Browsing by Subject "Fibrosis"

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Open Access
Identification of the fibrogenic cells in the liver
(2022-03) Bozlar, Müge
The liver is an important multitasking organ with diverse functions to maintain body homeostasis. Adjustment of glucose level, detoxification, secretion of blood proteins, and synthesis of bile acids are the among these crucial functions. The fibrosis is one of the most common liver injury types. Although liver has a high capacity for regeneration via the proliferation of cells, this regeneration ability is directly dependent on the severity of fibrosis. One of the proliferative cells during fibrosis is cholangiocytes located around the portal triad. The cholangiocytes contain a primary cilium, a non-motile, ciliated organelle. The response of primary cilium to fibrosis is not well understood. The aim of this thesis was to characterize responses of cholangiocytes to liver fibrosis. For this purpose, TAA induced fibrosis model was generated for different weeks of samples, 16 weeks, 20 weeks, and 24 weeks in mice. IHC stainings and data collection from confocal microscopy were performed. According to our study, we observed that around the portal vein, big cell masses are formed and these cells are migrating between portal veins by following a path upon fibrosis. Moreover, several cells are escaping from this path and expanding through the central vein. The main contributors of these giant cell masses are cholangiocytes which keep their primary cilia under fibrotic conditions. Furthermore, we identified a new subpopulation of Arl13b+ cholangiocytes which are undergoing partial EMT by expressing mesenchymal markers such as Vimentin, CD34, and Pdgfra in these cell masses. During this process, they gain novel mesenchymal features at the same time they maintain their epithelial character. As a following goal, we wanted to identify components of these big cell masses, so niche components of partial EMT undergoing Arl13b+ cholangiocytes. We identified that Vimentin, CD34, and Pdgfra mesenchymal cells and Thy1+ portal fibroblasts (minority) are components of these massive cells in addition to homeostatic and partial EMT undergoing Arl13b+ cholangiocytes. As a final objective, we identified novel, possible markers for partial EMT undergoing Arl13b+ cholangiocytes and their niche components. To conclude, the novelties of this thesis were the demonstration of the primary cilium response against fibrosis, the identification of a new Arl13b+ cholangiocyte subpopulation undergoing partial EMT, and the characterization of components of the migratory big cell masses under fibrotic conditions.
Open Access
Increased SGK1 activity potentiates mineralocorticoid/NaCl-induced kidney injury
(American Physiological Society, 2021-04-08) Sierra-Ramos, Catalina; Velazquez-Garcia, Silvia; Keskus, Ayşe Gökçe; Vastola-Mascolo, Arianna; Rodríguez-Rodríguez, Ana E.; Luis-Lima, Sergio; Hernández, Guadalberto; Navarro-González, Juan F.; Porrini, Esteban; Konu, Özlen; Alvarez de la Rosa, Diego
Serum and glucocorticoid-regulated kinase 1 (SGK1) stimulates aldosterone-dependent renal Na reabsorption and modulates blood pressure. In addition, genetic ablation or pharmacological inhibition of SGK1 limits the development of kidney inflammation and fibrosis in response to excess mineralocorticoid signaling. In this work, we tested the hypothesis that a systemic increase in SGK1 activity would potentiate mineralocorticoid/salt-induced hypertension and kidney injury. To that end, we used a transgenic mouse model with increased SGK1 activity. Mineralocorticoid/salt-induced hypertension and kidney damage was induced by unilateral nephrectomy and treatment with deoxycorticosterone acetate and NaCl in the drinking water for 6 wk. Our results show that although SGK1 activation did not induce significantly higher blood pressure, it produced a mild increase in glomerular filtration rate, increased albuminuria, and exacerbated glomerular hypertrophy and fibrosis. Transcriptomic analysis showed that extracellular matrix-and immune response-related terms were enriched in the downregulated and upregulated genes, respectively, in transgenic mice. In conclusion, we propose that systemically increased SGK1 activity is a risk factor for the development of mineralocorticoid-dependent kidney injury in the context of low renal mass and independently of blood pressure. NEW and NOTEWORTHY Increased activity of the protein kinase serum and glucocorticoid-regulated kinase 1 may be a risk factor for accelerated renal damage. Serum and glucocorticoid-regulated kinase 1 expression could be a marker for the rapid progression toward chronic kidney disease and a potential therapeutic target to slow down the process. © 2021 American Physiological Society. All rights reserved.
Open Access
Therapeutic potential of an immunosuppressive oligodeoxynucleotide encapsulated within liposomes on bleomycin-induced mouse model of lung inflammation and fibrosis
(2019-05) Kılıç, Gizem
Systemic sclerosis (SSc) is an autoimmune/autoinflammatory disease with unknown etiology. It is characterized by vascular dysfunction, inflammation and disseminated fibrosis of skin or internal organs. Although its prevalence is low, development of fibrosis on internal organs and lack of a curative treatment result in high morbidity. Current therapies targeting specific symptoms such as interstitial lung disease, Raynaud’s phenomenon and pulmonary arterial hypertension are inefficient, and at best, temporarily relieves the symptoms throughout the course of the treatment. Herein, we investigated the therapeutic potential of an immunosuppressive oligodeoxynucleotide expressing TTAGGG telomeric repeats which is known as the “A151 ODN” on bleomycin-induced mouse model of systemic sclerosis. A151 ODN is the single stranded synthetic form of the telomeric repeat sequence expressed on mammalian chromosome, and it contains four repeats of “TTAGGG” motif. In order to enhance the therapeutic effectivity while protecting its digestion from nuclease activity following administration, we encapsulated A151 ODN within anionic liposomes. Since pattern recognition receptors and their signaling pathways were demonstrated to initiate inflammation in SSc, we first explored the immunosuppressive capacity of A151 ODN by analyzing in vitro cytokine productions and surface marker expression levels. Similar with the previous findings, A151 ODN was highly potent to abolish cytokine production in response to TLR9 induction. Although A151 ODN by itself was not very effective to suppress cytokine secretion following TLR1/2 and TLR4 induction, encapsulation within anionic liposomes further improved the immunosuppressive potential in response to TLR engagement. Furthermore, flow cytometry analyses revealed that A151 ODN decreased antigen presentation capacity and activation of bone-marrow derived macrophages (BMDMs) in response to TLR stimulation which was demonstrated by the reduction in levels of surface MHCII and co-stimulatory molecules as well as proteins having role on macrophage adherence and migration. A151 ODN also inhibited transcription of two major genes known to play a critical role on the development of fibrosis, TGFβ and Col1a1, from fibroblasts. Following these promising results on A151 ODN’s immunosuppressive and anti-fibrotic potential, we tested its therapeutic role on bleomycin-induced lung inflammation and fibrosis on mice which reflects different phases of systemic sclerosis. First in vivo experiment that A151 ODN was used prior to bleomycin administration revealed that A151 ODN could prevent development of systemic sclerosis by reducing immune cell recruitment into alveolar space and suppressing the secretion of inflammatory cytokines. After that, we investigated if A151 ODN could abolish established lung inflammation triggered by bleomycin instillation. For that, we treated animals with an A151 ODN either in free form or encapsulated within anionic liposomes after lung inflammation was initiated following bleomycin instillation. Data indicated that A151 ODN reduced macrophage activation marker expressions, monocyte and neutrophil infiltration into alveolar space. Moreover, suppression on immune cells activation in bronchoalveolar lavage fluid (BALF) correlated with the inhibited cytokine production. As a result of reduced inflammation, pro-fibrotic gene expressions were less in A151 ODN-treated mice. Of note, liposomal encapsulation provided reduced gene expressions while failed to further enhance the immunosuppressive potential on surface marker expression or cytokine secretion of A151 ODN. Lastly, we tested whether treatment with liposome-encapsulated A151 ODN is still effective to regress fibrosis once it has been developed; therefore, we treated mice with single injection of liposomal A151 on different time points. Unfortunately, single instillation was insufficient to decrease fibrosis and macrophage activation as well as cytokine production. Taken together, our findings indicated that liposome-encapsulated A151 ODN is very potent to attenuate the lung inflammation whereas single injection was ineffective to regress established lung fibrosis.