Ma Martín-Piedra

Average cell viability levels of human dental pulp stem cells: an accurate combinatorial index for quality control in tissue engineering

BACKGROUND AIMS One of the most important issues in tissue engineering (TE) is the search for a suitable stem cell reservoir with optimal cell viability levels for the development of new tissues relevant for therapeutic needs. The aim of this study was to evaluate the cell viability levels of 10 sequential cell passages of human dental pulp stem cells (hDPSC) to determine their potential for TE techniques. METHODS To assess the average cell viability levels of hDPSC, four cell viability assays were used in a combinatorial approach: trypan blue exclusion test, water-soluble tetrazolium 1 assay, live/dead assay and electron probe x-ray microanalysis. RESULTS The results showed that cell viability as determined by trypan blue staining and live/dead assays was greater than 85%, with a significant decrease at the second passage (P < 0.05) and a significant increase at the ninth passage (P < 0.05). Electron probe x-ray microanalysis showed that the highest cell viability corresponded to the ninth passage, with the lowest K/Na values found at the third passage. No statistical differences were found among the different passages for the water-soluble tetrazolium 1 assay (P = 0.219). CONCLUSIONS Assessment of average cell viability levels showed that the highest viability of hDPSC was reached after nine passages, suggesting that this passage would be the most adequate for use in TE protocols.

An early and late cytotoxicity evaluation of lidocaine on human oral mucosa fibroblasts

Local anesthetic drugs are extensively used in dentistry. However, the cytotoxic effects of these pharmaceutical compounds remain unclear. In this work, we have evaluated the cell viability and cell function of human oral mucosa fibroblasts exposed to different concentrations of lidocaine for increasing incubation times, using a global screening methods including structural, metabolic and microanalytical analyses. Our results demonstrate that lidocaine is able to alter cell viability and function even at low concentrations and times, although the effect of lidocaine concentration was more important than the incubation time. First, the structural analysis methods revealed that ≥5% concentrations of lidocaine are able to significantly reduce cell viability. Then, the metabolic 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and water-soluble tetrazolium salt (WST-1) assays suggest that concentrations starting from 1% were able to significantly hinder cell physiology. Finally, electron-probe X-ray microanalysis confirmed the deleterious effects of lidocaine and allowed us to demonstrate that these effects are associated to an apoptosis process of cell death. Therefore, care should be taken when lidocaine is clinically used, and the lowest efficient concentrations should always be used. Furthermore, these results suggest that the comprehensive evaluation method used in this work is accurate and efficient for screening of local anesthetics.

Histological and immunohistochemical changes in the rat oral mucosa used as an autologous urethral graft

PURPOSE The purpose of this study was to determine the histological and functional (immunohistochemical) changes that take place in oral mucosa grafts implanted in the rat urethra. METHODS Urethroplasty was performed in 26 male Wistar rats weighing 250 g. All animals received autologous oral mucosa urethra grafting under general anesthesia. Samples were analyzed 10, 20, 30, 40, 50, 60, 90, and 120 days after surgery using light and scanning electron microscopy and immunofluorescence for the determination of the expression of epithelial markers (pancytokeratin, cytokeratin 1, 4, 13, and filaggrin). RESULTS Grafted oral mucosa tissues were subjected to significant histological changes from the beginning with the formation of a well-developed epithelium whose structure was comparable to the native urethra from day 60 of the surgical implant. The immunofluorescence analysis demonstrated that the cytokeratin expression profile tended to mimic the pattern of the native urethra. These data suggest that the oral mucosa is able to efficiently transdifferentiate to the urethral environment. CONCLUSIONS The efficient transdifferentiation process of the grafted oral mucosa at both the histological and immunofluorescence levels, and the absence of local complications confirm the clinical usefulness of this type of tissues for the repair of the urethra.

Generation and evaluation of novel stromal cell-containing tissue engineered artificial stromas for the surgical repair of abdominal defects†

Repair of abdominal wall defects is one of the major clinical challenges in abdominal surgery. Most biomaterials are associated to infection and severe complications, making necessary safer and more biocompatible approaches. In the present work, the adequate mechanical properties of synthetic polymer meshes with tissue-engineered matrices containing stromal mesenchymal cells is combined to generate a novel cell-containing tissue-like artificial stroma (SCTLAS) for use in abdominal wall repair. SCTLAS consisting on fibrin-agarose hydrogels seeded with stromal cells and reinforced with commercial surgical meshes (SM) are evaluated in vitro and in vivo in animal models of abdominal wall defect. Inflammatory cells, collagen, and extracellular matrix (ECM) components are analyzed and compared with grafted SM. Use of SCTLAS results in less inflammation and less fibrosis than SM, with most ECM components being very similar to control abdominal wall tissues. Cell migration and ECM remodeling within SCTLAS is comparable to control tissues. The use of SCTLAS could contribute to reduce the side-effects associated to currently available SM and regenerated tissues are more similar to control abdominal wall tissues. Bioengineered SCTLAS could contribute to a safer treatment of abdominal wall defects with higher biocompatibility than currently available SM.

Conceptions of learning factors in postgraduate health sciences master students: a comparative study with non-health science students and between genders

BackgroundThe students’ conceptions of learning in postgraduate health science master studies are poorly understood. The aim of this study was to compare the factors influencing conceptions of learning in health sciences and non-health sciences students enrolled in postgraduate master programs in order to obtain information that may be useful for students and for future postgraduate programs.MethodsA modified version of the Learning Inventory Conception Questionnaire (COLI) was used to compare students’ conception learning factors in 131 students at the beginning of their postgraduate studies in health sciences, experimental sciences, arts and humanities and social sciences.ResultsThe present study demonstrates that a set of factors may influence conception of learning of health sciences postgraduate students, with learning as gaining information, remembering, using, and understanding information, awareness of duty and social commitment being the most relevant. For these students, learning as a personal change, a process not bound by time or place or even as acquisition of professional competences, are less relevant. According to our results, this profile is not affected by gender differences.ConclusionsOur results show that the overall conceptions of learning differ among students of health sciences and non-health sciences (experimental sciences, arts and humanities and social sciences) master postgraduate programs. These finding are potentially useful to foster the learning process of HS students, because if they are metacognitively aware of their own conception or learning, they will be much better equipped to self-regulate their learning behavior in a postgraduate master program in health sciences.