Empirical studies of open source evolution

Copyright @ 2008 Springer-VerlagThis chapter presents a sample of empirical studies of Open Source Software (OSS) evolution. According to these studies, the classical results from the studies of proprietary software evoltion, such as Lehman’s laws of software evolution, might need to be revised, if not fully, at least in part, to account for the OSS observations. The book chapter also summarises what appears to be the empirical status of each of Lehman’s laws with respect to OSS and highlights the threads to validity that frequently emerge in these empirical studies. The chapter also discusses related topics for further research

Generation of a human control iPS cell line (ESi080‐A) from a donor with no rheumatic diseases

[Abstract] Here, we report the establishment of the human iPS cell line N1-FiPS4F#7 generated from skin cells of a patient with no rheumatic diseases, thus obtaining an appropriate control iPS cell line for researchers working in the field of rheumatic diseases. The reprogramming factors Oct4, Sox2, Klf4 and c-Myc were introduced using a non-integrating reprogramming strategy involving Sendai Virus.Instituto de Salud Carlos III; PI17/0219

Versatility of Induced Pluripotent Stem Cells (iPSCs) for Improving the Knowledge on Musculoskeletal Diseases

[Abstract] Induced pluripotent stem cells (iPSCs) represent an unlimited source of pluripotent cells capable of di erentiating into any cell type of the body. Several studies have demonstrated the valuable use of iPSCs as a tool for studying the molecular and cellular mechanisms underlying disorders a ecting bone, cartilage and muscle, as well as their potential for tissue repair. Musculoskeletal diseases are one of the major causes of disability worldwide and impose an important socio-economic burden. To date there is neither cure nor proven approach for e ectively treating most of these conditions and therefore new strategies involving the use of cells have been increasingly investigated in the recent years. Nevertheless, some limitations related to the safety and di erentiation protocols among others remain, which humpers the translational application of these strategies. Nonetheless, the potential is indisputable and iPSCs are likely to be a source of di erent types of cells useful in the musculoskeletal field, for either disease modeling or regenerative medicine. In this review, we aim to illustrate the great potential of iPSCs by summarizing and discussing the in vitro tissue regeneration preclinical studies that have been carried out in the musculoskeletal field by using iPSCs.Instituto de Salud Carlos III; PI17/02197Xunta de Galicia; R2016/036Xunta de Galicia; R2014/050Xunta de Galicia; CN2012/142Xunta de Galicia; GPC2014/04

Usefulness of mesenchymal cell lines for bone and cartilage regeneration research

[Abstract] The unavailability of sufficient numbers of human primary cells is a major roadblock for in vitro repair of bone and/or cartilage, and for performing disease modelling experiments. Immortalized mesenchymal stromal cells (iMSCs) may be employed as a research tool for avoiding these problems. The purpose of this review was to revise the available literature on the characteristics of the iMSC lines, paying special attention to the maintenance of the phenotype of the primary cells from which they were derived, and whether they are effectively useful for in vitro disease modeling and cell therapy purposes. This review was performed by searching on Web of Science, Scopus, and PubMed databases from 1 January 2015 to 30 September 2019. The keywords used were ALL = (mesenchymal AND (“cell line” OR immortal*) AND (cartilage OR chondrogenesis OR bone OR osteogenesis) AND human). Only original research studies in which a human iMSC line was employed for osteogenesis or chondrogenesis experiments were included. After describing the success of the immortalization protocol, we focused on the iMSCs maintenance of the parental phenotype and multipotency. According to the literature revised, it seems that the maintenance of these characteristics is not guaranteed by immortalization, and that careful selection and validation of clones with particular characteristics is necessary for taking advantage of the full potential of iMSC to be employed in bone and cartilage-related research.Xunta de Galicia; R2016/036Deputación da Coruña; BINV-CS/2016Xunta de Galicia; R2014/050Xunta de Galicia; CN2012/142Xunta de Galicia; GPC2014/04

Immortalizing mesenchymal stromal cells from aged donors while keeping their essential features

[Abstract] Human bone marrow-derived mesenchymal stromal cells (MSCs) obtained from aged patients are prone to senesce and diminish their differentiation potential, therefore limiting their usefulness for osteochondral regenerative medicine approaches or to study age-related diseases, such as osteoarthiritis (OA). MSCs can be transduced with immortalizing genes to overcome this limitation, but transduction of primary slow-dividing cells has proven to be challenging. Methods for enhancing transduction efficiency (such as spinoculation, chemical adjuvants, or transgene expression inductors) can be used, but several parameters must be adapted for each transduction system. In order to develop a transduction method suitable for the immortalization of MSCs from aged donors, we used a spinoculation method. Incubation parameters of packaging cells, speed and time of centrifugation, and valproic acid concentration to induce transgene expression have been adjusted. In this way, four immortalized MSC lines (iMSC#6, iMSC#8, iMSC#9, and iMSC#10) were generated. These immortalized MSCs (iMSCs) were capable of bypassing senescence and proliferating at a higher rate than primary MSCs. Characterization of iMSCs showed that these cells kept the expression of mesenchymal surface markers and were able to differentiate towards osteoblasts, adipocytes, and chondrocytes. Nevertheless, alterations in the CD105 expression and a switch of cell fate-commitment towards the osteogenic lineage have been noticed. In conclusion, the developed transduction method is suitable for the immortalization of MSCs derived from aged donors. The generated iMSC lines maintain essential mesenchymal features and are expected to be useful tools for the bone and cartilage regenerative medicine research.Xunta de Galicia; R2016/036Xunta de Galicia; R2014/050Xunta de Galicia; CN2012/142Xunta de Galicia; GPC2014/048Deputación da Coruña; BINV-CS/2016Instituto de Salud Carlos III; PI17/0219

Generation of Mesenchymal Cell Lines Derived from Aged Donors

[Abstract] Background: Mesenchymal stromal cells (MSCs) have the capacity for self-renewal and multi-differentiation, and for this reason they are considered a potential cellular source in regenerative medicine of cartilage and bone. However, research on this field is impaired by the predisposition of primary MSCs to senescence during culture expansion. Therefore, the aim of this study was to generate and characterize immortalized MSC (iMSC) lines from aged donors. Methods: Primary MSCs were immortalized by transduction of simian virus 40 large T antigen (SV40LT) and human telomerase reverse transcriptase (hTERT). Proliferation, senescence, phenotype and multi-differentiation potential of the resulting iMSC lines were analyzed. Results: MSCs proliferate faster than primary MSCs, overcome senescence and are phenotypically similar to primary MSCs. Nevertheless, their multi-differentiation potential is unbalanced towards the osteogenic lineage. There are no clear differences between osteoarthritis (OA) and non-OA iMSCs in terms of proliferation, senescence, phenotype or differentiation potential. Conclusions: Primary MSCs obtained from elderly patients can be immortalized by transduction of SV40LT and hTERT. The high osteogenic potential of iMSCs converts them into an excellent cellular source to take part in in vitro models to study bone tissue engineering.This research was carried out thanks to the funding from Rede Galega de Terapia Celular 2016 (R2016/036) and Grupos con Potencial de Crecemento 2020 (ED431B 2020/55) from Xunta de Galicia, Proyectos de Investigación 2017 (PI17/02197) from Instituto de Salud Carlos III and the Biomedical Research Network Center (CIBER). The Biomedical Research Network Center (CIBER) is an initiative from Instituto de Salud Carlos III (ISCIII). MPR and SRF were granted a predoctoral fellowship from Xunta de Galicia and European Union (European Social Fund)Xunta de Galicia; R2016/036Xunta de Galicia; ED431B 2020/5

Tips and Tricks for Successfully Culturing and Adapting Human Induced Pluripotent Stem Cells [Protocol]

Protocol[Abstract] Reprogramming somatic cells toward pluripotency became possible over a decade ago. Since then, induced pluripotent stem cells (iPSCs) have served as a versatile and powerful tool not only for basic research but also with the long-term goal of using them in human cell transplantation after differentiation. Nonetheless, downstream applications are frequently blurred by the difficulties that researchers have to face when working with iPSCs, such as trouble with clonal selection, in vitro culture and cryopreservation, adaptation to feeder-free conditions, or expansion of the cells. Therefore, in this article we aim to provide other researchers with practical and detailed information to successfully culture and adapt iPSCs. Specifically, we (1) describe the most common problems when in-vitro culturing iPSCs onto feeder cells as well as its possible troubleshooting, and (2) compare different matrices and culture media for adapting the iPSCs to feeder-free conditions. We believe that the troubleshooting and recommendations provided in this article can be of use to other researchers working with iPSCs and who may be experiencing similar issues, hopefully enhancing the appeal of this promising cell source to be used for biomedical investigations, such as tissue engineering or regenerative medicine applications.We thank the laboratory staff from INIBIC-CHUAC, the Radio Physics department from the Oncology Center of Galicia and the Servicio de Xenética (CHUAC) for their assistance. We also thank the staff from Oza University Library (University of A Coruña) for their collaboration. We also thank our funding agencies: Instituto de Salud Carlos III-General Subdirection of Assessment and Promotion of the Research—European Regional Development Fund (FEDER) “A way of making Europe” (PI17/02197 and PI20/00933); Rede Galega deTerapia Celular and Grupos con Potencial de Crecemento, Xunta de Galicia (R2016/036, R2014/050, CN2012/142, ED431B 2020/55, and GPC2014/048); the University of A Coruña; M.P.-R. and S.R.-F. are granted by a predoctoral fellowship from Xunta de Galicia and European Union (European Social Fund) and C.S.-R. was beneficiary of a postdoctoral fellowship from Xunta de GaliciaXunta de Galicia; R2016/036Xunta de Galicia; R2014/050Xunta de Galicia; CN2012/142Xunta de Galicia; ED431B 2020/55Xunta de Galicia; GPC2014/04

Generation of osteoarthritic mesenchymal stromal cell lines.

ResumenXunta de Galicia; R2016/036Xunta de Galicia; R2014/050Xunta de Galicia; CN2012/142Xunta de Gaicia; GPC2014/048Instituto de salud Carlos III; PI17/0219

Generation and Characterization of Human Induced Pluripotent Stem Cells (iPSCs) From Hand Osteoarthritis Patient-Derived Fibroblasts

[Abstract] Knowledge and research results about hand osteoarthritis (hOA) are limited due to the lack of samples and animal models of the disease. Here, we report the generation of two induced pluripotent stem cell (iPSC)-lines from patients with radiographic hOA. Furthermore, we wondered whether these iPSC-lines carried single nucleotide polymorphisms (SNPs) within genes that have been associated with hOA. Finally, we performed chondrogenic differentiation of the iPSCs in order to prove their usefulness as cellular models of the disease. We performed a non-integrative reprogramming of dermal fibroblasts obtained from two patients with radiographic rhizarthrosis and non-erosive hOA by introducing the transcriptional factors Oct4, Sox2, Klf4 and c-Myc using Sendai virus. After reprogramming, embryonic stem cell-like colonies emerged in culture, which fulfilled all the criteria to be considered iPSCs. Both iPSC-lines carried variants associated with hOA in the four studied genes and showed differences in their chondrogenic capacity when compared with a healthy control iPSC-line. To our knowledge this is the first time that the generation of iPSC-lines from patients with rhizarthrosis and non-erosive hOA is reported. The obtained iPSC-lines might enable us to model the disease in vitro, and to deeper study both the molecular and cellular mechanisms underlying hOA.This study was carried out thanks to the funding from Fundación Española de Reumatología (Proyectos 2014), Proyectos de Investigación 2016 (PI16/02124) and 2017 (PI17/02197) from Instituto de Salud Carlos III-General Subdirection of Assesment and Promotion of the Research – European Regional Development Fund (FEDER) “A way of making Europe”, Rede Galega de Terapia Celular and Grupos con Potencial de Crecemento, Xunta de Galicia (R2016/036, R2014/050, CN2012/142 and GPC2014/048); Deputación da Coruña (BINV-CS/2015); University of A Coruña; Centro de Investigación Biomédica en Red-Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN). Rocío Castro-Viñuelas, María Piñeiro-Ramil and Silvia Rodríguez-Fernández are granted by a predoctoral fellowship from Xunta de Galicia and European Union (European Social Fund) and Clara Sanjurjo-Rodríguez is beneficiary of a postdoctoral fellowship from Xunta de GaliciaXunta de Galicia; R2016/036Deputación da Coruña; BINV-CS/2015Xunta de Galicia; R2014/050Xunta de Galicia; CN2012/142Xunta de Galicia; GPC2014/04

Autophagy activation by resveratrol reduces severity of experimental rheumatoid arthritis

Instituto de Salud Carlos III; RETIC‐RIER RD16/0012/0002Instituto de Salud Carlos III; PI12/02771Instituto de Salud carlos III; AGRUP2015/05Instituto de Salud Carlos III; AGRUP2018/0