Silk fibroin based multifunctional materials

Silk fibroin is a commonly available natural biopolymer produced in specialized glands of arthropods, such as silkworms or spiders, scorpions, mites, bees and flies. This biopolymer has a long history of use in textile production and also as sutures or treatment of skin wounds. Silk fibroin has been increasingly explored in other areas of biomedical science where we can find a higher morphological diversification of silk biomaterials like films, electrospun fibers, 3D porous scaffolds or nanoparticles. In recent years it has been demonstrated that fibroin is an excellent material for active components in optical devices. This new application opens the way towards the development of multifunctional optoelectronic devices, which in perspective can be made fully biocompatible and eventually bioresorbable. Moreover, fibroin can be added to other biocomponents in order to modify the biomaterial properties leading to optimized and total different functions. These improvements can go from higher cell adhesion in tissue engineering or enhanced optical transparency, smoothness or flexibility in optoelectronic devices. The tuning and completely understanding of silk fibers physicochemical properties and interaction with other elements are of crucial importance for the improvement of already existent silk-based materials and the basis for the development of new products.

Multifunctional Ti–Me (Me=Al, Cu) thin film systems for biomedical sensing devices

Ti-Me binary intermetallic thin films based on a titanium matrix doped with increasing amounts of Me (Me = Al, Cu) were prepared by magnetron sputtering (under similar conditions), aiming their application in biomedical sensing devices. The differences observed on the composition and on the micro(structural) features of the films, attributed to changes in the discharge characteristics, were correlated with the electrical properties of the intermetallic systems (Ti-Al and Ti-Cu). For the same Me exposed areas placed on the Ti target (ranging from 0.25 cm2 to 20 cm2) the Cu content increased from 3.5 at.% to 71.7 at.% in the Ti-Cu system and the Al content, in Ti-Al films, ranged from 11 to 45 at.%. The structural characterization evidenced the formation of metastable Ti-Me intermetallic phases for Al/Ti atomic ratios above 0.20 and for Cu/Ti ratios above 0.25. For lower Me concentrations, the effect of the α-Ti(Me) structure domains the overall structure. With the increase amount of the Me into Ti structure a clear trend for amorphization was observed. For both systems it was observed a significant decrease of the electrical resistivity with increasing Me/Ti atomic ratios (higher than 0.5 for Al/Ti atomic ratio and higher than 1.3 for Cu/Ti atomic ratio). Although similar trends were observed in the resistivity evolution for both systems, the Ti-Cu films presented lower resistivity values in comparison to Ti-Al system.

Semipermeable capsules wrapping a multifunctional and self-regulated co-culture microenvironment for osteogenic differentiation

A new concept of semipermeable reservoirs containing co-cultures of cells and supporting microparticles is presented, inspired by the multi-phenotypic cellular environment of bone. Based on the deconstruction of the â stem cell nicheâ , the developed capsules are designed to drive a self-regulated osteogenesis. PLLA microparticles functionalized with collagen I, and a co-culture of adipose stem (ASCs) and endothelial (ECs) cells are immobilized in spherical liquified capsules. The capsules are coated with multilayers of poly(L-lysine), alginate, and chitosan nano-assembled through layer-by-layer. Capsules encapsulating ASCs alone or in a co-culture with ECs are cultured in endothelial medium with or without osteogenic differentiation factors. Results show that osteogenesis is enhanced by the co-encapsulation, which occurs even in the absence of differentiation factors. These findings are supported by an increased ALP activity and matrix mineralization, osteopontin detection, and the up regulation of BMP-2, RUNX2 and BSP. The liquified co-capsules also act as a VEGF and BMP-2 cytokines release system. The proposed liquified capsules might be a valuable injectable self-regulated system for bone regeneration employing highly translational cell sources.

HOXA9 as a key regulator in glioma initiation, aggressiveness and response to therapy

Tese de Doutoramento em Ciências da Saúde

Development of multifunctional coatings deposited on polymers based sensors for biomedical applications

Tese de Doutoramento (Programa Doutoral em Engenharia de Materiais)

Antibacterial properties of multifunctional coatings Ag-ZrCN for biomedical devices: a novel approach

PhD in Sciences Specialty in Physics

apoptosis-regulator c-FLIP : functional role in urothelial carcinoma and autoimmunity and identification of novel CD95 DISC-interacting proteins

Magdeburg, Univ., Fak. für Naturwiss., Diss., 2013

TRAIP is a regulator of the spindle assembly checkpoint.

Accurate chromosome segregation during mitosis is temporally and spatially coordinated by fidelity-monitoring checkpoint systems. Deficiencies in these checkpoint systems can lead to chromosome segregation errors and aneuploidy, and promote tumorigenesis. Here, we report that the TRAF-interacting protein (TRAIP), a ubiquitously expressed nucleolar E3 ubiquitin ligase important for cellular proliferation, is localized close to mitotic chromosomes. Its knockdown in HeLa cells by RNA interference (RNAi) decreased the time of early mitosis progression from nuclear envelope breakdown (NEB) to anaphase onset and increased the percentages of chromosome alignment defects in metaphase and lagging chromosomes in anaphase compared with those of control cells. The decrease in progression time was corrected by the expression of wild-type but not a ubiquitin-ligase-deficient form of TRAIP. TRAIP-depleted cells bypassed taxol-induced mitotic arrest and displayed significantly reduced kinetochore levels of MAD2 (also known as MAD2L1) but not of other spindle checkpoint proteins in the presence of nocodazole. These results imply that TRAIP regulates the spindle assembly checkpoint, MAD2 abundance at kinetochores and the accurate cellular distribution of chromosomes. The TRAIP ubiquitin ligase activity is functionally required for the spindle assembly checkpoint control.

Influence of short time exposure to an insect growth regulator, hexaflumuron, on mortality and adult emergence of vector mosquitoes

Hexaflumuron, an insect growth regulator (IGR), was found to greatly affect the development of immatures and emergence of adults of three species of vector mosquitoes, Culex quinquefasciatus, Aedes aegypti and Anopheles stephensi, when larvae were subjected to short time exposure of < or = 1h. This IGR could completely prevent adult emergence even at a minimum exposure time of 10 min at 0.001, 0.01 and 0.1 mg/l. On treatment, larval and pupal mortality as well as varying degrees of morphogenetic abnormalities were induced in immatures and adults of the three species. Four weeks of control achieved in a slow moving sullage canal breeding Culex quinquefasciatus indicates that this IGR can be of use in such breeding habitats.

Mini-review: Specificity and expression of CIITA, the master regulator of MHC class II genes.

The class II transactivator (CIITA) has been referred to as the "master control factor" for the expression of MHC class II (MHCII) genes. As our knowledge on the specificity and function of CIITA grows, it is becoming increasingly evident that this sobriquet is entirely justified. First, despite extensive investigations, the major target genes of CIITA remain those implicated in the presentation of antigenic peptides by MHCII molecules. Although other putative target genes have been reported, the contribution of CIITA to their expression remains indirect, controversial or comparatively minor relative to its decisive role as a regulator of MHCII and related genes. Second, the most important parameter dictating MHCII expression is by far the expression pattern of the gene encoding CIITA (MHC2TA). The vast majority of signals that activate or repress MHCII expression under physiological and pathological situations converge on one or more of the three alternative promoters that drive transcription of the MHC2TA gene. In short, with respect to its specificity and its exquisitely controlled pattern of expression, CIITA is by a long stretch the single most important transcription factor for the regulation of genes required for MHCII-restricted antigen-presentation.

EGFR signalling as a negative regulator of Notch1 gene transcription and function in proliferating keratinocytes and cancer.

The Notch1 gene has an important role in mammalian cell-fate decision and tumorigenesis. Upstream control mechanisms for transcription of this gene are still poorly understood. In a chemical genetics screen for small molecule activators of Notch signalling, we identified epidermal growth factor receptor (EGFR) as a key negative regulator of Notch1 gene expression in primary human keratinocytes, intact epidermis and skin squamous cell carcinomas (SCCs). The underlying mechanism for negative control of the Notch1 gene in human cells, as well as in a mouse model of EGFR-dependent skin carcinogenesis, involves transcriptional suppression of p53 by the EGFR effector c-Jun. Suppression of Notch signalling in cancer cells counteracts the differentiation-inducing effects of EGFR inhibitors while, at the same time, synergizing with these compounds in induction of apoptosis. Thus, our data reveal a key role of EGFR signalling in the negative regulation of Notch1 gene transcription, of potential relevance for combinatory approaches for cancer therapy.

Glut9 is a major regulator of urate homeostasis and its genetic inactivation induces hyperuricosuria and urate nephropathy.

Elevated plasma urate levels are associated with metabolic, cardiovascular, and renal diseases. Urate may also form crystals, which can be deposited in joints causing gout and in kidney tubules inducing nephrolithiasis. In mice, plasma urate levels are controlled by hepatic breakdown, as well as, by incompletely understood renal processes of reabsorption and secretion. Here, we investigated the role of the recently identified urate transporter, Glut9, in the physiological control of urate homeostasis using mice with systemic or liver-specific inactivation of the Glut9 gene. We show that Glut9 is expressed in the basolateral membrane of hepatocytes and in both apical and basolateral membranes of the distal nephron. Mice with systemic knockout of Glut9 display moderate hyperuricemia, massive hyperuricosuria, and an early-onset nephropathy, characterized by obstructive lithiasis, tubulointerstitial inflammation, and progressive inflammatory fibrosis of the cortex, as well as, mild renal insufficiency. In contrast, liver-specific inactivation of the Glut9 gene in adult mice leads to severe hyperuricemia and hyperuricosuria, in the absence of urate nephropathy or any structural abnormality of the kidney. Together, our data show that Glut9 plays a major role in urate homeostasis by its dual role in urate handling in the kidney and uptake in the liver.

Transcriptional control of the hydrogen cyanide biosynthetic genes hcnABC by the anaerobic regulator ANR and the quorum-sensing regulators LasR and RhlR in Pseudomonas aeruginosa.

Virulence factors of Pseudomonas aeruginosa include hydrogen cyanide (HCN). This secondary metabolite is maximally produced at low oxygen tension and high cell densities during the transition from exponential to stationary growth phase. The hcnABC genes encoding HCN synthase were identified on a genomic fragment complementing an HCN-deficient mutant of P. aeruginosa PAO1. The hcnA promoter was found to be controlled by the FNR-like anaerobic regulator ANR and by the quorum-sensing regulators LasR and RhlR. Primer extension analysis revealed two transcription starts, T1 and T2, separated by 29 bp. Their function was confirmed by transcriptional lacZ fusions. The promoter sequence displayed an FNR/ANR box at -42.5 bp upstream of T2 and a lux box centered around -42.5 bp upstream of T1. Expression of the hcn genes was completely abolished when this lux box was deleted or inactivated by two point mutations in conserved nucleotides. The lux box was recognized by both LasR [activated by N-(oxododecanoyl)-homoserine lactone] and RhlR (activated by N-butanoyl-homoserine lactone), as shown by expression experiments performed in quorum-sensing-defective P. aeruginosa mutants and in the N-acyl-homoserine lactone-negative heterologous host P. fluorescens CHA0. A second, less conserved lux box lying 160 bp upstream of T1 seems to account for enhanced quorum-sensing-dependent expression. Without LasR and RhlR, ANR could not activate the hcn promoter. Together, these data indicate that expression of the hcn promoter from T1 can occur under quorum-sensing control alone. Enhanced expression from T2 appears to rely on a synergistic action between LasR, RhlR, and ANR.