3D Bioprinted Osteogenic Tissue Models for In Vitro Drug Screening.

Metabolic bone disease affects hundreds of millions of people worldwide, and as a result, in vitro models of bone tissue have become essential tools to help analyze bone pathogenesis, develop drug screening, and test potential therapeutic strategies. Drugs that either promote or impair bone formation are in high demand for the treatment of metabolic bone diseases. These drugs work by targeting numerous signaling pathways responsible for regulating osteogenesis such as Hedgehog, Wnt/ß-catenin, and PI3K-AKT. In this study, differentiated bone marrow-derived mesenchymal stem cell (BM-MSC) scaffold-free 3D bioprinted constructs and 2D monolayer cultures were utilized to screen four drugs predicted to either promote (Icariin and Purmorphamine) or impair osteogenesis (PD98059 and U0126). Osteogenic differentiation capacity was analyzed over a four week culture period by evaluating mineralization, alkaline phosphatase (ALP) activity, and osteogenesis related gene expression. Responses to drug treatment were observed in both 3D differentiated constructs and 2D monolayer cultures. After four weeks in culture, 3D differentiated constructs and 2D monolayer cultures treated with Icariin or Purmorphamine showed increased mineralization, ALP activity, and the gene expression of bone formation markers (BGLAP, SSP1, and COL1A1), signaling molecules (MAPK1, WNT1, and AKT1), and transcription factors (RUNX2 and GLI1) that regulate osteogenic differentiation relative to untreated. 3D differentiated constructs and 2D monolayer cultures treated with PD98059 or U0126 showed decreased mineralization, ALP activity, and the expression of the aforementioned genes BGLAP, SPP1, COL1A1, MAPK1, AKT1, RUNX2, and GLI1 relative to untreated. Differences in ALP activity and osteogenesis related gene expression relative to untreated cells cultured in a 2D monolayer were greater in 3D constructs compared to 2D monolayer cultures. These findings suggest that our bioprinted bone model system offers a more sensitive, biologically relevant drug screening platform than traditional 2D monolayer in vitro testing platforms.

Crucial role for lung iron level and regulation in the pathogenesis and severity of asthma.

Accumulating evidence highlights links between iron regulation and respiratory disease. Here, we assessed the relationship between iron levels and regulatory responses in clinical and experimental asthma.We show that cell-free iron levels are reduced in the bronchoalveolar lavage (BAL) supernatant of severe or mild-moderate asthma patients and correlate with lower forced expiratory volume in 1 s (FEV1). Conversely, iron-loaded cell numbers were increased in BAL in these patients and with lower FEV1/forced vital capacity (FVC) ratio. The airway tissue expression of the iron sequestration molecules divalent metal transporter 1 (DMT1) and transferrin receptor 1 (TFR1) are increased in asthma, with TFR1 expression correlating with reduced lung function and increased Type-2 (T2) inflammatory responses in the airways. Furthermore, pulmonary iron levels are increased in a house dust mite (HDM)-induced model of experimental asthma in association with augmented Tfr1 expression in airway tissue, similar to human disease. We show that macrophages are the predominant source of increased Tfr1 and Tfr1+ macrophages have increased Il13 expression. We also show that increased iron levels induce increased pro-inflammatory cytokine and/or extracellular matrix (ECM) responses in human airway smooth muscle (ASM) cells and fibroblasts ex vivo and induce key features of asthma in vivo, including airway hyper-responsiveness (AHR) and fibrosis, and T2 inflammatory responses.Together these complementary clinical and experimental data highlight the importance of altered pulmonary iron levels and regulation in asthma, and the need for a greater focus on the role and potential therapeutic targeting of iron in the pathogenesis and severity of disease.

The regulation of ZCT1, a transcriptional repressor of monoterpenoid indole alkaloid biosynthetic genes in Catharanthus roseus.

Cys2/His2-type (C2H2) zinc finger proteins, such as ZCT1, are an important class of transcription factors involved in growth, development, and stress responses in plants. In the medicinal plant Catharanthus roseus, the zinc finger Catharanthus transcription factor (ZCT) family represses monoterpenoid indole alkaloid (MIA) biosynthetic gene expression. Here, we report the analysis of the ZCT1 promoter, which contains several hormone-responsive elements. ZCT1 is responsive to not only jasmonate, as was previously known, but is also induced by the synthetic auxin, 1-naphthalene acetic acid (1-NAA). Through promoter deletion analysis, we show that an activation sequence-1-like (as-1-like)-motif and other motifs contribute significantly to ZCT1 expression in seedlings. We also show that the activator ORCA3 does not transactivate the expression of ZCT1 in seedlings, but ZCT1 represses its own promoter, suggesting a feedback mechanism by which the expression of ZCT1 can be limited.

Rationale and design of MUSIC OS-EU: an international observational study of the treatment of postmenopausal women for osteoporosis in Europe and Canada.

OBJECTIVES: The Medication Use Patterns, Treatment Satisfaction, and Inadequate Control of Osteoporosis Study (MUSIC OS-EU) was designed to better understand the rate and burden of gastrointestinal (GI) events on clinical and health care outcomes among postmenopausal women with osteoporosis. METHODS: MUSIC OS-EU is a prospective, multinational, observational cohort study of postmenopausal women ≥50 years of age diagnosed with osteoporosis and enrolled in physician clinics in six countries: France, Italy, the Netherlands, Sweden, the United Kingdom, and Canada. The MUSIC OS-EU study has three components: (i) a physician survey to describe their management of osteoporotic patients with GI events; (ii) a retrospective chart survey to describe the receipt and type of osteoporosis medication prescribed; and (iii) a prospective cohort study including untreated and treated patients diagnosed with osteoporosis to investigate the rate of GI events and association with osteoporosis medication use patterns, health-related quality of life, treatment satisfaction and resource utilisation among postmenopausal women with osteoporosis. RESULTS: Physicians at 97 sites completed the physician questionnaire and data for 716 patients were abstracted for the retrospective chart review. Enrolment and the baseline data collection for the prospective cohort study were conducted between March 2012 and June 2013 for 292 untreated and 2,959 treated patients, of whom 684 were new users and 2,275 were experienced users of oral osteoporosis medications. CONCLUSIONS: The results of MUSIC OS-EU will illuminate the association of GI events with the management of osteoporosis and with patient-reported outcomes among postmenopausal women with osteoporosis in Europe and Canada.