Publications by authors named "Sinem Yilmaz Ozcan"

6 Publications

  • Page 1 of 1

Widespread brain parenchymal HMGB1 and NF-κB neuroinflammatory responses upon cortical spreading depolarization in familial hemiplegic migraine type 1 mice.

Neurobiol Dis 2021 08 10;156:105424. Epub 2021 Jun 10.

Institute of Neurological Sciences and Psychiatry, Hacettepe University, Ankara, Turkey. Electronic address:

Neuroinflammatory changes involving neuronal HMGB1 release and astrocytic NF-κB nuclear translocation occur following cortical spreading depolarization (CSD) in wildtype (WT) mice but it is unknown to what extent this occurs in the migraine brain. We therefore investigated in familial hemiplegic migraine type 1 (FHM1) knock-in mice, which express an intrinsic hyperexcitability phenotype, the extent of neuroinflammation without and after CSD. CSD was evoked in one hemisphere by pinprick (single CSD) or topical KCl application (multiple CSDs). Neuroinflammatory (HMGB1, NF-κB) and neuronal activation (pERK) markers were investigated by immunohistochemistry in the brains of WT and FHM1 mutant mice without and after CSD. Effects of NMDA receptor antagonism on basal and CSD-induced neuroinflammatory changes were examined by, respectively, systemically administered MK801 and ifenprodil or topical MK801 application. In FHM1 mutant mice, CSD caused enhanced neuronal HMGB1 release and astrocytic NF-κB nuclear translocation in the cortex and subcortical areas that were equally high in both hemispheres. In WT mice such effects were only pronounced in the hemisphere in which CSD was induced. Neuroinflammatory responses were associated with pERK expression indicating neuronal activation. Upon CSD, contralateral cortical and striatal HMGB1 release was reduced by topical application of MK801 in the hemisphere contralateral to the one in which CSD was induced. This study reveals that neuroinflammatory activation after CSD is widespread and extends to the contralateral hemisphere, particularly in brains of FHM1 mutant mice. Effective blockade of CSD-induced neuroinflammatory responses in the contralateral hemisphere in FHM1 mice by local NMDA receptor antagonism suggests that neuronal hyperexcitability-related neuroinflammation is relevant in migraine pathophysiology, but possibly also other neurological disorders in which spreading depolarization is involved.
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http://dx.doi.org/10.1016/j.nbd.2021.105424DOI Listing
August 2021

F-actin polymerization contributes to pericyte contractility in retinal capillaries.

Exp Neurol 2020 10 29;332:113392. Epub 2020 Jun 29.

Hacettepe University, Institute of Neurological Sciences and Psychiatry, Ankara, Turkey. Electronic address:

Although it has been documented that central nervous system pericytes are able to contract in response to physiological, pharmacological or pathological stimuli, the underlying mechanism of pericyte contractility is incompletely understood especially in downstream pericytes that express low amounts of alpha-smooth muscle actin (α-SMA). To study whether pericyte contraction involves F-actin polymerization as in vascular smooth muscle cells, we increased retinal microvascular pericyte tonus by intravitreal injection of a vasoconstrictive agent, noradrenaline (NA). The contralateral eye of each mouse was used for vehicle injection. The retinas were rapidly extracted and fixed within 2 min after injections. Polymeric/filamentous (F-actin) and monomeric/globular (G-actin) forms of actin were labeled by fluorescently-conjugated phalloidin and deoxyribonuclease-I, respectively. We studied 108 and 83 pericytes from 6 NA- and 6 vehicle-treated retinas and, found that F/G-actin ratio, a microscopy-based index of F-actin polymerization, significantly increased in NA-treated retinas [median (IQR): 4.2 (3.1) vs. 3.5 (2.1), p = .006], suggesting a role for F-actin polymerization in pericyte contractility. Shift from G-actin monomers to polymerized F-actin was more pronounced in 5th and 6th order contracted pericytes compared to non-contracted ones [7.6 (4.7) vs. 3.2 (1.2), p < .001], possibly due to their dependence on de novo F-actin polymerization for contractile force generation because they express α-SMA in low quantities. Capillaries showing F-actin polymerization had significantly reduced diameters compared to the ones that did not exhibit increased F/G-actin ratio in pericytes [near soma / branch origin diameter; 0.67 (0.14) vs. 0.81 (0.34), p = .005]. NA-responsive capillaries generally did not show nodal constrictions but a tide-like diameter decrease, reaching a maximum near pericyte soma. These findings suggest that pericytes on high order downstream capillaries have F-actin-mediated contractile capability, which may contribute to the vascular resistance and blood flow regulation in capillary bed.
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http://dx.doi.org/10.1016/j.expneurol.2020.113392DOI Listing
October 2020

Retinal ischemia induces α-SMA-mediated capillary pericyte contraction coincident with perivascular glycogen depletion.

Acta Neuropathol Commun 2019 08 20;7(1):134. Epub 2019 Aug 20.

Institute of Neurological Sciences and Psychiatry, Hacettepe University, Sihhiye, 06100, Ankara, Turkey.

Increasing evidence indicates that pericytes are vulnerable cells, playing pathophysiological roles in various neurodegenerative processes. Microvascular pericytes contract during cerebral and coronary ischemia and do not relax after re-opening of the occluded artery, causing incomplete reperfusion. However, the cellular mechanisms underlying ischemia-induced pericyte contraction, its delayed emergence, and whether it is pharmacologically reversible are unclear. Here, we investigate i) whether ischemia-induced pericyte contractions are mediated by alpha-smooth muscle actin (α-SMA), ii) the sources of calcium rise in ischemic pericytes, and iii) if peri-microvascular glycogen can support pericyte metabolism during ischemia. Thus, we examined pericyte contractility in response to retinal ischemia both in vivo, using adaptive optics scanning light ophthalmoscopy and, ex vivo, using an unbiased stereological approach. We found that microvascular constrictions were associated with increased calcium in pericytes as detected by a genetically encoded calcium indicator (NG2-GCaMP6) or a fluoroprobe (Fluo-4). Knocking down α-SMA expression with RNA interference or fixing F-actin with phalloidin or calcium antagonist amlodipine prevented constrictions, suggesting that constrictions resulted from calcium- and α-SMA-mediated pericyte contractions. Carbenoxolone or a Cx43-selective peptide blocker also reduced calcium rise, consistent with involvement of gap junction-mediated mechanisms in addition to voltage-gated calcium channels. Pericyte calcium increase and capillary constrictions became significant after 1 h of ischemia and were coincident with depletion of peri-microvascular glycogen, suggesting that glucose derived from glycogen granules could support pericyte metabolism and delay ischemia-induced microvascular dysfunction. Indeed, capillary constrictions emerged earlier when glycogen breakdown was pharmacologically inhibited. Constrictions persisted despite recanalization but were reversible with pericyte-relaxant adenosine administered during recanalization. Our study demonstrates that retinal ischemia, a common cause of blindness, induces α-SMA- and calcium-mediated persistent pericyte contraction, which can be delayed by glucose driven from peri-microvascular glycogen. These findings clarify the contractile nature of capillary pericytes and identify a novel metabolic collaboration between peri-microvascular end-feet and pericytes.
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http://dx.doi.org/10.1186/s40478-019-0761-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6701129PMC
August 2019

Non-viral delivery of chemically modified mRNA to the retina: Subretinal versus intravitreal administration.

J Control Release 2019 08 29;307:315-330. Epub 2019 Jun 29.

Ghent Research Group on Nanomedicines, Laboratory of General Biochemistry and Physical Pharmacy, Ghent University, Ottergemsesteenweg 460, 9000 Ghent, Belgium; Cancer Research Institute Ghent (CRIG), Ghent University Hospital, De Pintelaan 185, 9000 Ghent, Belgium. Electronic address:

mRNA therapeutics have recently experienced a new wave of interest, mainly due to the discovery that chemical modifications to mRNA's molecular structure could drastically reduce its inherent immunogenicity and perceived instability. On this basis, we aimed to explore the potential of chemically stabilized mRNA for ocular applications. More specifically, we investigated the behavior of mRNA-loaded lipid-based carriers in human retinal cells (in vitro), in bovine retinal explants (ex vivo) and in mouse retinas (in vivo). We demonstrate a clear superiority of mRNA over pDNA to induce protein expression in different retinal cell types, which was further enhanced by chemical modification of the mRNA, providing up to ~1800-fold higher reporter gene expression compared to pDNA. Moreover, transgene expression could be detected for at least 20 days after a single administration of chemically modified mRNA in vitro. We furthermore determined the localization and extent of mRNA expression depending on the administration route. After subretinal (SR) administration, mRNA expression was observed in vivo and ex vivo. By contrast, intravitreal (IVT) administration resulted in limited expression in vivo. Using ex vivo bovine explants with an intact vitreoretinal (VR) interface we could attribute this to the inner limiting membrane (ILM), which presents a large barrier for non-viral delivery of mRNA, trapping mRNA complexes at the vitreal side. When the vitreous was removed, which compromises the ILM, mRNA expression was apparent and seemed to colocalize with Müller cells or photoreceptors after respectively IVT or SR administration. Taken together, this study represents a first step towards mRNA-mediated therapy for retinal diseases.
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http://dx.doi.org/10.1016/j.jconrel.2019.06.042DOI Listing
August 2019

Capillary pericytes express α-smooth muscle actin, which requires prevention of filamentous-actin depolymerization for detection.

Elife 2018 03 21;7. Epub 2018 Mar 21.

Institute of Neurological Sciences and Psychiatry, Hacettepe University, Ankara, Turkey.

Recent evidence suggests that capillary pericytes are contractile and play a crucial role in the regulation of microcirculation. However, failure to detect components of the contractile apparatus in capillary pericytes, most notably α-smooth muscle actin (α-SMA), has questioned these findings. Using strategies that allow rapid filamentous-actin (F-actin) fixation (i.e. snap freeze fixation with methanol at -20°C) or prevent F-actin depolymerization (i.e. with F-actin stabilizing agents), we demonstrate that pericytes on mouse retinal capillaries, including those in intermediate and deeper plexus, express α-SMA. Junctional pericytes were more frequently α-SMA-positive relative to pericytes on linear capillary segments. Intravitreal administration of short interfering RNA (α-SMA-siRNA) suppressed α-SMA expression preferentially in high order branch capillary pericytes, confirming the existence of a smaller pool of α-SMA in distal capillary pericytes that is quickly lost by depolymerization. We conclude that capillary pericytes do express α-SMA, which rapidly depolymerizes during tissue fixation thus evading detection by immunolabeling.
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http://dx.doi.org/10.7554/eLife.34861DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5862523PMC
March 2018

Epigenetic mechanisms underlying the dynamic expression of cancer-testis genes, PAGE2, -2B and SPANX-B, during mesenchymal-to-epithelial transition.

PLoS One 2014 17;9(9):e107905. Epub 2014 Sep 17.

Department of Molecular Biology and Genetics, Bilkent University, Ankara, Turkey.

Cancer-testis (CT) genes are expressed in various cancers but not in normal tissues other than in cells of the germline. Although DNA demethylation of promoter-proximal CpGs of CT genes is linked to their expression in cancer, the mechanisms leading to demethylation are unknown. To elucidate such mechanisms we chose to study the Caco-2 colorectal cancer cell line during the course of its spontaneous differentiation in vitro, as we found CT genes, in particular PAGE2, -2B and SPANX-B, to be up-regulated during this process. Differentiation of these cells resulted in a mesenchymal-to-epithelial transition (MET) as evidenced by the gain of epithelial markers CDX2, Claudin-4 and E-cadherin, and a concomitant loss of mesenchymal markers Vimentin, Fibronectin-1 and Transgelin. PAGE2 and SPAN-X up-regulation was accompanied by an increase in Ten-eleven translocation-2 (TET2) expression and cytosine 5-hydroxymethylation as well as the disassociation of heterochromatin protein 1 and the polycomb repressive complex 2 protein EZH2 from promoter-proximal regions of these genes. Reversal of differentiation resulted in down-regulation of PAGE2, -2B and SPANX-B, and induction of epithelial-to-mesenchymal transition (EMT) markers, demonstrating the dynamic nature of CT gene regulation in this model.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0107905PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4168264PMC
December 2015
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