Osteogenesis Imperfecta (OI) is a heritable bone disorder characterized by bone fragility and often caused by mutations in the Type I collagen-encoding genes COL1A1 and COL1A2. The pathophysiology of OI, particularly at the cellular level, is still not well understood. This contributes to the lack of a cure for this disorder as well as an effective preventive or management options of its complications. In the bone environment, mesenchymal stem cells (MSCs) and osteoblasts (Ob) exert their function, at least partially, through the secretion of extracellular vesicles (EV). EV is a heterogeneous group of nanosized membrane-enclosed vesicles that carry/transfer a cargo of proteins, lipid and nucleic acids from the secreting cell to its target cells. Our objective is to characterize EVs secreted by human control (HC)- and OI-MSCs and their derived Obs, with focus on their protein content. We hypothesize that there will be differences in the protein content of EVs secreted by OI-Obs compared to HC-Ob, which may indicate a deviation from healthy Ob behavior and, thus, a role in OI pathophysiology.

MSCs were harvested from the adipose tissue of four COL1A1-OI and two HC patients. They were proliferated in an EV-depleted media, then induced to differentiate to extracellular matrix (ECM)-producing osteoblasts, which then gets mineralized. EVs secreted by MSCs (MSC-EV) and Obs (Ob-EV) were then purified and concentrated.

Using liquid chromatography- tandem mass spectrometry, proteomic analysis of the EV groups was done. A total of 384 unique proteins were identified in all EVs, 373 were found in Vesiclepedia indicating a good enrichment of our samples with EV proteins. 67 proteins of the total 384 were exclusively or significantly upregulated (p-value < 0 .05) in OI-Ob-EV and 28 proteins in the HC-Ob-EVs, relative to each other. These two groups of differentially expressed proteins were compared by Gene Ontology (GO) analysis of their cellular compartment, molecular functions and biological processes. We observed that there were differences in the cellular origin of EV-proteins, which may indicate heterogeneity of the isolated EVs. Molecular function and biological process analyses of the HC-Ob-EV proteins showed, as expected, predominantly calcium-related activities such as extracellular matrix (ECM) mineralization. OI-Ob-EV proteins were still predominantly exhibiting ECM organization and formation functions. Annexins A1,2,4,5 and 6 were differentially and significantly upregulated by the HC-Ob-EVs. Fibronectin (FN), Fibulin-1 and −2, and Laminins (α4 & γ1), which are amongst the early non-collagenous proteins to form the ECM, were differentially and significantly upregulated in the OI-Ob-EVs.

We concluded that the persistent expression of Fibronectin (FN), Fibulin-1 and −2, and Laminins in OI-Ob-EVs might indicate the presence of an immature ECM that the OI-Obs are trying to organize. ECM mineralization is largely dependent on the presence of an organized mature ECM, and this being compromised in OI bone environment, may be a contributor to the bone fragility seen in these patients. Annexins, which are calcium-binders that are vital for ECM mineralization, were significantly downregulated in the OI-Ob-EVs and this may be a further contributor to ECM mineralization impairment and bone fragility.

The different pathways by which bone morphogenetic protein 7 (BMP-7) could exert its osteogenic function in distraction osteogenesis (DO) were investigated. Using immunohistochemistry, the temporal and spatial expression of markers for angiogenesis, cell proliferation, Indian hedgehog pathway, osteogenic growth factors and their receptors were investigated in a rabbit model of DO. Our results showed that local injection of BMP-7 at the lengthened site caused up-regulation of expression of growth factors and their receptors, cell proliferation and vascular markers and Indian hedgehog gene in a temporal fashion. By knowing these pathways, manipulation of DO by pharmaceutical agents may be possible.

Based on preliminary data, BMP-7 can accelerate the consolidation of newly formed bone if locally injected early in the distraction phase; however, the exact mechanism remains unknown.

The purpose of this study was to investigate the different pathways through which BMP-7 exerts its effects in DO.

The right tibia of twenty-four rabbits was lengthened 2.0 cms. The rabbits were divided into three groups : control, placebo and treated groups. The rabbits received no injection (control), buffer (placebo) and 75 micro grams BMP7 (treated) in the distracted zone one week after the start of distraction. The rabbits were sacrificed ten minutes, one day, two days and two weeks following the injections. Using immunohistochemistry, the different pathways of bone formation were assessed by analysing the expression of markers for angiogenesis (VGEF, Vascular Endothelial Growth Factor and PECAM , platelet endothelial cell adhesion molecule) , cell proliferation markers (PCNA, proliferation cell nuclear antigen), osteogenic growth factors (TGFβ, IGF, FGF and their receptors) and Indian hedgehog gene as part of the parathyroid hormone related peptide pathway.

BMP-7 may stimulate bone formation through several pathways in a temporal fashion early after local injection, by up-regulating the expression of numerous osteogenic growth factors and their receptors and Indian hedgehog, and late two weeks after the injection, by up-regulating cell proliferation and vascular markers.

Our results showed the possible mechanisms of action of BMP-7 in DO and more importantly the various pathways through which pharmacological agents could be used in the manipulation of DO.

In osteogenesis imperfecta (OI) because of bone fragility, deformities in load bearing regions of the body such as femoral neck and proximal femur are expected. The purpose of this study was to determine the prevalence and clinical presentation of coxa vara in two hundred and ninety-two patients with different types of OI. More than half of the patients were OI type III (55%) and the highest prevalence of coxa vara was seen in OI type VI (44,5%). The children suffering from coxa vara had also a significant limitation of range of motion in their hips.

The charts and x-rays of one hundred and fifty-four girls and one hundred and thirty-eight boys with OI were reviewed. The patients were classified according to the Sillence classification modified by Glorieux: eighty-seven Type I, sixty-nine Type IV, sixty-two Type III, eighteen Type V, nine type VI, four types VII, and forty-three unclassified. The mean age was nine, four years (0, 3–23, 3).

Twenty-nine patients (9, 9%) had coxa vara (twenty-three left, twenty right). 55% of them were type III, 17% type IV, 13, 8% type VI and three, 4% each of types I, V, VII and unclassified OI. The prevalence of coxa vara was 1% in type I, 5,5% in type V, 7 % in type IV, 25% in type VII , 26% in type III and 44,5% in type VI (p< 0,001 for difference between types I, III and IV). Coxa vara was less frequent in patients with blue sclera (p=0,007). The mean neck-shaft angle was 99° (80°–110°) and the mean Hilgenreiner-epiphyseal angle was 68° (40°–90°). Twenty-five of coxa vara patients (thirty-six hips) had femoral rodding before diagnosis and six hips (all type III) had no history of rodding; however, 26 % of five hundred and thirty-one hips without coxa vara, had previous history of femoral rodding (p=0,004).

Abduction, extension and internal rotation were restricted in the hips with deformity. The abductors and extensors of the hips were weak in some that resulted in limping and Trendelenburg gait.

Special attention including clinical and radiological follow-up should be given to type III and VI patients particularly in the presence of previous femoral rodding.