Stromal vascular fraction cell sheets angiogenic potential for tissue engineering and regenerative medicine applications

One of the biggest concerns in the Tissue Engineering field is the correct vascularization of engineered constructs. Strategies involving the use of endothelial cells are promising but adequate cell sourcing and neo-vessels stability are enduring challenges. In this work, we propose the hypoxic pre-conditioning of the stromal vascular fraction (SVF) of human adipose tissue to obtain highly angiogenic cell sheets (CS). For that, SVF was isolated after enzymatic dissociation of adipose tissue and cultured until CS formation in normoxic (pO2=21%) and hypoxic (pO2=5%) conditions for 5 and 8 days, in basal medium. Immunocytochemistry against CD31 and CD146 revealed the presence of highly branched capillary-like structures, which were far more complex for hypoxia. ELISA quantification showed increased VEGF and TIMP-1 secretion in hypoxia for 8 days of culture. In a Matrigel assay, the formation of capillary-like structures by endothelial cells was more prominent when cultured in conditioned medium recovered from the cultures in hypoxia. The same conditioned medium increased the migration of adipose stromal cells in a scratch assay, when compared with the medium from normoxia. Histological analysis after implantation of 8 days normoxic- and hypoxic-conditioned SVF CS in a hindlimb ischemia murine model showed improved formation of neo-blood vessels. Furthermore, Laser Doppler results demonstrated that the blood perfusion of the injured limb after 30 days was enhanced for the hypoxic CS group. Overall, these results suggest that SVF CS created under hypoxia can be used as functional vascularization units for tissue engineering and regenerative medicine.

Adoção e difusão de aplicações das tecnologias da informação: iniciativas lançadas no âmbito de processos de implantação de aplicações em organizações – estudo de práticas profissionais e proposta de ferramenta de suporte

Tese de Doutoramento Tecnologias e Sistemas de Informação

Enteroscopia por cápsula e doença de Crohn: Caminhos para a otimização

Tese de Doutoramento em Medicina

Engineering modular viral scaffolds for targeted bacterial population editing

Bacteria are central to human health and disease, but existing tools to edit microbial consortia are limited. For example, broad-spectrum antibiotics are unable to precisely manipulate bacterial communities. Bacteriophages can provide highly specific targeting of bacteria, but assembling well-defined phage cocktails solely with natural phages can be a time-, labor- and cost-intensive process. Here, we present a synthetic biology strategy to modulate phage host ranges by engineering phage genomes in Saccharomyces cerevisiae. We used this technology to redirect Escherichia coli phage scaffolds to target pathogenic Yersinia and Klebsiella bacteria, and conversely, Klebsiella phage scaffolds to target E. coli by modular swapping of phage tail components. The synthetic phages achieved efficient killing of their new target bacteria and were used to selectively remove bacteria from multi-species bacterial communities with cocktails based on common viral scaffolds. We envision this approach accelerating phage biology studies and enabling new technologies for bacterial population editing.

Seismic retrofit of masonry-to-timber connections in historical constructions

Tese de Doutoramento em Engenharia Civil.

Light-distortion analysis as a possible indicator of visual quality after refractive lens exchange with diffractive multifocal intraocular lenses

METHODS: Refractive lens exchange was performed with implantation of an AT Lisa 839M (trifocal) or 909MP (bifocal toric) IOL, the latter if corneal astigmatism was more than 0.75 diopter (D). The postoperative visual and refractive outcomes were evaluated. A prototype light-distortion analyzer was used to quantify the postoperative light-distortion indices. A control group of eyes in which a Tecnis ZCB00 1-piece monofocal IOL was implanted had the same examinations. RESULTS: A trifocal or bifocal toric IOL was implanted in 66 eyes. The control IOL was implanted in 18 eyes. All 3 groups obtained a significant improvement in uncorrected distance visual acuity (UDVA) (P < .001) and corrected distance visual acuity (CDVA) (P Z .001). The mean uncorrected near visual acuity (UNVA) was 0.123 logMAR with the trifocal IOL and 0.130 logMAR with the bifocal toric IOL. The residual refractive cylinder was less than 1.00 D in 86.7% of cases with the toric IOL. The mean light-distortion index was significantly higher in the multifocal IOL groups than in the monofocal group (P < .001), although no correlation was found between the light-distortion index and CDVA. CONCLUSIONS: The multifocal IOLs provided excellent UDVA and functional UNVA despite increased light-distortion indices. The light-distortion analyzer reliably quantified a subjective component of vision distinct from visual acuity; it may become a useful adjunct in the evaluation of visual quality obtained with multifocal IOLs.

Biomimetic strategies to engineer mineralised human tissues

In the last few years, many reports have been describing promising biocompatible and biodegradable materials that can mimic in a certain extent the multidimensional hierarchical structure of bone, while are also capable of releasing bioactive agents or drugs in a controlled manner. Despite these great advances, new developments in the design and fabrication technologies are required to address the need to engineer suitable biomimetic materials in order tune cells functions, i.e. enhance cell-biomaterial interactions, and promote cell adhesion, proliferation, and differentiation ability. Scaffolds, hydrogels, fibres and composite materials are the most commonly used as biomimetics for bone tissue engineering. Dynamic systems such as bioreactors have also been attracting great deal of attention as it allows developing a wide range of novel in vitro strategies for the homogeneous coating of scaffolds and prosthesis with ceramics, and production of biomimetic constructs, prior its implantation in the body. Herein, it is overviewed the biomimetic strategies for bone tissue engineering, recent developments and future trends. Conventional and more recent processing methodologies are also described.

Hind limb ischemia in type 1 diabetic mice as useful tool to evaluate the neovascularization of tissue engineering constructs

Hind-limb ischemia has been used in type 1 diabetic mice to evaluate treatments for peripheral arterial disease or mechanisms of vascular impairment in diabetes [1]. Vascular deficiency is not only a pathophysiological condition, but also an obvious circumstance in tissue regeneration and in tissue engineering and regenerative medicine (TERM) strategies. We performed a pilot experiment of hind-limb ischemia in streptozotocin(STZ)-induced type 1 diabetic mice to hypothesise whether diabetes influences neovascularization induced by biomaterials. The dependent variables included blood flow and markers of arteriogenesis and angiogenesis. Type 1 diabetes was induced in 8-week-old C57BL/6 mice by an i.p. injection of STZ (50 mg/kg daily for 5 days). Hind-limb ischemia was created under deep anaesthesia and the left femoral artery and vein were isolated, ligated, and excised. The contralateral hind limb served as an internal control within each mouse. Non-diabetic ischaemic mice were used as experiment controls. At the hind-limb ischemia surgical procedure, different types of biomaterials were placed in the blood vessels gap. Blood flow was estimated by Laser Doppler perfusion imager, right after surgery and then weekly. After 28 days of implantation, surrounding muscle was excised and evaluated by histological analysis for arteriogenesis and angiogenesis. The results showed that implanted biomaterials were promote faster restoration of blood flow in the ischemic limbs and improved neovascularization in the diabetic mice. Therefore, we herein demonstrate that the combined model of hind-limb ischemia in type 1 diabetes mice is suitable to evaluate the neovascularization potential of biomaterials and eventually tissue engineering constructs.  

Study of the of the concept of community buildings and its importance for land use efficiency

Nowadays cities are facing several environmental problems due to the population migration to urban areas, which is causing urban sprawl. This way, it is very important to define solutions to improve Land Use Efficiency (LUE). This article proposes the use of community buildings features as a solution to increase land use efficiency. Community buildings consider the design of shared building spaces to reduce the floor area of buildings. This work tests the performance of some case-study buildings regarding LUE to analyse its possible pros and cons. A quantifiable method is used to assess buildingsâ LUE, which considers the number of occupants, the gross floor area, the functional area, the implantation area and the allotment area. Buildings with higher values for this index have reduced environmental impacts because they use less construction materials, produce less construction and demolition wastes and require less energy for building operation. The results showed that the use of community building features can increase Land Use Efficiency of buildings.

High-aspect-ratio tridimensional electrode arrays for neural applications

Tese de Doutoramento em Engenharia Biomédica.

Development of advanced cell/tissue culture systems, based on enhanced polymeric scaffolds and sophisticated bioreactors, for tissue engineering applications

Programa Doutoral em Engenharia Biomédica

Development of folate-targeted liposomes for rheumatoid arthritis therapy

Tese de Doutoramento em Biologia Molecular e Ambiental (área de especialização em Biologia Celular e Saúde).

Improving diagnostic yield in obscure gastrointestinal bleeding - how virtual chromoendoscopy may be the answer

Background and aim: A significant proportion of patients presenting with obscure gastrointestinal bleeding (OGIB) have negative small bowel capsule endoscopy (SBCE) examinations, and yet remain at risk of rebleeding. We aimed to evaluate whether a second-look review of SBCE images using flexible spectral color enhancement (FICE) may improve the detection of potentially bleeding lesions. Materials and methods: This was a retrospective, single-center study including consecutive patients with OGIB subjected to SBCE, whose standard white light examination was nondiagnostic. Each SBCE was reviewed using FICE 1. New findings were labeled as either P1 or P2 lesions according to bleeding potential. Patients were followed up to assess the incidence of rebleeding. Results: A total of 42 consecutive patients were included. Sixteen patients (38%) experienced rebleeding after a mean follow-up of 26 months. Review of SBCE images using FICE 1 enabled the identification of previously unrecognized P2 lesions, mainly angioectasias, in nine patients (21%) and P1 lesions, mainly erosions, in 26 patients (62%). Among patients who experienced rebleeding, 13/16 (81%) were diagnosed with P1 lesions with FICE 1 (P=0.043), whereas 3/16 (19%) had confirmed nondiagnostic SBCE and only 1/16 (6%) had newly diagnosed P2 (plus P1) lesions. An alternative source of bleeding outside the small bowel was found in only 3/16 (19%) patients with rebleeding during the follow-up. Conclusion: In a significant proportion of patients with OGIB, FICE 1 may detect potentially bleeding lesions previously missed under conventional white light SBCE. Review of nondiagnostic SBCE with FICE 1 may be a valuable strategy to obviate the need for further investigations in patients with OGIB, particularly for those who experience rebleeding.

PillCam COLON 2© in Crohn's disease: A new concept of pan-enteric mucosal healing assessment

AIM To evaluate mucosal healing in patients with small bowel plus colonic Crohn's disease (CD) with a single non-invasive examination, by using PillCam COLON 2 (PCC2). METHODS Patients with non-stricturing nonpenetrating small bowel plus colonic CD in sustained corticosteroid-free remission were included. At diagnosis, patients had undergone ileocolonoscopy to identify active CD lesions, such as ulcers and erosions, and small bowel capsule endoscopy to assess the Lewis Score (LS). After = 1 year of follow-up, patients underwent entire gastrointestinal tract evaluation with PCC2. The primary endpoint was assessment of CD mucosal healing, defined as no active colonic CD lesions and LS < 135. RESULTS Twelve patients were included (7 male; mean age: 32 years), and mean follow-up was 38 mo. The majority of patients (83.3%) received immunosuppressive therapy. Three patients (25%) achieved mucosal healing in both the small bowel and the colon, while disease activity was limited to either the small bowel or the colon in 5 patients (42%). It was possible to observe the entire gastrointestinal tract in 10 of the 12 patients (83%) who underwent PCC2. CONCLUSION Only three patients in sustained corticosteroid-free clinical remission achieved mucosal healing in both the small bowel and the colon, highlighting the limitations of clinical assessment when stratifying disease activity, and the need for pan-enteric endoscopy to guide therapeutic modification.

Lewis Score - Prognostic value in patients with isolated small bowel Crohn's disease

Background and aims: Small bowel capsule endoscopy (SBCE) allows mapping of small bowel inflammation in Crohn’s disease (CD). We aimed to assess the prognostic value of the severity of inflammatory lesions, quantified by the Lewis score (LS), in patients with isolated small bowel CD. Methods: A retrospective study was performed in which 53 patients with isolated small bowel CD were submitted to SBCE at the time of diagnosis. The Lewis score was calculated and patients had at least 12 months of follow-up after diagnosis. As adverse events we defined disease flare requiring systemic corticosteroid therapy, hospitalization and/or surgery during follow-up. We compared the incidence of adverse events in 2 patient subgroups, i.e. those with moderate or severe inflammatory activity (LS =790) and those with mild inflammatory activity (135 = LS < 790). Results: The LS was =790 in 22 patients (41.5%), while 58.5% presented with LS between 135 and 790. Patients with a higher LS were more frequently smokers (p = 0.01), males (p = 0017) and under immunosuppressive therapy (p = 0.004). In multivariate analysis, moderate to severe disease at SBCE was independently associated with corticosteroid therapy during follow-up, with a relative risk (RR) of 5 (p = 0.011; 95% confidence interval [CI] 1.5–17.8), and for hospitalization, with an RR of 13.7 (p = 0 .028; 95% CI 1.3–141.9). Conclusion: In patients with moderate to severe inflammatory activity there were higher prevalences of corticosteroid therapy demand and hospitalization during follow-up. Thus, stratifying the degree of small bowel inflammatory activity with SBCE and LS calculation at the time of diagnosis provided relevant prognostic value in patients with isolated small bowel CD.