice were sacrificed after 7 or 15 days and kidney tissues were analyzed by immunohistochemistry (IHC), western blot, ELISA, Sirius Red (SR) staining, and hydroxyproline (Hyp) quantification. RNA was extracted from frozen kidney tissues and sequenced on an Illumina HiSeq4000 platform. After 15 days the ligated bemcentinib-treated kidneys showed less fibrosis compared to the ligated vehicle-treated kidneys in SR analyses and Hyp quantification. Reduced IHC staining for Vimentin (VIM) and alpha smooth muscle actin (alpha SMA), as well as reduced mRNA abundance of key regulators of fibrosis such as transforming growth factor (Tgf beta), matrix metalloproteinase 2 (Mmp2), Smad2, Smad4, myofibroblast activation (Aldh1a2, Crlf1), and EMT (Snai1,2, Twist), in ligated bemcentinib-treated kidneys was compatible with reduced (partial) EMT induction. Furthermore, less F4/80 positive cells, less activity of pathways related to the immune system and lower abundance of MCP1, MCP3, MCP5, and TARC in ligated bemcentinib-treated kidneys was compatible with reduction in inflammatory infiltrates by bemcentinib treatment. The AXL RTK pathway represents a promising target for pharmacologic therapy of kidney fibrosis.
  • Bakour, Meryem; Laaroussi, Hassan; Ousaaid, Driss; El Ghouizi, Asmae; Es-Safi, Imane; Mechchate, Hamza; Lyoussi, Badiaa (2022)
    Bee bread is a natural product obtained from the fermentation of bee pollen mixed with bee saliva and flower nectar inside the honeycomb cells of a hive. Bee bread is considered a functional product, having several nutritional virtues and various bioactive molecules with curative or preventive effects. This paper aims to review current knowledge regarding the chemical composition and medicinal properties of bee bread, evaluated in vitro and in vivo, and to highlight the benefits of the diet supplementation of bee bread for human health. Bee bread extracts (distilled water, ethanol, methanol, diethyl ether, and ethyl acetate) have been proven to have antioxidant, antifungal, antibacterial, and antitumoral activities, and they can also inhibit alpha-amylase and angiotensin I-converting enzyme in vitro. More than 300 compounds have been identified in bee bread from different countries around the world, such as free amino acids, sugars, fatty acids, minerals, organic acids, polyphenols, and vitamins. In vivo studies have revealed the efficiency of bee bread in relieving several pathological cases, such as hyperglycemia, hyperlipidemia, inflammation, and oxidative stress.
  • Maliniemi, Pilvi; Laukkanen, Kirsi; Vakeva, Liisa; Dettmer, Katja; Lipsanen, Tuomas; Jeskanen, Leila; Bessede, Alban; Oefner, Peter J.; Kadin, Marshall E.; Ranki, Annamari (2017)
    Indoleamine 2,3-deoxygenase 1 (IDO1) induces immune tolerance in the tumor microenvironment (TME) and is recognized as a potential therapeutic target. We studied the expression of both IDO1 and the related tryptophan 2,3-dioxygenase (TDO) in several different subtypes of cutaneous T-cell lymphoma (CTCL), and evaluated the kynurenine (KYN) pathway in the local TME and in patient sera. Specimens from the total of 90 CTCL patients, including mycosis fungoides (MF, n = 37), lymphomatoid papulosis (LyP, n = 36), primary cutaneous anaplastic large cell lymphoma (pcALCL, n = 4), subcutaneous panniculitis-like T-cell lymphoma (SPTCL n = 13), and 10 patients with inflammatory lichen ruber planus (LRP), were analyzed by immunohistochemistry (IHC), immunofluorescence (IF), quantitative PCR, and/or liquid chromatography-tandem mass spectrometry (LC-MS/MS). Three CTCL cell lines also were studied. Expression of both IDO1 and TDO was upregulated in CTCL. In MF specimens and in the MF cell line MyLa2000, IDO1 expression exceeded that of TDO, whereas the opposite was true for LyP, ALCL, and corresponding Mac1/2A cell lines. The spectrum of IDO1-expressing cell types differed among CTCL subtypes and was reflected in the clinical behavior. In MF, SPTCL, and LyP, IDO1 was expressed by malignant cells and by CD33(+) myeloid-derived suppressor cells, whereas in SPTCL CD163(+) tumor-associated macrophages also expressed IDO1. Significantly elevated serum KYN/Trp ratios were found in patients with advanced stages of MF. Epacadostat, an IDO1 inhibitor, induced a clear decrease in KYN concentration in cell culture. These results show the importance of IDO1/TDO-induced immunosuppression in CTCL and emphasize its role as a new therapeutic target.
  • Mäkitie, Riikka E.; Kämpe, Anders; Costantini, Alice; Alm, Jessica J.; Magnusson, Per; Mäkitie, Outi (2020)
    Recent advancements in genetic research have uncovered new forms of monogenic osteoporosis, expanding our understanding of the molecular pathways regulating bone health. Despite active research, knowledge on the pathomechanisms, disease-specific biomarkers, and optimal treatment in these disorders is still limited. Mutations in WNT1, encoding a WNT/beta-catenin pathway ligand WNT1, and PLS3, encoding X chromosomally inherited plastin 3 (PLS3), both result in early-onset osteoporosis with prevalent fractures and disrupted bone metabolism. However, despite marked skeletal pathology, conventional bone markers are usually normal in both diseases. Our study aimed to identify novel bone markers in PLS3 and WNT1 osteoporosis that could offer diagnostic potential and shed light on the mechanisms behind these skeletal pathologies. We measured several parameters of bone metabolism, including serum dickkopf-1 (DKK1), sclerostin, and intact and C-terminal fibroblast growth factor 23 (FGF23) concentrations in 17 WNT1 and 14 PLS3 mutation-positive subjects. Findings were compared with 34 healthy mutation-negative subjects from the same families. Results confirmed normal concentrations of conventional metabolic bone markers in both groups. DKK1 concentrations were significantly elevated in PLS3 mutation-positive subjects compared with WNT1 mutation-positive subjects (p <.001) or the mutation-negative subjects (p = .002). Similar differences were not seen in WNT1 subjects. Sclerostin concentrations did not differ between any groups. Both intact and C-terminal FGF23 were significantly elevated in WNT1 mutation-positive subjects (p = .039 and p = .027, respectively) and normal in PLS3 subjects. Our results indicate a link between PLS3 and DKK1 and WNT1 and FGF23 in bone metabolism. The normal sclerostin and DKK1 levels in patients with impaired WNT signaling suggest another parallel regulatory mechanism. These findings provide novel information on the molecular networks in bone. Extended studies are needed to investigate whether these biomarkers offer diagnostic value or potential as treatment targets in osteoporosis. (c) 2020 American Society for Bone and Mineral Research.