scholarly journals Ovariectomy-induced bone loss in TNFα and IL6 gene knockout mice is regulated by different mechanisms

2018 ◽  
Vol 60 (3) ◽  
pp. 185-198 ◽  
Author(s):  
Siyi Zhu ◽  
Hongchen He ◽  
Chengfei Gao ◽  
Guojing Luo ◽  
Ying Xie ◽  
...  
Keyword(s):  
Bone Loss ◽  
Bone Metabolism ◽  
Gene Knockout ◽  
High Turnover ◽  
Trabecular Thickness ◽  
Gene Knockout Mice ◽  
Trabecular Number ◽  
Il6 Gene

We examined the effects of tumor necrosis factor-α (TNFα) and interleukin-6 (IL6) gene knockout in preserving the bone loss induced by ovariectomy (OVX) and the mechanisms involved in bone metabolism. Twenty female wild-type (WT), TNFα-knockout (TNFα−/−) or IL6-knockout (IL6−/−) mice aged 12 weeks were sham-operated (SHAM) or subjected to OVX and killed after 4 weeks. Bone mass and skeletal microarchitecture were determined using micro-CT. Bone marrow stromal cells (BMSCs) from all three groups (WT, TNFα−/− and IL6−/−) were induced to differentiate into osteoblasts or osteoclasts and treated with 17-β-estradiol. Bone metabolism was assessed by histological analysis, serum analyses and qRT-PCR. OVX successfully induced a high turnover in all mice, but a repair effect was observed in TNFα−/− and IL6−/− mice. The ratio of femoral trabecular bone volume to tissue volume, trabecular number and trabecular thickness were significantly decreased in WT mice subjected to OVX, but increased in TNFα−/− mice (1.62, 1.34, 0.27-fold respectively; P < 0.01) and IL6−/− mice (1.34, 0.80, 0.22-fold respectively; P < 0.01). Furthermore, we observed a 29.6% increase in the trabecular number in TNFα−/− mice when compared to the IL6−/− mice. Both, TNFα−/− and IL6−/− BMSCs exhibited decreased numbers of TRAP-positive cells and an increase in ALP-positive cells, with or without E2 treatment (P < 0.05). While the knockout of TNFα or IL6 significantly upregulated mRNA expressions of osteoblast-related genes (Runx2 and Col1a1) and downregulated osteoclast-related mRNA for TRAP, MMP9 and CTSK in vivo and in vitro, TNFα knockout appeared to have roles beyond IL6 knockout in upregulating Col1a1 mRNA expression and downregulating mRNA expressions of WNT-related genes (DKK1 and Sost) and TNF-related activation-induced genes (TRAF6). TNFα seemed to be more potentially invasive in inhibiting bone formation and enhancing TRAF6-mediated osteoclastogenesis than IL6, implying that the regulatory mechanisms of TNFα and IL6 in bone metabolism may be different.

Tenascin-X expression in tumor cells and fibroblasts: glucocorticoids as negative regulators in fibroblasts

1996 ◽  
Vol 109 (8) ◽  
pp. 2069-2077 ◽  
Author(s):  
T. Sakai ◽  
Y. Furukawa ◽  
R. Chiquet-Ehrismann ◽  
M. Nakamura ◽  
S. Kitagawa ◽  
...  
Keyword(s):  
Tumor Cells ◽  
Gene Knockout ◽  
Tumor Stroma ◽  
Carcinoma Cells ◽  
Mrna Levels ◽  
X Protein ◽  
Tenascin C ◽  
Gene Knockout Mice

Tenascin-X has recently been shown to be a novel member of the tenascin family and its distribution is often reciprocal to that of tenascin-C in the developing mouse embryo. We have investigated the expression of tenascin-X in fibroblasts and carcinoma cells in culture. Tenascin-X protein was secreted in vitro in the conditioned media at an apparent molecular mass of approximately 450 kDa. In addition fibroblasts contained a major tenascin-X isoform of 220 kDa. On northern blots, a single major transcript with a size of approximately 13 kb was detected. No overexpression of tenascin-X protein was found in primary fibroblasts of the tenascin-C-gene knockout mice. Steroid hormone glucocorticoids, were found to downregulate tenascin-X mRNA levels and protein synthesis in fibroblasts but not carcinoma cells at physiological concentrations. None of the growth factors or cytokines examined affected the expression level of tenascin-X. As in vivo study, carcinoma cells were transplanted into nude mice. In contrast to the ubiquitous presence of tenascin-X in adult skin, expression of tenascin-X protein during tumorigenesis was found to be down-regulated considerably not only in tumor cells themselves but also in tumor stroma. These findings provide evidence that the expression of tenascin-X can be influenced by stromal-epithelial interactions. We have identified glucocorticoids as physiological inhibitors of tenascin-X and suggest that glucocorticoids may in part participate in the downregulation of tenascin-X in fibroblasts in vivo.

scholarly journals The Impact of Microgravity on Bone Metabolism in vitro and in vivo

2001 ◽  
Vol 12 (3) ◽  
pp. 252-261 ◽  
Author(s):  
Peter M. Loomer
Keyword(s):  
Bone Loss ◽  
Bone Mass ◽  
Bone Metabolism ◽  
Bone Cells ◽  
Mechanical Load ◽  
Space Environment ◽  
In Vivo Studies ◽  
The Impact

Exposure to microgravity has been associated with several physiological changes in astronauts and cosmonauts, including an osteoporosis-like loss of bone mass. In-flight measures used to counteract this, including intensive daily exercise regimens, have been only partially successful in reducing the bone loss and in the process have consumed valuable work time. If this bone loss is to be minimized or, preferably, prevented, more effective treatment strategies are required. This, however, requires a greater understanding of the mechanisms through which bone metabolism is affected by microgravity. Various research strategies have been used to examine this problem, including in vitro studies using bone cells and in vivo studies on humans and rats. These have been conducted both in flight and on the ground, by strategies that produce weightlessness to mimic the effects of microgravity. Overall, the majority of the studies have found that marked decreases in gravitation loading result in the loss of bone mass. The processes of bone formation and bone resorption become uncoupled, with an initial transitory increase in resorption accompanied by a prolonged decrease in formation. Loss of bone mass is not uniform throughout the skeleton, but varies at different sites depending on the type of bone and on the mechanical load received. It appears that the skeletal response is a physiologic adaptation to the space environment which, after long space flights or repeated shorter ones, could eventually lead to significant reductions in the ability of the skeletal tissues to withstand the forces of gravity and increased susceptibility to fracture.

scholarly journals miR-203 Regulates Nociceptive Sensitization after Incision by Controlling Phospholipase A2 Activating Protein Expression

2012 ◽  
Vol 117 (3) ◽  
pp. 626-638 ◽  
Author(s):  
Yuan Sun ◽  
Xiang-Qi Li ◽  
Peyman Sahbaie ◽  
Xiao-You Shi ◽  
Wen-Wu Li ◽  
...  
Keyword(s):  
Substance P ◽  
Phospholipase A2 ◽  
Mechanical Allodynia ◽  
Gene Knockout ◽  
Skin Inflammation ◽  
Local Effect ◽  
Intraplantar Injection ◽  
Gene Knockout Mice

Background After incision keratinocytes in the epidermis become activated to produce a range of pain-related mediators. microRNA 203 (miR-203) is known to be involved in keratinocyte growth, differentiation, and skin inflammation. We hypothesized that one or more of these mediators might be under the control of miR-203. Methods The expression of miR-203 and its target gene, phospholipase A2 activating protein (PLAA), were examined after hind paw incision in mice. We investigated the local effect of intraplantar PLAA peptide injection in normal mice and the effects of a selective secretory phospholipase A2 inhibitor (HK064) on PLAA or incision-induced mechanical allodynia. Last, we investigated the role of substance P signaling in regulating miR-203 and PLAA expression in vitro and in vivo. Results Levels of miR-203 were strongly down-regulated in keratinocytes after incision. Informatics-based approaches identified PLAA as a likely candidate for regulation by miR-203. PLAA caused mechanical allodynia and conditioned place aversion but not thermal sensitization. HK064 reduced mechanical allodynia after incision and after intraplantar injection of PLAA. Using preprotachykinin gene knockout mice or with neurokinin-1 selective antagonist LY303870 treatment, we observed that substance P-mediated signaling was also required for miR-203 and PLAA regulation after incision. Finally, using the rat epidermal keratinocyte cell line, we observed that a miR-203 mimic molecule could block the substance P-induced increase in PLAA expression observed under control conditions. Conclusions miR-203 may regulate expression of the novel nociceptive mediator PLAA after incision. Furthermore, the regulation of miR-203 and PLAA levels is reliant upon intact substance P signaling.

Mammalian multidrug-resistance proteins (MRPs)

2011 ◽  
Vol 50 ◽  
pp. 179-207 ◽  
Author(s):  
Andrew J. Slot ◽  
Steven V. Molinski ◽  
Susan P.C. Cole
Keyword(s):  
Multidrug Resistance ◽  
Gene Knockout ◽  
Multidrug Resistance Proteins ◽  
Resistance Proteins ◽  
Gene Knockout Mice ◽  
Membrane Spanning ◽  
Endogenous Metabolites ◽  
Membrane Spanning Domains

Subfamily C of the human ABC (ATP-binding cassette) superfamily contains nine proteins that are often referred to as the MRPs (multidrug-resistance proteins). The ‘short’ MRP/ABCC transporters (MRP4, MRP5, MRP8 and ABCC12) have a typical ABC structure with four domains comprising two membrane-spanning domains (MSD1 and MSD2) each followed by a nucleotide-binding domain (NBD1 and NBD2). The ‘long’ MRP/ABCCs (MRP1, MRP2, MRP3, ABCC6 and MRP7) have five domains with the extra domain, MSD0, at the N-terminus. The proteins encoded by the ABCC6 and ABCC12 genes are not known to transport drugs and are therefore referred to as ABCC6 and ABCC12 (rather than MRP6 and MRP9) respectively. A large number of molecules are transported across the plasma membrane by the MRPs. Many are organic anions derived from exogenous sources such as conjugated drug metabolites. Others are endogenous metabolites such as the cysteinyl leukotrienes and prostaglandins which have important signalling functions in the cell. Some MRPs share a degree of overlap in substrate specificity (at least in vitro), but differences in transport kinetics are often substantial. In some cases, the in vivo substrates for some MRPs have been discovered aided by studies in gene-knockout mice. However, the molecules that are transported in vivo by others, including MRP5, MRP7, ABCC6 and ABCC12, still remain unknown. Important differences in the tissue distribution of the MRPs and their membrane localization (apical in contrast with basolateral) in polarized cells also exist. Together, these differences are responsible for the unique pharmacological and physiological functions of each of the nine ABCC transporters known as the MRPs.

scholarly journals Effects of ANG II on bradykinin receptor gene expression in cardiomyocytes and vascular smooth muscle cells

2001 ◽  
Vol 281 (4) ◽  
pp. H1778-H1783 ◽  
Author(s):  
Ekaterina Kintsurashvili ◽  
Irena Duka ◽  
Irene Gavras ◽  
Conrado Johns ◽  
Dimitrios Farmakiotis ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gene Knockout ◽  
Receptor Gene ◽  
Ang Ii ◽  
Wild Type ◽  
Receptor Mrna ◽  
Gene Knockout Mice ◽  
Receptor Gene Expression

Bradykinin has vasodilatory and tissue-protective effects exerted via its B2 type receptor, whereas the B1 receptor is constitutively absent but inducible by inflammation and toxins. In previous studies, we found that B2 receptor gene knockout mice exhibit overexpression of the B1 receptor, which assumes a vasodilatory function and is further upgraded in renovascular hypertension. The present study was designed to explore the effects of excess angiotensin II (ANG II) on B1 receptor and B2 receptor gene expression in mouse cardiomyocytes and rat vascular smooth muscle cells (VSMC) in vivo (after a 3-day infusion of 30 ng/min ANG II in 11 wild-type and in 13 genetically engineered mice with deleted B2 receptor gene) and in vitro (ANG II added in rat VSMC culture in the presence or absence of AT1 or AT2 receptor antagonist). Expression of B1 and B2 receptor mRNA was assessed by reverse transcriptase-polymerase chain reaction. ANG II infusion caused upregulation by 30% of the already significantly overexpressed B1 receptors in cardiomyocytes of the B2receptor gene knockout mice, but in the wild-type mice it upregulated only the B2 receptor mRNA by 47%. The addition of ANG II in VSMC culture produced a time-dependent induction of B1and upregulation of B2 receptor gene expression, maximal at 3 h (by fivefold), declining almost to baseline by 24 h. The addition of losartan completely blocked this effect, whereas the AT2 blocker PD-123319 made no difference, indicating that this is an AT1-mediated effect of ANG II. The data indicate that excess ANG II in subpressor doses in vivo upregulates expression of the B2 receptor, but in its absence, the already overexpressed B1 receptor is further upregulated, evidently assuming a counterregulatory response; in vitro, it transiently upregulates both bradykinin receptors.

scholarly journals Eleutherococcus sessiliflorus Inhibits Receptor Activator of Nuclear Factor Kappa-B Ligand (RANKL)-Induced Osteoclast Differentiation and Prevents Ovariectomy (OVX)-Induced Bone Loss

Molecules ◽  
2021 ◽  
Vol 26 (7) ◽  
pp. 1886
Author(s):  
Sang-Yong Han ◽  
June-Hyun Kim ◽  
Eun-Heui Jo ◽  
Yun-Kyung Kim
Keyword(s):  
Bone Loss ◽  
Transmembrane Protein ◽  
Osteoclast Differentiation ◽  
Mapk Signaling ◽  
Root Bark ◽  
Nuclear Factor ◽  
Mineral Density ◽  
Trabecular Thickness

The aim of this study was to evaluate the effects of root bark of Eleutherococcus sessiliflorus (ES) on osteoclast differentiation and function in vitro and in vivo. In vitro, we found that ES significantly inhibited the RANKL-induced formation of TRAP-positive multinucleated osteoclasts and osteoclastic bone resorption without cytotoxic effects. ES markedly downregulated the expression of nuclear factor of activated T cells cytoplasmic 1 (NFATc1); c-Fos; and osteoclast-related marker genes, such as TRAP, osteoclast-associated receptor (OSCAR), matrix metalloproteinase-9 (MMP-9), calcitonin receptor, cathepsin K, the 38 kDa d2 subunit of the vacuolar H+-transporting lysosomal ATPase (Atp6v0d2), dendritic cell-specific transmembrane protein (DC-STAMP), and osteoclast-stimulatory transmembrane protein (OC-STAMP). These effects were achieved by inhibiting the RANKL-mediated activation of MAPK signaling pathway proteins, including p38, ERK, and JNK. In vivo, ES attenuated OVX-induced decrease in bone volume to tissue volume ratio (BV/TV), trabecular thickness (Tb.Th), trabecular number (Tb.N), and bone mineral density, but increased trabecular separation (Tb.Sp) in the femur. Collectively, our findings showed that ES inhibited RANKL-activated osteoclast differentiation in bone marrow macrophages and prevented OVX-mediated bone loss in rats. These findings suggest that ES has the potential to be used as a therapeutic agent for bone-related diseases, such as osteoporosis.

scholarly journals Curculigoside Protects against Excess-Iron-Induced Bone Loss by Attenuating Akt-FoxO1-Dependent Oxidative Damage to Mice and Osteoblastic MC3T3-E1 Cells

2019 ◽  
Vol 2019 ◽  
pp. 1-14 ◽  
Author(s):  
Quanlong Zhang ◽  
Lu Zhao ◽  
Yi Shen ◽  
Yuqiong He ◽  
Gang Cheng ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Antioxidant Enzymes ◽  
Bone Loss ◽  
Iron Overload ◽  
Cell Viability ◽  
Bone Metabolism ◽  
Primary Osteoporosis ◽  
Excess Iron

Summary. The present investigation found that curculigoside (CUR) can prevent excess-iron-induced bone loss in mice and cells through antioxidation and inhibiting excess-iron-induced phosphorylation of the Akt-FoxO1 pathway. CUR can attenuate the decreasing of cell viability, enhance autophagy, potentiate the antioxidant effect, and reduce apoptosis in MC3T3-E1 cells treated with excess iron through regulating the expression of FoxO1 target gene. Introduction. Oxidative stress induced by iron overload is an important factor involved in primary osteoporosis disease and iron overload-related diseases. Curculigoside (CUR), a phenolic glycoside found abundantly in Curculigo orchioides Gaertn., has been demonstrated to possess antioxidant and antiosteoporotic properties. The aim of the present study is to explore the underlying molecular mechanism of CUR on excess-iron-induced bone loss in mice and osteoblastic MC3T3-E1 cells. Methods. An iron-overload mice model was used to study the protective effects of CUR on bone loss induced by oxidative stress. Serum bone metabolism markers and antioxidant enzymes were also measured. To explore the antioxidant mechanism of CUR, the MC3T3-E1 osteoblastic cell line was used. Results. In vivo studies showed that BMD and microarchitectural parameters were improved after a 3-month administration of CUR. CUR improved the biochemical parameters related to bone metabolism and the expressions of Runx2, OCN, and type 1 collagen and increased the formation of bone-mineralized nodules in vitro. CUR also inhibited ROS generation and increased the activities of antioxidant enzymes both in vivo and in vitro treated with excess iron. CUR can upregulate the level of FoxO1 and Nrf2, downregulate the level of p53 and the phosphorylation level of FoxO1, improve nuclear translocation of FoxO1, probably by inhibiting the IGFR/AKT signaling pathway, then increased cell viability and autophagy, and reduced apoptosis of MC3T3-E1 cells treated with excess iron by regulating the expression of FoxO1 target genes MnSOD, Gadd45a, Bim, FasL, and Rab7. Conclusions. These results demonstrated that CUR was able to alleviate bone loss induced by oxidative stress resulting from iron overload, suggesting its potential use for the treatment of primary osteoporosis and bone loss in iron-overload-related diseases.

scholarly journals Reversal of Proinflammatory Responses by Ligating the Macrophage Fcγ Receptor Type I

1998 ◽  
Vol 188 (1) ◽  
pp. 217-222 ◽  
Author(s):  
Fayyaz S. Sutterwala ◽  
Gary J. Noel ◽  
Padmini Salgame ◽  
David M. Mosser
Keyword(s):  
Gene Knockout ◽  
Reciprocal Inhibition ◽  
Type I ◽  
In Vivo Models ◽  
Inflammatory Stimuli ◽  
Proinflammatory Responses ◽  
Gene Knockout Mice ◽  
Macrophage Cytokine

Macrophages can respond to a variety of infectious and/or inflammatory stimuli by secreting an array of proinflammatory cytokines, the overproduction of which can result in shock or even death. In this report, we demonstrate that ligation of macrophage Fcγ receptors (FcγR) can lead to a reversal of macrophage proinflammatory responses by inducing an upregulation of interleukin (IL)-10, with a reciprocal inhibition of IL-12 production. IL-10 upregulation was specific to FcγR ligation, since the ligation of the Mac-1 receptor did not alter IL-10 production. The identification of the specific FcγR subtype responsible for IL-10 upregulation was determined in gene knockout mice. Macrophages from mice lacking the FcR γ chain, which is required for assembly and signaling by FcγRI and FcγRIII, failed to upregulate IL-10 in response to immune complexes. However, mice lacking either the FcγRII or the FcγRIII were fully capable of upregulating IL-10 production, implicating FcγRI in this process. The biological consequences of FcγRI ligation were determined in both in vitro and in vivo models of inflammation and sepsis. In all of the models tested, the ligation of FcγR promoted the production of IL-10 and inhibited the secretion of IL-12. This reciprocal alteration in the pattern of macrophage cytokine production illustrates a potentially important role for FcγR-mediated clearance in suppressing macrophage proinflammatory responses.

Effects of TSH and calcitriol on bone metabolism: in vivo and in vitro study

2014 ◽  
Author(s):  
Ivo Dumic-Cule ◽  
Dunja Rogic ◽  
Damir Jezek ◽  
Lovorka Grgurevic ◽  
Slobodan Vukicevic
Keyword(s):  
Bone Metabolism ◽  
In Vitro Study ◽  
Vitro Study

scholarly journals A novel BRD4 inhibitor suppresses osteoclastogenesis and ovariectomized osteoporosis by blocking RANKL-mediated MAPK and NF-κB pathways

2021 ◽  
Vol 12 (7) ◽  
Author(s):  
Ying Liu ◽  
Wenjie Liu ◽  
Ziqiang Yu ◽  
Yan Zhang ◽  
Yinghua Li ◽  
...  
Keyword(s):  
Bone Loss ◽  
Osteoporosis Treatment ◽  
Inhibitory Effect ◽  
Specific Gene ◽  
Actin Ring ◽  
Treatment Target ◽  
Significant Inhibitory Effect ◽  
Promising Treatment

AbstractBromodomain-containing protein 4 (BRD4) has emerged as a promising treatment target for bone-related disorders. (+)-JQ1, a thienotriazolodiazepine compound, has been shown to inhibit pro-osteoclastic activity in a BRD4-dependent approach and impede bone loss caused by ovariectomy (OVX) in vivo. However, clinical trials of (+)-JQ1 are limited because of its poor druggability. In this study, we synthesized a new (+)-JQ1 derivative differing in structure and chirality. One such derivative, (+)-ND, exhibited higher solubility and excellent inhibitory activity against BRD4 compared with its analogue (+)-JQ1. Interestingly, (-)-JQ1 and (-)-ND exhibited low anti-proliferative activity and had no significant inhibitory effect on RANKL-induced osteoclastogenesis as compared with (+)-JQ1 and (+)-ND, suggesting the importance of chirality in the biological activity of compounds. Among these compounds, (+)-ND displayed the most prominent inhibitory effect on RANKL-induced osteoclastogenesis. Moreover, (+)-ND could inhibit osteoclast-specific gene expression, F‐actin ring generation, and bone resorption in vitro and prevent bone loss in OVX mice. Collectively, these findings indicated that (+)-ND represses RANKL‐stimulated osteoclastogenesis and averts OVX-triggered osteoporosis by suppressing MAPK and NF-κB signalling cascades, suggesting that it may be a prospective candidate for osteoporosis treatment.

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