<p>This thesis aimed at studying mechanisms involved in control of tissue fluid homeostasis during inflammation.</p><p>The interstitial fluid pressure (P_IF) is of importance for control of tissue fluid balance. A lowering of P_IF <i>in vivo</i> will result in a transport of fluid from the circulation into the tissue, leading to edema. Loose connective tissues that surround blood vessels have an intrinsic ability to take up fluid and swell. The connective tissue cells exert a tension on the fibrous network of the tissues, thereby preventing the tissues from swelling. Under normal homeostasis, the interactions between the cells and the fibrous network are mediated by β1 integrins. Connective tissue cells are in this way actively controlling P_IF.</p><p>Here we show a previously unrecognized function for the integrin αVβ3, namely in the control of P_IF. During inflammation the β1 integrin function is disturbed and the connective tissue cells release their tension on the fibrous network resulting in a lowering of P_IF. Such a lowering can be restored by platelet-derived growth factor (PDGF) -BB. We demonstrated that PDGF-BB restored P_IF through a mechanism that was dependent on integrin αVβ3. This was shown by the inability of PDGF-BB to restore a lowered P_IF in the presence of anti-integrin β3 IgG or a peptide inhibitor of integrin αVβ3. PDGF-BB was in addition unable to normalize a lowered P_IF in β3 null mice. Furthermore, we demonstrated that extracellular proteins from <i>Streptococcus equi</i> modulated αVβ3-mediated collagen gel contraction. Because of the established concordance between collagen gel contraction <i>in vitro</i> and control of P_IF <i>in vivo</i>, a potential role for these proteins in control of tissue fluid homeostasis during inflammation could be assumed. Sepsis and septic shock are severe, and sometimes lethal, conditions. Knowledge of how bacterial components influence P_IF and the mechanisms for tissue fluid control during inflammatory reactions is likely to be of clinical importance in treating sepsis and septic shock.</p>

This thesis aimed at studying mechanisms involved in control of tissue fluid homeostasis during inflammation. The interstitial fluid pressure (PIF) is of importance for control of tissue fluid balance. A lowering of PIF in vivo will result in a transport of fluid from the circulation into the tissue, leading to edema. Loose connective tissues that surround blood vessels have an intrinsic ability to take up fluid and swell. The connective tissue cells exert a tension on the fibrous network of the tissues, thereby preventing the tissues from swelling. Under normal homeostasis, the interactions between the cells and the fibrous network are mediated by β1 integrins. Connective tissue cells are in this way actively controlling PIF. Here we show a previously unrecognized function for the integrin αVβ3, namely in the control of PIF. During inflammation the β1 integrin function is disturbed and the connective tissue cells release their tension on the fibrous network resulting in a lowering of PIF. Such a lowering can be restored by platelet-derived growth factor (PDGF) -BB. We demonstrated that PDGF-BB restored PIF through a mechanism that was dependent on integrin αVβ3. This was shown by the inability of PDGF-BB to restore a lowered PIF in the presence of anti-integrin β3 IgG or a peptide inhibitor of integrin αVβ3. PDGF-BB was in addition unable to normalize a lowered PIF in β3 null mice. Furthermore, we demonstrated that extracellular proteins from Streptococcus equi modulated αVβ3-mediated collagen gel contraction. Because of the established concordance between collagen gel contraction in vitro and control of PIF in vivo, a potential role for these proteins in control of tissue fluid homeostasis during inflammation could be assumed. Sepsis and septic shock are severe, and sometimes lethal, conditions. Knowledge of how bacterial components influence PIF and the mechanisms for tissue fluid control during inflammatory reactions is likely to be of clinical importance in treating sepsis and septic shock.

All blood vessels in the microvasculature are embedded in loose connective tissue, which regulates the transport of fluid to and from tissues. The intersti-tial fluid pressure (IFP) is one of the forces that control this transport. A lowering of IFP in vivo results in an increased transport of fluid from the circulation into the underhydrated connective tissues, resulting in edema formation. During homeostasis, contractile connective tissue cells exert a tension on the connective tissue fibrous network by binding with β1 in-tegrins, thereby actively controlling IFP. During inflammation, the IFP is lowered but platelet-derived growth factor (PDGF)-BB induces an IFP nor-malization dependent on integrin αVβ3. We demonstrate that extracellular proteins from Streptococcus equi subspecies equi modulated cell-mediated and integrin αVβ3-directed collagen gel contraction in vitro. One of these proteins, the collagen- and fibronectin binding FNE, stimulated contraction by a process dependent on fibronectin synthesis. This study identified a pos-sible novel virulence mechanism for bacteria based on the ability of bacteria to modulate the edema response. Another protein, the collagen-binding pro-tein CNE, inhibited contraction and this led to the identification of sites in collagen monomers that potentially are involved in connecting αVβ3 to the collagen network. PDGF-BB and prostaglandin E1 (PGE1) stimulate and inhibit collagen gel contraction in vitro and normalize and lower IFP, respec-tively. We showed that these agents affected both similar and different sets of actin-binding proteins. PDGF-BB stimulated actin cytoskeleton dynamics whereas PGE1 inhibited processes dependent on cytoskeletal motor and adhesive functions, suggesting that these different activities may partly ex-plain the contrasting effects of PGE1 and PDGF-BB on contraction and IFP. Mutation of the phosphatidylinositol 3’-kinase (PI3K), but not phospholipase C (PLC)γ activation site, rendered cells unable to respond to PDGF-BB in contraction and in activation of the actin binding and severing protein cofilin. Ability to activate cofilin after PDGF-BB stimulation correlated with ability to respond to PDGF-BB in contraction, suggesting a role for cofilin in this process downstream of PDGF receptor-activated PI3K. Many proteins can modulate contraction either by affecting the extracellular matrix and cell adhesions or by altering cytoskeletal dynamics. Knowledge on how these proteins might influence IFP is likely to be of clinical importance for treat-ment of inflammatory conditions including anaphylaxis, septic shock and also carcinoma growth.

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This thesis emerged from a series of journeys / ‘percorsi’, conducted between 2007 and 2009 along the course of the aqueducts and railway lines in the south-east of Rome. During this time, my focus shifted from figure to background, from the archaeological fragments to the territorial and urban contexts encountered along their passage. This zone offers a unique cross sectional experience of the city, spanning for approximately seven kilometers - from the Aqueduct Park (Parco degli Acquedotti) at the outer edges of the city to the Aurelian Walls at Porta Maggiore – exposing a variety of conditions ranging from the openness of Roman Campagna to the dense agglomeration of Rome’s urban periphery. This is a place of contrasts, of industrial and archaeological, in a state of abandonment and in a constant state of flux. It is as difficult to grasp conceptually as it is to traverse physically, lacking territorial continuity, legibility and ease of access. It is physically fragmented and separated from the rest of the city by infrastructural arteries and property boundaries. This thesis proposes reversing the marginal aspect of this area by re-establishing its relationship with its urban context. It suggests a reading of this landscape as ‘interstitial tissue’ a connective element rather than an inert zone of separation. Using the biological connotation of the interstitial (“the fine connective tissue lying between the cells of other tissue”) - the thesis focuses on the relational potential of this in-between zone, its ability to connect rather than separate places and neighbourhoods along it. It proposes identifying spatial continuities as well as establishing temporal connections between the past, present, and future of this area. The thesis further proposes a design intervention along a site situated between the aqueducts and via del Mandrione, a historical road running in parallel to the railway lines. Although this site is currently enclosed and separated from its context by property boundaries, the thesis proposes reconfiguring this site as a threshold, a point of intensity between the city and its margins.

A brief introduction to cancer biology and treatment is presented with a focus on current clinical advances in the delivery of chemotherapy and antiangiogenic therapies. Mathematical oncology is then surveyed with summaries of various models of tumor growth, tumor angiogenesis and other relevant biological entities such as angiogenic growth factors. Both strictly time-dependent ordinary differential equation (ODE)-based and spatial partial differential equation (PDE)-based models are considered. These biological models are first developed into an ODE model where various treatment options can be compared including different combinations of drugs and dosage schedules. This model gives way to a PDE model that includes the spatially heterogeneous blood vessel distribution found in tumors, as well as angiogenic growth factor imbalances. This model is similarly analyzed and implications are summarized. Finally, including the effects of interstitial fluid pressure into an angiogenic activity model is performed. This model displays the importance of factor convection on the angiogenic behaviour of tumours.

Fibroblasts continuously strive to organize and compact the surrounding extracellular matrix (ECM). Recent data suggest that this cellular contractility controls interstitial fluid homeostasis in loose connective tissues (CT). The aim of this thesis was to study the effects of prostaglandins on fibroblast contractility and to investigate whether fibroblasts in the interstitial CT surrounding the human endolymphatic duct (ED) can modulate inner ear fluid pressure and endolymph resorption. Paper I shows that prostaglandin E1 (PGE1) and prostacyclin inhibit fibroblast-mediated collagen matrix compaction in vitro and lower the interstitial fluid pressure in vivo in rat dermis. Paper II demonstrates that the inhibition of collagen matrix compaction by PGE1 is protein kinase A-dependent. Furthermore, PGE1 induces a complete but reversible actin depolymerization in human dermal fibroblasts by affecting the phosphorylation state of regulatory actin-binding proteins. Paper III describes that the cells of the interstitial CT encompassing the human ED are organized in a network based on intercellular- and cell-ECM contacts. Paper IV shows that two distinct cell phenotypes populate this interstitial CT: one expressing the lymph endothelial marker podoplanin and the other a fibroblast marker. Furthermore, CT cells isolated from human ED tissues exhibited the same tissue compacting properties in vitro as dermal fibroblasts. In conclusion, PGE1 inhibits fibroblast contractility by interfering with the stability and dynamics of the actin cytoskeleton, which leads to a loss of integrin-mediated adhesion to the ECM. These mechanisms are supposedly involved in edema formation in skin during inflammation and might be involved in the formation of endolymphatic hydrops in the inner ear of patients with Ménière’s disease.

This thesis emerged from a series of journeys / ‘percorsi’, conducted between 2007 and 2009 along the course of the aqueducts and railway lines in the south-east of Rome. During this time, my focus shifted from figure to background, from the archaeological fragments to the territorial and urban contexts encountered along their passage. This zone offers a unique cross sectional experience of the city, spanning for approximately seven kilometers - from the Aqueduct Park (Parco degli Acquedotti) at the outer edges of the city to the Aurelian Walls at Porta Maggiore – exposing a variety of conditions ranging from the openness of Roman Campagna to the dense agglomeration of Rome’s urban periphery. This is a place of contrasts, of industrial and archaeological, in a state of abandonment and in a constant state of flux. It is as difficult to grasp conceptually as it is to traverse physically, lacking territorial continuity, legibility and ease of access. It is physically fragmented and separated from the rest of the city by infrastructural arteries and property boundaries. This thesis proposes reversing the marginal aspect of this area by re-establishing its relationship with its urban context. It suggests a reading of this landscape as ‘interstitial tissue’ a connective element rather than an inert zone of separation. Using the biological connotation of the interstitial (“the fine connective tissue lying between the cells of other tissue”) - the thesis focuses on the relational potential of this in-between zone, its ability to connect rather than separate places and neighbourhoods along it. It proposes identifying spatial continuities as well as establishing temporal connections between the past, present, and future of this area. The thesis further proposes a design intervention along a site situated between the aqueducts and via del Mandrione, a historical road running in parallel to the railway lines. Although this site is currently enclosed and separated from its context by property boundaries, the thesis proposes reconfiguring this site as a threshold, a point of intensity between the city and its margins.

A brief introduction to cancer biology and treatment is presented with a focus on current clinical advances in the delivery of chemotherapy and antiangiogenic therapies. Mathematical oncology is then surveyed with summaries of various models of tumor growth, tumor angiogenesis and other relevant biological entities such as angiogenic growth factors. Both strictly time-dependent ordinary differential equation (ODE)-based and spatial partial differential equation (PDE)-based models are considered. These biological models are first developed into an ODE model where various treatment options can be compared including different combinations of drugs and dosage schedules. This model gives way to a PDE model that includes the spatially heterogeneous blood vessel distribution found in tumors, as well as angiogenic growth factor imbalances. This model is similarly analyzed and implications are summarized. Finally, including the effects of interstitial fluid pressure into an angiogenic activity model is performed. This model displays the importance of factor convection on the angiogenic behaviour of tumours.