Proliferation and differentiation potential of canine synovial fluid cells

The aim of this study was to determine whether synovial fluid (SF) of dogs contains cells that have characteristics of MSCs and to describe their differentiation potential. SF adherent cells from 5 young German shepherd dogs (average 3.8 +/- 0.9 years) were expanded (37 degrees C, 5% CO2, humidified atmosphere) three weeks before their phenotype was characterized by flow-cytometry for the presence of CD90 and CD34. Population doubling time (PDT), number of CFU-F and adipogenic, osteogenic and chondrogenic potentials have been determined in vitro. In early passages PTD was 31 +/- 10 hours and expansion fold after 3 sub cultivations (9 days) theoretically could be 372 +/- 134. At P1, 0.55 +/- 0.05% of SF cells had the ability to form CFU-F. Sixty-six percent of cells expressed CD90 and none of the cells expressed markers of hematopoietic cells. Oil Red O staining has shown accumulation of fat droplets in cells grown in adipogenic medium, while deposits of calcium in the osteogenic medium were evidenced with Alizarin red staining. SF cultured in hondrogenic and control medium in three-dimensional conditions formed a cartilage-like tissue. Alcian blue staining of pellets' slides have shown a significant amount of glycosaminoglycans (GAGs) and immunohistochemistry analysis documented collagen type II expression. The amount of GAGs in pellets grown in both conditions showed no difference. SF cells in vitro exhibited osteogenic, adipogenic and chondrogenic differentiation potentials, suggesting the presence of different mesenchymal progenitors. These results also demonstrated that SF cells have a spontaneous chondrogenic potential that should be further explored for possible tissue engineering protocols

Effect of subclinical and overt form of rat maternal hypothyroidism on offspring endochondral bone formation

Maternal hypothyroidism in its overt form affects skeletal development of the offspring, but these data are not available for the subclinical form which is becoming very frequent among pregnant women. We hypothesized that the subclinical form of hypothyroidism in rat dams, influences the process of offspring endochondral ossification affecting proliferation and differentiation of chondrocytes, osteoclasts and osteoblasts in pups. Seven-day-old male pups (n=18) derived from control dams and dams treated with a low dose (1.5 mg/L) or high dose (150 mg/L) of propylthiouracil in drinking water during pregnancy and lactation were used. Histomorphometric analysis of pups tibia proximal growth plate, expression of mRNA, immunohistochemical and histochemical visualization of extracellular matrix components was performed. The length of the tibia was reduced in hypothyroid pups. Secretion of type 2 and 10 collagens in the subclinical and overt form were lower while the amount of glycosaminoglycans was higher when compared with controls. Down-regulated tartrate resistant acid phosphatase mRNA indicated altered osteoclasts function while lower expression of dentin matrix acid protein-1 mRNA and reduced synthesis of type 1 collagen accentuated a compromised bone formation in the overt form of hypothyroidism. The subclinical form of maternal hypothyroidism had a negative effect on the differentiation of hypertrophic chondrocytes and calcified cartilage removal in 7-day-old pups. In addition, overt hypothyroidism had a negative effect on the proliferation of chondrocytes and deposition of osteoid. Both forms of hypothyroidism resulted in a decrease of tibia length due to changes in growth plate formation

Slc20a2, Encoding the Phosphate Transporter PiT2, Is an Important Genetic Determinant of Bone Quality and Strength.

Osteoporosis is characterized by low bone mineral density (BMD) and fragility fracture and affects over 200 million people worldwide. Bone quality describes the material properties that contribute to strength independently of BMD, and its quantitative analysis is a major priority in osteoporosis research. Tissue mineralization is a fundamental process requiring calcium and phosphate transporters. Here we identify impaired bone quality and strength in Slc20a2-/- mice lacking the phosphate transporter SLC20A2. Juveniles had abnormal endochondral and intramembranous ossification, decreased mineral accrual, and short stature. Adults exhibited only small reductions in bone mass and mineralization but a profound impairment of bone strength. Bone quality was severely impaired in Slc20a2-/- mice: yield load (-2.3 SD), maximum load (-1.7 SD), and stiffness (-2.7 SD) were all below values predicted from their bone mineral content as determined in a cohort of 320 wild-type controls. These studies identify Slc20a2 as a physiological regulator of tissue mineralization and highlight its critical role in the determination of bone quality and strength. © 2019 The Authors. Journal of Bone and Mineral Research Published by Wiley Periodicals Inc

Co-evolution of segregation guide DNA motifs and the FtsK translocase in bacteria: identification of the atypical Lactococcus lactis KOPS motif

Bacteria use the global bipolarization of their chromosomes into replichores to control the dynamics and segregation of their genome during the cell cycle. This involves the control of protein activities by recognition of specific short DNA motifs whose orientation along the chromosome is highly skewed. The KOPS motifs act in chromosome segregation by orienting the activity of the FtsK DNA translocase towards the terminal replichore junction. KOPS motifs have been identified in γ-Proteobacteria and in Bacillus subtilis as closely related G-rich octamers. We have identified the KOPS motif of Lactococcus lactis, a model bacteria of the Streptococcaceae family harbouring a compact and low GC% genome. This motif, 5′-GAAGAAG-3, was predicted in silico using the occurrence and skew characteristics of known KOPS motifs. We show that it is specifically recognized by L. lactis FtsK in vitro and controls its activity in vivo. L. lactis KOPS is thus an A-rich heptamer motif. Our results show that KOPS-controlled chromosome segregation is conserved in Streptococcaceae but that KOPS may show important variation in sequence and length between bacterial families. This suggests that FtsK adapts to its host genome by selecting motifs with convenient occurrence frequencies and orientation skews to orient its activity

Production of transgenic pear plants expressing ferritin gene with the aim to reduce fire blight susceptibility

International audienceFire blight caused by the bacteria Erwinia amylovora is one of the most important diseases of pear. This causal agent produces a siderophore (desferrioxamine), which was identified as one of its virulence factors. This protein enables the bacteria to overcome conditions of iron limitation encountered in host tissues, and may also protect the bacteria against active oxygen species. Previous experiments indicate that the use of an iron chelator protein, encoded by the bovine lactoferrin gene, reduces fire blight symptoms in some transgenic pear clones. The aim of the present work is to test the ability of a more efficient iron chelator, plant ferritin, to reduce fire blight susceptibility in pear. In the literature, ferritin genes have been overexpressed under the control of a constitutive promoter in different transgenic plant species for various purposes. In several cases, the constitutive expression of ferritin produced negative effects such as reduced growth and chlorophyll content. Therefore, we decided to place the exogenous ferritin gene from pea under the control of a pathogen inducible promoter (sgd24) in comparison with a constitutive promoter (CaMV 35S). Two pear varieties, 'Conference' (CF) and 'Passe-Crassane' (PC) were transformed using both constructs. Transformation rates depended on variety and construct. They were respectively of 12 and 4.3% for CF and PC using the sgd24-ferritin construct. Only PC was transformed with the 35S-ferritin construct, with a transformation rate of 2%. First analyses of the transgenic clones by RT-PCR showed the expression of pea ferritin in both constructs and in all clones. The transgenic clones were acclimatized in greenhouse and exhibited normal growth. Quantification of ferritin gene expression, ferritin protein accumulation, and evaluation of fire blight resistance are underway

Porous beta tricalcium phosphate scaffolds used as a BMP-2 delivery system for bone tissue engineering

Macroporous beta tricalcium phosphate (β-TCP) scaffolds were evaluated as potential carriers and delivery systems for bone morphogenetic protein-2 (BMP-2). Chemical etching was performed to increase the available surface and thus the protein loading. X-ray diffraction and infrared spectrocopy analyses confirmed the preparation of pure β-TCP scaffolds. Scanning electron microscopy revealed interconnected porosity (64%) and a microporous surface after chemical etching. Scaffolds loaded with 30 and 15 μg of BMP-2 were implanted respectively into the back muscles and into femoral defects (condyle and diaphysis) of rabbits for 4 weeks. Histological observations confirmed the activity of the BMP-2 released from the scaffolds. Intramuscularly, bone was formed within the BMP-2-loaded scaffold pores. In the bone defects, the effect of released BMP-2 was similarly noticeable, as evaluated by histomorphometry. The incorporation of BMP-2 resulted in an amount of newly formed bone that was 1.3 times higher than with unloaded scaffolds. The implant site, however, did not have an effect on bone formation as no statistical differences were measured between cortical (diaphysis) and trabecular (condyle) defects. These results indicate the suitability of chemically etched β-TCP scaffolds as BMP-2 carriers, in the context of bone regeneration

Combination of a transgene-mediated defence mechanism with a natural resistance gene increases apple scab resistance

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Giant canine with dentine anomalies in oculo-facio-cardio-dental syndrome

International audienceRadiculomegaly affecting incisors, canines or premolars is a rare radiological finding (Maden et al., 2010) but is pathognomomic of a rare x-linked dominant syndrome called oculo-facio-cardio-dental syndrome (OFCDS). As this syndrome includes cardiac malformations and can lead to blindness due to congenital glaucoma, oral and maxillofacial surgeons should be aware of the somatic anomalies potentially associated with radiculomegaly. We report a typical case of OFCDS and provide the first description of the microscopic dental anomalies associated with this syndrome

Bone regeneration using porous titanium particles vs. bovine hydroxyapatite: a sinus lift study in rabbits

Background: The use of porous titanium particles as space filling biomaterials for intraoral bone regeneration might be of interest because of both the mechanical stability and nonresorbable properties. Titanium particles were recently described for bone regeneration in sinus lifts and in the treatment of periimplantitis. Nevertheless, tissue integration and 3D bone regeneration with titanium particles were poorly explored in these previous reports. Aim: The first objective of this study was to qualitatively and quantitatively assess the bone formation process, particularly the long-term behavior and 3D volume stability of subsinusal bone regeneration, using titanium or bovine hydroxyapatite granules, in a rabbit model. The second objective was to evaluate the effect of the hydration of the BHA particles with a therapeutic concentration of doxycycline solution on the osteogenesis and biomaterial resorption. Methods: Rabbits underwent a double sinus lift procedure using one of three materials: grade 1 porous titanium particles (Ti), bovine hydroxyapatite (BHA) or chemically modified bovine hydroxyapatite (BHATTC). Animals were sacrificed after 1 week, 5 weeks or 6 months. Samples were analyzed using mCT and nondecalcified histology. Results: The materials used in each of the three groups allowed an optimal bone formation; bone quantities and densities were not statistically different between the three groups. At 6 months, more stable 3D volume stability was found with TI and BHATTC (P¼0.0033). At 5 weeks and 6 months, bone to material contact (BMC) corroborating osteoconduction was significantly higher with BHA and BHATTC than with Ti (P < 0.0001). Conclusions and clinical implications: Even though the studied biomaterials displayed different architectures, they are relevant candidates for sinus lift bone augmentation before dental implants because they allow adequate 3D stability and osteogenesis. However, to recommend the clinical use of Ti, both an observation on the drilling effects of Ti particles and clinical trials are needed

A comparison of three calcium phosphate based space fillers in sinus elevation: a study in rabbits

Aim: Sub-sinusal bone regeneration procedures are reliable procedures that are known to be efficient with the use of biomaterials alone. Nevertheless, many types of biomaterials are available, and their efficacy in terms of bone formation and resorption rate have rarely been compared. This study aimed to compare the bone formation, resorption rate, osteoconductivity and 3D volume changes of three biomaterials often used for alveolar ridge augmentation. Materials and Methods: Rabbits underwent a double sinus lift procedure using three different types of space fillers: bovine hydroxyapatite (BHA), beta calcium phosphate (ß-TCP) and biphasic calcium phosphate (BCP). Animals were sacrificed at 1 week, 5 weeks and 6 months. Samples were subjected to X-ray microtomography and histology. Qualitative analysis was performed on non-decalcified sections, and quantitative histomorphometric analysis was conducted using SEM. Volume variations of bone augmentations were calculated at different time points. Results: All three particulated biomaterials proved to be effective in promoting osteogenesis. At 6 months, biomaterial resorption rates were significantly different across the 3 groups (p<0.0001). The highest resorption rate was found with β-TCP, in which only 6.7% of the baseline particle surface remained. At 6 months, bone was in close contact with the BHA particles, constituting a composite network; in contrast, BCP particles were often surrounded by soft tissues. In each group, no significant difference in volume variations were found at the different time point. Conclusions and clinical implications: Despite the limitations of the study, the three particulated space-filling biomaterials proved to be efficient to promote osteogenesis. High resorption rates and complete replacement of the biomaterials by bone seemed to withstand intrasinusal pressure. Further investigations in humans should consider longer follow-up periods