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Abstract: Bone marrow derived mesenchymal stem/stromal cells (MSC) contribute to skeletal tissue formation and the regulation of haematopoiesis. The Eph/ephrin family of receptor tyrosine kinases is potentially important in the maintenance of the stem cell niche within neural, intestinal and dental tissues and has recently been shown to play a role in regulating bone homeostasis. However, the contribution of EphB/ephrin-B molecules in human MSC function remains to be determined. In the present study, EphB and ephrin-B molecules were expressed by ex vivo expanded human MSC populations and within human bone marrow trephine samples. To elucidate the contribution of EphB/ephrin-B molecules in MSC recruitment, we performed functional spreading and migration assays and showed that reverse ephrin-B signalling inhibited MSC attachment and spreading by activating Src-, PI3Kinase- and JNK-dependent signalling pathways. In contrast, forward EphB2 signalling promoted MSC migration by activating the Src kinase- and Abl-dependent signalling pathways. Furthermore, activation of ephrin-B1 and/or ephrin-B2 molecules expressed by MSC was found to increase osteogenic differentiation, while ephrin-B1 activation promoted chondrogenic differentiation. These observations suggest that EphB/ephrin-B interactions may mediate the recruitment, migration and differentiation of MSC during bone repair. [Copyright &y& Elsevier]

Four decades after the first isolation and characterization of clonogenic bone marrow stromal cells or mesenchymal stem cells (MSC) in the laboratory of Dr. Alexander Friedenstien, the therapeutic application of their progeny following ex vivo expansion are only now starting to be realized in the clinic. The multipotency, paracrine effects, and immune-modulatory properties of MSC present them as an ideal stem cell candidate for tissue engineering and regenerative medicine. In recent years it has come to light that MSC encompass plasticity that extends beyond the conventional bone, adipose, cartilage, and other skeletal structures, and has expanded to the differentiation of liver, kidney, muscle, skin, neural, and cardiac cell lineages. This review will specifically focus on the skeletal regenerative capacity of bone marrow derived MSC alone or in combination with growth factors, biocompatible scaffolds, and following genetic modification. J. Cell. Physiol. 218: 237–245, 2009. © 2008 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

The chemokine CXCL12 (variously termed stromal-derived factor 1 or B cell-stimulating factor) is a highly conserved chemotactic cytokine belonging to the large family of CXC chemokines. CXCL12 has crucial roles in the formation of multiple organ systems during embryogenesis and in the regulation of bone marrow haematopoiesis and immune function in the postnatal organism. Although considered an important factor in normal bone metabolism, recent studies implicate CXCL12 in the pathogenesis of several diseases involving the skeleton, including rheumatoid arthritis and cancers that metastasize to bone. Recent studies have highlighted an emerging role for CXCL12 in the processes of physiological and pathological bone remodelling. [Copyright &y& Elsevier]

Examines the molecular and cellular characterization of highly purified human bone marrow stromal stem cells. Use of cell separation techniques; Absence of phenotypic characteristics of leukocytes and mature stromal elements; Expression of elomerase activity in vivo.

Simple Summary: Like in solid cancers, the process of dissemination is a critical feature of disease progression in the blood cancer multiple myeloma. At diagnosis, myeloma patients have cancer that has spread throughout the bone marrow, with patients with more disseminatory myeloma having worse outcomes for their disease. In this review, we discuss the current understanding of the mechanisms that underpin the dissemination process in multiple myeloma. Furthermore, we discuss the potential for the use of therapies that target the dissemination process as a novel means of improving outcomes for multiple myeloma patients. Multiple myeloma (MM) is a plasma cell (PC) malignancy characterised by the presence of MM PCs at multiple sites throughout the bone marrow. Increased numbers of peripheral blood MM PCs are associated with rapid disease progression, shorter time to relapse and are a feature of advanced disease. In this review, the current understanding of the process of MM PC dissemination and the extrinsic and intrinsic factors potentially driving it are addressed through analysis of patient-derived MM PCs and MM cell lines as well as mouse models of homing and dissemination. In addition, we discuss how patient cytogenetic subgroups that present with highly disseminated disease, such as t(4;14), t(14;16) and t(14;20), suggest that intrinsic properties of MM PC influence their ability to disseminate. Finally, we discuss the possibility of using therapeutic targeting of tumour dissemination to slow disease progression and prevent overt relapse. [ABSTRACT FROM AUTHOR]

The side effects of cancer therapy continue to cause significant health and cost burden to the patient, their friends and family, and governments. A major barrier in the way in which these side effects are managed is the highly siloed mentality that results in a fragmented approach to symptom control. Increasingly, it is appreciated that many symptoms are manifestations of common underlying pathobiology, with changes in the gastrointestinal environment a key driver for many symptom sequelae. Breakdown of the mucosal barrier (mucositis) is a common and early side effect of many anti-cancer agents, known to contribute (in part) to a range of highly burdensome symptoms such as diarrhoea, nausea, vomiting, infection, malnutrition, fatigue, depression, and insomnia. Here, we outline a rationale for how, based on its already documented effects on the gastrointestinal microenvironment, medicinal cannabis could be used to control mucositis and prevent the constellation of symptoms with which it is associated. We will provide a brief update on the current state of evidence on medicinal cannabis in cancer care and outline the potential benefits (and challenges) of using medicinal cannabis during active cancer therapy. [ABSTRACT FROM AUTHOR]

[Display omitted] • The neurobiology of the developing brain is unique. • This may leave the paediatric brain more vulnerable to chemotherapy-induced cognitive impairment. • Age-matched preclinical models are necessary to develop relevant findings for childhood cancer survivors. Childhood is recognised as a period of immense physical and emotional development, and this, in part, is driven by underlying neurophysiological transformations. These neurodevelopmental processes are unique to the paediatric brain and are facilitated by augmented rates of neuroplasticity and expanded neural stem cell populations within neurogenic niches. However, given the immaturity of the developing central nervous system, innate protective mechanisms such as neuroimmune and antioxidant responses are functionally naïve which results in periods of heightened sensitivity to neurotoxic insult. This is highly relevant in the context of paediatric cancer, and in particular, the neurocognitive symptoms associated with treatment, such as surgery, radio- and chemotherapy. The vulnerability of the developing brain may increase susceptibility to damage and persistent symptomology, aligning with reports of more severe neurocognitive dysfunction in children compared to adults. It is therefore surprising, given this intensified neurocognitive burden, that most of the pre-clinical, mechanistic research focuses exclusively on adult populations and extrapolates findings to paediatric cohorts. Given this dearth of age-specific research, throughout this review we will draw comparisons with neurodevelopmental disorders which share comparable pathways to cancer treatment related side-effects. Furthermore, we will examine the unique nuances of the paediatric brain along with the somatic systems which influence neurological function. In doing so, we will highlight the importance of developing in vitro and in vivo paediatric disease models to produce age-specific discovery and clinically translatable research. [ABSTRACT FROM AUTHOR]

Summary: Expression of myeloperoxidase (MPO), a key inflammatory enzyme restricted to myeloid cells, is negatively associated with the development of solid tumours. Activated myeloid cell populations are increased in multiple myeloma (MM); however, the functional consequences of myeloid‐derived MPO within the myeloma microenvironment are unknown. Here, the role of MPO in MM pathogenesis was investigated, and the capacity for pharmacological inhibition of MPO to impede MM progression was evaluated. In the 5TGM1‐KaLwRij mouse model of myeloma, the early stages of tumour development were associated with an increase in CD11b+ myeloid cell populations and an increase in Mpo expression within the bone marrow (BM). Interestingly, MM tumour cell homing was increased towards sites of elevated myeloid cell numbers and MPO activity within the BM. Mechanistically, MPO induced the expression of key MM growth factors, resulting in tumour cell proliferation and suppressed cytotoxic T‐cell activity. Notably, tumour growth studies in mice treated with a small‐molecule irreversible inhibitor of MPO (4‐ABAH) demonstrated a significant reduction in overall MM tumour burden. Taken together, our data demonstrate that MPO contributes to MM tumour growth, and that MPO‐specific inhibitors may provide a new therapeutic strategy to limit MM disease progression. [ABSTRACT FROM AUTHOR]

Background: This study aimed to examine the performance of machine learning algorithms for the prediction of discharge within 12 and 24 h to produce a measure of readiness for discharge after general surgery. Methods: Consecutive general surgery patients at two tertiary hospitals, over a 2‐year period, were included. Observation and laboratory parameter data were stratified into training, testing and validation datasets. Random forest, XGBoost and logistic regression models were evaluated. Each ward round note time was taken as a different event. Primary outcome was classification accuracy of the algorithmic model able to predict discharge within the next 12 h on the validation data set. Results: 42 572 ward round note timings were included from 8826 general surgery patients. Discharge occurred within 12 h for 8800 times (20.7%), and within 24 h for 9885 (23.2%). For predicting discharge within 12 h, model classification accuracies for derivation and validation data sets were: 0.84 and 0.85 random forest, 0.84 and 0.83 XGBoost, 0.80 and 0.81 logistic regression. For predicting discharge within 24 h, model classification accuracies for derivation and validation data sets were: 0.83 and 0.84 random forest, 0.82 and 0.81 XGBoost, 0.78 and 0.79 logistic regression. Algorithms generated a continuous number between 0 and 1 (or 0 and 100), representing readiness for discharge after general surgery. Conclusions: A derived artificial intelligence measure (the Adelaide Score) successfully predicts discharge within the next 12 and 24 h in general surgery patients. This may be useful for both treating teams and allied health staff within surgical systems. [ABSTRACT FROM AUTHOR]

Saethre–Chotzen syndrome (SCS), associated with TWIST‐1 mutations, is characterized by premature fusion of cranial sutures. TWIST‐1 haploinsufficiency, leads to alterations in suture mesenchyme cellular gene expression patterns, resulting in aberrant osteogenesis and craniosynostosis. We analyzed the expression of the TWIST‐1 target, Tyrosine kinase receptor c‐ros‐oncogene 1 (C‐ROS‐1) in TWIST‐1 haploinsufficient calvarial cells derived from SCS patients and calvaria of Twist‐1del/+ mutant mice and found it to be highly expressed when compared to TWIST‐1 wild‐type controls. Knock‐down of C‐ROS‐1 expression in TWIST‐1 haploinsufficient calvarial cells derived from SCS patients was associated with decreased capacity for osteogenic differentiation in vitro. Furthermore, treatment of human SCS calvarial cells with the tyrosine kinase chemical inhibitor, Crizotinib, resulted in reduced C‐ROS‐1 activity and the osteogenic potential of human SCS calvarial cells with minor effects on cell viability or proliferation. Cultured human SCS calvarial cells treated with Crizotinib exhibited a dose‐dependent decrease in alkaline phosphatase activity and mineral deposition, with an associated decrease in expression levels of Runt‐related transcription factor 2 and OSTEOPONTIN, with reduced PI3K/Akt signalling in vitro. Furthermore, Crizotinib treatment resulted in reduced BMP‐2 mediated bone formation potential of whole Twist‐1del/+ mutant mouse calvaria organotypic cultures. Collectively, these results suggest that C‐ROS‐1 promotes osteogenic differentiation of TWIST‐1 haploinsufficient calvarial osteogenic progenitor cells. Furthermore, the aberrant osteogenic potential of these cells is inhibited by the reduction of C‐ROS‐1. Therefore, targeting C‐ROS‐1 with a pharmacological agent, such as Crizotinib, may serve as a novel therapeutic strategy to alleviate craniosynostosis associated with aberrant TWIST‐1 function. [ABSTRACT FROM AUTHOR]

Multiple myeloma (MM) is a disease of older people, yet factors relating to comorbidity and frailty may threaten treatment tolerability for many of this heterogenous group. There has been increasing interest in defining specific and clinically relevant frailty assessment tools within the MM population, with the goal of using these frailty scores, not just as a prognostic instrument, but also as a predictive tool to allow for a frailty‐adapted treatment approach. This paper reviews the various frailty assessment frameworks used in the evaluation of patients with MM, including the International Myeloma Working Group Frailty Index (IMWG‐FI), the Mayo Frailty Index and the simplified frailty scale. While the IMWG‐FI remains the most widely accepted tool, the simplified frailty scale is the most user‐friendly in busy day‐to‐day clinics based on its ease of use. This paper summarises the recommendations from the Myeloma Scientific Advisory Group (MSAG) of Myeloma Australia, on the use of frailty assessment tools in clinical practice and proposes a frailty‐stratified treatment algorithm to aid clinicians in tailoring therapy for this highly heterogeneous patient population. [ABSTRACT FROM AUTHOR]

Background Context: Neural compression associated with lumbar disc herniation is usually managed surgically by microdiscectomy. However, 10%-20% of patients re-present with debilitating back pain, and approximately 15% require further surgery.Purpose: Using an ovine model of microdiscectomy, the present study investigated the relative potential of pentosan polysulfate-primed mesenchymal progenitor cells (pMPCs) or MPC alone implanted into the lesion site to facilitate disc recovery.Study Design: An ovine model of lumbar microdiscectomy was used to compare the relative outcomes of administering MPCs or pMPCs to the injury site postsurgery.Methods: At baseline 3T magnetic resonance imaging (MRI) of 18 adult ewes was undertaken followed by annular microdiscectomy at two lumbar disc levels. Sheep were randomized into three groups (n=6). The injured controls received no further treatment. Defects of the treated groups were implanted with a collagen sponge and MPC (5×105 cells) or pMPC (5×105 cells). After 6 months, 3T MRI and standard radiography were performed. Spinal columns were dissected, individual lumbar discs were sectioned horizontally, and nucleus pulposus (NP) and annulus fibrosus (AF) regions were assessed morphologically and histologically. The NP and AF tissues were dissected into six regions and analyzed biochemically for their proteoglycans (PGs), collagen, and DNA content.Results: Both the MPC- and pMPC-injected groups exhibited less reduction in disc height (p<.05) and lower Pfirrmann grades (p≤.001) compared with the untreated injury controls, but morphologic scores for the pMPC-injected discs were lower (p<.05). The PG content of the AF injury site region (AF1) of pMPC discs was higher than MPC and injury control AF1 (p<.05). At the AF1 and contralateral AF2 regions, the DNA content of pMPC discs was significantly lower than injured control discs and MPC-injected discs. Histologic and birefringent microscopy revealed increased structural organization and reduced degeneration in pMPC discs compared with MPC and the injured controls.Conclusions: In an ovine model 6 months after administration of pMPCs to the injury site disc PG content and matrix organization were improved relative to controls, suggesting pMPCs' potential as a postsurgical adjunct for limiting progression of disc degeneration after microdiscectomy. [ABSTRACT FROM AUTHOR]

The proliferation, differentiation, adhesion, and migration of hematopoietic stem and progenitor cells (HSPCs) are dependent upon bone marrow stromal cells (BMSCs). In this study, we found that human primitive HSPCs (CD34 + CD38 − ), but not lineage-committed hematopoietic cell populations, express the tyrosine kinase receptors EphA5 and EphA7. Moreover, we found that the ephrinA5 ligand, the high-affinity binding partner of EphA5 and EphA7, is highly expressed by primary human BMSCs. Previous studies have reported that interactions between EphA and ephrinA play important roles in hematopoietic cell trafficking; however, their role in BMSC support of hematopoiesis had not been described previously. Herein, we show that stimulating EphA5 and/or EphA7 forward signaling in HSPCs using soluble ephrinA5-Fc molecules promoted human HSPC-derived colony formation significantly and was associated with increased expression of granulocyte macrophage colony-stimulating factor receptor on HSPCs. Studies using functional blocking peptides to EphA5/7 found that disruption of EphA5/ephrinA5 and/or EphA7/ephrinA5 interactions inhibited HSPC function in BMSC-dependent long-term culture-initiating cell assays. Furthermore, the adhesion and migration of HSPCs was increased significantly in the presence of ephrinA5-Fc molecules compared with human immunoglobulin G-treated controls. Conversely, blocking EphA5 activation led to a reduction of HSPC adhesion, whereas inhibiting EphA5 and/or EphA7 activation hindered HSPC migration. Analysis of HSPC cultured in the presence of ephrinA5-Fc showed that EphA forward signaling stimulated Rac1 gene and protein expression and the Rac1 target molecule WAVE1. Moreover, a significant reduction of ephrinA5-mediated HSPC adhesion and migration was observed in the presence of Rac1 inhibitor. These findings suggest that interactions between EphA and ephrinA5 are important in maintaining the HSPC niche mediated in part by activation of Rac1 signaling. [ABSTRACT FROM AUTHOR]

The TWIST-1 gene encodes a basic helix-loop-helix (bHLH) transcription factor important in mediating skeletal and head mesodermal tissue development. Bone marrow-derived mesenchymal stem/stromal cells (BMSC), express high levels of TWIST-1, which is down regulated during ex vivo expansion. Cultured BMSC over-expressing TWIST-1 display decreased capacity for osteogenic differentiation and an enhanced capacity to undergo adipogenesis, suggesting that TWIST-1 is a mediator of lineage commitment. However, little is known regarding the mechanism(s) by which TWIST-1 mediates cell fate determination. In this study, microarray analysis was used to identify a novel downstream TWIST-1 target, tyrosine kinase receptor c-ros-oncogene 1 (C-ROS-1), which was down regulated in TWIST-1 over-expressing BMSC. Chromatin immunoprecipitation analysis showed that TWIST-1 directly bound to two E -box binding sites on the proximal C-ROS-1 promoter. Knock-down of C-ROS-1 in human BMSC and cranial bone cells resulted in a decreased capacity for osteogenic differentiation in vitro. Conversely, suppression of C-ROS-1 in BMSC resulted in an enhanced capacity to undergo adipogenesis. Furthermore, reduced C-ROS-1 levels led to activation of different components of the PI3K/AKT/mTORC1 signalling pathway during osteogenic and adipogenic differentiation. Collectively, these data suggest that C-ROS-1 is involved in BMSC fate switching between osteogenesis and adipogenesis, mediated via PI3K/AKT/mTORC1 signalling. [ABSTRACT FROM AUTHOR]

Prophylaxis and the treatment of surgical site infections (SSIs) with antibiotics frequently fail due to the antibiotic resistance of bacteria and the ability of bacteria to reside in biofilms (i.e., bacterial clusters in a protective matrix). Therefore, alternative antibacterial treatments are required to combat biofilm infections. The combination of diethyldithiocarbamate (DDC−) and copper ions (Cu2+) exhibited antibiofilm activity against the staphylococci species associated with SSIs; however, the formation of a water-insoluble Cu(DDC)2 complex limits its application to SSIs. Here, we describe the development and antibiofilm activity of an injectable gel containing a liposomal formulation of Cu(DDC)2 and Cu2+ (lipogel). Lyophilized liposomes were incorporated into a mixture of chitosan (CS) and beta-glycerophosphate (βGP), and the thermosensitive gelling properties of CS-βGP and the lipogel were determined. The liposomes remained stable after lyophilization over six months at 4–6 °C and −20 °C. The sol-gel transition of the gel and lipogel occurred between 33 and 39 °C, independently of sterilization or storage at −20 °C. CS-βGP is biocompatible and the liposomes were released over time. The lipogel prevented biofilm formation over 2 days and killed 98.7% of the methicillin-resistant Staphylococcus aureus and 99.9% of the Staphylococcus epidermidis biofilms. Therefore, the lipogel is a promising new prophylaxis and treatment strategy for local application to SSIs. [ABSTRACT FROM AUTHOR]

Multiple myeloma (MM) is an incurable haematological malignancy, caused by the uncontrolled proliferation of plasma cells within the bone marrow (BM). Obesity is a known risk factor for MM, however, few studies have investigated the potential of dietary intervention to prevent MM progression. Calorie restriction (CR) is associated with many health benefits including reduced cancer incidence and progression. To investigate if CR could reduce MM progression, dietary regimes [30% CR, normal chow diet (NCD), or high fat diet (HFD)] were initiated in C57BL/6J mice. Diet-induced changes were assessed, followed by inoculation of mice with Vk*MYC MM cells (Vk14451-GFP) at 16 weeks of age. Tumour progression was monitored by serum paraprotein, and at endpoint, BM and splenic tumour burden was analysed by flow cytometry. 30% CR promoted weight loss, improved glucose tolerance, increased BM adiposity and elevated serum adiponectin compared to NCD-fed mice. Despite these metabolic changes, CR had no significant effect on serum paraprotein levels. Furthermore, endpoint analysis found that dietary changes were insufficient to affect BM tumour burden, however, HFD resulted in an average two-fold increase in splenic tumour burden. Overall, these findings suggest diet-induced BM changes may not be key drivers of MM progression in the Vk14451-GFP transplant model of myeloma. [ABSTRACT FROM AUTHOR]

The proteasome inhibitor bortezomib is one of the primary therapies used for the haematological malignancy multiple myeloma (MM). However, intrinsic or acquired resistance to bortezomib, via mechanisms that are not fully elucidated, is a barrier to successful treatment in many patients. Our previous studies have shown that elevated expression of the chemokine receptor CCR1 in MM plasma cells in newly diagnosed MM patients is associated with poor prognosis. Here, we hypothesised that the poor prognosis conferred by CCR1 expression is, in part, due to a CCR1-mediated decrease in MM plasma cell sensitivity to bortezomib. In order to investigate the role of CCR1 in MM cells, CCR1 was knocked out in human myeloma cell lines OPM2 and U266 using CRISPR-Cas9. Additionally, CCR1 was overexpressed in the mouse MM cell line 5TGM1. The effect of bortezomib on CCR1 knockout or CCR1-overexpressing cells was then assessed by WST-1 assay, with or without CCL3 siRNA knockdown or addition of recombinant human CCL3. NSG mice were inoculated intratibially with OPM2-CCR1KO cells and were treated with 0.7 mg/kg bortezomib or vehicle twice per week for 3 weeks and GFP+ tumour cells in the bone marrow were quantitated by flow cytometry. The effect of CCR1 overexpression or knockout on unfolded protein response pathways was assessed using qPCR for ATF4 , HSPA5 , XBP1 , ERN1 and CHOP and Western blot for IRE1α and p-Jnk. Using CCR1 overexpression or CRIPSR-Cas9-mediated CCR1 knockout in MM cell lines, we found that CCR1 expression significantly decreases sensitivity to bortezomib in vitro , independent of the CCR1 ligand CCL3. In addition, CCR1 knockout rendered the human MM cell line OPM2 more sensitive to bortezomib in an intratibial MM model in NSG mice in vivo. Moreover, CCR1 expression negatively regulated the expression of the unfolded protein response receptor IRE1 and downstream target gene XBP1 , suggesting this pathway may be responsible for the decreased bortezomib sensitivity of CCR1-expressing cells. Taken together, these studies suggest that CCR1 expression may be associated with decreased response to bortezomib in MM cell lines. • Elevated expression of CCR1 in multiple myeloma cells is associated with poor prognosis in newly diagnosed patients • CCR1 expression in myeloma cell lines significantly decreases sensitivity to the proteasome inhibitor bortezomib in vitro • This effect is independent of endogenous expression of the CCR1 ligand CCL3 • CCR1 expression negatively regulated the expression of proteins involved in the unfolded protein response [ABSTRACT FROM AUTHOR]

The aim of this investigation wasto understand and engineer theinteractions between endothelial cells and the electrospun (ES) polyvinylidenefluoride-co-hexafluoropropylene (PVDF-HFP) nanofibersurfaces and evaluate their potential for endothelialization. ElastomericPVDF-HFP samples were electrospun to evaluate their potential useas small diameter artificial vascular graft scaffold (SDAVG) and comparedwith solvent cast (SC) PVDF-HFP films. We examined the consequencesof fibrinogen adsorption onto the ES and SC samples for endothelialisation.Bone marrow derived endothelial cells (BMEC) of human origin wereincubated with the test and control samples and their attachment,proliferation, and viability were examined. The nature of interactionof fibrinogen with SC and ES samples was investigated in detail usingELISA, XPS, and FTIR techniques. The pristine SC and ES PVDF-HFP samplesdisplayed hydrophobic and ultrahydrophobic behavior and accordingly,exhibited minimal BMEC growth. Fibrinogen adsorbed SC samples didnot significantly enhance endothelial cell binding or proliferation.In contrast, the fibrinogen adsorbed electrospun surfaces showed aclear ability to modulate endothelial cell behavior. This system alsorepresents an ideal model system that enables us to understand thenatural interaction between cells and their extracellular environment.The research reported shows potential of ES surfaces for artificialvascular graft applications. [ABSTRACT FROM AUTHOR]

Imaging modalities for multiple myeloma (MM) have evolved to enable earlier detection of disease. Furthermore, the diagnosis of MM requiring therapy has recently changed to include disease prior to bone destruction, specifically the detection of focal bone lesions. Focal lesions are early, abnormal areas in the bone marrow, which may signal the development of subsequent lytic lesions that typically occur within the next 18–24 months. Cross‐sectional imaging modalities are more sensitive for the detection and monitoring of bone and bone marrow disease and are now included in the International Myeloma Working Group current consensus criteria for initial diagnosis and treatment response assessment. The aim of this consensus practice statement is to review the evidence supporting these modalities. A more detailed Position Statement can be found on the Myeloma Australia website. [ABSTRACT FROM AUTHOR]

The article reviews research on mesenchymal stromal/stem cells (MSCs), noting MSCs have the potential to promote neo-angiogenesis in cardiovascular therapy. The article notes the benefits of traditional plastic adherence-isolate MSCs and strategies for optimizing MSC therapy. The discussion focuses on cell-targeted methods for preparing MSC cells ex vivo, methods for delivering MSC to the heart, and substrate-directed techniques for supporting MSCs interaction with the host myocardium.

Human mesenchymal stem cells (hMSC) are currently being introduced for cell therapy, yet, antibodies specific for native and differentiated MSCs are required for their identification prior to clinical use. Herein, high quality antibodies against MSC surface proteins were developed by immunizing mice with hMSC, and by using a panel of subsequent screening methods. Flow cytometry analysis revealed that 83.5, 1.1, and 8.5% of primary cultures of hMSC were double positive for STRO-1 and either of DJ 3, 9, and 18, respectively. However, none of the three DJ antibodies allowed enrichment of clonogenic hMSC from BMMNCs as single reagents. Using mass-spectrometric analysis, we identified the antigen recognised by DJ3 as CD44, whereas DJ9 and DJ18 recognized HLA-DRB1 and Collagen VI, respectively. The identified proteins were highly expressed throughout in vitro osteogenic- and adipogenic differentiation. Interestingly, undifferentiated cells revealed a sole cytoplasmic distribution pattern of Collagen VI, which however changed to an extracellular matrix appearance upon osteogenic- and adipogenic differentiation. In relation to this, we found that STRO-1/Collagen VI sorted hMSC contained fewer differentiated alkaline phosphatase cells compared to STRO-1/Collagen VI hMSC, suggesting that Collagen VI on the cell membrane exclusively defines differentiated MSCs. In conclusion, we have generated a panel of high quality antibodies to be used for characterization of MSCs, and in addition our results may suggest that the DJ18 generated antibody against Collagen VI can be used for negative selection of cultured undifferentiated MSCs. [ABSTRACT FROM AUTHOR]

Since discovery, significant interest has been generated in the potential application of mesenchymal stem cells or multipotential stromal cells (MSC) for tissue regeneration and repair, due to their proliferative and multipotential capabilities. Although the sheep is often used as a large animal model for translating potential therapies for musculoskeletal injury and repair, the characteristics of MSC from ovine bone marrow have been inadequately described. Histological and gene expression studies have previously shown that ovine MSC share similar properties with human and rodents MSC, including their capacity for clonogenic growth and multiple stromal lineage differentiation. In the present study, ovine bone marrow derived MSCs positively express cell surface markers associated with MSC such as CD29, CD44 and CD166, and lacked expression of CD14, CD31 and CD45. Under serum-deprived conditions, proliferation of MSC occurred in response to EGF, PDGF, FGF-2, IGF-1 and most significantly TGF-α. While subcutaneous transplantation of ovine MSC in association with a ceramic HA/TCP carrier into immunocomprimised mice resulted in ectopic osteogenesis, adipogenesis and haematopoietic-support activity, transplantation of these cells within a gelatin sponge displayed partial chondrogenesis. The comprehensive characterisation of ovine MSC described herein provides important information for future translational studies involving ovine MSC. J. Cell. Physiol. 219: 324–333, 2009. © 2008 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Abstract: Injured growth plate cartilage is often repaired by bony tissue resulting in impaired bone growth in children. Using a rat injury model, our previous studies show that following the injury-induced initial inflammatory response, an influx of mesenchymal-like cells occurs within the growth plate injury site prior to formation of bony tissue. As platelet-derived growth factor (PDGF-BB) is a potent chemotactic factor of mesenchymal cells during skeletal tissue repair, we examined its role during the early fibrogenic response and the subsequent bony repair of injured growth plate. Following growth plate injury, rats received daily injection of the PDGF receptor (PDGFR) inhibitor, Imatinib, for 7 days. Immunohistochemical analysis of injured growth plate at day 1 showed the presence of PDGF-BB expression in some inflammatory cells, while at day 4 PDGFR was expressed by a proportion of the infiltrating mesenchymal cells at the injury site. By day 4, PDGFR inhibition reduced mesenchymal infiltrate (P <0.05); by day 14, Imatinib-treated rats exhibited less bony trabeculae and cartilaginous repair tissues, fewer osteoclasts and less bone marrow (BM) at the injury site, compared to vehicle controls (P <0.01). In vitro “scratch” migration assays with rat BM mesenchymal cells revealed that recombinant PDGF-BB increased cell migration into the “wound” (P <0.05), while Imatinib inhibited this chemotactic response. Quantitative RT-PCR analysis showed that Imatinib treatment decreased expression of the cartilage and bone related genes, Col2a1 and osteocalcin, respectively. These results suggest that PDGF-BB contributes to growth plate injury repair by promoting mesenchymal progenitor cell infiltration, the chondrogenic and osteogenic responses, and remodelling of the repair tissues. [Copyright &y& Elsevier]

Mesenchymal stern/stromal cells (MSC) are an accessible source of precursor cells that can be expanded in vitro and used for tissue regeneration for different clinical applications. The advent of microarray technology has enabled the monitoring of individual and global gene expression patterns across multiple cell populations. Thus, genomic profiling has fundamentally changed our capacity to characterize MSCs, identify potential biomarkers and determined key molecules regulating biological processes involved in stem cell survival, growth and development. Numerous studies have now examined the genomic profiles of MSCs derived from different tissues that exhibit varying levels of differentiation and proliferation potentials. The knowledge gained from these studies will help improve our understanding of the cellular signalling pathways involved in MSC growth, survival and differentiation, and may aid in the development of strategies to improve the tissue regeneration potential of MSCs for different clinical indications. The present review summarizes studied characterizing the gene expression profile of MSCs. [ABSTRACT FROM AUTHOR]

Abstract: Fragility fractures, including neck of femur fractures, result from reductions in the amount, quality and architecture of bone. The aim of this study was to compare the cancellous bone structure, and static indices of bone turnover, in female patients, who had sustained fragility fracture at the femoral neck, with age-matched females without fragility fracture. Bone samples were taken from the intertrochanteric region of the proximal femur of female patients undergoing hip arthroplasty surgery for a subcapital fragility fracture of the femoral neck (#NOF) or from age-matched female control individuals at routine autopsy. Contiguous bone samples were analyzed for undecalcified histomorphometry and for mRNA expression. The histomorphometric data, which were normally distributed, indicated no difference between the mean values for any of the structural parameters in control and fracture samples. In particular, the bone volume (BV/TV) values were not different and did not change significantly with age in these cohorts of individuals aged >65 years. The static indices of bone turnover, eroded surface (ES/BS) and osteoid surface (OS/BS), were positively correlated with age in the >65-year-old control group (p <0.055 and p <0.03, respectively). The median values for these indices were not different between the fracture and control groups. However, both the median and the range of OS/BS values were increased for >65-year-old controls compared with a group of younger females aged <65 years, suggesting an increase in bone formation surface in older females in the proximal femur after 65 years of age. When the data were further interrogated, a reduction in the percentage osteoid surface to eroded surface quotient (OS/ES) was found for the fracture group compared with the age-matched control group suggesting a reduced adaptive modeling drift capability in the fracture group. In contiguous bone samples, increased median values for receptor activator of nuclear factor kappa β (RANK) and interleukin-6 (IL-6) mRNA expression were observed in the fracture group. Study of cultured human osteoblasts showed that recombinant human IL-6 (rhIL-6) inhibited osteoblast differentiation, as measured by an increase in the immature osteoblast marker, STRO-1 and concomitantly decreased expression of the osteoblast maturation marker, alkaline phosphatase. Importantly, cells cultured in the presence of IL-6 showed significantly less mineral deposition in vitro compared with control cultures. These data suggest that perturbations in bone formation surface, relative to resorption surface, are potentially important in producing bone in the proximal femur with increased propensity to fracture. [Copyright &y& Elsevier]

Summary Previous studies have shown that the antibody 7H9D6 identifies CD44, a glycoprotein receptor for hyaluronic acid. 7H9D6 recognizes an epitope of CD44 that is not always present on CD44 molecules. The 7H9D6 antibody bound to the hyaluronic acid binding domain of CD44 and inhibited cell adhesion to immobilized hyaluronic acid. However, the expression of the 7H9D6 epitope was not sufficient for hyaluronic acid binding. Immunofluorescent staining with 7H9D6 revealed a punctate surface staining pattern, suggesting that CD44 molecules recognized by 7H9D6 are located in clusters on the cell surface. In contrast, other CD44 antibodies produced a uniform staining pattern. Early bone marrow B cells were negative for 7H9D6 but reactive with other CD44 monoclonal antibodies. In contrast, leukaemic cells from 65% of patients (28 of 43) with B lineage acute lymphoblastic leukaemia bound 7H9D6. Patients expressing the 7H9D6 epitope on their leukaemic cells had an increased risk of death (HR 3.5 95% CI 1.1-10.9, P = 0.029) and of disease relapse (HR 3.2 95% CI 1.2-8.5, P = 0.017) when corrected for white cell count. This antibody may be useful for the detection of residual disease in B lineage acute lymphoblastic leukaemia and as a prognostic indicator and for the study of CD44 function. [ABSTRACT FROM AUTHOR]

Summary: Disease relapse is the greatest cause of treatment failure in paediatric B‐cell acute lymphoblastic leukaemia (B‐ALL). Current risk stratifications fail to capture all patients at risk of relapse. Herein, we used a machine‐learning approach to identify B‐ALL blast‐secreted factors that are associated with poor survival outcomes. Using this approach, we identified a two‐gene expression signature (CKLF and IL1B) that allowed identification of high‐risk patients at diagnosis. This two‐gene expression signature enhances the predictive value of current at diagnosis or end‐of‐induction risk stratification suggesting the model can be applied continuously to help guide implementation of risk‐adapted therapies. [ABSTRACT FROM AUTHOR]

The protein SAMSN1 was recently identified as a putative tumor suppressor in multiple myeloma, with re‐expression of Samsn1 in the 5TGM1/KaLwRij murine model of myeloma leading to a near complete abrogation of intramedullary tumor growth. Here, we sought to clarify the mechanism underlying this finding. Intratibial administration of 5TGM1 myeloma cells into KaLwRij mice revealed that Samsn1 had no effect on primary tumor growth, but that its expression significantly inhibited the metastasis of these primary tumors. Notably, neither in vitro nor in vivo migration was affected by Samsn1 expression. Both knocking‐out SAMSN1 in the RPMI‐8226 and JJN3 human myeloma cell lines, and retrovirally expressing SAMSN1 in the LP‐1 and OPM2 human myeloma cell lines had no effect on either cell proliferation or migration in vitro. Altering SAMSN1 expression in these human myeloma cells did not affect the capacity of the cells to establish either primary or metastatic intramedullary tumors when administered intratibially into immune deficient NSG mice. Unexpectedly, the tumor suppressive and anti‐metastatic activity of Samsn1 in 5TGM1 cells were not evidenced following cell administration either intratibially or intravenously to NSG mice. Crucially, the growth of Samsn1‐expressing 5TGM1 cells was limited in C57BL/6/Samsn1−/− mice but not in C57BL/6 Samsn1+/+ mice. We conclude that the reported potent in vivo tumor suppressor activity of Samsn1 can be attributed, in large part, to graft‐rejection from Samsn1−/− recipient mice. This has broad implications for the design and interpretation of experiments that utilize cancer cells and knockout mice that are mismatched for expression of specific proteins. [ABSTRACT FROM AUTHOR]

Previous studies of global binding patterns identified the epigenetic factor, EZH2, as a regulator of the homeodomain-only protein homeobox (HOPX) gene expression during bone marrow stromal cell (BMSC) differentiation, suggesting a potential role for HOPX in regulating BMSC lineage specification. In the present study, we confirmed that EZH2 direct binds to the HOPX promoter region, during normal growth and osteogenic differentiation but not under adipogenic inductive conditions. HOPX gene knockdown and overexpression studies demonstrated that HOPX is a promoter of BMSC proliferation and an inhibitor of adipogenesis. However, functional studies failed to observe any affect by HOPX on BMSC osteogenic differentiation. RNA-seq analysis of HOPX overexpressing BMSC during adipogenesis, found HOPX function to be acting through suppression of adipogenic pathways associated genes such as ADIPOQ, FABP4, PLIN1 and PLIN4. These findings suggest that HOPX gene target pathways are critical factors in the regulation of fat metabolism. [ABSTRACT FROM AUTHOR]

Multipotent bone marrow-derived mesenchymal stem/stromal cells (BMSCs) exhibit a finite life span after ex vivo expansion leading to cellular senescence. Many factors can contribute to this. Recently, our group has identified for the first time expression of the chemokine-like factor superfamily 8 (CMTM8) gene in cultured human BMSCs. In this study, we examine the role of CMTM8 in BMSC proliferation, migration, and differentiation. Functional studies using siRNA-mediated knockdown of CMTM8 in human BMSCs resulted in decreased capacity to undergo proliferation and migration and an increased capacity for osteogenic differentiation in vitro. Furthermore, reduced CMTM8 levels led to a decrease in the epidermal growth factor receptor (EGFR) signaling pathway during BMSC proliferation and migration, respectively. Supportive studies using retroviral mediated enforced expression of CMTM8 in BMSC resulted in an increased capacity for proliferation and migration but a decreased osteogenic differentiation potential. Collectively, these data suggest that CMTM8 promotes BMSC proliferation and BMSC migration through the EGFR/ERK1/2 pathway. This study provides insight into novel regulatory mechanisms of human BMSC growth and cell fate determination. [ABSTRACT FROM AUTHOR]

The Eph receptor tyrosine kinase ligand, ephrinB1 (EfnB1) is important for correct skeletal and cartilage development, however, the role of EfnB1 in fracture repair is unknown. This study investigated the role of EfnB1 during fracture repair where EfnB1 expression increased significantly at 1 and 2 weeks post fracture in C57Bl/6 wildtype mice, coinciding with the haematoma, soft callus formation/remodelling stages, respectively. To investigate the specific role of EfnB1 within the osteogenic lineage during fracture repair, male mice with a conditional deletion of EfnB1 in the osteogenic lineage (EfnB1 OB fl / O ), driven by the Osterix (Osx) promoter, and their male Osx : Cre counterparts were subject to a femoral fracture with internal fixation. Two weeks post fracture micro computed tomography (μCT) analysis revealed that EfnB1 OB fl / O mice displayed a significant decrease in bone volume relative to tissue volume within the fracture callus. This was attributed to an alteration in the distribution of osteoclasts within the fracture site, a significant elevation in cartilaginous tissue and reduction in the osteoprogenitor population and calcein labelled bone within the fracture site of EfnB1 OB fl / O mice. Supportive in vitro studies demonstrated that under osteogenic conditions, cultured EfnB1 OB fl / O stromal cells derived from the 2 week fracture site exhibited a reduced capacity to produce mineral and decreased expression of the osteogenic gene, Osterix , when compared to Osx : Cre controls. These findings suggest that the loss of EfnB1 delays the fracture repair process. The present study confirmed that EFNB1 activation in human BMSC, following stimulation with soluble-EphB2 resulted in de-phosphorylation of TAZ, demonstrating similarities in EfnB1 signalling between human and mouse stromal populations. Overall, the present study provides evidence that loss of EfnB1 in the osteo/chondrogenic lineages delays the soft callus formation/remodelling stages of the fracture repair process. • The loss of ephrinB1 in osteoprogenitors delays bone repair at the early phases of fracture healing. • The loss of ephrinB1 in osteoprogenitors alters osteoclast distribution during the remodeling of the softy callus. • Human osteoprogenitors utilize TAZ signaling to activate osteogenic differentiation as previously shown in mouse. [ABSTRACT FROM AUTHOR]

N‐cadherin is a homophilic cell‐cell adhesion molecule that plays a critical role in maintaining vascular stability and modulating endothelial barrier permeability. Pre‐clinical studies have shown that the N‐cadherin antagonist peptide, ADH‐1, increases the permeability of tumor‐associated vasculature thereby increasing anti‐cancer drug delivery to tumors and enhancing tumor response. Small molecule library screens have identified a novel compound, LCRF‐0006, that is a mimetic of the classical cadherin His‐Ala‐Val sequence‐containing region of ADH‐1. Here, we evaluated the vascular permeability‐enhancing and anti‐cancer properties of LCRF‐0006 using in vitro vascular disruption and cell apoptosis assays, and a well‐established pre‐clinical model (C57BL/KaLwRij/5TGM1) of the hematological cancer multiple myeloma (MM). We found that LCRF‐0006 disrupted endothelial cell junctions in a rapid, transient and reversible manner, and increased vascular permeability in vitro and at sites of MM tumor in vivo. Notably, LCRF‐0006 synergistically increased the in vivo anti‐MM tumor response to low‐dose bortezomib, a frontline anti‐MM agent, leading to regression of disease in 100% of mice. Moreover, LCRF‐0006 and bortezomib synergistically induced 5TGM1 MM tumor cell apoptosis in vitro. Our findings demonstrate the potential clinical utility of LCRF‐0006 to significantly increase bortezomib effectiveness and enhance the depth of tumor response in patients with MM. [ABSTRACT FROM AUTHOR]

Approximately 15% of patients with multiple myeloma (MM) harbour the t(4;14) chromosomal translocation, leading to the overexpression of the histone methyltransferase NSD2. Patients with this translocation display increased tumour dissemination, accelerated disease progression and rapid relapse. Using publicly available gene expression profile data from NSD2high (n = 135) and NSD2low (n = 878) MM patients, we identified 39 epithelial-mesenchymal transition (EMT)-associated genes which are overexpressed in NSD2high MM plasma cells. In addition, our analyses identified Twist-1 as a key transcription factor upregulated in NSD2high MM patients and t(4;14)-positive cell lines. Overexpression and knockdown studies confirmed that Twist-1 is involved in driving the expression of EMT-associated genes in the human MM cell line KMS11 and promoted the migration of myeloma cell lines in vitro. Notably, Twist-1 overexpression in the mouse MM cell line 5TGM1 significantly increased tumour dissemination in an intratibial tumour model. These findings demonstrate that Twist-1, downstream of NSD2, contributes to the induction of an EMT-like signature in t(4;14)-positive MM and enhances the dissemination of MM plasma cells in vivo, which may, in part, explain the aggressive disease features associated with t(4;14)-positive MM. [ABSTRACT FROM AUTHOR]

Multiple myeloma, a plasma cell malignancy, is a genetically heterogeneous disease and the genetic factors that contribute to its development and progression remain to be fully elucidated. The tumour suppressor gene GLIPR1 has previously been shown to be deleted in approximately 10% of myeloma patients, to inhibit the development of plasma cell tumours in ageing mice and to have reduced expression levels in the plasma cells of patients with light-chain amyloidosis, a myeloma-related malignancy. Therefore, we hypothesised that GLIPR1 may have tumour suppressor activity in multiple myeloma. In this study, we demonstrate that plasma cell expression of GLIPR1 is reduced in the majority of myeloma patients and Glipr1 expression is lost in the 5TGM1 murine myeloma cell line. However, overexpression of GLIPR1 in a human myeloma cell line did not affect cell proliferation in vitro. Similarly, re-expression of Glipr1 in 5TGM1 cells did not significantly reduce their in vitro proliferation or in vivo growth in C57BL/KaLwRij mice. In addition, using CRISPR-Cas9 genome editing, we generated C57BL/Glipr1-/- mice and showed that loss of Glipr1 in vivo did not affect normal haematopoiesis or the development of monoclonal plasma cell expansions in these mice up to one year of age. Taken together, our results suggest that GLIPR1 is unlikely to be a potent tumour suppressor in multiple myeloma. However, it remains possible that the down-regulation of GLIPR1 may cooperate with other genetic lesions to promote the development of myeloma. [ABSTRACT FROM AUTHOR]

Multiple myeloma is a fatal plasma cell malignancy that is reliant on the bone marrow microenvironment. The bone marrow is comprised of numerous cells of mesenchymal and hemopoietic origin. Of these, macrophages have been implicated to play a role in myeloma disease progression, angiogenesis, and drug resistance; however, the role of macrophages in myeloma disease establishment remains unknown. In this study, the antimyeloma efficacy of clodronate-liposome treatment, which globally and transiently depletes macrophages, was evaluated in the well-established C57BL/KaLwRijHsd murine model of myeloma. Our studies show, for the first time, that clodronate-liposome pretreatment abrogates myeloma tumor development in vivo. Clodronate-liposome administration resulted in depletion of CD169+ bone marrow–resident macrophages. Flow cytometric analysis revealed that clodronate-liposome pretreatment impaired myeloma plasma cell homing and retention within the bone marrow 24 hours postmyeloma plasma cell inoculation. This was attributed in part to decreased levels of macrophage-derived insulin-like growth factor 1. Moreover, a single dose of clodronate-liposome led to a significant reduction in myeloma tumor burden in KaLwRij mice with established disease. Collectively, these findings support a role for CD169-expressing bone marrow–resident macrophages in myeloma disease establishment and progression and demonstrate the potential of targeting macrophages as a therapy for myeloma patients. [ABSTRACT FROM AUTHOR]

The bone marrow stromal microenvironment contributes to the maintenance and function of hematopoietic stem/progenitor cells (HSPCs). The Eph receptor tyrosine kinase family members have been implicated in bone homeostasis and stromal support of HSPCs. The present study examined the influence of EfnB1-expressing osteogenic lineage on HSPC function. Mice with conditional deletion of EfnB1 in the osteogenic lineage (EfnB1 OB –/–), driven by the Osterix promoter, exhibited a reduced prevalence of osteogenic progenitors and osteoblasts, correlating to lower numbers of HSPCs compared with Osx:Cre mice. Long-term culture-initiating cell (LTC-IC) assays confirmed that the loss of EfnB1 within bone cells hindered HSPC function, with a significant reduction in colony formation in EfnB1 OB –/– mice compared with Osx:Cre mice. Human studies confirmed that activation of EPHB2 on CD34+ HSPCs via EFNB1-Fc stimulation enhanced myeloid/erythroid colony formation, whereas functional blocking of either EPHB1 or EPHB2 inhibited the maintenance of LTC-ICs. Moreover, EFNB1 reverse signaling in human and mouse stromal cells was found to be required for the activation of the HSPC-promoting factor CXCL12. Collectively, the results of this study confirm that EfnB1 contributes to the stromal support of HSPC function and maintenance and may be an important factor in regulating the HSPC niche. [ABSTRACT FROM AUTHOR]

Key transcription factors, which activate or repress master gene regulators and signaling pathways, tightly regulate self-renewal and cell lineage differentiation of bone marrow-derived stromal cells (BMSC). Among these factors is the basic helix-loop-helix transcription factor Twist-related protein 1 (TWIST-1), which is important in BMSC self-renewal, life span, and differentiation. Another layer of gene regulation comes from microRNAs (miRNAs). miRNAs are short noncoding RNAs that interfere with translation of specific target mRNAs and thereby regulate diverse biological processes, including BMSC lineage commitment. However, little is known of how TWIST-1-regulated miRNAs control osteogenic commitment, and influence the fate of bone precursor cells. In this study, we have discovered a novel TWIST-1-regulated miRNA, miR-376c-3p. Reduced miR-376c-3p expression by a miR-376c-3p inhibitor or due to TWIST-1 haploinsufficiency promotes alkaline phosphatase (ALP) activity, mineral deposition, and expression of osteoblast-associated genes in BMSC and calvarial cells. Conversely, overexpression of miR-376c-3p using a miR-376c-3p mimic inhibited BMSC proliferation and the osteogenic potential of BMSC and TWIST-1 haploinsufficient calvarial cells. This was demonstrated by a decrease in insulin growth factor 1 receptor (IGF1R) levels, Akt signaling, ALP activity, mineral deposition, and expression of osteoblast-associated genes. Thus, miR-376c-3p reduces IGF1R/Akt signaling in BMSC and is one mechanism by which osteogenesis may be inhibited. Overall, we have identified miR-376c-3p as a TWIST-1-regulated miRNA, which plays an important role in the osteogenesis of bone precursor cells and can mediate TWIST-1 inhibition of osteogenesis. Furthermore, overexpression of miRNA-376c-3p in TWIST-1 haploinsufficient calvarial cells can decrease the aberrant osteogenesis of these cells, which contributes to increased calvarial bone volume and premature fusion of the coronal sutures. [ABSTRACT FROM AUTHOR]

In many types of solid tumours, the aberrant expression of the cell adhesion molecule N-cadherin is a hallmark of epithelial-to-mesenchymal transition, resulting in the acquisition of an aggressive tumour phenotype. This transition endows tumour cells with the capacity to escape from the confines of the primary tumour and metastasise to secondary sites. In this review, we will discuss how N-cadherin actively promotes the metastatic behaviour of tumour cells, including its involvement in critical signalling pathways which mediate these events. In addition, we will explore the emerging role of N-cadherin in haematological malignancies, including bone marrow homing and microenvironmental protection to anti-cancer agents. Finally, we will discuss the evidence that N-cadherin may be a viable therapeutic target to inhibit cancer metastasis and increase tumour cell sensitivity to existing anti-cancer therapies. [ABSTRACT FROM AUTHOR]

Recurrent oncogenic fusion genes play a critical role in the development of various cancers and diseases and provide, in some cases, excellent therapeutic targets. To date, analysis tools that can identify and compare recurrent fusion genes across multiple samples have not been available to researchers. To address this deficiency, we developed Co-occurrence Fusion (Co-fuse), a new and easy to use software tool that enables biologists to merge RNA-seq information, allowing them to identify recurrent fusion genes, without the need for exhaustive data processing. Notably, Co-fuse is based on pattern mining and statistical analysis which enables the identification of hidden patterns of recurrent fusion genes. In this report, we show that Co-fuse can be used to identify 2 distinct groups within a set of 49 leukemic cell lines based on their recurrent fusion genes: a multiple myeloma (MM) samples-enriched cluster and an acute myeloid leukemia (AML) samples-enriched cluster. Our experimental results further demonstrate that Co-fuse can identify known driver fusion genes (e.g., IGH-MYC, IGH-WHSC1) in MM, when compared to AML samples, indicating the potential of Co-fuse to aid the discovery of yet unknown driver fusion genes through cohort comparisons. Additionally, using a 272 primary glioma sample RNA-seq dataset, Co-fuse was able to validate recurrent fusion genes, further demonstrating the power of this analysis tool to identify recurrent fusion genes. Taken together, Co-fuse is a powerful new analysis tool that can be readily applied to large RNA-seq datasets, and may lead to the discovery of new disease subgroups and potentially new driver genes, for which, targeted therapies could be developed. The Co-fuse R source code is publicly available at https://github.com/sakrapee/co-fuse. [ABSTRACT FROM AUTHOR]

The skeleton has recently emerged as a critical insulin target tissue that regulates whole body glucose metabolism and male reproductive function. While our understanding of these new regulatory axes remains in its infancy, the bone‐specific protein, osteocalcin, has been shown to be centrally involved. Undercarboxylated osteocalcin acts as a secretagogue in a feed‐forward loop to stimulate pancreatic β‐cell proliferation and insulin secretion, improve insulin sensitivity, and promote testosterone production. Importantly, dysregulation of insulin signaling in bone causes a reduction in serum osteocalcin levels that is associated with elevated blood glucose and reduced serum insulin levels, suggesting that the skeleton may play a significant role in the development of diet‐induced insulin resistance. Insulin signaling is negatively regulated by the mammalian target of rapamycin complex 1 (mTORC1) which becomes hyper‐activated in response to nutrient overload. Loss‐ and gain‐of function models suggest that mTORC1 function in bone is essential for normal skeletal development; however, the role of this complex in the regulation of glucose metabolism remains to be determined. This review highlights our current understanding of the role played by osteocalcin in the skeletal regulation of glucose metabolism and fertility. In particular, it examines data emerging from transgenic mouse models which have revealed a pancreas‐bone‐testis regulatory axis and discusses recent human studies which seek to corroborate findings from mouse models with clinical observations. Moreover, we review recent studies which suggest dysregulation of insulin signaling in bone leads to the development of insulin resistance and discuss the potential role of mTORC1 signaling in this process. [ABSTRACT FROM AUTHOR]

Background: The pharmaceutical agent pentosan polysulfate (PPS) is known to induce proliferation and chondrogenesis of mesenchymal progenitor cells (MPCs) in vitro and in vivo. However, the mechanism(s) of action of PPS in mediating these effects remains unresolved. In the present report we address this issue by investigating the binding and uptake of PPS by MPCs and monitoring gene expression and proteoglycan biosynthesis before and after the cells had been exposed to limited concentrations of PPS and then re-established in culture in the absence of the drug (MPC priming). Methods: Immuno-selected STRO-1+ mesenchymal progenitor stem cells (MPCs) were prepared from human bone marrow aspirates and established in culture. The kinetics of uptake, shedding, and internalization of PPS by MPCs was determined by monitoring the concentration-dependent loss of PPS media concentrations using an enzyme-linked immunosorbent assay (ELISA) and the uptake of fluorescein isothiocyanate (FITC)-labelled PPS by MPCs. The proliferation of MPCs, following pre-incubation and removal of PPS (priming), was assessed using the Wst-8 assay method, and proteoglycan synthesis was determined by the incorporation of 35SO4 into their sulphated glycosaminoglycans. The changes in expression of MPC-related cell surface antigens of non-primed and PPS-primed MPCs from three donors was determined using flow cytometry. RNA sequencing of RNA isolated from non-primed and PPS-primed MPCs from the same donors was undertaken to identify the genes altered by the PPS priming protocol. Results: The kinetic studies indicated that, in culture, PPS rapidly binds to MPC surface receptors, followed by internalisation and localization within the nucleus of the cells. Following PPS-priming of MPCs and a further 48 h of culture, both cell proliferation and proteoglycan synthesis were enhanced. Reduced expression of MPC-related cell surface antigen expression was promoted by the PPS priming, and RNA sequencing analysis revealed changes in the expression of 42 genes. Conclusion: This study has shown that priming of MPCs with low concentrations of PPS enhanced chondrogenesis and MPC proliferation by modifying their characteristic basal gene and protein expression. These findings offer a novel approach to re-programming mesenchymal stem cells for clinical indications which require the repair or regeneration of cartilaginous tissues such as in osteoarthritis and degenerative disc disease. [ABSTRACT FROM AUTHOR]

Multiple myeloma (MM) is a hematological malignancy resulting from the uncontrolled proliferation of antibodyproducing plasma cells in the bone marrow. At diagnosis, independent plasma cell tumors are found throughout the skeleton. The recirculation of mutant plasma cells from the initial lesion and their recolonization of distant marrow sites are thought to occur by a process similar to solid tumor metastasis. However, the efficiency of this bone marrow homing process and the proportion of disseminated cells that actively divide and contribute to new tumor growth in MM are both unknown. We used the C57BL/KaLwRij mouse model of myeloma, lentiviralmediated DNA barcoding of 5TGM1 myeloma cells, and next-generation sequencing to investigate the relative efficiency of plasma cell migration to, and growth within, the bone marrow. This approach revealed three major findings: firstly, establishment of metastasis within the bone marrow was extremely inefficient, with approximately 0.01% of circulating myeloma cells becoming resident long term in the bone marrow of each long bone; secondly, the individual cells of each metastasis exhibited marked differences in their proliferative fates, with the majority of final tumor burden within a bone being attributable to the progeny of between 1 and 8 cells; and, thirdly, the proliferative fate of individual clonal plasma cells differed at each bone marrow site in which the cells "landed." These findings suggest that individual myeloma plasma cells are subjected to vastly different selection pressures within the bone marrow microenvironment, highlighting the importance of niche-driven factors, which determine the disease course and outcome. [ABSTRACT FROM AUTHOR]

Tumor hypoxia is a major cause of treatment failure for a variety of malignancies. However, hypoxia also leads to treatment opportunities as demonstrated by the development of compounds that target regions of hypoxia within tumors. Evofosfamide is a hypoxia-activated prodrug that is created by linking the hypoxia-seeking 2-nitroimidazole moiety to the cytotoxic bromo-isophosphoramide mustard (Br- IPM). When evofosfamide is delivered to hypoxic regions of tumors, the DNA cross-linking toxin, Br- IPM, is released leading to cell death. This study assessed the anticancer efficacy of evofosfamide in combination with the Proapoptotic Receptor Agonists ( PARAs) dulanermin and drozitumab against human osteosarcoma in vitro and in an intratibial murine model of osteosarcoma. Under hypoxic conditions in vitro, evofosfamide cooperated with dulanermin and drozitumab, resulting in the potentiation of cytotoxicity to osteosarcoma cells. In contrast, under the same conditions, primary human osteoblasts were resistant to treatment. Animals transplanted with osteosarcoma cells directly into their tibiae developed mixed osteosclerotic/osteolytic bone lesions and consequently developed lung metastases 3 weeks post cancer cell transplantation. Tumor burden in the bone was reduced by evofosfamide treatment alone and in combination with drozitumab and prevented osteosarcoma-induced bone destruction while also reducing the growth of pulmonary metastases. These results suggest that evofosfamide may be an attractive therapeutic agent, with strong anticancer activity alone or in combination with either drozitumab or dulanermin against osteosarcoma. [ABSTRACT FROM AUTHOR]

Multiple Myeloma ( MM) is a haematological malignancy characterised by the clonal expansion of plasma cells ( PCs) within the bone marrow. Despite advances in therapy, MM remains a largely incurable disease with a median survival of 6 years. In almost all cases, the development of MM is preceded by the benign PC condition Monoclonal Gammopathy of Undetermined Significance ( MGUS). Recent studies show that the transformation of MGUS to MM is associated with complex genetic changes. Understanding how these changes contribute to evolution will present targets for clinical intervention. We discuss three models of MM evolution; the linear, the expansionist and the intraclonal heterogeneity models. Of particular interest is the intraclonal heterogeneity model. Here, distinct populations of MM PCs carry differing combinations of genetic mutations. Acquisition of additional mutations can contribute to subclonal lineages where 'driver' mutations may influence selective pressure and dominance, and 'passenger' mutations are neutral in their effects. Furthermore, studies show that clinical intervention introduces additional selective pressure on tumour cells and can influence subclone survival, leading to therapy resistance. This review discusses how Next Generation Sequencing approaches are revealing critical insights into the genetics of MM development, disease progression and treatment. MM disease progression will illuminate possible mechanisms underlying the tumour. [ABSTRACT FROM AUTHOR]

We have discovered that a small cationic molecule, GW4869, is cytotoxic to a subset of myeloma cell lines and primary myeloma plasma cells. Biochemical analysis revealed that GW4869 binds to anionic phospholipids such as phosphatidylserine - a lipid normally confined to the intracellular side of the cell membrane. However, interestingly, phosphatidylserine was expressed on the surface of all myeloma cell lines tested ( n = 12) and 9/15 primary myeloma samples. Notably, the level of phosphatidylserine expression correlated well with sensitivity to GW4869. Inhibition of cell surface phosphatidylserine exposure with brefeldin A resulted in resistance to GW4869. Finally, GW4869 was shown to delay the growth of phosphatidylserine-high myeloma cells in vivo. To the best of our knowledge, this is the first example of using a small molecule to target phosphatidylserine on malignant cells. This study may provide the rationale for the development of phosphatidylserine-targeting small molecules for the treatment of surface phosphatidylserine-expressing cancers. [ABSTRACT FROM AUTHOR]

The mammalian target of rapamycin complex 1 (mTORC1) is activated by extracellular factors that control bone accrual. However, the direct role of this complex in osteoblast biology remains to be determined. To investigate this question, we disrupted mTORC1 function in preosteoblasts by targeted deletion of Raptor (Rptor) in Osterix-expressing cells. Deletion of Rptor resulted in reduced limb length that was associated with smaller epiphyseal growth plates in the postnatal skeleton. Rptor deletion caused a marked reduction in pre- and postnatal bone accrual, which was evident in skeletal elements derived from both intramembranous and endochondrial ossification. The decrease in bone accrual, as well as the associated increase in skeletal fragility, was due to a reduction in osteoblast function. In vitro, osteoblasts derived from knockout mice display a reduced osteogenic potential, and an assessment of bone-developmental markers in Rptor knockout osteoblasts revealed a transcriptional profile consistent with an immature osteoblast phenotype suggesting that osteoblast differentiation was stalled early in osteogenesis. Metabolic labeling and an assessment of cell size of Rptor knockout osteoblasts revealed a significant decrease in protein synthesis, a major driver of cell growth. These findings demonstrate that mTORC1 plays an important role in skeletal development by regulating mRNA translation during preosteoblast differentiation. [ABSTRACT FROM AUTHOR]