Theses supervised by Prof. Dr. Fikrettin Şahin
47 theses · Yeditepe University
The effect of wheat (Triticum aestivum L.) derived exosomes on wound healing
Wound healing is the main physiological process which undergoes homeostasis, inflammation, proliferation, migration and tissue remodeling. Acute injuries usually heal in a short period of time but chronic illnesses associated with infection and prolonged inflammation, may resulted with the life-threatening infections, limb amputations that presents a serious challenge for the current technology. Although wound healing has been studied for decades, improved drugs are not still effective enough due to their high cost, low efficiency, having patient specific response, requiring wound care facilities and severe side effects. In this sense, developing new, safe, self-applicable, effective and cheap wound care products is still an attractive area of international research. Cell to cell communication is compulsory in order to sustain tissue homeostasis but most communication mechanisms between cells requires a coordination of the receptors and the signaling molecules. Recently, nano vesicle mediated cell communication become prominence because of the ability to shuttle different molecules from producing cells to the target cells via extracellular vesicles called exosomes. Previous studies reported that, exosomes released from plants such as grape, grapefruit, ginger and carrot have anti-inflammatory and anti-cancer properties. Although the underlying mechanisms of wound healing process of plant exosomes are still unknown. For this reason, in this study, for the first time, rate limiting factors of wound regeneration such as cell proliferation, migration, angiogenesis, gene and growth factor expression cell migration and proliferation was stimulated by using wheat derived exosomes by using both in vitro and in vivo approaches. According to the results, wheat exosomes increased cell proliferation, migration and angiogenesis, have anti-inflammatory effect, increased essential growth factor, and gene expression levels of HDF, HUVEC and HACAT cells in vitro. Moreover, wheat exosome-containing hydrogel formulations improved wound healing and histopathological rates in vivo. All in all, this research indicates a brand-new scope for wound healing therapeutics using wheat derived exosomes for the first time.
Determination and comparison of the therapeutic effectiveness of septal cartilage stem cells and adipose stem cells in house mite- allergic rat model
Allergic rhinitis (AR) can be simply defined as the presence of at least one of the following "symptoms after exposure to an airborne allergen: rhinorrhea, sneezing, itching and nasal congestion. AR is defined as a chronic inflammatory disease mediated by the antibodies of IgE-type. The most important step in AR management is to avoid allergens. Pharmacotherapy options are used for symptomatic treatment. Subcutaneous or sublingual allergy immunotherapy can be applied in moderate or severe AR, especially when unresponsive to standard treatments. Alternative treatment methods such as, acupuncture, homeopathy, intranasal air filter, cellulose and CO2 are being explored in various trials, but none of these methods have received widespread acceptance or approval. The implementation of Mesenchymal stem cells (MSCs), which had been reported to have immunomodulatory characteristics seem to be promising for AR treatment. This study was aimed to determine and compare the effects of systemic administrations of in vitro produced septal cartilage stem cells (SCs) and adipose SCs on rats with allergic rhinitis (AR). House dust mite allergens were isolated and administered to 3-4-month-old albino rats. Adipose and cartilage SCs that were produced in vitro were isolated and injected intravenously to rats. Several allergic inflammation parameters and histopathological analysis of nasal mucosal thickness of inferior turbinates were analyzed. Cartilage SC recipients had more reduction in mucosal thickness in the inferior turbinates compared to recipients of adipose SC. The cartilage SC group had greater reduction in serum IgE, TNF-α and IL-6 levels compared to the adipose SC group, however IL-13 level remained unaffected. Mesenchymal SC therapy, especially cartilage-derived SC therapy, was effective in decreasing allergic inflammation in an experimental model of AR in rats. The date in the present study showed that mesenchymal SC therapy has the potential to be an alternative option in the treatment of AR.
The effects of Triticum Aestivum Lin(Wheat) and Mentha Piperita(Peppermint) derived exosomes on adipogenic differentiation of human adipose stem cells
In living organisms, when food intake is excessive, this excess energy must be stored, and multicellular organisms have specialized cells or organs to store this excess energy as lipids. The enzymes, hormones, growth factors, and cytokines secreted by adipose tissue play a vital role in the regulation of biological activities and energy metabolism in animals. Adipogenesis is defined as the transformation of preadipocytes into mature fat cells. In this study, adipogenesis was done by using human-derived adipose stem cells ordered from ATCC, and adipogenesis differentiation of these cells. In order to see whether they play a role in cellular fat breakdown, the exosomes isolated from Triticum aestivum Lin and Mentha piperita were characterized by size analysis on the NTA device. In order to determine whether exosomes have a toxic effect on human adipose stem cells, the MTS cytotoxicity experiment was carried out over six doses, and as a result, the experiments were continued by choosing a optimum dose. The optimum dose effect of exosomes from peppermint and wheatgrass on adipogenically differentiated cells were observed with Oil-Red-O staining. Adipogenic markers and their expressions in obesity markers were examined in the cells treated with mint exosome, in addition to these, the results were supported by ELISA and immunocytochemistry experiments. In line with these results, it was observed that peppermint exosome upregulated adipogenesis and induced obesity in cells treated with it. In addition, in cells treated with wheatgrass juice exome, it was observed that it downregulated adipogenesis and played a supportive role in the destruction of intracellular fat, as seen in the result of Oil-Red O staining. The findings of this study are encouraging for further research into the possible positive impacts of plant exosomes on adipose biology. In this sense, our findings guide for future research into plant exosome-based chemicals for the prevention and treatment of obesity, a modern pandemic with a strong relationship to comorbidities that is common globally.
Quantum carbon dot-gold nanoparticle hybrid nanomaterials as new generation contrast agents in bioimaging
"daha sonra doldurulacCarbon dot-gold nanoparticle hybrid systems [CD/AuNP) show great potential for biological imaging of cancer due to the enhanced photoluminescence capability arising from the synergistic coupling between the carbon dots and the gold nanoparticles called fluorescence resonance energy transfer (FRET). Compared to conventional dyes, CD/AuNP hybrid systems do not exhibit bleaching effects, so their fluorescence capacity does not decrease over time. Also, this hybrid system does not exhibit cellular cytotoxicity. Traditionally, CD/AuNP hybrids are prepared separately by wet chemical methods and then mixed together. Namely, both nanoparticle solutions (CD and AuNP) are prepared in different reaction containers and then mixed together to obtain the hybrid. However, in the proposed solid-state synthesis method, it is possible to prepare the hybrid in the same reaction container, which shortens the time and increases the photoluminescence yield of the nanoparticles. The aim of this study is to investigate the photoluminescence properties of CD/AuNP hybrid systems obtained with different reaction parameters via a one-pot solid-state synthesis method. The photoluminescent properties of the synthesized particles were investigated by using fluorescent spectroscopy. The particles were used as cellular imaging agents on different cancer cell lines such as A549, Panc02, and MCF -7. HDF was used as a control. Afterward, the cellular proliferation capacity, cellular cytotoxicity, and bio-imaging properties of the CD /AuNP systems were analyzed to find the most effective system for further studies.aktır"
Investigation of the potential of post-translational modifications as biomarkers and therapeutic targets for amyotrophic lateral sclerosis disease
Amyotrophic lateral sclerosis (ALS) is a neurodegenerative and fatal disease that affects upper and lower motor neurons with damage to the motor neurons in the cortex, brainstem, and spinal cord regions. ALS results in death due to painless and progressive muscle weakness and respiratory failure after progression to paralysis from atrophy. Although there is an increased knowledge about ALS generated in a series of genetic and molecular pathological researches, the aetiology of many ALS cases is still unknown. The diagnosis of ALS is established through a comprehensive evaluation of the patient's medical history and clinical symptoms. These diagnostic methods can take a long time and be quite costly. Thus, the discovery of new biomarkers for the diagnosis of ALS is needed. Some recent studies have shown that post-translational modifications in several proteins may be indicative as biomarkers in the course of neurodegenerative diseases. This study aimed to investigate the post-translational modifications, such as ubiquitination and sumoylation II/III, of the proteins in the central nervous system (CNS) of B6SJL-Tg (SOD1G93A)1Gur /J) transgenic mouse model of ALS, as candidate biomarkers for the diagnosis of ALS. To compare the in vivo findings with the in vitro model, NSC-34 cells were mutated into SOD1G93A after neural differentiation. Proteome profiling via the 2D-PAGE method showed that there was a significant difference between proteins from 105-day-old transgenic mouse brains and wild-type mouse brains. As a result of LC-MS/MS analysis, this protein was determined to be a Rho-related GTP protein (Rhob). Post-translational modifications of proteins obtained from the in vivo and in vitro models were analyzed by the Simple Wes method and were found to have no ubiquitination and sumoylation II/III modifications within the range determined according to the target protein.
Increasing the differentiation potential of adipose derived stem cell into chondrocyte by plant derived exosomes: A Possible therapeutic strategy for cartilage diseases
Stem cells are cells that have potential to differentiate into different cell types depending on the signals and self-renewal. Mesenchymal Stem Cells (MSCs) are multipotent cell that can differentiate with limited capacity. Adipose tissue contains MSCs that can be isolated and used in in vitro studies. Adipose derived stem cells have potential to differentiate into neurons, osteocyte, chondrocyte, myocyte and adipocytes. MSCs are used as a therapeutic agent in order to treat cartilage injuries due to its chondrogenic differentiation potential. Extracellular vesicules has an important role in cell homeostasis. Especially, exosomes responsible from cell to cell communication are nano sized messengers capable of transporting proteins, miRNAs, lipids and mRNAs to other cells. This message can determine the fate of a cell depending of inner biomolecules. Previous studies showed that plant derived exosomes have the same properties like mammalians. Until now, exosomes were isolated from a large number of plants and used for different applications. In this study, for the first time, plant derived exosomes from wheat (Triticum aestivum), beet (Beta vulgaris), radish (Raphanus sativus), tomato (Solanum lycopersicum) and lemon (Citrus limon) will be used to trigger chondrogenic differentiation of hASCs.
The effect of radish (raphanus sativus) derived exosomes on osteoarthritis
Osteoarthritis, characterized by articular cartilage loss, is a well-established cause of impairment. Given the underlying pathology of OA, it is clear that injured cartilage has a low ability for regeneration. This means the proposed action is essential. Deterioration of articular cartilage, alterations in subchondral bone, and synovitis are hallmarks of this complex illness. Current OA treatments focus mostly on alleviating symptoms like pain and inflammation rather than stopping the disease itself from progressing. There is a significant knowledge gap when it comes to the mechanism that contribute to the OA development. Articular cartilage's extracellular matrix homeostasis is kept in check by a delicate balancing act between anabolic and catabolic pathways; abnormalities in this equilibrium are linked to OA. Chondrocytes are the special cells that make up cartilage in joints. Collagens, proteoglycans, and non-collagen proteins are all synthesized by these cells to form the extracellular matrix. Chondrocyte responses to molecular cues vary between areas and stages of illness, suggesting that mechanical stress and biochemical variables play major roles in disease development. In this regard, the utilization of plant-based products in the treatment of medical disorders is gradually becoming a preferred option due to the fewer negative effects associated with these items. In this respect, objective of this study to examine potential effects of Raphanus sativus derived exosomes on osteoarthritis treatment by defining proliferative, anti-inflammation potential and upregulating cartilage specific activities on protein and gene expression levels for osteoarthritis. Our results demonstrated that, Raphanus sativus derived exosomes has anti-inflammation effect on OA while decreasing expression of IL-1β and IL-6 levels and inducing TGF-β. Also, these exosomes has proliferative effect on OA chondryocyte cells while promoting main extracellular matrix components. Based on these findings, Raphanus sativus derived exosomes appear as a potential therapeutic for osteoarthritis treatment.
Development of high efficacy and low toxicity glioblastoma multiforme (GBM) treatment models
Glioblastoma (GBM), en çok karşılaşılan primer beyin tümörlerinden biridir ve ortalama sağ kalım süresinin 15 ay gibi son derece kısa olmasından kaynaklanan agresif fenotipiyle bilinir. Standart GBM tedavisi tümör cerrahisini takip eden radyasyon ve klinikte kullanılan altın standart olan Temozolomid (TMZ) tedavilerini içerir. Ayrıca, GBM genetik fenotipinin biyolojik karmaşıklığı ve TMZ'ye karşı gelişen direnç, etkili tedaviyi sınırlayan ve dolayısıyla terapötik sonuçlara ulaşılmasını engelleyen iki yaygın zorluktur. Çeşitli kanser türlerinde yaşam kalitesini iyileştirme ve sağkalımı uzatmadaki rolünü bildiren son klinik araştırma çalışmalarına dayalı olarak farmakolojik askorbata büyük ilgi gösterildi. Benzer şekilde, bor bileşikleri da kanser önleyici özellikleriyle giderek daha fazla tanınmaktadır. Mevcut çalışmada, IDH-MUT-U87 ve U373MG hücre hatlarının in vitro GBM modellerinde Vitamin-C, sodyum pentaborat pentahidrat (NaB) ve bunların kombinasyonunun anti-kanser etkisi incelenmiştir. Bu bileşikler, apoptoz, otofaji, hücre göçü, hücre döngüsü ve DNA Hasar Tepkisi (DDR) dahil olmak üzere çeşitli hücresel süreçler üzerindeki etkilerini araştırılmıştır. Ayrıca, çalışmanın ikinci bölümünde Olaparib ve Vitamin-C kombinasyonunun hücre toksisitesi ölçümlerinde sinerjistik bir etki gösterdiği belirlendi. Çarpıcı bir şekilde, Vitamin-C ve NaB kombinasyonunun, IDH-MUT-U87 hücre hattında DDR sinyalinin aktivasyonu, otofajinin indüklenmesi ve apoptoz mekanizması yoluyla bir anti-tümorijenik etkiyi desteklediğini de gözlemledik. Son olarak, bu bileşiklerin çeşitli kombinasyonları, U373MG hücre hattında DDR ve ROS'a bağlı sinyalleşme yoluyla bir anti-tümorijenik etkiyi desteklediği gözlenmiştir. Son olarak, bu bileşiklerin çeşitli kombinasyonları, U373MG hücre hattında DDR ve ROS'a bağlı sinyalleşme yoluyla anti kanser etkisi sergilediği belirlenmiştir. Bu çalışmada GBM kanser modellerinde NaB ve Vitamin-C kombinasyonunun anti-kanser etkisi vurgulanmıştır. Bu çalışmanın sonuçları, NaB ve Vitamin-C kombinasyonunun kanseri tedavi etmek için bir ilaç olarak potansiyel kullanımına ilişkin bir tahmin sunabilir
A novel seed coating formulation for the management of seed-borne diseases
Almost 90% of the world's food crops are grown from seeds. Seed quality is affected by abiotic and biotic factors. Abiotic factors include environmental conditions, while biotic factors arise from bacteria, fungi, viruses, insects, and nematodes. Seed-borne microorganisms cause serious yield losses (approx. 27-42%) and affect large agricultural areas by transmitting them to seasons/regions via contaminated seeds. Therefore, several physical, biological, and chemical seed treatment methods have been developed and used to control seed-borne diseases up to now. Indeed, seed treatment can become the most effective way to eliminate or reduce seed-borne pathogens. However, none of the existing methods has effectively controlled seed-borne endophytic and epiphytic pathogens. This study aims to develop a new seed-coating formulation that eliminates all types of plant pathogens (bacterial and fungal pathogens) from seeds without harming germination. In this study, piroctone olamine (PO) based on a novel chemical formulation developed and used for coating wheat, tomato, and corn seeds. Different doses including 0.4, 0.1, 0.08, 0.05, 0.03, and 0.01% of PO-based formulation were tested against CMM, PSS, PST bacteria and two fungal pathogens like Aspergillus niger and Penicillium sp. in vitro conditions. According to antimicrobial activity test results, seed coating formulations containing different concentrations of PO, except 0.01%, were effective against all microorganisms tested. It was also observed that all endophytic and epiphytic bacteria and fungi species were eliminated on/in seeds treated with PO seed coating formulations. It was observed that the application PO seed coating formulations at rates of 0.4% and 0.1% was almost 100% effective in pathogen control. On the other hand, it was determined that 0.1% PO formulation did not have any negative effects on root and shoot development of treated seeds. This is the first study to show that 0.1% PO-based seed coating formulation may have great potential to eradicate seed-borne pathogens of different crops in agriculture. Therefore, further study is required to evaluate the effect of PO based formulations against viral pathogens in different crop seeds/seeds in vitro and on yield and quality parameters.
The effect of extracellular vesicles of rhus Coriaria L. on wound healing
Wounds are injuries, disruptions, or damage to living tissue that occur as a direct result of trauma or due to underlying disease processes such as diabetes, venous/arterial insufficiency, or immunological diseases. Wounds are classified as acute and chronic. Acute wounds are injuries that usually heal within 3-4 weeks. During this time, chronic wounds do not heal. Wound healing is a complex biological and physiological process in which the body uses two different mechanisms to heal, including tissue regeneration and tissue repair. The history of wound healing treatment dates back to ancient times. Medicinal plants have been used in wound treatment among the people in Turkey for many years. Rhus coriaria L. (Anacardiaceae), known as sumac, is traditionally used in the treatment of wounds and burns in the Southeastern Anatolia Region of Turkey, especially in Gaziantep. The wound healing effect of sumac extract has been well documented in previous studies, but sumac extracellular vesicles have not been studied yet. Therefore, the objectives of this study are to develop an optimized protocol for the isolation and identification of extracellular vesicles from sumac plant and then evaluate the wound healing effect of SECVs based on in vitro tests including cell viability and proliferation, cell migration, wound healing assay, tube formation assay, and gene expression analysis. According to the results, it is observed that SECVs enhance cell migration, proliferation, and healing capacity of wound-associated cells. On the other hand, it is found that SECVs do not induce the angiogenesis of the endothelial cells. Furthermore, qPCR results showed that genes related cell migration and anti inflammation were upregulated as a result of SECV treatment. Therefore, all the results indicate that SECVs have a promising potential in terms of providing successful wound healing process. In conclusion, the presented study will be a valuable precursor in searching SECVs and the potential effects on wound healing. With new developing techniques, the mechanisms might understood better and the application of SECVs' effect might reinforced via in vivo studies.
In vitro screening for natural quorum sensing inhibitor molecules
Increasing antimicrobial resistance in pathogenic bacteria has created the need for the development of novel therapeutic agents. Having a role in various mechanisms of gene regulation such as biofilm formation and production of virulence factors; quorum sensing (QS), cell-density dependent bacterial intercellular signaling mechanism, has become a target for developing next generation antimicrobials. Recent studies have shown that disrupting the communication between bacteria reduces their pathogenicity and, therefore, attenuate the possible infections. Both Gram negative and positive bacteria use this mechanism to regulate such physiological processes. Among all of them, Pseudomonas aeruginosa, being an opportunistic human pathogen, attracted the greatest focus on QS system investigations.In this study, we aimed to screen some natural compounds (rose absolute, rose oil, clove oil, cinnamon oil and pine turpentine) for their inhibitory effects on Pseudomonas aeruginosa QS system. Therefore, we initially detected these molecules whether they have QS inhibition activity or not. The results showed that all of these compounds were potential QS inhibitors. To test them further, the molecules were probed with a QS reporter system that allow gathering a dose-response relationship. According to our data; rose absolute was found to be the most effective compound on the inhibition of QS. This is the first study demonstrated that rose absolute has inhibitory effects on QS and it can be potential candidate to be used as a therapeutic agent to fight with bacterial infections in the future.
Effect of F68, F127 and P85 pluronic block copolymers on odontogenic differentiation of human tooth germ stem cells (hTGSCs)
Mesenchymal Stem Cells are non-embryonic adult stem cells, they can differentiate into a variety of tissues, including bone, tooth, cartilage, tendon, muscle and adipose in an optimum conditions. This potential of MSCs can be used in tissue engineering applications such as bone, tooth, skin and cartilage tissue repair. Stem cells isolated from 3rd molar tooth germs as hTGSCs, were recently shown to be pluripotent and these stem cells can be used in tissue engineering as well. Biomaterials are indispensible tools in tissue engineering and among them, pluronics have gained great interest in recent years. Pluronics, also known as ?poloxamers? which consist of hydrophilic poly (ethylene oxide) and hydrophobic poly (propylene oxide) chains are one of the most attractive polymers used as vehicles for therapeutic agents such as drugs, chemicals and growth factors. In this study, we tested the effects of three pluronics F68, F127 and P85 on odontogenic differentiation of mesenchymal stem cells (MSCs) from human tooth germ stem cells (hTGSCs) taken from the 3rd molar tooth. The results showed that none of the pluronics used in this study were toxic for the cells except fornthe P85 at high concentration. Long term treatment of P85 can cause toxicity. F68 remarkably increased the cell viability and the differentiation efficiency of MSCs into odontogenic cell types. P85 showed toxic effect for cells during the differentiation process. It was found that F127 has not significantly altered the cell viability and differentiation of MSCs. These data suggest, for the first time, that F68 has a great potential to increase odontogenic differentiation of MSCs which might lead to the development of new tissue engineering strategies for regenerative medicine.
The role of pluronic F68 and F127 combined growth factors and stem cells in reducing ketamine induced neurotoxicity
Ketamine is an uncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist which is used as human and animal medicine as an injectable anesthetic. Recently it was shown that Ketamine gives rise to neurotoxicity and cell death during surgical operations. Furthermore, long-term Ketamine administration increases the number of hyperphosphorylated tau positive cells, which is a hallmark of Alzheimer?s Disease. In this study, we aimed to decrease the Ketamine induced neurotoxicity on SH-SY5Y neuroblastoma and primary mix brain culture cells applying several growth factors, namely; Fibroblast Growth Factor-2 (FGF-2) Insulin-Like Growth Factor-1 (IGF-1), Brain-Derived Neurotrophic Factor (BDNF), Nerve Growth Factor (NGF), Vascular Endothelial Growth Factor (VEGF) and stem cell conditioned medium in the presence of Pluronic F68 and F127. Moreover the effect of stem cells on this toxicity was investigated using stem cell co-culture system. Pluronics used in this study were reported to act as drug delivery reagents and membrane stabilizers which are important agents in the treatment neurodegenerative diseases. The stem cells used in this study were derived from Human Tooth Germs and not only comprise mesenchymal stem cell properties but also possess the capacity to differentiate into neurogenic cells. Our results demonstrated that the administration of either stem cell conditioned medium or direct stem cells in co-culture system protect cells from the Ketamine induced toxicity. The growth factors tested were also protective on SH-SY5Y and primary mix brain culture cells to some extent. Both stem cells and growth factors administered in combination with the Pluronics F68 and F127 were much more effective in reducing the neurotoxicity. These data suggested that stem cells and growth factors play an important role in ameliorating the adverse effects of Ketamine and the administration of Pluronics might increase the protection and healing of cells from the Ketamine induced stress. Pluronics can also provide an alternate approach for the treatment of other neurodenerative diseases.
Dual role of human tooth germ derived mesenchymal stem cells on cancer cells
Mesencyhmal stem cells (MSCs), important components of the tumor microenvironment, could modulate tumor growth and development in many ways. MSCs have been used as targeted-delivery vehicles for cancer gene therapy because of their potential of tumor tropism. In this study, we investigated the effects of Human Tooth Germ (HTG) derived MSCs on MCF-7 (Human breast cancer cell line) and SH-SY5Y (Human metastatic neuroblastoma cell line) cells treated with doxorubicin and paclitaxel, anticancer drugs. We also investigated the in vitro effect of genetically modified MSCs secreting apoptosis inducers (Tumor necrosis factors related apoptosis inducing ligand (TRAIL), Dickkopf-related protein-1(Dkk-1)) on SH-SY5Y cells.In the first part, conditioned medium (CM) of hTGSCs was collected when the cells reached to 70% confluency. Effects of this CM on MCF-7 and SH-SY5Y treated doxorubicin and paclitaxel were measured by MTS (cell viability assay) assay and real time PCR analysis of apoptotic markers caspase3 and p53.In the second part, TRAIL and Dkk-1 genes tagged with pkh67 (Green Fluorescent Cell Linker Kit) were transferred into MSCs by electroporation either alone or in combination. A GFP gene transfected group was used as a control. Genetically modified MSCs were co-cultured with SH-SY5Y cells for 24 hours. Thereafter the cell survival of SH-SY5Y cells was determined by using flow cytometry. The expression of Bcl-2, Bax, Stat3 (proliferation genes), FADD, caspase3 (apoptotic genes) in SH-SY5Y cells were assessed by real time PCR.The results showed CM of hTGSCs increased the survival of MCF-7 and SH-SY5Y cells treated with doxorubicin and paclitaxel by 30% and also CM reduced doxorubicin and paclitaxel induced apoptosis. On the other side, genetically modified MSCs secreting TRAIL and Dkk-1 reduced the cell survival of cancer cells by inducing apoptosis and inhibiting the proliferation. Our findings demonstrated that MSCs play roles in growth of tumor cells by decreasing the effect of anti-cancer drugs on cancer cells. On the other hand genetically modified HTG derived MSCs expressing Trail and Dkk-1 might be a potential gene and cell therapy approach to treat certain types of cancer.
Effect of melatonin on osteogenic, neurogenic, adipogenic, chondrogenic, myogenic and odontogenic differentiation of human tooth germ stem cells
Stem cell based applications have become a popular and promising approach for therapy of a number of disorders including neurodegenerative disorders, diabetes, degenerative muscle diseases and osteoporosis as well as trauma, inflammations, burns and injuries. Human tooth germ stem cells are an adult stem cell source, which have mesenchymal stem cell properties and show high proliferative and differentiation capacity. Melatonin, a hormone produced mainly by pineal gland in humans, are demonstrated to regulate differentiation of human and mouse mesenchymal stem cells into various cell lineages besides its other functions in the body. In this study, the effects of melatonin on osteogenic, neurogenic, adipogenic, chondrogenic, myogenic and odontogenic differentiation of human tooth germ stem cells have been investigated. The results showed that melatonin increases the viability of cells. It significantly increases osteogenic, neurogenic, chondrogenic, myogenic and odontogenic of the cells whereas it decreases adipogenic differentiation capability. These results suggest that melatonin has a great potential to increase differentiation capacity of human tooth germ stem cells and might be used in regenerative therapy applications involving stem cell differentiations besides defining potential treatments for obesity because of its suppressor effects on adipogenesis.
P2X7 reseptörlerinin beyin felci üzerine etkileri ve rolü
P2X7 receptors are cationic-selective ion channels gated by extracellular ATP and expressed throughout the peripheral and central nervous systems. P2X7 receptors are involved in the regulation of receptor trafficking, inflammation and ATP-mediated cell death. In the present study, we aimed to examine the impacts of P2X7 receptors in the development of neuronal cell death after middle cerebral artery occlusion (MCAo) in mice. In the present studies, we show that the activation of P2X7 receptors with 250 µM BzATP does not increase the cellular damage significantly, as compared with the vehicle-treated control animals after MCAo in which slow and rapid neuronal cell death are observed. However, inhibition of P2X7 receptors by 10 mM Brilliant Blue G (BBG) improved neuronal survival. Furthermore, inhibition of P2X7 receptors decreased infarct volume, brain swelling and neurological scores 90 min after MCAo and 24 hours reperfusion. In addition, inhibition of P2X7 receptors decreased DNA fragmentation and increased neuronal survival after 30 min of MCAo which was associated with increased phosphorylation of survival kinases. Here, we provide evidence for a potential role of P2X7 receptors in mediation of neuronal cell death. We predict that clinical implementation of P2X7 receptor antagonists can be beneficial after neurodegenerative disorders.
The differential effect of USP7 on breast cancer cell lines
THE DIFFERENTIAL EFFECT OF USP7 ON BREAST CANCER CELL LINES Breast cancer has the highest mortality rate among women compared to other various types of cancer. As a multi-stage and complicated cancer type, breast cancer might be initiated by various stimuli. Most of the breast cancer patients have been diagnosed with a familial predisposition due to the poor genetical heritage. Investigation of mutated proteins and affected pathways are bare essential to develop an effective cure for targeted therapy. In the current study, potential anti-cancer activity of USP7 gene supression has been evaluated by in vitro cell proliferation analysis, the examination of alterations in gene and protein expression profile, three dimensional sphere formation capacity and colony forming unit assays. The small molecule inhibitor of USP7 enzyme in addition to small interfering RNA (siRNA) and lentiviral short hairpin vector (shRNA) has been used as targeting strategies against USP7. MCF7 and T47D human breast cancer cell lines, which maintain different P53 gene status, have been used to determine whether the anti-cancer effect of USP7 enzyme is p53 dependent or in-dependent. Overall, a significant anticancer activity of USP7 inhibition has been detected in vitro for the breast cancer cell lines. Many molecular, physiological and morphological changes have been observed upon USP7 inactivation in all of the three different experimental strategy indicating the potential anti-cancer activity of USP7 blockage. USP7 inactivation inhibited the cancer cell proliferation, invasion and metastasis in MCF7 and T47D cell lines. However, further investigation should also be conducted with the purpose of comprehending the exact molecular mechanism which underlies in the total effect of USP7 on tumorigenic progression. This is the first study in literature which offers a critical examination of the differential effects of targeting USP7 enzyme by both at protein and gene levels in MCF7 and T47D cell lines.
Role of ananas comosus derived exosomes in scarless cutaneous wound repair
Wound healing is a complex and dynamic process involving a great number of cellular interactions based on growth factors, cytokine, and chemokine release. Although many new therapeutic agents are being introduced to the market for their remarkable wound healing capabilities, the problem of why some organisms heal perfectly while others heal with a pathological connective tissue known as a scar remains a mystery. Scar formation is a natural process that occurs after the healing process due to dysfunctionalities in the remodeling phase that may occur either in the epithelial epidermis or the dermis. Reducing scar formation is lately gaining interest as a new area of scientific research. Following the discovery of scarless healing of the fetal skin, an explosive number of studies have highlighted striking differences between fetal and adult human skin in terms of the differences in their extracellular matrix, growth factor, and cytokine characteristics. Despite of the accumulating evidence to mitigate the scar formation, gain or loss of function of no single pathway or cell type has been shown to be responsible for scar formation therefore mimicking the scarless healing remains as a challenging issue. Nonetheless, findings from recent studies have shed light to certain aspects of scarless healing. In this respect, the use of plant-based products in the treatment medical conditions is becoming a matter of preference due to their lower side effects. Taking a parallel approach, this study aimed to evaluate the potential of scar reducing effects of Ananas comosus derived exosomes by determining their proliferative, migratory, and angiogenetic activities on the gene and protein expression levels of key factors of wound healing. Our results indicate that these exosomes activate ERK/MAPK and PI3K/AKT pathways and induce cellular proliferation and migration while increasing the ratio of transforming growth factor-β3 (TGF-β3) to TGF-β1, and certain MMPs, which is an indication for suppression of scar formation. Additionally, it has been shown that these exosomes also promote extracellular matrix reconstruction. Based on these results A. comosus exosomes emerge as a promising agent that facilitate cutaneous wound healing suppressing formation of scars.
The effect of exosomes isolated from human foreskin stem cells on wound healing
Wound healing is one of the important processes in the body. The wounds are separated into two groups when considering the healing time and process: acute wounds and chronic wounds. Side effects of natural and chemical wound healing compounds to the human body are well known. Due to these side effects, attempts to create new drugs are of interest. Therefore, the use of stem cells has augmented. However, both difficulties in collection and risks such as rejection and teratoma in the recipient body have limited the use of stem cells. The extracellular vesicles secreted by stem cells have recently become prominent agents. With the use of these agents, the potential content of the stem cells can be transferred to the recipient cells without an immune response, and since it is easier to obtain than the stem cells, it has been used as a new generation wound healing agent. In this study, the wound healing effect of exosomes derived from foreskin stem cells was investigated on both keratinocyte and endothelial cells in vitro. Cell migration, new vessel formation, inflammation response, quantitative polymerase chain reaction and western blot experiments were performed to investigate the potential positive contribution to gene and protein levels and the underlying mechanisms. Overall, cell proliferation, cell invasion and cell migration were accelerated in exosome treated groups compared to the control group and new vessel formation was inducted in exosome treated groups. Increase in gene levels of growth factors related to proliferation and inflammation, and change in related signaling pathway proteins in exosome treated groups compared to the control group were shown. Possible molecular mechanisms underlying cell movements were associated with mitogen-activated protein kinase (MAPK) pathway. We found that foreskin stem cell exosomes can heal the wound in a short period of time so that it could be a promising wound healing agent. The current study was the first study in the literature representing the wound healing activity of foreskin stem cell-derived exosomes.
An alternative and therapeutic approach for hepatocellular carcinoma
Hepatocellular carcinoma is among of the most frequently forms of liver cancer, with a high death rate, emphasizing the need for improved treatment techniques. Current research has focused on the anti-cancer effects of natural compounds. The most common therapy for HCC varies according to stage: patients in the early stages may require surgical resection or liver transplantation, whereas intermediate-stage patients may get Transarterial chemoembolization (TACE). In advanced stages, immunotherapy combined with targeted therapies, especially Atezolizumab-Bevacizumab, has emerged as the standard. CBD (cannabidiol), a non-psychoactive cannabinoid, has gained attention in cancer research because of its anti-inflammatory, anti-proliferative, anti-angiogenic, and apoptosis-inducing properties. Studies suggest CBD may inhibit tumor growth, induce cell death, and reduce metastasis. Additionally, CBD is being explored as a complementary treatment for alleviating chemotherapy side effects. Vitamin D is also recognized for its potential function in HCC treatment, given its impact on immune regulation and cell growth control. Vitamin D insufficiency has been associated to HCC progress, emphasizing its potential therapeutic value. The current research tested the anti-cancer impacts of single and combination treatments of Vitamin D and CBD in HEP3B and HUH7 cell lines, focusing on apoptosis, autophagy, cell cycle regulation, and DDR. The combination treatment of Vitamin D and CBD demonstrated a synergistic effect, significantly reducing cell survival in both HEP3B and HUH7 cell lines. Notably, the combination induced apoptosis and autophagy in HEP3B cells and inhibited the cell cycle in HUH7 cells, suggesting anti-tumorigenic and anti-cancer effects. These findings indicate the possibility of Vitamin D and CBD as a novel treatment approach against HCC, requiring additional in vivo studies and clinical trials to assess efficacy, safety, and mechanisms in more complex biological systems.
Comparison of two-dimensional and three-dimensional triple negative breast cancer cell culture models on treatment response
Triple negative breast cancer (TNBC) has a dismal prognosis and few available treatments, making it a major clinical challenge. This study assesses the effectiveness of cannabidiol (CBD), which has pro-apoptotic and anti-proliferative qualities, and olaparib (OLAP), a PARP inhibitor that targets DNA repair, as single and combination therapies in TNBC and estrogen receptor positive (ER+) breast cancer cell lines. In two-dimensional (2D) and three- dimensional (3D) culture systems, analyses were conducted on MDA-MB-231 (TNBC), HCC-70 (TNBC), and MCF-7 (ER+) cell lines. The results demonstrated that in 3D culture models, increased efficacy was attained with lower doses, and that OLAP and CBD caused synergistic cytotoxic effects, particularly in TNBC cells. In MDA-MB-231 cells with G2/M phase arrest, increased apoptosis was seen, as demonstrated by flow cytometry and caspase 3/7 tests. DNA damage response and cell cycle-related genes (ATM, ATR, BRCA1/2, RAD51, and CDK1/2/4/6) were down-regulated by the combined treatment, according to gene expression analysis. Specifically, 3D spheroid models under metabolic and hypoxic stress demonstrated heightened drug sensitivity. These results imply that 3D culture methods provide more physiologic and sensitive models for evaluating medication efficacy, and that OLAP and CBD together offer a possible therapeutic approach for aggressive breast cancer subtypes.
Anti-cancer effect of postbiotics of novel probiotic strains isolated from fermented food
Kolorektal kanser, dünya genelinde en yaygın ve ölümcül kanser türlerinden biri olarak kabul edilmektedir. Son yıllarda, bağırsak mikrobiyotası ve bu mikroorganizmaların metabolitlerinin hem kanserin gelişiminde hem de önlenmesinde oynadığı rol üzerine artan bir ilgi gözlemlenmektedir. Bu tez çalışması iki ana aşamada gerçekleştirilmiştir. İlk aşamada, çeşitli probiyotik suşları izole edilerek ayrıntılı bir şekilde karakterize edilmiştir. Bu karakterizasyon sürecinde morfolojik ve biyokimyasal özellikler, enzim aktiviteleri (örneğin, proteaz, ve lipaz) ve antioksidan kapasiteleri değerlendirilmiştir. Özellikle Lactobacillus ve Bacillus türlerine odaklanılmış olup, bu türlerin gastrointestinal sistemde sağlık üzerinde olumlu etkiler gösterdiği bilinmektedir. İkinci aşamada ise bu probiyotik suşlardan elde edilen postbiyotiklerin HCT116 ve HT29 kolon kanseri hücreleri üzerindeki biyolojik etkileri değerlendirilmiştir. Postbiyotikler, canlı olmayan bakteri metabolitleri ve hücre bileşenleri olarak tanımlanmakta ve potansiyel terapötik etkileri ile dikkat çekmektedir. Hücre kültürü tabanlı deneylerde, çeşitli hücresel süreçler incelenmiştir. MTS testi ile hücre canlılığı ve metabolik aktivite değerlendirilmiş, akış sitometrisi ile hücre döngüsü ve apoptoz analizleri yapılmıştır. Ayrıca, apoptoz (Caspase 3, 7, 8, BAX), hücre döngüsü düzenlenmesi (CDK1, CDK2, CDK4, CDK6) ile ilişkili genlerin ekspresyon düzeyleri RT-PCR ile analiz edilmiştir. Bu araştırma, seçilen probiyotik suşların ürettiği postbiyotiklerin kanser hücrelerinin çoğalmasını baskılayabildiğini, hücre ölümünü indükleyebildiğini ve hücre döngüsüne müdahale edebildiğini göstermektedir. Elde edilen veriler, probiyotik kaynaklı postbiyotiklerin kolorektal kanser hücrelerinde kritik hücresel ve moleküler süreçleri modüle edebileceğini ortaya koyarak, yeni önleyici ve tedavi edici stratejilere kapı aralamaktadır. Bu kapsamda, bağırsak sağlığını destekleyici ve kanser riskini azaltıcı fonksiyonel gıda ürünleri veya biyoterapötikler geliştirilmesi mümkün görünmektedir. Bu çalışmada, laboratuvar ortamında elde edilen sonuçların klinik uygulamalara dönüştürülmesinde karşılaşılabilecek zorluklara da dikkat çekilmektedir. Bu etkilerin fizyolojik koşullar altında doğrulanması için ileri düzey in vivo çalışmalar ve klinik araştırmalar gereklidir.
The role of plant-derived extracellular vesicles on the in vitro Alzheimer's disease model
Alzheimer's disease is a progressive and irreversible neurodegenerative disorder, in which the accumulation of amyloid beta (Aβ) peptides and the hyperphosphorylation of tau proteins play a pivotal role in its pathogenesis. Current treatment methods mostly aim to alleviate symptoms, but are insufficient in stopping disease progression or targeting the underlying mechanisms. Extracellular vesicles (EVs) are nanovesicles that provide intercellular communication, have a lipid bilayer membrane structure, and can carry biomolecules such as proteins, lipids, and RNA. Lately, EVs have gained significant interest as therapeutic tools due to their effects such as regulating gene expression, suppressing inflammation, and triggering cellular repair processes in disease models. Plant-derived EVs (PDEVs) stand out as important biotherapeutic candidates due to their natural origin, low immunogenicity, economical production, and cross-species biological activity. In addition, their anti- inflammatory and antioxidant properties make EVs attractive candidates for targeting neurodegenerative pathways. This study aims to investigate the effects of extracellular vesicles isolated from tobacco (Nicotiana tabacum) (TEV) suspension culture and celery (Apium graveolens) extract (CEV) in an in vitro AD model. In the study, Aβ accumulation and hyperphosphorylation of tau proteins were induced in differentiated SH-SY5Y cells and an AD-like model was created by treating neurons with TEV, while astrocytes were treated with CEV in a gliotic model created by applying Aβ peptide. Molecular and cellular analyses performed after the application showed that TEV application reduced neuronal degeneration, while CEV application suppressed Aβ-induced astrogliosis responses. In particular, a significant decrease was observed in Aβ and phospho-tau accumulations and protein aggregate load. The findings suggest that PDEVs may affect AD pathophysiology and have neuroprotective potential on both neuronal and glial targets. This study is the first to explore the therapeutic potential of PDEVs in AD, laying an important foundation for future research in this field.
Discovery of functional characteristics of plant cell suspension-derived extracellular vesicles on the in vitro cerebrovascular inflammation
Extracellular vesicles (EVs) are essential biomolecule carriers responsible for intercellular communication. They play vital roles in many physiological events. Thanks to their bilayer lipid membrane, biomolecules can be protected from the harsh extracellular environment. One of living organisms in which EVs consistently function is plants. Plant-derived EVs (PDEVs) have a role in physiological events and defense against pathogens. Besides, these nanovesicles showed therapeutic potential in antioxidant, anti-inflammatory, and immunomodulatory activities. On the other hand, cerebrovascular inflammation (CVI) is a chronic inflammatory disorder characterized by cerebrovascular dysfunction, proinflammatory phenotype, and vascular hyperpermeability. Based on their therapeutic activities, PDEVs can become a promising tool for cerebrovascular disorders. Our study aims to investigate the functionality of plant cell suspension extracellular vesicles (PCSEVs) and their therapeutic effects on the in vitro CVI model. To that end, these nanovesicles were purified from the media of Stevia rebaudiana, Vaccaria hispanica and characterized. The CVI pathology was modelled and the effects of PCSEVs were investigated. In the results, first, the PCSEVs were successfully purified and characterized in the sense of physical and molecular properties. Besides, the CVI pathology was successfully modelled in the cells and it exhibited critical features of its pathologies. Lastly, it was observed that PCSEVs led to a protective response in the CVI model, and these effects mainly occurred by modulating adhesion molecule expression via the NRF2 pathway. In addition, further analysis revealed that these effects of PCSEVs were mediated through the phosphorylation of key signaling pathways. However, further studies are needed to transform these in vitro results into clinical applications. Future studies should include in vivo experiments in animal models to confirm the efficacy and safety of the PCSEVs. By demonstrating the therapeutic effects of PCSEVs on cerebrovascular inflammation, our study paves the way for innovative, plant-derived nanomedicine solutions for vascular diseases.