Epidermal growth factor receptor

The epidermal growth factor receptor (EGFR; ErbB-1; HER1 in humans) is a transmembrane protein that is a receptor for members of the epidermal growth factor family (EGF family) of extracellular protein ligands. The epidermal growth factor receptor is a member of the ErbB family of receptors, a subfamily of four closely related receptor tyrosine kinases: EGFR (ErbB-1), HER2/neu (ErbB-2), Her 3 (ErbB-3) and Her 4 (ErbB-4). In many cancer types, mutations affecting EGFR expression or activity could result in cancer. Epidermal growth factor and its receptor was discovered by Stanley Cohen of Vanderbilt University. Cohen shared the 1986 Nobel Prize in Medicine with Rita Levi-Montalcini for their discovery of growth factors. Deficient signaling of the EGFR and other receptor tyrosine kinases in humans is associated with diseases such as Alzheimer's, while over-expression is associated with the development of a wide variety of tumors. Interruption of EGFR signalling, either by block

In vitro lymphatic endothelial morphogenesis

Carolyn Yong Pullin

Cell organization into functional multicellular three-dimensional structures is a long-standing challenge. In particular, engineering of vascularized tissues requires both blood and lymphatic neovascular network formation in a simultaneous and coordinated fashion. While extensive in vitro investigations in blood and lymphangiogenesis have identified a vast array of cellular phenomena and key cellular and extracellular bioactive substances, the dynamic natural cellular environment and its regulatory role in governing cellular response and interactions remain largely unclear. Furthermore, the wealth of existing knowledge describing the molecular regulation of vessel morphogenesis is heavily biased towards the angiogenic growth of blood capillaries, despite the importance of the lymphatic system. This thesis first addresses the differential gene regulation of isolated microvascular lymphatic endothelial cells (LECs) in response to the lymphatic growth factor VEGF-C and to fluid shear stress using gene arrays and QRT-PCR. First, characteristic of a response to a growth factor stimulus, the largest numbers of differentially expressed genes in response to VEGF-C stimulation were transcription factors and cell cycle related. A number of genes known to be important in angiogenesis, tumorigenesis and tumor invasion, and transport of proteins, solutes, and lipids were also affected. Interestingly, a number of genes related to lipid metabolism as well as neurogenesis were also responsive to VEGF-C stimulation. Further analysis of these genes may not only provide insight into the molecular mechanisms underlying lymphangiogenesis and associated pathogenesis, but may also identify other important roles of VEGF-C. The ability of lymphatic endothelium to sense and actively regulate drainage function is poorly understood, and shear stress is likely a key indicator of lymphatic drainage function. Thus, LECs were exposed to 0, 2 and 20 dyn/cm2 shear stress as representative of chronic lymphedema, normal, and acute inflammatory conditions, respectively. Important adaptive responses to both low and high shear stress conditions that were correlated to lymphatic function, including changes in gene expression patterns related to water and solute transport, cell-cell and cell-matrix adhesion, inflammatory cytokines and cell cycle were found. These changes were consistent with increased transport function, both active and passive, in high shear conditions and indicate that during lymphedema, lymphatic endothelium shuts down transport functions. These data demonstrate a functional-adaptive response of lymphatic endothelium to flow conditions, thus indicating that the lymphatic endothelium plays an active role in regulating their drainage function. It has been previously shown in our lab that in vitro blood and lymphatic morphogenesis are differentially enhanced by interstitial flow. Using a three-dimensional tissue culture model, the molecular mechanisms orchestrating endothelial cell morphogenesis, specifically gene regulation of the lymphatic microvascular system was investigated, focusing on genes implicated in both cellular morphogenesis and lymphatic development. Finally, concurrent angiogenesis and lymphangiogenesis were studied in vitro to examine the influences and crosstalk between blood and lymphatic microvascular development. The data suggest that both blood endothelial cell (BEC) and LEC in vitro capillary morphogenesis are enhanced by the presence of BEC-secreted factors, and these effects are, in part, mediated through proliferative and migratory responses of which the latter was induced by VEGFR-2 and VEGFR-3 signaling for BECs and LECs, respectively. When maintained in mixed three-dimensional cultures, both homotypic and heterotypic interactions between LECs and BECs were observed. Furthermore, capillary morphogenesis and interactions were enhanced and distinct when under the additional influence of interstitial flow. In conclusion, the novelties of the work presented in this dissertation include studies of 1) overall LEC gene response to its primary growth factor VEGF-C and biophysical factor, fluid flow and 2) the genetic regulation underlying the morphogenetic processes of LECs under interstitial flow in vitro, by using unique approaches which combine bioengineering principles to address fundamental biological questions and thereby contribute to this novel niche which has so far been neglected. Furthermore, the importance of biophysical processes along with crucial biochemical signal regulation is demonstrated in the concurrent induction of angiogenesis and lymphangiogenesis. Such are the requisites for the understanding of microvascular development and the determination of useful design principles for the in vitro vascularization of engineered tissues.

EPFL2011

From the early steps of whisker formation to its evolutionary disappearance: the (curious) case of Prdm1 and its regulation

Pierluigi Giuseppe Manti

Whiskers (also known as vibrissae) are sensory organs that are thought to have first appeared in Therapsida, a group of synapsids that includes mammals and evolutionary ancestors like the Thrinaxodon(1). They are highly conserved among mammals to the notable exception of the higher primates and are important for behavior, especially for nocturnal animals and the ones living in burrows(2). Furthermore, the mechanoreceptors of the vibrissae signal to the primary somatosensory cortex, where a discrete and well-defined structure in layer 4 represents each whisker(3). This structure, known as barrel cortex, occupies a large portion of the rodent brain. Strikingly, Prdm1 conditional KO (cKO) mice are one of the rare transgenic strains that do not develop whiskers while forming a pelage; on the other hand, the other head structures expressing Prdm1 (including pelage hair follicles and teeth) grow normally(4). In the developing whisker pad, Prdm1 expression is confined to the mesenchyme underlying the embryonic epidermis that will later form the whisker epidermal placode; it lasts until dermal papilla formation, when a âswitchâ occurs to the precursors of the inner root sheath. The Prdm1 ablation study we conducted revealed the absence of the patterned up-regulation of Bmp2 (marker of placode formation), Shh (a protein secreted by the epidermal placode responsible for the first epidermal signal) and Bmp4 (expressed in the pre-follicle mesenchyme). On the other hand, Lef1 is expressed uniformly in the mesenchyme of the whisker pad, thus pointing out the integrity of the Î²Catenin-mediated first mesenchymal signal. Those results suggest the role of Prdm1 as a master gene in whisker development. Prdm1 expressing cells are mainly non-proliferative; however, a small subpopulation at the periphery is cycling. Being Sox2 coexpressed with Prdm1 in the whisker mesenchyme, we performed lineage tracing with Sox2CreERT2 and ROSAYFP transgenic strains. Those experiments revealed that Sox2/Prdm1 expressing cells contribute to the formation of the dermal papilla, dermal sheath and putative pericytes of the whisker follicle and thus constitute a population of embryonic progenitors crucial for whisker formation. We subsequently investigated the development of the barrel cortex in our cKO mice. Intriguingly, a disorganized pattern was observed in cKO mice, suggesting that the loss of expression of a single gene, and consequently the disappearance of vibrissae, drastically impacts the development of the whole somatosensory system. Finally, we hypothesized that the loss of Prdm1 expression might explain the disappearance of whisker follicles in the human during evolution. Prdm1 being no longer expressed in the human lineage during hair follicle development(5), we reasoned that the loss of regulatory elements of its upstream genes might contribute to this phenomenon; we demonstrated that Lef1 acts upstream of Prdm1 during whisker development and are currently testing the role of its potential enhancers (Leaf) during this process. Bibliography: 1: (http://en.wikipedia.org/wiki/therapsid) 2: Petersen, C. C. H. The Functional Organization of the Barrel Cortex. Neuron 56, 339â355 (2007) 3: Woolsey, T. A. & Van der Loos, H. The structural organization of layer IV in the somatosensory region (SI) of mouse cerebral cortex. The description of a cortical field composed of discrete cytoarchitectonic units. Brain Res. 17, 205â242 (1970) 4: Robertson, E. J. et al. Blimp1 regulates development of the posterior forelimb, caudal pharyngeal arches, heart and sensory vibrissae in mice. Development 134, 4335â4345 (2007). 5: Sellheyer, K. & Krahl, D. Blimp-1: a marker of terminal differentiation but not of sebocytic progenitor cells. J. Cutan. Pathol. 37, 362â370 (2010).

EPFL2016

Photoresponsive Nanocarriers Based on Lithium Niobate Nanoparticles for Harmonic Imaging and On-Demand Release of Anticancer Chemotherapeutics

Luigi Bonacina, Dario Diviani, Sandrine Gerber, Adrian Stefan Gheata, Simon Kaiser, Jérémy Vuilleumier

Nanoparticle-based drug delivery systems have the potential for increasing the efficiency of chemotherapeutics by enhancing the drug accumulation at specific target sites, therebyreducing adverse side effects and mitigating patient acquired resistance. In particular, photo-responsive nanomaterials have attracted much interest due to their ability to release molecularcargos on demand upon light irradiation. In some settings, they can also provide complementary information by optical imaging on the (sub)cellular scale. We herein present a system based on lithium niobate harmonic nanoparticles (LNO HNPs) for the decoupled multi-harmonic cell imaging and near-infrared light-triggered delivery of an erlotinib derivative (ELA) for the treatment of epidermal growth factor receptor (EGFR)-overexpressing carcinomas. The ELA cargo was covalently conjugated to the surface of silica-coated LNO HNPs through a coumarinyl photo-cleavable linker, achieving a surface loading of the active molecule of 27 nmol/mg NPs. The resulting nanoconjugates (LNO-CM-ELA NPs) were successfully imaged upon pulsed laser excitation at 1250 nm in EGFR-overexpressing human prostate cancer cells DU145 by detecting the second harmonic emission at 625 nm, in the tissue transparency window. Tuning the laser at 790 nm resulted in the uncaging of the ELA cargo as a result of the second harmonic emission of the inorganic HNP core at 395 nm. This protocol induced a significant growth inhibition in DU145 cells, which was only observed upon specific irradiation at 790 nm, highlighting the promising capabilities of LNO-CM-ELA NPs for theranostic applications.

2022