Beyond lectures and practical courses: Teaching pharmacology using imaginative pedagogical tools.

Pharmacology has broadened its scope considerably in recent decades. Initially, it was of interest to chemists, doctors and pharmacists. In recent years, however, it has been incorporated into the teaching of biologists, molecular biologists, biotechnologists, chemical engineers and many health professionals, among others. Traditional teaching methods, such as lectures or laboratory work, have been superseded by the use of new pedagogical approaches to enable a better conceptualization and understanding of the discipline. In this article, we present several new methods that have been used in Spanish universities. Firstly, we describe a teaching network that has allowed the sharing of pedagogical innovations in Spanish universities. A European experience to improve prescribing safety is described in detail. The use of popular films and medical TV series in biomedical students shows how these audiovisual resources can be helpful in teaching pharmacology. The use of virtual worlds is detailed to introduce this new approach to teaching. The increasingly important area of the social aspects of pharmacology is also considered in two sections, one devoted to social pharmacology and the other to the use of learning based on social services to improve understanding of this important area. Finally, the use of Objective Structured Clinical Evaluation in pharmacology allows to know how this approach can help to better evaluate clinical pharmacology students. In conclusion, this article allows to know new pedagogical methods resources used in some Spanish universities that may help to improve the teaching of pharmacology.

Impact of TGF-ß family-related growth factors on chondrogenic differentiation of adipose-derived stem cells isolated from lipoaspirates and infrapatellar fat pads of osteoarthritic patients.

The success of cell-based approaches for the treatment of cartilage defects requires an optimal autologous cell source with chondrogenic differentiation ability that maintains its differentiated properties and stability following implantation. The objective of this study was to compare the chondrogenic capacity of mesenchymal stem cells (MSCs) isolated from lipoaspirates (ASCs) and the infrapatellar fat pad (IFPSCs) of osteoarthritic patients and treated with transforming growth factor (TGF)-ß family-related growth factors. Cells were cultured for 6 weeks in a 3D pellet culture system with the chimeric activin A/bone morphogenic protein (BMP)-2 ligand (AB235), the chimeric nodal/BMP-2 ligand (NB260) or BMP-2. To investigate the stability of the new cartilage, ASCs-treated pellets were transplanted subcutaneously into severe combined immunodeficiency (SCID) mice. Histological and immunohistochemical assessment confirmed that the growth factors induced cartilage differentiation in both isolated cell types. However, reverse transcription-quantitative PCR results showed that ASCs presented a higher chondrogenic potential than IFPSCs. In vivo results revealed that AB235-treated ASCs pellets were larger in size and could form stable cartilage-like tissue as compared to NB260-treated pellets, while BMP-2-treated pellets underwent calcification. The chondrogenic induction of ASCs by AB235 treatment was mediated by SMAD2/3 activation, as proved by immunofluorescence analysis. The results of this study indicated that the combination of ASCs and AB235 might lead to a cell-based cartilage regeneration treatment.

Utility and cost of low-vacuum reinfusion drains in patients undergoing surgery for subcapital hip fracture repair. A before and after cohort study.

BACKGROUND: Postoperative blood loss may be a risk factor for allogeneic blood transfusion (ABT) in patients undergoing subcapital hip fracture (SHF) repair. We investigated the utility and costs of using a low-vacuum reinfusion drain (Bellovac ABT) within a blood management protocol for reducing ABT requirements in consecutive SHF. METHODS: The blood management protocol consisted of the application of a restrictive transfusion trigger (Hb < 8 g/dl), the peri-operative administration of IV iron sucrose (3 × 200 mg/48 h) ± recombinant erythropoietin (1 × 40 000 IU sc) and the use of Bellovac ABT (Group 2, n = 117). An immediate previous SHF series managed without Bellovac ABT served as control (Group 1, n = 138). RESULTS: Overall, 72 out of 255 (28%) received at least one ABT unit (2·1 ± 1·0 U/transfused patient) without differences between groups. However, in the subgroup of patients with admission Hb < 13 g/dl, the use of Bellovac ABT reduced postoperative ABT rates (16% vs. 46%, for groups 2 and 1, respectively; P = 0·001), although only 3 were reinfused, and was cost-saving. The use of Bellovac ABT also resulted in fewer wound bleeding complications, but there were no differences in Hb at postoperative days 7 and 30 between groups. CONCLUSIONS: In SHF patients with admission Hb < 13 g/dl and managed with peri-operative IV iron ± recombinant erythropoietin plus restrictive transfusion indication, the use of Bellovac ABT was associated with reduced ABT requirements, without increasing postoperative complications, and cost-savings.

Chondrocytes extract from patients with osteoarthritis induces chondrogenesis in infrapatellar fat pad-derived stem cells.

OBJECTIVE: Infrapatellar fat pad of patients with osteoarthritis (OA) contains multipotent and highly clonogenic adipose-derived stem cells that can be isolated by low invasive methods. Moreover, nuclear and cytoplasmic cellular extracts have been showed to be effective in induction of cell differentiation and reprogramming. The aim of this study was to induce chondrogenic differentiation of autologous mesenchymal stem cells (MSCs) obtained from infrapatellar fat pad (IFPSCs) of patients with OA using cellular extracts-based transdifferentiation method. DESIGN: IFPSCs and chondrocytes were isolated and characterized by flow cytometry. IFPSCs were permeabilized with Streptolysin O and then exposed to a cell extract obtained from chondrocytes. Then, IFPSCs were cultured for 2 weeks and chondrogenesis was evaluated by morphologic and ultrastructural observations, immunologic detection, gene expression analysis and growth on 3-D poly (dl-lactic-co-glycolic acid) (PLGA) scaffolds. RESULTS: After isolation, both chondrocytes and IFPSCs displayed similar expression of MSCs surface makers. Collagen II was highly expressed in chondrocytes and showed a basal expression in IFPSCs. Cells exposed to chondrocyte extracts acquired a characteristic morphological and ultrastructural chondrocyte phenotype that was confirmed by the increased proteoglycan formation and enhanced collagen II immunostaining. Moreover, chondrocyte extracts induced an increase in mRNA expression of chondrogenic genes such as Sox9, L-Sox5, Sox6 and Col2a1. Interestingly, chondrocytes, IFPSCs and transdifferentiated IFPSCs were able to grow, expand and produce extracellular matrix (ECM) on 3D PLGA scaffolds. CONCLUSIONS: We demonstrate for the first time that extracts obtained from chondrocytes of osteoarthritic knees promote chondrogenic differentiation of autologous IFPSCs. Moreover, combination of transdifferentiated IFPSCs with biodegradable PLGA 3D scaffolds can serve as an efficient system for the maintenance and maturation of cartilage tissue. These findings suggest its usefulness to repair articular surface in OA.

Pore geometry influences growth and cell adhesion of infrapatellar mesenchymal stem cells in biofabricated 3D thermoplastic scaffolds useful for cartilage tissue engineering.

The most pressing need in cartilage tissue engineering (CTE) is the creation of a biomaterial capable to tailor the complex extracellular matrix of the tissue. Despite the standardized used of polycaprolactone (PCL) for osteochondral scaffolds, the pronounced stiffness mismatch between PCL scaffold and the tissue it replaces remarks the biomechanical incompatibility as main limitation. To overcome it, the present work was focused in the design and analysis of several geometries and pore sizes and how they affect cell adhesion and proliferation of infrapatellar fat pad-derived mesenchymal stem cells (IPFP-MSCs) loaded in biofabricated 3D thermoplastic scaffolds. A novel biomaterial for CTE, the 1,4-butanediol thermoplastic polyurethane (b-TPUe) together PCL were studied to compare their mechanical properties. Three different geometrical patterns were included: hexagonal (H), square (S), and, triangular (T); each one was printed with three different pore sizes (PS): 1, 1.5 and 2 mm. Results showed differences in cell adhesion, cell proliferation and mechanical properties depending on the geometry, porosity and type of biomaterial used. Finally, the microstructure of the two optimal geometries (T1.5 and T2) was deeply analyzed using multiaxial mechanical tests, with and without perimeters, µCT for microstructure analysis, DNA quantification and degradation assays. In conclusion, our results evidenced that IPFP-MSCs-loaded b-TPUe scaffolds had higher similarity with cartilage mechanics and T1.5 was the best adapted morphology for CTE.

Efficiency of gene transfection reagents in NG108-15, SH-SY5Y and CHO-K1 cell lines.

Several gene delivery reagents were analyzed for their transfection efficiency. Genes studied belonged to the class of mammalian proteins termed regulators of G-protein signaling (RGS), ranged in size up to 2.2 Kb long and were transfected into the NG108-15, SH-SY5Y and CHO-K1 cell lines. Prior to transfection, genes were cloned into a nonviral vector pcDNA 6.2/EmGFP, so as to express a green fluorescent protein tag at the 3' end. Flow cytometry was used to analyze cell fluorescent activity and thereby transfection efficiency. Gene delivery reagents Lipofectamine 2000 and ExGen 500 produced more effective transfection in NG108-15 cells whereas Lipofectamine 2000, ExGen 500 and TurboFectin 8.0 were more effective in CHO-K1 cells. In both these cell lines, transfection efficiency reached 60-80%. In SH-SY5Y cells, TurboFectin 8.0 produced the best transfection result; however efficiency level was only 5%. Gene size had no effect on transfection efficiency. Unlike Lipofectamine 2000, cells transfected using ExGen 500 showed morphological deformation. Our results suggest that Lipofectamine 2000 is the most suitable transfection medium for gene delivery to NG108-15 and CHO-K1 cells.

The S1P mimetic fingolimod phosphate regulates mitochondrial oxidative stress in neuronal cells.

Fingolimod is one of the few oral drugs available for the treatment of multiple sclerosis (MS), a chronic, inflammatory, demyelinating and neurodegenerative disease. The mechanism of action proposed for this drug is based in the phosphorylation of the molecule to produce its active metabolite fingolimod phosphate (FP) which, in turns, through its interaction with S1P receptors, triggers the functional sequestration of T lymphocytes in lymphoid nodes. On the other hand, part if not most of the damage produced in MS and other neurological disorders seem to be mediated by reactive oxygen species (ROS), and mitochondria is one of the main sources of ROS. In the present work, we have evaluated the anti-oxidant profile of FP in a model of mitochondrial oxidative damage induced by menadione (Vitk3) on neuronal cultures. We provide evidence that incubation of neuronal cells with FP alleviates the Vitk3-induced toxicity, due to a decrease in mitochondrial ROS production. It also decreases regulated cell death triggered by imbalance in oxidative stress (restore values of advanced oxidation protein products and total thiol levels). Also restores mitochondrial function (cytochrome c oxidase activity, mitochondrial membrane potential and oxygen consumption rate) and morphology. Furthermore, increases the expression and activity of protective factors (increases Nrf2, HO1 and Trx2 expression and GST and NQO1 activity), being some of these effects modulated by its interaction with the S1P receptor. FP seems to increase mitochondrial stability and restore mitochondrial dynamics under conditions of oxidative stress, making this drug a potential candidate for the treatment of neurodegenerative diseases other than MS.

Addition of a short central extension to surface cemented tibial trays in primary TKA: an in vitro study of the effect on initial fixation stability and its relationship to supporting bone density.

BACKGROUND: Short central extensions which do not enter the tibial medullary canal are incorporated to cemented tibial components to increase initial stability in primary total knee arthroplasty. Their role in tibiae of differing preoperative mechanical quality has been little studied. METHODS: Twelve embalmed cadaveric tibiae were paired and divided into two groups, receiving a similar cemented tibial component with or without a non-cemented short central extension (10 mm diameter, 35 mm length). The specimens were subjected to 6000 cycles of a medially applied 1350 N load. Relative bone-tray displacements were measured and the evolution of inducible and permanent micromotions were computed. The apparent density of the cancellous bone under the tibial tray and at the area to support the extension was computed from computed tomography images of each specimen. FINDINGS: No significant differences between groups were detected for any parameters. For the group with extension, a significant negative linear correlation (P = 0.009, r(2) = 0.849) was found between the inducible tilt of the tray and the bone density at the zone of the extension. Also a trend towards a negative linear relation (P = 0.07, r(2) = 0.59) was observed for the same group between maximum subsidence and apparent density at the zone of the extension. INTERPRETATION: The study did not find that the addition of a non-cemented short central extension provides any overall improvement of the initial fixation stability. However, it was found that short extensions may enhance tilting stiffness of the bone-implant construct if bone of sufficient mechanical quality is located around its supporting area.

IGF-II promotes neuroprotection and neuroplasticity recovery in a long-lasting model of oxidative damage induced by glucocorticoids.

Insulin-like growth factor-II (IGF-II) is a naturally occurring hormone that exerts neurotrophic and neuroprotective properties in a wide range of neurodegenerative diseases and ageing. Accumulating evidence suggests that the effects of IGF-II in the brain may be explained by its binding to the specific transmembrane receptor, IGFII/M6P receptor (IGF-IIR). However, relatively little is known regarding the role of IGF-II through IGF-IIR in neuroprotection. Here, using adult cortical neuronal cultures, we investigated whether IGF-II exhibits long-term antioxidant effects and neuroprotection at the synaptic level after oxidative damage induced by high and transient levels of corticosterone (CORT). Furthermore, the involvement of the IGF-IIR was also studied to elucidate its role in the neuroprotective actions of IGF-II. We found that neurons treated with IGF-II after CORT incubation showed reduced oxidative stress damage and recovered antioxidant status (normalized total antioxidant status, lipid hydroperoxides and NAD(P) H:quinone oxidoreductase activity). Similar results were obtained when mitochondria function was analysed (cytochrome c oxidase activity, mitochondrial membrane potential and subcellular mitochondrial distribution). Furthermore, neuronal impairment and degeneration were also assessed (synaptophysin and PSD-95 expression, presynaptic function and FluoroJade B® stain). IGF-II was also able to recover the long-lasting neuronal cell damage. Finally, the effects of IGF-II were not blocked by an IGF-IR antagonist, suggesting the involvement of IGF-IIR. Altogether these results suggest that, in or model, IGF-II through IGF-IIR is able to revert the oxidative damage induced by CORT. In accordance with the neuroprotective role of the IGF-II/IGF-IIR reported in our study, pharmacotherapy approaches targeting this pathway may be useful for the treatment of diseases associated with cognitive deficits (i.e., neurodegenerative disorders, depression, etc.).

Activin A/BMP2 chimera AB235 drives efficient redifferentiation of long term cultured autologous chondrocytes.

Autologous chondrocyte implantation (ACI) depends on the quality and quantity of implanted cells and is hindered by the fact that chondrocytes cultured for long periods of time undergo dedifferentiation. Here we have developed a reproducible and efficient chondrogenic protocol to redifferentiate chondrocytes isolated from osteoarthritis (OA) patients. We used morphological, histological and immunological analysis together with a RT-PCR detection of collagen I and collagen II gene expression to show that chondrocytes isolated from articular cartilage biopsies of patients and subjected to long-term culture undergo dedifferentiation and that these cells can be redifferentiated following treatment with the chimeric Activin A/BMP2 ligand AB235. Examination of AB235-treated cell pellets in both in vitro and in vivo experiments revealed that redifferentiated chondrocytes synthesized a cartilage-specific extracellular matrix (ECM), primarily consisting of vertically-orientated collagen fibres and cartilage-specific proteoglycans. AB235-treated cell pellets also integrated into the surrounding subcutaneous tissue following transplantation in mice as demonstrated by their dramatic increase in size while non-treated control pellets disintegrated upon transplantation. Thus, our findings describe an effective protocol for the promotion of redifferentiation of autologous chondrocytes obtained from OA patients and the formation of a cartilage-like ECM that can integrate into the surrounding tissue in vivo.

Arthroscopic repair of the posterior cruciate ligament: two techniques.

SUMMARY: Isolated posterior cruciate ligament injuries are rare and their treatment is controversial. These lesions have commonly been treated by open reduction and internal fixation using a posterior approach. However, this approach makes it difficult to explore other combined injuries of the knee joint. We report 2 cases of posterior cruciate ligament avulsion of the tibia that were arthroscopically reduced and fixed using 2 different methods, cannulated screws and tension band wire.

[Diagnosis and localization of hyperparathyroidism by nuclear medicine procedures]. / Contribución de las técnicas de medicina nuclear al diagnóstico y localización del hiperparatiroidismo.

Primary hyperparathryoidism is a PTH hypersecretion caused by the parathyroid glands. In most cases (85%), the origin is to be due to the existence of a parathyroid adenoma, despite the intrinsic difficulty in being localized under certain circumstances. From some time now, we can count with the invaluable help of a nuclear medicine technique, namely the parathyroid scintigraphy with Technetium 99m-sestamibi (Tc99m-MIBI), a technique which is easy to perform, cheap and with excellent results, and which additionally can provide us with the above mentioned necessary information regarding location. We present here the case of a patient suffering from primary hyperparatyiroidism, in whom both the disease and the precise location of the hyperfunctioning tissue were identified by means of the parathyroid scintigraphy. Another nuclear medicine procedure, the one known as bone scintigraphy, also contributed meaningfully to the correct diagnosis in the same patient.

Perioperative intravenous iron: an upfront therapy for treating anaemia and reducing transfusion requirements.

Perioperative anaemia, with iron deficiency being its leading cause, is a frequent condition among surgical patients, and has been linked to increased postoperative morbidity and mortality, and decreased quality of life. Postoperative anaemia is even more frequent and is mainly caused by perioperative blood loss, aggravated by inflammation-induced blunting of erythropoiesis. Allogenic transfusion is commonly used for treating acute perioperative anaemia, but it also increases the rate of morbidity and mortality in surgical and critically ill patients. Thus, overall concerns about adverse effects of both preoperative anaemia and allogeneic transfusion have prompted the review of transfusion practice and the search for safer and more biologically rational treatment options. In this paper, the role of intravenous iron therapy (mostly with iron sucrose and ferric carboxymaltose), as a safe and efficacious tool for treating anaemia and reducing transfusion requirements in surgical patients, as well as in other medical areas, has been reviewed. From the analysis of published data and despite the lack of high quality evidence in some areas, it seems fair to conclude that perioperative intravenous iron administration, with or without erythropoiesis stimulating agents, is safe, results in lower transfusion requirements and hastens recovery from postoperative anaemia. In addition, some studies have reported decreased rates of postoperative infection and mortality, and shorter length of hospital stay in surgical patients receiving intravenous iron.