First Mouse Model for Combined Osteogenesis Imperfecta and Ehlers-Danlos Syndrome

Authors

Frieda Chen Ph.D., Ruolin Guo Ph.D., Shousaku Itoh D.D.S., Ph.D., Luisa Moreno Ph.D., Esther Rosenthal, Tanya Zappitelli, Ralph A. Zirngibl Ph.D., Ann Flenniken Ph.D., William Cole M.D., Marc Grynpas Ph.D., Lucy R. Osborne Ph.D., Wolfgang Vogel Ph.D., Lee Adamson Ph.D., Janet Rossant Ph.D., Jane E. Aubin Ph.D.

Abstract

Using a genome-wide N-ethyl-N-nitrosourea (ENU)-induced dominant mutagenesis screen in mice, we identified a founder with low bone mineral density (BMD). Mapping and sequencing revealed a T to C transition in a splice donor of the collagen alpha1 type I (Col1a1) gene resulting in the skipping of exon 9 and a predicted 18 amino acid deletion within the N-terminal region of the triple helical domain of Col1a1. Col1a1Jrt/+ mice were smaller in size, had lower BMD associated with decreased bone volume/tissue volume (BV/TV) and reduced trabecular number and furthermore exhibited mechanically weak and brittle, fracture-prone bones, a hallmark of Osteogenesis Imperfecta (OI). Several markers of osteoblast differentiation were upregulated in mutant bone and histomorphometry showed that the proportion of trabecular bone surfaces covered by activated osteoblasts (Ob.S/BS and N.Ob/BS) was elevated but bone surfaces undergoing resorption (Oc.S/BS and N.Oc/BS) were not. The number of bone marrow stromal osteoprogenitors (CFU-ALP) was unaffected, but mineralization was decreased in cultures from young Col1a1Jrt/+ versus +/+ mice. Total collagen and type I collagen content of matrices deposited by Col1a1Jrt/+ dermal fibroblasts in culture was ∼40% and 30% respectively that of +/+ cells, suggesting that mutant collagen chains exerted a dominant negative effect on type I collagen biosynthesis. Mutant collagen fibrils were also markedly smaller in diameter than +/+ fibrils in bone, tendon and extracellular matrices deposited by dermal fibroblasts in vitro. Col1a1Jrt/+ mice also exhibited traits associated with Ehlers-Danlos syndrome (EDS): their skin had reduced tensile properties, tail tendon appeared more frayed and a third of the young adult mice had noticeable curvature of the spine. Col1a1Jrt/+ is the first reported model of combined OI/EDS and will be useful for exploring aspects of OI and EDS pathophysiology and treatment.

Link To Article

http://dx.doi.org/10.1002/jbmr.2177

Evolution of mesoporous bioactive glass scaffold implanted in rat femur based on 45Ca labelling, tracing and histological analysis

Authors

Baiyan Sui, Gaoren Zhong, and Jiao Sun

Abstract

Mesoporous bioactive glass (MBG) as a biodegradable scaffold with a nanostructure has attracted significant attention. However, the in vivo evolution of MBG, which includes in-situ degradation, the local effect induced by degradation and the disposition of degradation products, remains unclear. In this study, we performed in situ labelling and synthesis of MBG scaffold for the first time using 45CaCl2. The obtained 45Ca-MBG scaffolds possessed as mesoporous-macroporous cross-linked structure. These 45Ca-MBG scaffolds were implanted in critical-sized rat femur defects (3×3 mm) for 1 day and for 1, 4, 8 and 12 weeks and analyzed by isotopic quantitative tracing. The results illustrated MBG scaffolds gradually degraded over time and persisted at a local level of approximately 9.63% at week 12. This finding suggests that only a very small amount of MBG-released calcium ions may have been transformed into calcium components of the new bone matrix. The research also confirmed that the active ingredients derived from the degradation of MBG scaffolds could actively regulate the mRNA expression levels of osteoblast-related genes in rat bone marrow-derived mesenchymal stem cells (rBMSCs) and promote bone regeneration in vivo. Moreover, through isotopic tracing of the entire body, 45Ca, which disappeared in situ after implantation, could be detected in the heart, lungs, spleen, kidneys, intestines and brain via the blood and mainly accumulated in distal bone tissue, including the radius and cranium. However, 45Ca radioactivity in the body tissues significantly decreased or disappeared after 12 weeks. Systemic toxicological studies on MBG scaffolds demonstrated the degradation products that spread to major organs did not cause abnormal histopathological changes. The above discoveries comprehensively address crucial issues regarding the application of MBG in vivo, and these findings provide a scientific basis for introducing a material with mesoporous structure into clinical applications.

Link To Article

http://dx.doi.org/10.1021/am4056886

Induced ablation of Bmp1 and Tll1 produces osteogenesis imperfecta in mice

Authors

Alison M. Muir, Yinshi Ren, Delana Hopkins Butz, Nicholas A. Davis, Robert D. Blank, David E. Birk, Se-Jin Lee, David Rowe, Jian Q. Feng and Daniel S. Greenspan

Abstract

Osteogenesis imperfecta (OI), or brittle bone disease, is most often caused by dominant mutations in the collagen I genes COL1A1/COL1A2, whereas rarer recessive OI is often caused by mutations in genes encoding collagen I-interacting proteins. Recently, mutations in the gene for the proteinase bone morphogenetic 1 (BMP1) were reported in two recessive OI families. BMP1 and the closely related proteinase mammalian tolloid-like 1 (mTLL1) are co-expressed in various tissues, including bone, and have overlapping activities that include biosynthetic processing of procollagen precursors into mature collagen monomers. However, early lethality of Bmp1- and Tll1-null mice has precluded use of such models for careful study of in vivo roles of their protein products. Here we employ novel mouse strains with floxed Bmp1 and Tll1 alleles to induce postnatal, simultaneous ablation of the two genes, thus avoiding barriers of Bmp1−/− and Tll1−/− lethality and issues of functional redundancy. Bones of the conditionally null mice are dramatically weakened and brittle, with spontaneous fractures—defining features of OI. Additional skeletal features include osteomalacia, thinned/porous cortical bone, reduced processing of procollagen and dentin matrix protein 1, remarkably high bone turnover and defective osteocyte maturation that is accompanied by decreased expression of the osteocyte marker and Wnt-signaling inhibitor sclerostin, and by marked induction of canonical Wnt signaling. The novel animal model presented here provides new opportunities for in-depth analyses of in vivo roles of BMP1-like proteinases in bone and other tissues, and for their roles, and for possible therapeutic interventions, in OI.

Link To Article

http://dx.doi.org/10.1093/hmg/ddu013

Vascular expression of the chemokine CX3CL1 promotes osteoclast recruitment and exacerbates bone resorption in an irradiated murine model

Authors

Ki Hoon Han, Jae Won Ryu, Kyung-Eun Lim, Soo-Han Lee, Yuna Kim, Chang Sun Hwang, Je-Yong Choi, Ki Ok Han

Abstract

Circulating osteoclast precursor cells highly express CX3C chemokine receptor 1 (CX3CR1), which is the only receptor for the unique CX3C membrane-anchored chemokine, fractalkine (CX3CL1). An irradiated murine model was used to evaluate the role of the CX3CL1–CX3CR1 axis in osteoclast recruitment and osteoclastogenesis. Ionizing radiation (IR) promoted the migration of circulating CD11b + cells to irradiated bones and dose-dependently increased the number of differentiated osteoclasts in irradiated bones. Notably, CX3CL1 was dramatically upregulated in the vascular endothelium after IR. IR-induced production of CX3CL1 by skeletal vascular endothelium promoted chemoattraction of circulating CX3CR1 +/CD11b + cells and triggered homing of these osteoclast precursor cells toward the bone remodeling surface, a specific site for osteoclast differentiation. CX3CL1 also increased the endothelium-derived expression of other chemokines including stromal cell-derived factor-1 (CXCL12) and macrophage inflammatory protein-2 (CXCL2) by activating the hypoxia-inducible factor-1 α pathway. These effects may further enhance osteoclastogenesis. A series of in vivo experiments confirmed that knockout of CX3CR1 in bone marrow-derived cells and functional inhibition of CX3CL1 using a specific neutralizing antibody significantly ameliorated osteoclastogenesis and prevented bone loss after IR. These results demonstrate that the de novo CX3CL1–CX3CR1 axis plays a pivotal role in osteoclast recruitment and subsequent bone resorption, and verify its therapeutic potential as a new target for anti-resorptive treatment.

Link To Article

http://dx.doi.org/10.1016/j.bone.2013.12.032

The Rho-GEF Kalirin regulates bone mass and the function of osteoblasts and osteoclasts

Authors

Su Huang, Pierre P. Elensite, Kornchanok Wayakanon, Prashant Mandela, Betty A. Eipper, Richard E. Mains, Matthew R. Allen, Angela Bruzzaniti

Abstract

Bone homeostasis is maintained by the balance between bone resorption by osteoclasts and bone formation by osteoblasts. Dysregulation in the activity of the bone cells can lead to osteoporosis, a disease characterized by low bone mass and an increase in bone fragility and risk of fracture. Kalirin is a novel GTP-exchange factor protein that has been shown to play a role in cytoskeletal remodeling and dendritic spine formation in neurons. We examined Kalirin expression in skeletal tissue and found that it was expressed in osteoclasts and osteoblasts. Furthermore, micro-CT analyses of the distal femur of global Kalirin knockout (Kal-KO) mice revealed significantly reduced trabecular and cortical bone parameters in Kal-KO mice, compared to WT mice, with significantly reduced bone mass in 8, 14 and 36 week-old female Kal-KO mice. Male mice also exhibited a decrease in bone parameters but not the level seen in female mice. Histomorphometric analyses also revealed decreased bone formation rate in 14 week-old female Kal-KO mice, as well as decreased osteoblast number/bone surface and increased osteoclast surface/bone surface. Consistent with our in vivo findings, the bone resorbing activity and differentiation of Kal-KO osteoclasts was increased in vitro. Although alkaline phosphatase activity by Kal-KO osteoblasts was increased in vitro, Kal-KO osteoblasts showed decreased mineralizing activity, as well as decreased secretion of OPG, which was inversely correlated with ERK activity. Taken together, our findings suggest that deletion of Kalirin directly affects osteoclast and osteoblast activity, leading to decreased OPG secretion which is likely to alter the RANKL/OPG ratio, promoting osteoclastogenesis. Therefore, Kalirin may play a role in paracrine and/or endocrine signaling events that control skeletal bone remodeling and the maintenance of bone mass.

Link To Article

http://dx.doi.org/10.1016/j.bone.2013.12.023

Biceps Detachment Decreases Joint Damage in a Rotator Cuff Tear Rat Model

Authors

Stephen J. Thomas PhD, ATC, Katherine E. Reuther BS, Jennica J. Tucker BS, Joseph J. Sarver PhD, Sarah M. Yannascoli MD, Adam C. Caro DVM, Pramod B. Voleti MD, Sarah I. Rooney MSE, David L. Glaser MD, Louis J. Soslowsky PhD

Abstract

Background

Pathology in the long head of the biceps tendon often occurs in patients with rotator cuff tears. Arthroscopic tenotomy is the most common treatment. However, the role of the long head of the biceps at the shoulder and the consequences of surgical detachment on the remaining shoulder structures remain unknown.

Questions/purposes

We hypothesized that detachment of the long head of the biceps, in the presence of supraspinatus and infraspinatus tears, would decrease shoulder function and decrease mechanical and histologic properties of both the subscapularis tendon and the glenoid articular cartilage.

Methods

We detached the supraspinatus and infraspinatus or the supraspinatus, infraspinatus, and long head of the biceps after 4 weeks of overuse in a rat model. Animals were gradually returned to overuse activity after detachment. At 8 weeks, the subscapularis and glenoid cartilage biomechanical and histologic properties were evaluated and compared.

Results

The group with the supraspinatus, infraspinatus, and long head of the biceps detached had greater medial force and decreased change in propulsion, braking, and vertical force. This group also had an increased upper and lower subscapularis modulus but without any differences in glenoid cartilage modulus. Finally, this group had a significantly lower cell density in both the upper and lower subscapularis tendons, although cartilage histology was not different.

Conclusions

Detachment of the long head of the biceps tendon in the presence of a posterior-superior cuff tear resulted in improved shoulder function and less joint damage in this animal model.

Clinical Relevance

This study provides evidence in an animal model that supports the use of tenotomy for the management of long head of the biceps pathology in the presence of a two-tendon cuff tear. However, long-term clinical trials are required.

Link To Article

http://dx.doi.org/10.1007/s11999-013-3422-8