The effects of young and aged, male and female megakaryocyte conditioned media on angiogenic properties of endothelial cells.

With aging, the risk of fractures and compromised healing increases. Angiogenesis plays a significant role in bone healing and is impaired with aging. We have previously shown the impact of megakaryocytes (MKs) in regulating bone healing.

Thrombopoietic agents enhance bone healing in mice, rats, and pigs.

Achieving bone union remains a significant clinical dilemma. The use of osteoinductive agents, specifically bone morphogenetic proteins (BMPs), has gained wide attention. However, multiple side effects, including increased incidence of cancer, has renewed interest in investigating alternatives that provide safer, yet effective bone regeneration.

A novel class of self-complementary AAV vectors with multiple advantages based on cceAAV lacking mutant ITR.

Self-complementary AAV vectors (scAAV) use a mutant inverted terminal repeat (mITR) for efficient packaging of complementary stranded DNA, enabling rapid transgene expression. However, inefficient resolution at the mITR leads to the packaging of monomeric or subgenomic AAV genomes. These noncanonical particles reduce transgene expression and may affect the safety of gene transfer.

Osteoclast-derived IGF1 induces RANKL production in osteocytes and contributes to pagetic lesion formation.

We previously reported that measles virus nucleocapsid protein (MVNP) expression in osteoclasts (OCLs) of patients with Paget disease (PD) or targeted to the OCL lineage in MVNP-transgenic mice (MVNP mice) increases IGF1 production in osteoclasts (OCL-IGF1) and leads to development of PD OCLs and pagetic bone lesions (PDLs). Conditional deletion of Igf1 in OCLs of MVNP mice fully blocked development of PDLs. In this study, we examined whether osteocytes (OCys), key regulators of normal bone remodeling, contribute to PD.

Chemical modification of AAV9 capsid with N-ethyl maleimide alters vector tissue tropism.

Although more adeno-associated virus AAV-based drugs enter the clinic, vector tissue tropism remains an unresolved challenge that limits its full potential despite that the tissue tropism of naturally occurring AAV serotypes can be altered by genetic engineering capsid vie DNA shuffling, or molecular evolution. To further expand the tropism and thus potential applications of AAV vectors, we utilized an alternative approach that employs chemical modifications to covalently link small molecules to reactive exposed Lysine residues of AAV capsids. We demonstrated that AAV9 capsid modified with N-ethyl Maleimide (NEM) increased its tropism more towards murine bone marrow (osteoblast lineage) while decreased transduction of liver tissue compared to the unmodified capsid.

Cryptic resolution sites in the vector plasmid lead to the heterogeneities in the rAAV vectors.

Recombinant adeno-associated virus (rAAV) vectors carry a cassette of interest retaining only the inverted terminal repeats (ITRs) from the wild-type virus. Conventional rAAV production primarily uses a vector plasmid as well as helper genes essential for AAV replication and packaging. Nevertheless, plasmid backbone related contaminants have been a major source of vector heterogeneity.

Genomic Designs of rAAVs Contribute to Pathological Changes in the Livers and Spleens of Mice.

Recombinant AAV (rAAV) gene therapy is being investigated as an effective therapy for several diseases including hemophilia B. Reports of liver tumor development in certain mouse models due to AAV treatment and genomic integration of the rAAV vector has raised concerns about the long-term safety and efficacy of this gene therapy. To investigate whether rAAV treatment causes cancer, we utilized two mouse models, inbred C57BL/6 and hemophilia B Balb/C mice (HemB), to test if injecting a high dose of various rAAV8 vectors containing or lacking hFIX transgene, a Poly-A sequence, or the CB or TTR promoter triggered liver fibrosis and/or cancer development over the course of the 6.

Adding recombinant AAVs to the cancer therapeutics mix.

Gene therapy is a powerful biological tool that is reshaping therapeutic landscapes for several diseases. Researchers are using both non-viral and viral-based gene therapy methods with success in the lab and the clinic. In the cancer biology field, gene therapies are expanding treatment options and the possibility of favorable outcomes for patients.

Fast and high-throughput LC-MS characterization, and peptide mapping of engineered AAV capsids using LC-MS/MS.

Adeno-associated virus (AAV) has emerged as a leading platform for gene therapy. With the skyrocketing rate of AAV research and the prevalence of many new engineered capsids being investigated in preclinical and clinical trials, capsid characterization plays a vital role in serotype confirmation and quality control. Further, peptide mapping the capsid proteins might inevitably be a future requirement by regulatory agencies since it is a critical step in good manufacturing practice (GMP) for biotherapeutic characterization.

Subgenomic particles in rAAV vectors result from DNA lesion/break and non-homologous end joining of vector genomes.

Recombinant adeno-associated virus (rAAV) vectors have been developed for therapeutic treatment of genetic diseases. Current rAAV vectors administered to affected individuals often contain vector DNA-related contaminants. Here we present a thorough molecular analysis of the configuration of non-standard AAV genomes generated during rAAV production using single-molecule sequencing.

GFI1-Dependent Repression of Increases Multiple Myeloma Cell Survival.

Multiple myeloma (MM) remains incurable for most patients due to the emergence of drug resistant clones. Here we report a p53-independent mechanism responsible for Growth Factor Independence-1 (GFI1) support of MM cell survival by its modulation of sphingolipid metabolism to increase the sphingosine-1-phosphate (S1P) level regardless of the p53 status. We found that expression of enzymes that control S1P biosynthesis, , dephosphorylation, and were differentially correlated with GFI1 levels in MM cells.

The effects of bone morphogenetic protein 2 and thrombopoietin treatment on angiogenic properties of endothelial cells derived from the lung and bone marrow of young and aged, male and female mice.

With an aging world population, there is an increased risk of fracture and impaired healing. One contributing factor may be aging-associated decreases in vascular function; thus, enhancing angiogenesis could improve fracture healing. Both bone morphogenetic protein 2 (BMP-2) and thrombopoietin (TPO) have pro-angiogenic effects.

Osteocyte Vegf-a contributes to myeloma-associated angiogenesis and is regulated by Fgf23.

Multiple Myeloma (MM) induces bone destruction, decreases bone formation, and increases marrow angiogenesis in patients. We reported that osteocytes (Ocys) directly interact with MM cells to increase tumor growth and expression of Ocy-derived factors that promote bone resorption and suppress bone formation. However, the contribution of Ocys to enhanced marrow vascularization in MM is unclear.

β2ARs stimulation in osteoblasts promotes breast cancer cell adhesion to bone marrow endothelial cells in an IL-1β and selectin-dependent manner.

Progression and recurrence of breast cancer, as well as reduced survival of patients with breast cancer, are associated with chronic stress, a condition known to impact the hypothalamic-pituitary axis and the autonomic nervous system. Preclinical and clinical evidence support the involvement of the sympathetic nervous system in the control of bone remodeling and in pathologies of the skeleton, including bone metastasis. In experimental mouse models of skeletal metastasis, administration of the βAR agonist isoproterenol (ISO), used as a surrogate of norepinephrine, the main neurotransmitter of sympathetic neurons, was shown to favor bone colonization of metastatic breast cancer cells via an increase bone marrow vascularity.

Dual targeting c-met and VEGFR2 in osteoblasts suppresses growth and osteolysis of prostate cancer bone metastasis.

Prostate cancer characteristically induces osteoblastic bone metastasis, for which no therapies are available. A dual kinase inhibitor of c-Met and VEGFR-2 (cabozantinib) was shown to reduce prostate cancer growth in bone, with evidence for suppressing osteoblastic activity. However, c-Met and VEGFR2 signaling in osteoblasts in the context of bone metastasis remain unclear.

Skeletal Colonization by Breast Cancer Cells Is Stimulated by an Osteoblast and β2AR-Dependent Neo-Angiogenic Switch.

The skeleton is a common site for breast cancer metastasis. Although significant progress has been made to manage osteolytic bone lesions, the mechanisms driving the early steps of the bone metastatic process are still not sufficiently understood to design efficacious strategies needed to inhibit this process and offer preventative therapeutic options. Progression and recurrence of breast cancer, as well as reduced survival of patients with breast cancer, are associated with chronic stress, a condition known to stimulate sympathetic nerve outflow.

The Bone Microenvironment: a Fertile Soil for Tumor Growth.

Bone metastatic disease remains a significant and frequent problem for cancer patients that can lead to increased morbidity and mortality. Unfortunately, despite decades of research, bone metastases remain incurable. Current studies have demonstrated that many properties and cell types within the bone and bone marrow microenvironment contribute to tumor-induced bone disease.

Synergistic effects of GDNF and VEGF on lifespan and disease progression in a familial ALS rat model.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the progressive loss of motor neurons in the brain and spinal cord. We have recently shown that human mesenchymal stem cells (hMSCs) modified to release glial cell line-derived neurotrophic factor (GDNF) decrease disease progression in a rat model of ALS when delivered to skeletal muscle. In the current study, we determined whether or not this effect could be enhanced by delivering GDNF in concert with other trophic factors.

Intermittent hypoxia and stem cell implants preserve breathing capacity in a rodent model of amyotrophic lateral sclerosis.

Rationale: Amyotrophic lateral sclerosis (ALS) is a devastating motor neuron disease causing paralysis and death from respiratory failure. Strategies to preserve and/or restore respiratory function are critical for successful treatment. Although breathing capacity is maintained until late in disease progression in rodent models of familial ALS (SOD1(G93A) rats and mice), reduced numbers of phrenic motor neurons and decreased phrenic nerve activity are observed.

Male-specific differences in proliferation, neurogenesis, and sensitivity to oxidative stress in neural progenitor cells derived from a rat model of ALS.

Amyotrophic Lateral Sclerosis (ALS) is a fatal neurodegenerative disease characterized by progressive motor dysfunction and the loss of large motor neurons in the spinal cord and brain stem. A clear genetic link to point mutations in the superoxide dismutase 1 (SOD1) gene has been shown in a small group of familial ALS patients. The exact etiology of ALS is still uncertain, but males have consistently been shown to be at a higher risk for the disease than females.

Gonadectomy and dehydroepiandrosterone (DHEA) do not modulate disease progression in the G93A mutant SOD1 rat model of amyotrophic lateral sclerosis.

Epidemiological studies have shown a higher incidence of amyotrophic lateral sclerosis (ALS) in men than women. Interestingly, there are clear gender differences in disease onset and progression in rodent models of familial ALS overexpressing mutated human superoxide dismutase-1 (SOD1-G93A). In the present study we sought to determine whether the alterations of serum steroid levels by gonadectomy or chronic treatment of neuroprotective neurosteroids can modulate disease onset and progression in a rat model of ALS (SOD1-G93A transgenic rats).