Impact of isolation procedures on the development of a preclinical synovial fibroblasts/macrophages in an in vitro model of osteoarthritis

There is a lack of in vitro models able to properly represent osteoarthritis (OA) synovial tissue (ST). We aimed to characterize OA ST and to investigate whether a mechanical or enzymatic digestion procedures influence synovial cell functional heterogeneity in vitro. Procedures using mechanical nondigested fragments (NDF), synovial digested fragments (SDF), and filtrated synovial digested cells (SDC) were compared. An immunophenotypic profile was performed to distinguish synovial fibroblasts (CD55, CD73, CD90, CD106), macrophages (CD14, CD68), M1-like (CD80, CD86), and M2-like (CD163, CD206) synovial macrophages. Pro-inflammatory (interleukin 6 IL6), tumor necrosis factor alpha (TNFα), chemokine C-C motif ligand 3 (CCL3/MIP1α), C-X- motif chemokine ligand 10 (CXCL10/IP10) and anti-inflammatory (interleukin 10 (IL10)), transforming growth factor beta 1 (TGFβ1), C-C motif chemokine ligand 18 (CCL18) cytokines were evaluated. CD68 and CD163 markers were higher in NDF and SDF compared to the SDC procedure, while CD80, CD86, and CD206 were higher only in NDF compared to the SDC procedure. Synovial fibroblast markers showed similar percentages. TNFα, CCL3/MIP1α, CXCL10/IP10, and CCL18 were higher in NDF compared to SDC, but not compared to SDF. IL10 and TGFβ1 were higher in NDF than SDC at the molecular level, while IL6 did not show differences among procedures. We demonstrated that NDF isolation procedures better preserved the heterogeneity of specific OA synovial populations (fibroblasts, macrophages), fostering their use for testing new cell therapies or drugs for OA, reducing or avoiding the use of animal models

Mesenchymal stromal cells from a progressive pseudorheumatoid dysplasia patient show altered osteogenic differentiation

Background: Progressive pseudorheumatoid dysplasia (PPRD) is a rare autosomal recessive non-inflammatory skeletal disease with childhood onset and is characterized by a progressive chondropathy in multiple joints, and skeletal abnormalities. To date, the etiopathological relationship between biological modification occurring in PPRD and genetic mutation remains an open issue, partially due to the limited availability of biological samples obtained from PPRD patients for experimental studies. Case presentation: We describe the clinical features of a PPRD patient and experimental results obtained from the biological characterization of PPRD mesenchymal stromal cells (MSCs) and osteoblasts (OBs) compared to normal cell populations. Phenotypic profile modifications were found in PPRD compared to normal subjects, essentially ascribed to decreased expression of CD146, osteocalcin (OC) and bone sialoprotein in PPRD MSCs and enhanced CD146, OC and collagen type I expression in PPRD OBs. Gene expression of Dickkopf-1, a master inhibitor of WNT signaling, was remarkably increased in PPRD MSCs compared to normal expression range, whereas PPRD OBs essentially exhibited higher OC gene expression levels. PPRD MSCs failed to efficiently differentiate into mature OBs, so showing a greatly impaired osteogenic potential. Conclusions: Since all regenerative processes require stem cell reservoirs, compromised functionality of MSCs may lead to an imbalance in bone homeostasis, suggesting a potential role of MSCs in the pathological mechanisms of PPRD caused by WNT1-inducible signaling pathway protein-3 (WISP3) mutations. In consideration of the lack of compounds with proven efficacy in such a rare disease, these data might contribute to better identify new specific and effective therapeutic approaches

Basic science of osteoarthritis

Osteoarthritis (OA) is a prevalent, disabling disorder of the joints that affects a large population worldwide and for which there is no definitive cure. This review provides critical insights into the basic knowledge on OA that may lead to innovative end efficient new therapeutic regimens. While degradation of the articular cartilage is the hallmark of OA, with altered interactions between chondrocytes and compounds of the extracellular matrix, the subchondral bone has been also described as a key component of the disease, involving specific pathomechanisms controlling its initiation and progression. The identification of such events (and thus of possible targets for therapy) has been made possible by the availability of a number of animal models that aim at reproducing the human pathology, in particular large models of high tibial osteotomy (HTO). From a therapeutic point of view, mesenchymal stem cells (MSCs) represent a promising option for the treatment of OA and may be used concomitantly with functional substitutes integrating scaffolds and drugs/growth factors in tissue engineering setups. Altogether, these advances in the fundamental and experimental knowledge on OA may allow for the generation of improved, adapted therapeutic regimens to treat human OA.(undefined

APPLICATION OF AUTOMATIC IMAGE SEGMENTATION TECHNIQUES TO REMOTE SENSING SURVEYS OF CULTURAL HERITAGE

In the field of Cultural Heritage, image analysis represents an indispensable practice for restorers and Institutions called to plan restoration interventions. Quantification of the extension of degraded areas is usually performed by manually tracing the decay zones. This procedure is cumbersome and time consuming; in addition it is subjective and requires expertise. Moreover, the recent development and widespread of technologies able to exploit different characteristics of light (i.e. infrared and x-ray spectroscopy) and therefore provide images of phenomena that are invisible to the naked eyes suggest applications that could be improved by automatic analysis in different fields of investigations. In addition to these aspects, as remote sensing technologies nowadays allow to provide reality-based models of artefacts, the possibility to automatically extract and easily manage information derived from 2d images in a 3d environment could enrich documentation about their state of preservation. The purpose of this contribution is to show the results of investigations held in order to provide a methodology for the automatic detection of decay areas within architectures and artefacts using colour images as a field of examination. Within our investigations, we selected images representing recurrent decays, as, for example, detachments, cracks and chromatic alterations and run them both to manual and to automatic recognition and selection tests, in order to subsequently compare the results obtained using the two approaches and evaluate the reliability of the automatic one. In particular, automatic detection was based on the analysis of the histograms of the three components of rgb images from which automatic selection of thresholding values were computed and morphological operators were applied. Following this analysis, a regularization step based on level set techniques was applied to optimize the detection of decays areas. Results comparison included computational and user time, quantification of the decay area error between manual and automatically detected zones in both percentage and overlapping terms as well as the Hausdorff distance between the detected contours. Additional parameters characterizing the type of decay (i.e. crack length, mean crack width, type of detachment) were also computed for each case study. Automatic analysis in all case studies resulted faster than the manual analysis (mean time: 19 sec vs 9 min). Mean area difference between the manual and automatic analysis was 0,66%; non overlapping area resulted in a mean value of 0,42. Mean Hausdorff distance was 1,8 cm. An example of the qualitative comparison of detected missing bricks is shown in figure 1 together with the segmentation of the 3d model using the automatic detection on 2d image as a source of selection of polygonal faces. Figure 2 shows the results obtained with automatic segmentation and decay completion of a frescoed wall. Comparison between the automatic and the manual procedure showed that the automatic detection is faster and reliable in all our selected case studies. In addition, our methodology showed evident improvements in the segmentation of reality-based models derived from remote sensing, with important consequences in the evaluation of the entity and extension of decay areas on 3d geometry

Chemoselective synthesis of pyrazole derivatives via beta-enamino keto esters

beta -Enamino keto ester (2a) reacts with hydroxylamine to give an isoxazole derivative (3). Compound (2a) reacts with hydrazine, alkyl- and arylhydrazines to give pyrazole derivatives (4a-e) as only reaction products. Methylation of compound (4a) afforded the two isomeric pyrazole derivatives (4b) and (5)

An integrated and automated segmentation approach to deteriorated regions recognition on 3D reality-based models of cultural heritage artifacts

In the field of Cultural Heritage, image analysis represents an indispensable practice for restorers to collect information about the state of preservation of monuments and artifacts and plan restoration interventions. In addition, during the last two decades, the wide spread of remote sensing technologies and the possibility to build 3D reality-based models of artifacts allow the extension of image analysis to 3D environments. In this context, the purpose of this contribution is to show the results of investigations held in order to provide a methodology for the automatic detection of deteriorated areas within architectures and artifacts using colour images as a field of examination. Using both 2D and 3D segmentation approaches, our methodology aims at speeding and efficiently performing the automatic detection of deteriorated zones within Cultural Heritage and therefore segment 3D digital models acquired using different survey technologies. Within our investigations, we selected case studies concerning recurrent deteriorations, such as, for example, detachments, cracks and chromatic alterations; we run them both to manual and to automatic recognition and selection tests, in order to compare the results obtained using these approaches and evaluate the reliability of the automatic one. Results comparison included computational and user time, quantification of the deteriorated area error between manual and automatically detected zones. Additional parameters characterizing the specific type of deteriorations were also computed for each case study. Comparison between the automatic and the manual procedure showed that the automatic detection is faster and reliable in all our selected case studies, with evident improvements in the efficient evaluation of the entity and extension of deteriorated areas on 3D geometry

A new route to the synthesis of pyrazole and pyrimidine C-nucleoside derivatives

A new route to the synthesis of pyrazole and pyrimidine C-nucleosides, which involves as the key step a metal promoted reaction of beta-D-ribofuranosyl ketoesters with alkyl cyanoformates is described. 2,3,5-Tri-O-benzoyl-beta-D-ribofuranosyl cyanide 1 reacts with alpha-bromoesters, in the presence of zinc dust, to give beta-D-ribofuranosyl-enaminoesters 2 which are hydrolysed with IN hydrochloric acid to beta-ketoesters 3. The reactions of beta-ketoesters 3 with alkyl cyanoformates, in the presence of tin(IV) chloride or of catalytic amounts of metal acetylacetonates, afford beta-D-ribofuranosyl enaminoketoesters 4. These compounds react with benzylhydrazine and acetamidine to give pyrazole and pyrimidine C-nucleosides (6,7). (C) 1999 Elsevier Science Ltd. All rights reserved