Cortical bone stem cell-derived exosomes' therapeutic effect on myocardial ischemia-reperfusion and cardiac remodeling.

Heart failure is the one of the leading causes of death in the United States. Heart failure is a complex syndrome caused by numerous diseases, including severe myocardial infarction (MI). MI occurs after an occlusion of a cardiac artery causing downstream ischemia. MI is followed by cardiac remodeling involving extensive remodeling and fibrosis, which, if the original insult is severe or prolonged, can ultimately progress into heart failure. There is no "cure" for heart failure because therapies to regenerate dead tissue are not yet available. Previous studies have shown that in both post-MI and post-ischemia-reperfusion (I/R) models of heart failure, administration of cortical bone stem cell (CBSC) treatment leads to a reduction in scar size and improved cardiac function. Our first study investigated the ability of mouse CBSC-derived exosomes (mCBSC-dEXO) to recapitulate mouse CBSCs (mCBSC) therapeutic effects in a 24-h post-I/R model. This study showed that injection of mCBSCs and mCBSC-dEXOs into the ischemic region of an infarct had a protective effect against I/R injury. mCBSC-dEXOs recapitulated the effects of CBSC treatment post-I/R, indicating exosomes are partly responsible for CBSC's beneficial effects. To examine if exosomes decrease fibrotic activation, adult rat ventricular fibroblasts (ARVFs) and adult human cardiac fibroblasts (NHCFs) were treated with transforming growth factor ß (TGFß) to activate fibrotic signaling before treatment with mCBSC- and human CBSC (hCBSC)-dEXOs. hCBSC-dEXOs caused a 100-fold decrease in human fibroblast activation. To further understand the signaling mechanisms regulating the protective decrease in fibrosis, we performed RNA sequencing on the NHCFs after hCBSC-dEXO treatment. The group treated with both TGFß and exosomes showed a decrease in small nucleolar RNA (snoRNA), known to be involved with ribosome stability.NEW & NOTEWORTHY Our work is noteworthy due to the identification of factors within stem cell-derived exosomes (dEXOs) that alter fibroblast activation through the hereto-unknown mechanism of decreasing small nucleolar RNA (snoRNA) signaling within cardiac fibroblasts. The study also shows that the injection of stem cells or a stem-cell-derived exosome therapy at the onset of reperfusion elicits cardioprotection, emphasizing the importance of early treatment in the post-ischemia-reperfusion (I/R) wounded heart.

Bmi1 Augments Proliferation and Survival of Cortical Bone-Derived Stem Cells after Injury through Novel Epigenetic Signaling via Histone 3 Regulation.

Ischemic heart disease can lead to myocardial infarction (MI), a major cause of morbidity and mortality worldwide. Multiple stem cell types have been safely transferred into failing human hearts, but the overall clinical cardiovascular benefits have been modest. Therefore, there is a dire need to understand the basic biology of stem cells to enhance therapeutic effects. Bmi1 is part of the polycomb repressive complex 1 (PRC1) that is involved in different processes including proliferation, survival and differentiation of stem cells. We isolated cortical bones stem cells (CBSCs) from bone stroma, and they express significantly high levels of Bmi1 compared to mesenchymal stem cells (MSCs) and cardiac-derived stem cells (CDCs). Using lentiviral transduction, Bmi1 was knocked down in the CBSCs to determine the effect of loss of Bmi1 on proliferation and survival potential with or without Bmi1 in CBSCs. Our data show that with the loss of Bmi1, there is a decrease in CBSC ability to proliferate and survive during stress. This loss of functionality is attributed to changes in histone modification, specifically histone 3 lysine 27 (H3K27). Without the proper epigenetic regulation, due to the loss of the polycomb protein in CBSCs, there is a significant decrease in cell cycle proteins, including Cyclin B, E2F, and WEE as well as an increase in DNA damage genes, including ataxia-telangiectasia mutated (ATM) and ATM and Rad3-related (ATR). In conclusion, in the absence of Bmi1, CBSCs lose their proliferative potential, have increased DNA damage and apoptosis, and more cell cycle arrest due to changes in epigenetic modifications. Consequently, Bmi1 plays a critical role in stem cell proliferation and survival through cell cycle regulation, specifically in the CBSCs. This regulation is associated with the histone modification and regulation of Bmi1, therefore indicating a novel mechanism of Bmi1 and the epigenetic regulation of stem cells.

Cortical bone stem cells modify cardiac inflammation after myocardial infarction by inducing a novel macrophage phenotype.

Acute damage to the heart, as in the case of myocardial infarction (MI), triggers a robust inflammatory response to the sterile injury that is part of a complex and highly organized wound-healing process. Cortical bone stem cell (CBSC) therapy after MI has been shown to reduce adverse structural and functional remodeling of the heart after MI in both mouse and swine models. The basis for these CBSC treatment effects on wound healing are unknown. The present experiments show that CBSCs secrete paracrine factors known to have immunomodulatory properties, most notably macrophage colony-stimulating factor (M-CSF) and transforming growth factor-ß, but not IL-4. CBSC therapy increased the number of galectin-3+ macrophages, CD4+ T cells, and fibroblasts in the heart while decreasing apoptosis in an in vivo swine model of MI. Macrophages treated with CBSC medium in vitro polarized to a proreparative phenotype are characterized by increased CD206 expression, increased efferocytic ability, increased IL-10, TGF-ß, and IL-1RA secretion, and increased mitochondrial respiration. Next generation sequencing revealed a transcriptome significantly different from M2a or M2c macrophage phenotypes. Paracrine factors from CBSC-treated macrophages increased proliferation, decreased α-smooth muscle actin expression, and decreased contraction by fibroblasts in vitro. These data support the idea that CBSCs are modulating the immune response to MI to favor cardiac repair through a unique macrophage polarization that ultimately reduces cell death and alters fibroblast populations that may result in smaller scar size and preserved cardiac geometry and function.NEW & NOTEWORTHY Cortical bone stem cell (CBSC) therapy after myocardial infarction alters the inflammatory response to cardiac injury. We found that cortical bone stem cell therapy induces a unique macrophage phenotype in vitro and can modulate macrophage/fibroblast cross talk.

Hypoxia-cultured mouse mesenchymal stromal cells from bone marrow and compact bone display different phenotypic traits.

It has been suggested that the bone marrow microenvironment harbors two distinct populations of mesenchymal stromal cells (MSC), one with a perivascular location and other present in the endosteum. A better understanding of the biology of these MSC subsets has been pursued in order to refine its clinical application. However, most comparative characterizations of mouse MSC have been performed in normoxia. This can result in misleading interpretations since mouse MSC subsets with low/defective p53 activity are known to be selected during culture in normoxia. Here, we report a comprehensive in vitro characterization of mouse MSC isolated from bone marrow (BM-MSC) and compact bone (CB-MSC) expanded and assayed under hypoxia for their morphology, clonogenic efficiency and differentiation capacity. We found that, under hypoxia, compact bone is richer in absolute numbers of MSC and isolation of MSC from compact bone is associated with a reduced risk of hematopoietic cell carryover. In addition, CB-MSC have higher in vitro osteogenic capacity than BM-MSC, while adipogenic differentiation potential is similar. These findings reinforce the hypothesis of the existence of MSC in bone marrow and compact bone representing functionally distinct cell populations and highlight the compact bone as an efficient source of murine MSC under physiological oxygen concentrations.

Comparison of small-angle neutron and X-ray scattering for studying cortical bone nanostructure.

In this study, we present a combined small-angle neutron and X-ray scattering (SANS and SAXS) study of the nanoscale structure of cortical bone specimens from three different species. The variation of the scattering cross section of elements across the periodic table is very different for neutrons and X-rays. For X-rays, it is proportional to the electron density while for neutrons it varies irregularly with the atomic number. Hence, combining the two techniques on the same specimens allows for a more detailed interpretation of the scattering patterns as compared to a single-contrast experiment. The current study was performed on bovine, porcine and ovine specimens, obtained in two perpendicular directions with respect to the main axis of the bone (longitudinal and radial) in order to maximise the understanding of the nanostructural organisation. The specimens were also imaged with high resolution micro-computed tomography (micro-CT), yielding tissue mineral density and microstructural orientation as reference. We show that the SANS and SAXS patterns from the same specimen are effectively identical, suggesting that these bone specimens can be approximated as a two-component composite material. Hence, the observed small-angle scattering results mainly from the mineral-collagen contrast, apart from minor features associated with the internal collagen structure.

Localization of DLL1- and NICD-positive osteoblasts in cortical bone during postnatal growth in rats.

The long bone midshaft expands by forming primary osteons at the periosteal surface of cortical bone in humans and rodents. Osteoblastic bone formation in the vascular cavity in the center of primary osteons is delayed during cortical bone development. The mechanisms of the formation of primary osteons is not fully understood, however. Focusing on NOTCH1 signaling, an inhibitory signaling on osteoblastic bone formation, our immunohistochemical analysis revealed Delta like1 (DLL1), a ligand of NOTCH1, and the NOTCH1 intracellular domain (NICD, an activated form of NOTCH1) immunoreactivity, in the cuboidal osteoblasts lining the bone surface in the vascular cavity of primary osteons during postnatal growth in rats. Interestingly, five days after treatment of primary osteoblasts with ascorbic acid and ß glycerophosphate, protein levels of both DLL1 and NICD increased transiently, indicating that DLL1 activates NOTCH1 in primary cultured osteoblasts. Thus, the results imply that DLL1-NOTCH1 signaling in osteoblasts is associated with primary osteonal bone formation.

Bone ongrowth and mechanical fixation of implants in cortical and cancellous bone.

BACKGROUND: What is the right surface for an implant to achieve biological fixation? Surface technologies can play important roles in encouraging interactions between the implant surface and the host bone to achieve osseointegration. Preclinical animal models provide important insight into in vivo performance related to bone ongrowth and implant fixation. METHODS: A large animal model was used to compare the in vivo response of HA and plasma-sprayed titanium coatings in a well-reported adult ovine model to evaluate bone ongrowth in terms of mechanical properties in cortical sites, and histology and histomorphometry in cortical and cancellous sites at 4 and 12 weeks. RESULTS: Titanium plasma-sprayed surfaces outperformed the HA-coated samples in push-out testing in cortical sites while both surfaces supported new bone ongrowth and remodeling in cortical and cancellous sites. CONCLUSIONS: While both HA and Ti plasma provided an osteoconductive surface for bone ongrowth, the Ti plasma provided a more robust bone-implant interface that ideally would be required for load transfer and implant stability in the longer term.

Skeletal Stem Cells for Bone Development and Repair: Diversity Matters.

PURPOSE OF REVIEW: Skeletal stem cells (SSCs) are considered to play important roles in bone development and repair. These cells have been historically defined by their in vitro potential for self-renewal and differentiation into "trilineage" cells; however, little is known about their in vivo identity. Here, we discuss recent progress on SSCs and how they potentially contribute to bone development and repair. RECENT FINDINGS: Bone is composed of diverse tissues, which include cartilage and its perichondrium, cortical bone and its periosteum, and bone marrow and its trabecular bone and stromal compartment. We are now at the initial stage of understanding the precise identity of SSCs in each bone tissue. The emerging concept is that functionally dedicated SSCs are encased by their own unique cellular and extracellular matrix microenvironment, and locally support its own compartment. Diverse groups of SSCs are likely to work in concert to achieve development and repair of the highly functional skeletal organ.

Effect of cortical bone micro-structure in fragility fracture patients on lamellar stress.

This work investigates how changes in cortical bone microstructure alter the risk of fragility fractures. The secondary osteons of non-osteoporotic (by DXA) women with fragility fractures have reduced lamellar width and greater areas of birefringent brightness in transverse sections, a pathological condition. We used hierarchical finite element (FE) models of the proximal femur of two women aged 67 and 88 (younger and older) during one-legged stance. At specific locations of the anterior-inferior neck (ROI), we analyzed micro-models containing osteons comprised of alternating birefringent extinct and bright lamellae. The plane of lamellar isotropy (XY) was transverse to the osteon longitudinal axis (Z) which was parallel to the femoral neck axis. To evaluate changes in fracture risk with changes in microstructure, we investigated principal and von Mises stresses, and planar stress measures that accounted for transverse isotropy. For both younger and older femurs, 48% to 100% of stress measures were larger in models with healthy architecture than in models with pathological architecture, while controlling for type of lamella and osteon. These findings suggest that bone adaptation reduces stress at most pathological lamellar sites. However, in the bright lamellae of the younger femur, the pathological tensile, compressive and distortional stresses in the transverse plane and distortional stress in the longitudinal planes were larger than the non-negligible corresponding stresses in 6 of the 28 osteon models with healthy architecture, in 5 of the 7 locations. Therefore, a minority of sites with pathological architecture present greater stress, and therefore, greater fracture risk.

What is the influence of two strain rates on the relationship between human cortical bone toughness and micro-structure?

Cortical bone fracture mechanisms are well studied under quasi-static loading. The influence of strain rate on crack propagation mechanisms needs to be better understood, however. We have previously shown that several aspects of the bone micro-structure are involved in crack propagation, such as the complete porosity network, including the Haversian system and the lacunar network, as well as biochemical aspects, such as the maturity of collagen cross-links. The aim of this study is to investigate the influence of strain rate on the toughness of human cortical bone with respect to its microstructure and organic non-collagenous composition. Two strain rates will be considered: quasi-static loading (10-4 s-1), a standard condition, and a higher loading rate (10-1 s-1), representative of a fall. Cortical bone samples were extracted from eight female donors (age 50-91 years). Three-point bending tests were performed until failure. Synchrotron radiation micro-computed tomography imaging was performed to assess bone microstructure including the Haversian system and the lacunar system. Collagen enzymatic cross-link maturation was measured using a high performance liquid chromatography column. Results showed that that under quasi-static loading, the elastic contribution of the fracture process is correlated to both the collagen cross-links maturation and the microstructure, while the plastic contribution is correlated only to the porosity network. Under fall-like loading, bone organization appears to be less linked to crack propagation.

Woven bone overview: structural classification based on its integral role in developmental, repair and pathological bone formation throughout vertebrate groups.

Cortical bone development is characterised by initial formation of woven bone followed by deposition of lamellar bone on the woven scaffold. This occurs in normal bone formation as an integral obligate self-assembly pattern throughout all vertebrate groups, with specific temporal and spatial features. It also occurs in repair bone, modified by the biophysical/mechanical environment, and in pathological bone, modified by the specific disorder and its severity. Two spatially distinct osteoblast cell populations synthesise woven and lamellar bone: mesenchymal osteoblasts surround themselves circumferentially with collagen in a random array to form woven bone; surface osteoblasts align themselves in a linear array on the woven bone surface (or adjacent lamellar bone) to synthesise parallel-fibred lamellar bone. Four specific stages of woven bone formation are defined: stage I, early differentiation of pre-osteoblasts from undifferentiated mesenchymal cells; stage II, mesenchymal osteoblasts surrounding themselves in a 360° arc with randomly oriented matrix fibres; stage III, woven matrix acting as a scaffold on which surface osteoblasts begin to synthesise bone in parallel-fibred lamellar conformation; stage IV, progressive relative diminution of woven bone in the woven bone/lamellar bone complex. Stages II and IV are further subdivided (in a, b and c) by shifting cell area/matrix area and woven bone/lamellar bone relationships. The under-appreciated biological significance of woven bone is that it initiates formation de novo at sites of no previous bone. This information allows for targeted assessment of molecular-biophysical mechanisms underlying woven bone formation and their utilisation for initiating enhanced bone formation.

Bone microstructure of mice after prolonged taurine treatment.

Taurine, a sulphur - containing amino acid, has been termed a functional nutrient. Its synthetic form is a common ingredient in supplements and energy drinks. There is no information concerning taurine impact on bone microstructure after prolonged supplemental use. Also, differences in bone parameters of mice following taurine exposure are unknown. In this study, a detailed microstructure of compact and trabecular bone tissues of mice subchronically exposed to taurine was determined. Animals (n=12) were segregated into three groups: E1 group - mice received 20 mg/kg b.w. of taurine per day during 8 weeks; E2 group - mice were fed by taurine at a dose of 40 mg/kg b.w. for 8 weeks and a control (C) group. Decreased density of secondary osteons, increased sizes of primary osteon's vascular canals (P<0.05) were observed in taurine - treated animals. Cortical bone thickness, trabecular thickness were decreased (P<0.05) in E1 group, and relative volume of trabecular bone was lower (P<0.05) in E2 group as compared to C group. According to our results, prolonged taurine exposure at the doses used in this study can negatively affect both compact and trabecular bone tissues microstructure.

No Signature of Osteocytic Osteolysis in Cortical Bone from Lactating NMRI Mice.

The roles of osteocytes in bone homeostasis have garnered increasing attention since it has been realized that osteocytes communicate with other organs. It has long been debated whether and/or to which degree osteocytes can break down the bone matrix surrounding them in a process called osteocytic osteolysis. Osteocytic osteolysis has been indicated to be induced by a number of skeletal challenges including lactation in CD1 and C57BL/6 mice, whereas immobilization-induced osteocytic osteolysis is still a matter of controversy. Motivated by the wish to understand this process better, we studied osteocyte lacunae in lactating NMRI mice, which is a widely used outbred mouse strain. Surprisingly, no trace of osteocytic osteolysis could be detected in tibial or femoral cortical bone either by 3D investigation by synchrotron nanotomography, by studies of lacunar cross-sectional areas using scanning electron microscopy, or by light microscopy. These results lead us to conclude that osteocytic osteolysis does not occur in NMRI mice as a response to lactation, in turn suggesting that osteocytic osteolysis may not play a generic role in mobilizing calcium during lactation.

Canalicular Junctions in the Osteocyte Lacuno-Canalicular Network of Cortical Bone.

The osteocyte lacuno-canalicular network (LCN) is essential for bone remodeling because osteocytes regulate cell recruitment. This has been proposed to occur through liquid-flow-induced shear forces in the canaliculi. Models of the LCN have thus far assumed that it contains canaliculi connecting the osteocyte lacunae. However, here, we reveal that enlarged spaces occur at places where several canaliculi cross; we name these spaces canalicular junctions. We characterize them in detail within mice cortical bone using synchrotron nanotomography at two length scales, with 50 and 130 nm voxel size, and show that canalicular junctions occur at a density similar to that of osteocyte lacunae and that canalicular junctions tend to cluster. Through confocal laser scanning microscopy, we show that canalicular junctions are widespread as we have observed them in cortical bone from several species, even though the number density of the canalicular junctions was not universal. Fluid flow simulations of a simple model system with and without a canalicular junction clearly show that liquid mass transport and flow velocities are altered by the presence of canalicular junctions. We suggest that these canalicular junctions may play an important role in osteocyte communication and possibly also in canalicular fluid flow. Therefore, we believe that they constitute an important component in the bone osteocyte network.

Ageing characteristics of bone indicated by transcriptomic and exosomal proteomic analysis of cortical bone cells.

BACKGROUND: Degenerative changes in the skeleton play an important role in ageing. As the foremost sensors and orchestrators of bone remodelling, osteocytes contribute significantly to the health of the skeleton. Embedded in a mineralized bone matrix, the osteocyte network and the surrounding lacunar canaliculae work together as a functional syncytium-the osteocytic lacunar-canalicular system (OLCS). However, changes in the OLCS during ageing and related mechanisms cannot be fully understood by using traditional histological analysis. METHODS: To link the phenotypes of aged osteocytes and their functional changes during ageing, we analysed the changes in the gene expression profiles of bone cells and the proteomic profiles of OLCS exosomes derived from aged and young cortical bone. RESULTS: Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of differentially expressed genes (DEGs) suggested that a decline in cell energy metabolism and an increased level of the proinflammatory state are major characteristics of bone ageing. Moreover, some DEGs were key regulators of bone mechanical sensation and bone remodelling, which are indicative of reduced bone-specific function with age. Further, the identified proteins in OLCS exosomes showed potential changes in the secretory function bone. Compared with young controls, the decreased functional proteins in aged OLCS exosomes were enriched mainly in GO terms that included regulating bone development and remodelling, cell-matrix adhesion, and cell clearance and homeostasis. Notably, several functions of exosomal proteins of the aged group revealed potential new roles, such as regulating innate and adaptive immunity, wound healing, and angiogenesis and eliminating oxidative stress. CONCLUSION: The information obtained from bone cells and OLCS exosomes will help us discover new features of bone ageing.

The in vitro behaviour of canine osteoblasts derived from different bone types.

BACKGROUND: Our understanding of the biology of osteoblasts is important as they underpin bone remodelling, fracture healing and processes such as osseointegration. Osteoblasts isolated from human humeral samples display distinctive biological activity in vitro, which relates to the samples' bone types (subchondral (S), trabecular (T), cortical (C)). Our aim was to isolate primary osteoblast cultures from different bone types from the proximal femur of a clinical population of dogs presented for total hip replacement and compare the behaviour of the osteoblasts derived from different bone types, to identify a preferred bone type for isolation. RESULTS: No differences were found for osteoblast doubling time (median for S = 2.9, T = 3.1 and C = 2.71 days, respectively; p = 0.33), final cell number (median for S = 54,849, T = 49,733, C = 61,390 cells/cm2; p = 0.34) or basal tissue non-specific alkaline phosphatase (TNAP) activity (median for S = 0.02, T = 0.02, C = 0.03 U/min/mg protein; p = 0.81) between bone types after 6 days of culture in basal media. There were no differences in mineralizing TNAP activity (S = 0.02, T = 0.02, C = 0.03 U/min/mg protein, p = 0.84) or in mineralized area (S = 0.05, T = 0.04, C = 0.04%, p = 0.92) among cells from different bone types. CONCLUSIONS: There is no significant difference in mean doubling time, basal or mineralizing TNAP activity or mineralized area in osteoblasts derived from subchondral, cortical, or trabecular bone types from the canine femoral head. However, there appears to be a high level of inter-animal variability in the studied parameters, which was independent of age, body mass, and sex. Trabecular isolate osteoblasts have the least variation of the bone types studied, and therefore should be considered a preferred source for primary osteoblast cultures. The work here provides baselines for canine osteoblast function, which has utility for future comparative studies.

Histomorphometry and cortical robusticity of the adult human femur.

Recent quantitative analyses of human bone microanatomy, as well as theoretical models that propose bone microstructure and gross anatomical associations, have started to reveal insights into biological links that may facilitate remodeling processes. However, relationships between bone size and the underlying cortical bone histology remain largely unexplored. The goal of this study is to determine the extent to which static indicators of bone remodeling and vascularity, measured using histomorphometric techniques, relate to femoral midshaft cortical width and robusticity. Using previously published and new quantitative data from 450 adult human male (n = 233) and female (n = 217) femora, we determine if these aspects of femoral size relate to bone microanatomy. Scaling relationships are explored and interpreted within the context of tissue form and function. Analyses revealed that the area and diameter of Haversian canals and secondary osteons, and densities of secondary osteons and osteocyte lacunae from the sub-periosteal region of the posterior midshaft femur cortex were significantly, but not consistently, associated with femoral size. Cortical width and bone robusticity were correlated with osteocyte lacunae density and scaled with positive allometry. Diameter and area of osteons and Haversian canals decreased as the width of cortex and bone robusticity increased, revealing a negative allometric relationship. These results indicate that microscopic products of cortical bone remodeling and vascularity are linked to femur size. Allometric relationships between more robust human femora with thicker cortical bone and histological products of bone remodeling correspond with principles of bone functional adaptation. Future studies may benefit from exploring scaling relationships between bone histomorphometric data and measurements of bone macrostructure.

Quantifying proliferative and surface marker heterogeneity in colony-founding connective tissue progenitors and their progeny using time-lapse microscopy.

Connective tissue progenitors (CTPs) are defined as the heterogeneous population of tissue-resident stem and progenitor cells that are capable of proliferating and differentiating into connective tissue phenotypes. The prevalence and variation in clonal progeny of CTPs can be characterized using a colony formation assay. However, colony assays do not directly assess the characteristics of the colony-founding CTP. We performed large, field-of-view, time-lapse microscopy to manually track colonies back to the founding cells. Image processing and analysis was used to characterize the colonies and their founding cells. We found that the traditional colony-forming unit (CFU) assay underestimates the number of founding cells as colonies can be formed by more than one founding cell. After 6 days in culture, colonies do not completely express CD73, CD90, and CD105. Heterogeneity in colony cells was characterized by two cell populations, proliferative and spread cells. Regression modelling of duration of lag phase and doubling time by cell marker suggests the presence of CD90 and CD105 in CTP subpopulations with different proliferative capabilities. From mathematical modelling of clonal colonies, we quantitatively characterized proliferation, migration, and cell marker expression rates to identify desirable clones for selection. Direct assessment of colony formation parameters led to more accurate assessment of CFU heterogeneity. Furthermore, these parameters can be used to quantify the diversity and hierarchy of stem and progenitor cells from a cell source or tissue for tissue engineering applications.

Trop2 Guarantees Cardioprotective Effects of Cortical Bone-Derived Stem Cells on Myocardial Ischemia/Reperfusion Injury.

Stem cell transplantation represents a promising therapeutic approach for myocardial ischemia/reperfusion (I/R) injury, where cortical bone-derived stem cells (CBSCs) stand out and hold superior cardioprotective effects on myocardial infarction than other types of stem cells. However, the molecular mechanism underlying CBSCs function on myocardial I/R injury is poorly understood. In a previous study, we reported that Trop2 (trophoblast cell-surface antigen 2) is expressed exclusively on the CBSCs membrane, and is involved in regulation of proliferation and differentiation of CBSCs. In this study, we found that the Trop2 is essential for the ameliorative effects of CBSCs on myocardial I/R-induced heart damage via promoting angiogenesis and inhibiting cardiomyocytes apoptosis in a paracrine manner. Trop2 is required for the colonization of CBSCs in recipient hearts. When Trop2 was knocked out, CBSCs largely lost their functions in lowering myocardial infarction size, improving heart function, enhancing capillary density, and suppressing myocardial cell death. Mechanistically, activating the AKT/GSK3ß/ß-Catenin signaling axis contributes to the essential role of Trop2 in CBSCs-rendered cardioprotective effects on myocardial I/R injury. In conclusion, maintaining the expression and/or activation of Trop2 in CBSCs might be a promising strategy for treating myocardial infarction, I/R injury, and other related heart diseases.

Unfocused shockwaves for osteoinduction in bone substitutes in rat cortical bone defects.

Bone substitutes are frequently used in clinical practice but often exhibit limited osteoinductivity. We hypothesized that unfocused shockwaves enhance the osteoinductivity of bone substitutes and improve osteointegration and angiogenesis. Three different bone substitutes, namely porous tricalcium phosphate, porous hydroxyapatite and porous titanium alloy, were implanted in a critical size (i.e. 6-mm) femoral defect in rats. The femora were treated twice with 1500 shockwaves at 2 and 4 weeks after surgery and compared with non-treated controls. The net volume of de novo bone in the defect was measured by microCT-scanning during 11-weeks follow-up. Bone ingrowth and angiogenesis in the bone substitutes was examined at 5 and 11 weeks using histology. It was shown that hydroxyapatite and titanium both had an increase of bone ingrowth with more bone in the shockwave group compared to the control group, whereas resorption was seen in tricalcium phosphate bone substitutes over time and this was insensitive to shockwave treatment. In conclusion, hydroxyapatite and titanium bone substitutes favour from shockwave treatment, whereas tricalcium phosphate does not. This study shows that osteoinduction and osteointegration of bone substitutes can be influenced with unfocused shockwave therapy, but among other factors depend on the type of bone substitute, likely reflecting its mechanical and biological properties.