| 1. |
- Ashrafzadeh, Parham
(författare)
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Exploring Cellular Dynamics : From Vesicle Tethering to Cell Migration
- 2016
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Cells in the body communicate with each other in order to cooperate efficiently. This communication is in part achieved by regulated secretion of signaling molecules, which when released from a cell may activate receptors present at the plasma membrane of an adjacent cell. Such signals affect both cell fate and behavior. Dysregulated signaling may lead to disease, including cancer. This thesis is focused on how exocytosis and subsequent activation and trafficking of receptors can be regulated, and what the consequences of this regulation may be for cell migration.Actin filaments are important transport structures for secretory vesicle trafficking. In Paper 1, actin polymerization was shown to induce formation of ordered lipid domains in the plasma membrane. Accordingly, actin filaments may thus create and stabilize specific membrane domains that enable docking of vesicles containing secretory cargo.The RhoGEF FGD5 regulates Cdc42 which can result in cytoskeletal rearrangements. In Paper II, FGD5 was shown to be selectively expressed in blood vessels and required for normal VEGFR2 signaling. FGD5 protected VEGFR2 from proteasome-mediated degradation and was essential for endothelial cells to efficiently respond to chemotactic gradients of VEGFA.The exocyst component EXOC7 is essential for tethering secretory vesicles to the plasma membrane prior to SNARE-mediated fusion. In Paper III, EXOC7 was required for trafficking of VEGFR2-containing vesicles to the inner plasma membrane and VEGFR2 presentation at the cell surface.The ability of tumor cells to escape the primary tumor and establish metastasis is in part dependent on their capacity to migrate. In Paper IV, a method based on time-lapse microscopy and fluorescent dyes was created to analyze single cancer cell migration in mixed cancer cell cultures, and in particular the influence of different types on neighboring cells was assessed.In conclusion, these studies have enhanced our understanding of the mechanisms behind cellular trafficking, and may be applied in the future to develop more specific therapeutics to treat cancer and other diseases associated with abnormal angiogenesis and cellular migration.
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| 2. |
- Dieterich, Lothar
(författare)
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Molecular Regulation of Inflammation and Angiogenesis in the Tumor Microenvironment
- 2011
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Tumor growth and progression not only depend on properties of the malignant cells but are strongly influenced by the tumor microenvironment. The tumor stroma consists of various cell types such as inflammatory cells, endothelial cells and fibroblasts, which can either inhibit or promote tumor growth. Consequently, therapeutic targeting of the tumor stroma is increasingly recognized as an important tool to fight cancer. Two particularly important processes that contribute to the pathology of most types of tumors are angiogenesis and inflammation. In order to target these processes specifically and efficiently, it is fundamental to identify and understand the factors and signaling pathways involved. This thesis initially describes the multiple functions of the small heat shock protein αB-crystallin in the tumor microenvironment. αB-crystallin was first identified in a screen of proteins specifically up-regulated in endothelial cells forming vessel-like structures. We found that αB-crystallin is expressed in a subset of tumor vessels and promotes angiogenesis by inhibiting endothelial apoptosis, suggesting that targeting of αB-crystallin might inhibit angiogenesis and thereby decrease tumor growth. However, we also discovered an important role of αB-crystallin in regulation of inflammatory processes. We show that αB-crystallin increases the surface levels of E-selectin, an important leukocyte-endothelial adhesion molecule. Thereby, αB-crystallin may alter leukocyte recruitment to inflamed tissues such as the tumor stroma. In addition, we found that αB-crystallin is expressed in immature myeloid cells that accumulate in the periphery and at the tumor site during tumor development. Importantly, lack of αB-crystallin resulted in increased accumulation of immature myeloid cells, which might increase tumor associated inflammation. Finally, through combining laser microdissection of vessels from human tissue and microarray analysis, we identified a gene expression signature specifically associated with vessels in high grade glioma. Blood vessels in malignant glioma are highly abnormal and contribute to the pathology of the disease. Thus, knowledge about the molecular set-up of these vessels might contribute to the development of future vascular normalizing treatments.
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| 3. |
- Ekman, Maria, 1979-
(författare)
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The role of Smad7 and TRAF6 in Prostate Cancer Cell Invasion, Migration and Survival
- 2011
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Transforming growth factor (TGF) β is a tumor suppressor during early tumor development, by inhibiting proliferation and inducing apoptosis. At later stages of cancer, it becomes a tumor promoter, and promotes tumor cell migration and invasion. TGFβ signals via its type II and type I receptors to several downstream signaling pathways. In the present work we have focused on the TRAF6 (tumor necrosis factor receptor-associated factor 6)/ TAK1 (TGFβ activated kinase 1) signaling pathway and the Smad7-dependent activation of p38 in prostate carcinoma cells (PC3U). We found that TGFβ-induced activation of the ubiquitin ligase TRAF6 was needed for cell invasion, by a mechanism that involves activation of the metalloproteinase TNFα converting enzyme (TACE), via protein kinase Cζ (PKCζ). TACE cleaves the TβRI, whereafter the intracellular domain (ICD) translocates to the nucleus, where it binds to the transcriptional co-activator p300 and regulates gene expression, promoting invasion. Interestingly, the translocation of the TβRI ICD was observed in several cancer cell lines and in sections of primary tumors, but not in primary prostate epithelial cells. We also found that Smad7 and adenomatous polyposis coli (APC) are important for TGFβ- and epidermal growth factor (EGF)-induced cell migration in PC3U cells. TGFβ induces the formation of a complex consisting of Smad7, p38, glycogene synthase kinase 3β (GSK-3β), APC and β-catenin, which localizes to the membrane ruffles in the leading edge of migrating cells. The complex links the TβRI to the microtubule system and promotes membrane ruffling and microtubule polarization, which are known to be important for cell migration. In the EGF signaling pathway, Smad7 was found to be important for phosphorylation of the EGF receptor at Tyr1068, for the activation of p38 and JNK, and for induction of membrane ruffles. Smad7 is required for TGFβ-induced activation of p38 and apoptosis. We found that Smad7 forms a complex with p38 and ataxia telangiectasia mutated (ATM), which is important for activation of p53 mediated apoptosis. Many tumor cells including the PC3U cells lack a functional p53, which is one of the reasons to why cancer cells can avoid the tumor suppressor effects of TGFβ.
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| 4. |
- Fuxe, Jonas
(författare)
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p16INK4A and p15INK4B in senescence, immortalization and cancer : gene transfer by adenovirus vectors
- 2001
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The astrocytic gliomas are the most common form of brain tumors in humans. The malignant progression from low grade (astrocytoma, grade II) to high-grade (glioblastoma multiforme, grade, IV) astrocytomas is well documented and is accompanied by an increasing number of genetic aberrations. Many of these genetic alterations center round genes that regulate the G 1/S transition of the cell cycle. This is particularly the case with the genes encoding the cyclin dependent kinase inhibitors (CDKIs), p16 INK4A (p16) and p15 INK4B (p15). They are homozygously deleted in 40-50% of primary gliomas and in an even higher percentage in glioma cell lines. In addition, the p16 gene is frequently inactivated by point mutations and by DNA methylation, and mice that are deficient for the p16 gene spontaneously develop tumors within their first year of life. While the role of p16 in senescence and tumor suppression has been amply documented, the role of p 15 remained less clear. Recombinant adenoviruses (Ad) expressing p15 (Adp15) or p16 (Adp16) were constructed and used to infect human glioma cells with different status of the p15, p16, and pRB genes. The results indicated that p15 is as potent as p16 in inducing cell cycle arrest and senescence-like changes in human glioma cells with an intact pRB. In addition, both p 15 and p 16 strongly inhibited telomerase activity in these cells. This shows the potential of p15 to function as a tumor suppressor and mediator of senescence. Together with the fact that p15 accumulates in T lymphocytes as they approach senescence and that p15 deficient mice display tumors of the hematopoietic cell system, these results suggest that p15 plays a role in regulating homeostasis in these (and perhaps other) cell systems. The mechanism behind the synthesis of an alternative isoform of p 15 was also elucidated. This protein, termed p15.5, was found to be an N-terminally extended variant of p15, initiated from an upstream GUG codon. It was shown to separately, or in combination with p15, interact with both CDK4 and CDK6, and to induce cell cycle arrest and a senescent-like phenotype when transfected into human glioma cells. The involvement of the p16/pRB pathway components in telomerase-mediated immortalization of bovine capillary endothelial (BCE) cells was determined. In these studies, transfection of the catalytic subunit of human telomerase (hTERT) alone was sufficient to immortalize BCE cells. Surprisingly, the telomere lengths in the hTERT-BCE cells were consistently shorter than in the senescent parental cells. pRB was hyperphosphorylated, and the expression of p16 (and p21CIP1) was repressed by promoter methylation. Reactivation of p16 by either Ad-mediated expression or by demethylation reversed the immortalized phenotype and induced senescence-like changes. These results suggest that the immortalization of BCE cells by hTERT is mediated through inactivation of crucial cellular senescence machineries including p161pR13 and p21CIP1, proteins that also by others have been implicated in inducing and maintaining a senescent phenotype in mammalian cells. The ability of type C Ad to infect human cells is dependent on the expression of the coxsackie- and adenovirus receptor, hCAR. Since very little is known about the expression of hCAR in primary human gliomas and since hCAR has been implicated to function as a tumor suppressor, such expression studies were performed. A great variation in hCAR expression was detected, both in glioma cell lines and in the primary tumors, and the expression correlated well with Ad infectability. A significant decrease in the mean CAR expression levels was detected in the grade IV tumors as compared to the values for the grade II and grade III tumors. Moreover, a mean 12-fold higher expression of hCAR was detected in xenografts derived from glioblastomas, compared to the parental tumors. Interestingly, the two xenografts that did not show any upregulation of hCAR expression grew much faster than the hCAR-expressing cells. This points to an inverse correlation between growth rate and hCAR expression, and suggests that hCAR may contribute to suppression of tumor growth.
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| 5. |
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| 6. |
- Georgoudaki, Anna-Maria, et al.
(författare)
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Reprogramming Tumor-Associated Macrophages by Antibody Targeting Inhibits Cancer Progression and Metastasis
- 2016
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Ingår i: Cell Reports. - : Elsevier BV. - 2211-1247. ; 15:9, s. 2000-2011
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Tidskriftsartikel (refereegranskat)abstract
- Tumors are composed of multiple cell types besides the tumor cells themselves, including innate immune cells such as macrophages. Tumor-associated macrophages (TAMs) are a heterogeneous population of myeloid cells present in the tumor microenvironment (TME). Here, they contribute to immunosuppression, enabling the establishment and persistence of solid tumors as well as metastatic dissemination. We have found that the pattern recognition scavenger receptor MARCO defines a subtype of suppressive TAMs and is linked to clinical outcome. An anti-MARCO monoclonal antibody was developed, which induces anti-tumor activity in breast and colon carcinoma, as well as in melanoma models through reprogramming-TAM-populations to a pro-inflammatory phenotype and increasing tumor immunogenicity. This anti-tumor activity is dependent on the inhibitory Fc-receptor, Fc gamma RIIB, and also enhances the efficacy of checkpoint therapy. These results demonstrate that immunotherapies using antibodies designed to modify myeloid cells of the TME represent a promising mode of cancer treatment.
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| 7. |
- He, Fei, et al.
(författare)
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Sex dimorphism in the tumor microenvironment : From bench to bedside and back
- 2022
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Ingår i: Seminars in Cancer Biology. - : Elsevier. - 1044-579X .- 1096-3650. ; 86:3, s. 166-179
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Forskningsöversikt (refereegranskat)abstract
- Cancer represents a significant cause of death and suffering in both the developed and developing countries. Key underlying issues in the mortality of cancer are delayed diagnosis and resistance to treatments. However, improvements in biomarkers represent one important step that can be taken for alleviating the suffering caused by malignancy. Precision-based medicine is promising for revolutionizing diagnostic and treatment strategies for cancer patients worldwide. Contemporary methods, including various omics and systems biology approaches, as well as advanced digital imaging and artificial intelligence, allow more accurate assessment of tumor characteristics at the patient level. As a result, treatment strategies can be specifically tailored and adapted for individual and/or groups of patients that carry certain tumor characteristics. This includes immunotherapy, which is based on characterization of the immunosuppressive tumor microenvironment (TME) and, more specifically, the presence and activity of immune cell subsets. Unfortunately, while it is increasingly clear that gender strongly affects immune regulation and response, there is a knowledge gap concerning differences in sex-specific immune responses and how these contribute to the immunosuppressive TME and the response to immunotherapy. In fact, sex dimorphism is poorly understood in cancer progression and is typically ignored in current clinical practice. In this review, we aim to survey the available literature and highlight the existing knowledge gap in order to encourage further studies that would contribute to understanding both gender-biased immunosuppression in the TME and the driver of tumor progression towards invasive and metastatic disease. The review highlights the need to include sex optimized/genderized medicine as a new concept in future medicine cancer diagnostics and treatments.
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| 8. |
- Heldin, Johan
(författare)
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Identification and Characterization of Proteins and MicroRNAs that Modulate Receptor Signaling, Vesicular Trafficking and Cell Migration in Vascular Cells
- 2014
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Blood vessels deliver nutrients and oxygen to tissues. Importantly, the functions and growth of blood vessels are commonly altered in disease. The inside of all blood vessels are lined with endothelium, a thin specialized layer of endothelial cells that separate the blood from other tissues. This thesis deals with the identification and functional characterization of proteins and microRNAs that have key roles as modulators of growth factor signaling and directed cell migration of endothelial cells and other vascular cells.A previously uncharacterized protein of the exocyst complex, Exocyst complex component 3-like 2 (ExoC3L2) was identified and shown to be highly expressed in endothelial cells of sprouting vessels. Suppression of ExoC3L2 resulted in reduced VEGF-A signaling together with reduced chemotaxis in response to VEGF-A gradients. VEGF-A-signaling via its receptor VEGFR-2 is thus modulated by the exocyst complex and ExoC3L2.Expression profiling of highly vascularized tissues were used to identify several microRNAs selectively expressed in blood vessels. miR-145, targeting the transcription factor Fli1, was shown to be expressed in pericytes and mural cells. Elevated levels of miR-145 reduced chemotaxis of both endothelial cells and fibroblasts in response to growth factor gradients. miR-145 depletion in fibroblasts was shown to inhibit chemotaxis in response to PDGF-BB.The guanine nucleotide exchange factor FGD5 was shown to be selectively expressed in endothelial cells and to regulate Cdc42 activity. FGD5 was shown to regulate the turnover of activated VEGF-receptors. Suppression of FGD5 impaired endothelial cell chemotaxis, suggesting that FGD5 is required for efficient and sustained VEGF-A signaling.Inactivation of RhoD, a regulator of endosomal trafficking, resulted in an increased pool of acetylated and stable microtubules. Knockdown of RhoD in human fibroblasts resulted in a loss of cell polarity. A link between PDGFR-β and RhoD was implicated by the finding that PDGF-BB was shown to trigger formation of GTP-bound RhoD. Chemotaxis towards PDGF-BB was severely inhibited in cells with reduced RhoD expression, suggesting a role for RhoD in chemotaxis via its regulation of microtubule dynamics.
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| 9. |
- Johnson, Jill R., et al.
(författare)
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Pericytes contribute to airway remodeling in a mouse model of chronic allergic asthma
- 2015
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Ingår i: American Journal of Physiology: Lung Cellular and Molecular Physiology. - : American Physiological Society. - 1522-1504 .- 1040-0605. ; 308:7, s. 658-671
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Tidskriftsartikel (refereegranskat)abstract
- Myofibroblast accumulation, subepithelial fibrosis, and vascular remodeling are complicating features of chronic asthma, but the mechanisms are not clear. Platelet-derived growth factors (PDGFs) regulate the fate and function of various mesenchymal cells and have been implicated as mediators of lung fibrosis. However, it is not known whether PDGF-BB signaling via PDGFR beta, which is critical for the recruitment of pericytes to blood vessels, plays a role in airway remodeling in chronic asthma. In the present study, we used a selective PDGFR beta inhibitor (CP-673451) to investigate the role of PDGFR beta signaling in the development of airway remodeling and lung dysfunction in an established mouse model of house dust mite-induced chronic allergic asthma. Unexpectedly, we found that pharmacological inhibition of PDGFR beta signaling in the context of chronic aeroallergen exposure led to exacerbated lung dysfunction and airway smooth muscle thickening. Further studies revealed that the inflammatory response to aeroallergen challenge in mice was associated with decreased PDGF-BB expression and the loss of pericytes from the airway microvasculature. In parallel, cells positive for pericyte markers accumulated in the subepithelial region of chronically inflamed airways. This process was exacerbated in animals treated with CP-673451. The results indicate that perturbed PDGF-BB/PDGFR beta signaling and pericyte accumulation in the airway wall may contribute to airway remodeling in chronic allergic asthma.
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| 10. |
- Nilchian, Azadeh, et al.
(författare)
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CXADR-Mediated Formation of an AKT Inhibitory Signalosome at Tight Junctions Controls Epithelial-Mesenchymal Plasticity in Breast Cancer.
- 2019
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Ingår i: Cancer Research. - : American Association for Cancer Research. - 0008-5472 .- 1538-7445. ; 79:1, s. 47-60
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Tidskriftsartikel (refereegranskat)abstract
- Tight junctions (TJ) act as hubs for intracellular signaling pathways controlling epithelial cell fate and function. Deregulation of TJ is a hallmark of epithelial-mesenchymal transition (EMT), which contributes to carcinoma progression and metastasis. However, the signaling mechanisms linking TJ to the induction of EMT are not understood. Here we identify a TJ-based signalosome, which controls AKT signaling and EMT in breast cancer. The coxsackie- and adenovirus receptor (CXADR), a TJ protein with an essential yet uncharacterized role in organogenesis and tissue homeostasis, was identified as a key component of the signalosome. CXADR regulated the stability and function of the phosphatases and AKT inhibitors PTEN and PHLPP2. Loss of CXADR led to hyper-activation of AKT and sensitized cells to TGF-β1-induced EMT. Conversely, restoration of CXADR stabilized PHLPP2 and PTEN, inhibited AKT, and promoted epithelial differentiation. Loss of CXADR in luminal A breast cancer correlated with loss of PHLPP2 and PTEN and poor prognosis. These results show that CXADR promotes the formation of an AKT-inhibitory signalosome at TJ and regulates epithelial-mesenchymal plasticity in breast cancer cells. Moreover, loss of CXADR might be used as a prognostic marker in luminal breast cancer.
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