Browsing by Author "Department of Anatomy & Cell Biology, IU School of Medicine"
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Item Alterations in microRNA expression profiles in inflamed and non-inflamed ascending colon mucosae of patients with active Crohn's disease(Wiley, 2017) Wu, Lu Yi; Ma, Xiao Peng; Shi, Yin; Bao, Chun Hui; Jin, Xiao Ming; Lu, Yuan; Zhao, Ji Meng; Zhou, Ci Li; Chen, Dai; Liu, Hui Rong; Department of Anatomy & Cell Biology, IU School of MedicineBackground and aims The miRNA expression profiles of the terminal ileum, sigmoid colon, and rectal mucosa of adult patients with active Crohn';s disease (CD) have been previously reported. The purpose of this study was to identify dysregulated miRNAs in the mucosa of the ascending colon. Methods Biopsy tissue samples were taken from the mucosae of inflammatory (iCD) or non-inflammatory (niCD) areas of the ascending colons of adult patients with active CD. miRNA and mRNA expression profiles were detected using microarray analyses. miRNAs and mRNAs demonstrating significant differences were validated via quantitative real-time PCR (qRT-PCR). Luciferase reporter genes were used to measure two miRNAs inhibition of potential target genes in human 293T cells in vitro. Results Compared with the HC group, the ascending colon miRNA expression profiles revealed that 43 miRNAs were significantly up-regulated and 35 were down-regulated in the iCD group. The mRNA expression profiles indicated that 3,370 transcripts were significantly differentially expressed in the ascending colon, with 2169 up-regulated and 1201 down-regulated mRNAs in the iCD group, and only 20 miRNAs demonstrated significant differential expression in the niCD group. In contrast, nearly 100 miRNAs significantly varied between the iCD and niCD groups. Finally, luciferase reporter gene assays showed that hsa-miR-16-1 directly regulated the human C10orf54 gene and that they were negatively correlated. Conclusions Our results indicated that the differentially expressed miRNAs and mRNAs were related to immune inflammation and intestinal flora. The data provide preliminary evidence that the occurrence of CD involves the inhibition of C10orf54 expression by hsa-miR-16-1.Item Antagonizing miR-218-5p attenuates Wnt signaling and reduces metastatic bone disease of triple negative breast cancer cells(Impact Journals, 2016-11-29) Taipaleenmäki, Hanna; Farina, Nicholas H.; van Wijnen, Andre J.; Stein, Janet L.; Hesse, Eric; Stein, Gary S.; Lian, Jane B.; Department of Anatomy & Cell Biology, IU School of MedicineWnt signaling is implicated in bone formation and activated in breast cancer cells promoting primary and metastatic tumor growth. A compelling question is whether osteogenic miRNAs that increase Wnt activity for bone formation are aberrantly expressed in breast tumor cells to support metastatic bone disease. Here we report that miR-218-5p is highly expressed in bone metastases from breast cancer patients, but is not detected in normal mammary epithelial cells. Furthermore, inhibition of miR-218-5p impaired the growth of bone metastatic MDA-MB-231 cells in the bone microenvironment in vivo. These findings indicate a positive role for miR-218-5p in bone metastasis. Bioinformatic and biochemical analyses revealed a positive correlation between aberrant miR-218-5p expression and activation of Wnt signaling in breast cancer cells. Mechanistically, miR-218-5p targets the Wnt inhibitors Sclerostin (SOST) and sFRP-2, which highly enhances Wnt signaling. In contrast, delivery of antimiR-218-5p decreased Wnt activity and the expression of metastasis-related genes, including bone sialoprotein (BSP/IBSP), osteopontin (OPN/SPP1) and CXCR-4, implicating a Wnt/miR-218-5p regulatory network in bone metastatic breast cancer. Furthermore, miR-218-5p also mediates the Wnt-dependent up-regulation of PTHrP, a key cytokine promoting cancer-induced osteolysis. Antagonizing miR-218-5p reduced the expression of PTHrP and Rankl, inhibited osteoclast differentiation in vitro and in vivo, and prevented the development of osteolytic lesions in a preclinical metastasis model. We conclude that pathological elevation of miR-218-5p in breast cancer cells activates Wnt signaling to enhance metastatic properties of breast cancer cells and cancer-induced osteolytic disease, suggesting that miR-218-5p could be an attractive therapeutic target for preventing disease progression.Item Antibacterial TAP-mimic electrospun polymer scaffold: effects on P. gingivalis-infected dentin biofilm(Springer, 2016-03) Albuquerque, Maria Tereza P.; Evans, Joshua D.; Gregory, Richard L.; Valera, Marcia C.; Bottino, Marco C.; Department of Anatomy & Cell Biology, IU School of MedicineOBJECTIVES: This study sought to investigate, in vitro, the effects of a recently developed triple antibiotic paste (TAP)-mimic polymer nanofibrous scaffold against Porphyromonas gingivalis-infected dentin biofilm. MATERIALS AND METHODS: Dentin specimens (4 × 4 × 1 mm(3)) were prepared from human canines. The specimens were sterilized, inoculated with P. gingivalis (ATCC 33277), and incubated for 1 week to allow for biofilm formation. Infected dentin specimens were exposed for 3 days to the following treatments: antibiotic-free polydioxanone scaffold (PDS, control), PDS + 25 wt% TAP [25 mg of each antibiotic (metronidazole, ciprofloxacin, and minocycline) per mL of the PDS polymer solution], or a saturated TAP-based solution (50 mg of each antibiotic per mL of saline solution). In order to serve as the negative control, infected dentin specimens were left untreated (bacteria only). To determine the antimicrobial efficacy of the TAP-mimic scaffold, a colony-forming unit (CFU) per milliliter (n = 10/group) measurement was performed. Furthermore, additional specimens (n = 2/group) were prepared to qualitatively study biofilm inhibition via scanning electron microscopy (SEM). Statistics were performed, and significance was set at the 5% level. RESULTS: Both the TAP-mimic scaffold and the positive control (TAP solution) led to complete bacterial elimination, differing statistically (p < 0.05) from the negative control group (bacteria only). No statistical differences were observed for CFU per milliliter data between antibiotic-free scaffolds (2.7 log10 CFU/mL) and the negative control (5.9 log10 CFU/mL). CONCLUSIONS: The obtained data revealed significant antimicrobial properties of the novel PDS-based TAP-mimic scaffold against an established P. gingivalis-infected dentin biofilm. CLINICAL RELEVANCE: Collectively, the data suggest that the proposed nanofibrous scaffold might be used as an alternative to the advocated clinical gold standard (i.e., TAP) for intracanal disinfection prior to regenerative endodontics.Item Antiremodeling Agents Influence Osteoblast Activity Differently in Modeling and Remodeling Sites of Canine Rib(Calcified Tissue International, 2006-10-10) Allen, Matthew R.; Follet, Helene; Khurana, M.; Sato, M.; Burr, David B.; Department of Anatomy & Cell Biology, IU School of MedicineAntiremodeling agents reduce bone loss in part through direct actions on osteoclasts. Their effects on osteoblasts and bone formation activity are less clear and may differ at sites undergoing modeling vs. remodeling. Skeletally mature intact beagles, 1–2 years old at the start of the study, were treated daily with clinically relevant doses of alendronate (0.10 or 0.20 mg/kg), risedronate (0.05 or 0.10 mg/kg), raloxifene (0.50 mg/kg), or vehicle (1 mL/kg). Dynamic bone formation parameters were histologically assessed on periosteal, endocortical/trabecular, and intracortical bone envelopes of the rib. Raloxifene significantly increased periosteal surface mineral apposition rate (MAR), a measure of osteoblast activity, compared to all other treatments (+108 to +175%, P < 0.02), while having no significant effect on MAR at either the endocortical/trabecular or intracortical envelope. Alendronate (both 0.10 and 0.20 doses) and risedronate (only the 0.10 dose) significantly (P ≤ 0.05) suppressed MAR on the endocortical/trabecular envelope, while none of the bisphosphonate doses significantly altered MAR at either the periosteal or intracortical envelopes compared to vehicle. Based on these results, we conclude that (1) at clinically relevant doses the two classes of antiremodeling agents, bisphosphonates and selective estrogen receptor modulators, exert differential effects on osteoblast activity in the canine rib and (2) this effect depends on whether modeling or remodeling is the predominant mechanism of bone formation.Item Apoptotic osteocytes and the control of targeted bone resorption(Springer US, 2014-03) Plotkin, Lilian I.; Department of Anatomy & Cell Biology, IU School of MedicineStudies from the 1950s and 1960s already recognize the fact that osteocytes, although long living cells, die, as evidenced by accumulation of osteocytic lacunae devoid of cells. More recently, it was demonstrated that these cells die by apoptosis. The rate of osteocyte apoptosis is regulated by the age of the bone, as well as by systemic hormones, local growth factors, cytokines, pharmacological agents, and mechanical forces. Apoptotic osteocytes, in turn, recruit osteoclasts to initiate targeted bone resorption. This results in the removal of “dead” bone and may improve the mechanical properties of the skeleton. However, the molecular regulators of osteocyte survival and targeted bone remodeling are not completely known. In this review, the current knowledge on the molecular mechanism that lead to osteocyte death or survival, and the signals that mediate targeted bone resorption is discussed.Item Assessment of regional bone tissue perfusion in rats using fluorescent microspheres(Elsevier, 2017-04-26) Aref, Mohammad W.; Akans, Ethan; Allen, Matthew R.; Department of Anatomy & Cell Biology, IU School of MedicineDisturbances in bone blood flow have been shown to have deleterious effects on bone properties yet there remain many unanswered questions about skeletal perfusion in health and disease, partially due to the complexity of measurement methodologies. The goal of this study was use fluorescent microspheres in rats to assess regional bone perfusion by adapting mouse-specific fluorescent microsphere protocol. Ten fifteen-week old Sprague Dawley rats were injected with fluorescent microspheres either via cardiac injection (n = 5) or via tail vein injection (n = 5). Femora and tibiae were harvested and processed to determine tissue fluorescence density (TFD) which is proportional to the number of spheres trapped in the tissue capillaries. Right and left total femoral TFD (2.77 ± 0.38 and 2.70 ± 0.24, respectively) and right and left tibial TFD (1.11 ± 0.26 and 1.08 ± 0.34, respectively) displayed bilateral symmetry in flow when assessed in cardiac injected animals. Partitioning of the bone perfusion into three segments along the length of the bone showed the distal femur and proximal tibia received the greatest amount of perfusion within their respective bones. Tail vein injection resulted in unacceptably low TFD levels in the tibia from 4 of the 5 animals. In conclusion this report demonstrates the viability of cardiac injection of fluorescent microspheres to assess bone tissue perfusion in rats.Item Avenanthramides Prevent Osteoblast and Osteocyte Apoptosis and Induce Osteoclast Apoptosis in Vitro in an Nrf2-Independent Manner(MDPI, 2016-07-11) Pellegrini, Gretel G.; Morales, Cynthya C.; Wallace, Taylor C.; Plotkin, Lilian I.; Bellido, Teresita; Department of Anatomy & Cell Biology, IU School of MedicineOats contain unique bioactive compounds known as avenanthramides (AVAs) with antioxidant properties. AVAs might enhance the endogenous antioxidant cellular response by activation of the transcription factor Nrf2. Accumulation of reactive oxygen species plays a critical role in many chronic and degenerative diseases, including osteoporosis. In this disease, there is an imbalance between bone formation by osteoblasts and bone resorption by osteoclasts, which is accompanied by increased osteoblast/osteocyte apoptosis and decreased osteoclast apoptosis. We investigated the ability of the synthethic AVAs 2c, 2f and 2p, to 1-regulate gene expression in bone cells, 2-affect the viability of osteoblasts, osteocytes and osteoclasts, and the generation of osteoclasts from their precursors, and 3-examine the potential involvement of the transcription factor Nrf2 in these actions. All doses of AVA 2c and 1 and 5 µM dose of 2p up-regulated collagen 1A expression. Lower doses of AVAs up-regulated OPG (osteoprotegerin) in OB-6 osteoblastic cells, whereas 100 μM dose of 2f and all concentrations of 2c down-regulated RANKL gene expression in MLO-Y4 osteocytic cells. AVAs did not affect apoptosis of OB-6 osteoblastic cells or MLO-Y4 osteocytic cells; however, they prevented apoptosis induced by the DNA topoisomerase inhibitor etoposide, the glucocorticoid dexamethasone, and hydrogen peroxide. AVAs prevented apoptosis of both wild type (WT) and Nrf2 Knockout (KO) osteoblasts, demonstrating that AVAs-induced survival does not require Nrf2 expression. Further, KO osteoclast precursors produced more mature osteoclasts than WT; and KO cultures exhibited less apoptotic osteoclasts than WT cultures. Although AVAs did not affect WT osteoclasts, AVA 2p reversed the low apoptosis of KO osteoclasts. These in vitro results demonstrate that AVAs regulate, in part, the function of osteoblasts and osteocytes and prevent osteoblast/osteocyte apoptosis and increase osteoclast apoptosis; further, these regulatory actions are independent of Nrf2.Item Bidirectional Notch signaling and osteocyte-derived factors in the bone marrow microenvironment promote tumor cell proliferation and bone destruction in multiple myeloma(American Association for Cancer Research, 2016-03-01) Delgado-Calle, Jesus; Anderson, Judith; Cregor, Meloney D.; Hiasa, Masahiro; Chirgwin, John M.; Carlesso, Nadia; Yoneda, Toshiyuki; Mohammad, Khalid S.; Plotkin, Lilian I.; Roodman, G. David; Bellido, Teresita; Department of Anatomy & Cell Biology, IU School of MedicineIn multiple myeloma, an overabundance of monoclonal plasma cells in the bone marrow induces localized osteolytic lesions that rarely heal due to increased bone resorption and suppressed bone formation. Matrix-embedded osteocytes comprise more than 95% of bone cells and are major regulators of osteoclast and osteoblast activity, but their contribution to multiple myeloma growth and bone disease is unknown. Here, we report that osteocytes in a mouse model of human MM physically interact with multiple myeloma cells in vivo, undergo caspase-3-dependent apoptosis, and express higher RANKL (TNFSF11) and sclerostin levels than osteocytes in control mice. Mechanistic studies revealed that osteocyte apoptosis was initiated by multiple myeloma cell-mediated activation of Notch signaling and was further amplified by multiple myeloma cell-secreted TNF. The induction of apoptosis increased osteocytic Rankl expression, the osteocytic Rankl/Opg (TNFRSF11B) ratio, and the ability of osteocytes to attract osteoclast precursors to induce local bone resorption. Furthermore, osteocytes in contact with multiple myeloma cells expressed high levels of Sost/sclerostin, leading to a reduction in Wnt signaling and subsequent inhibition of osteoblast differentiation. Importantly, direct contact between osteocytes and multiple myeloma cells reciprocally activated Notch signaling and increased Notch receptor expression, particularly Notch3 and 4, stimulating multiple myeloma cell growth. These studies reveal a previously unknown role for bidirectional Notch signaling that enhances MM growth and bone disease, suggesting that targeting osteocyte-multiple myeloma cell interactions through specific Notch receptor blockade may represent a promising treatment strategy in multiple myeloma.Item Biopsy proven medullary sponge kidney: clinical findings, histopathology, and role of osteogenesis in stone and plaque formation(John Wiley & Sons, Inc., 2015-05) Evan, Andrew P.; Worcester, Elaine M.; Williams, James C., Jr.; Sommer, Andre J.; Lingeman, James E.; Phillips, Carrie L.; Coe, Fredric L.; Department of Anatomy & Cell Biology, IU School of MedicineMedullary sponge kidney (MSK) is associated with recurrent stone formation, but the clinical phenotype is unclear because patients with other disorders may be incorrectly labeled MSK. We studied 12 patients with histologic findings pathognomonic of MSK. All patients had an endoscopically recognizable pattern of papillary malformation, which may be segmental or diffuse. Affected papillae are enlarged and billowy, due to markedly enlarged inner medullary collecting ducts (IMCD), which contain small, mobile ductal stones. Patients had frequent dilation of Bellini ducts, with occasional mineral plugs. Stones may form over white (Randall's) plaque, but most renal pelvic stones are not attached, and have a similar morphology as ductal stones, which are a mixture of calcium oxalate and apatite. Patients had no abnormalities of urinary acidification or acid excretion; the most frequent metabolic abnormality was idiopathic hypercalciuria. Although both Runx2 and Osterix are expressed in papillae of MSK patients, no mineral deposition was seen at the sites of gene expression, arguing against a role of these genes in this process. Similar studies in idiopathic calcium stone formers showed no expression of these genes at sites of Randall's plaque. The most likely mechanism for stone formation in MSK appears to be crystallization due to urinary stasis in dilated IMCD with subsequent passage of ductal stones into the renal pelvis where they may serve as nuclei for stone formation.Item Bone and skeletal muscle: Key players in mechanotransduction and potential overlapping mechanisms(Elsevier, 2015-11) Goodman, Craig A.; Hornberger, Troy A.; Robling, Alexander G.; Department of Anatomy & Cell Biology, IU School of MedicineThe development and maintenance of skeletal muscle and bone mass is critical for movement, health and issues associated with the quality of life. Skeletal muscle and bone mass are regulated by a variety of factors that include changes in mechanical loading. Moreover, bone mass is, in large part, regulated by muscle-derived mechanical forces and thus by changes in muscle mass/strength. A thorough understanding of the cellular mechanism(s) responsible for mechanotransduction in bone and skeletal muscle is essential for the development of effective exercise and pharmaceutical strategies aimed at increasing, and/or preventing the loss of, mass in these tissues. Thus, in this review we will attempt to summarize the current evidence for the major molecular mechanisms involved in mechanotransduction in skeletal muscle and bone. By examining the differences and similarities in mechanotransduction between these two tissues, it is hoped that this review will stimulate new insights and ideas for future research and promote collaboration between bone and muscle biologists.(1).