Osteoblast function in patients with idiopathic osteonecrosis of the femoral head

Aims To investigate whether idiopathic osteonecrosis of the femoral head (ONFH) is related to impaired osteoblast activities. Methods We cultured osteoblasts isolated from trabecular bone explants taken from the femoral head and the intertrochanteric region of patients with idiopathic ONFH, or from the intertrochanteric region of patients with osteoarthritis (OA), and compared their viability, mineralization capacity, and secretion of paracrine factors. Results Osteoblasts from the intertrochanteric region of patients with ONFH showed lower alkaline phosphatase (ALP) activity and mineralization capacity than osteoblasts from the same skeletal site in age-matched patients with OA, as well as lower messenger RNA (mRNA) levels of genes encoding osteocalcin and bone sialoprotein and higher osteopontin expression. In addition, osteoblasts from patients with ONFH secreted lower osteoprotegerin (OPG) levels than those from patients with OA, resulting in a higher receptor activator of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) ligand (RANKL)-to-OPG ratio. In patients with ONFH, osteoblasts from the femoral head showed reduced viability and mineralized nodule formation compared with osteoblasts from the intertrochanteric region. Notably, the secretion of the pro-resorptive factors interleukin-6 and prostaglandin E2 as well as the RANKL-to-OPG ratio were markedly higher in osteoblast cultures from the femoral head than in those from the intertrochanteric region. Conclusion Idiopathic ONFH is associated with a reduced mineralization capacity of osteoblasts and increased secretion of pro-resorptive factors. Cite this article: Bone Joint Res 2021;10(9):619–628.

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Breast Cancer Cells Interact with Osteoblasts to Support Osteoclast Formation1

Endocrinology ◽

10.1210/endo.140.10.7037 ◽

1999 ◽

Vol 140 (10) ◽

pp. 4451-4458 ◽

Cited By ~ 254

Author(s):

Rachel J. Thomas ◽

Theresa A. Guise ◽

Juan Juan Yin ◽

Jan Elliott ◽

Nicole J. Horwood ◽

...

Keyword(s):

Breast Cancer ◽

Messenger Rna ◽

Cancer Metastasis ◽

Osteoclast Differentiation ◽

Osteoclast Formation ◽

Breast Cancer Cell Lines ◽

Mrna Levels ◽

Breast Cancers ◽

Receptor Activator ◽

Mcf 7

Abstract Breast cancers commonly cause osteolytic metastases in bone, a process that is dependent upon osteoclast-mediated bone resorption. Recently the osteoclast differentiation factor (ODF), better termed RANKL (receptor activator of NF-κB ligand), expressed by osteoblasts has been cloned as well as its cognate signaling receptor, receptor activator of NFκB (RANK), and a secreted decoy receptor osteoprotegerin (OPG) that limits RANKL’s biological action. We determined that the breast cancer cell lines MDA-MB-231, MCF-7, and T47D as well as primary breast cancers do not express RANKL but express OPG and RANK. MCF-7, MDA-MB-231, and T47D cells did not act as surrogate osteoblasts to support osteoclast formation in coculture experiments, a result consistent with the fact that they do not express RANKL. When MCF-7 cells overexpressing PTH-related protein (PTHrP) were added to cocultures of murine osteoblasts and hematopoietic cells, osteoclast formation resulted without the addition of any osteotropic agents; cocultures with MCF-7 or MCF-7 cells transfected with pcDNAIneo required exogenous agents for osteoclast formation. When MCF-7 cells overexpressing PTHrP were cultured with murine osteoblasts, osteoblastic RANKL messenger RNA (mRNA) levels were enhanced and osteoblastic OPG mRNA levels diminished; MCF-7 parental cells had no effect on RANKL or OPG mRNA levels when cultured with osteoblastic cells. Using a murine model of breast cancer metastasis to bone, we established that MCF-7 cells that overexpress PTHrP caused significantly more bone metastases, which were associated with increased osteoclast formation, elevated plasma PTHrP concentrations and hypercalcaemia compared with parental or empty vector controls.

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Messenger RNA levels of enzymes involved in glycerolipid synthesis in the brain of the mouse and its alterations in Agpat2-/- and db/db mice

10.21203/rs.3.rs-128675/v1 ◽

2020 ◽

Author(s):

Lila Gonzalez-Hodar ◽

Anil K. Agarwal ◽

Víctor Cortés

Keyword(s):

Mouse Brain ◽

Messenger Rna ◽

Brain Regions ◽

Mrna Levels ◽

Expression Of Genes ◽

Genes Encoding ◽

Glycerolipid Synthesis ◽

Altered Gene ◽

The Brain

Abstract Background and Aims: Expression of genes encoding enzymes involved in glycerolipid and monoacylglycerol pathways in specific brain regions is poorly known and their alterations in insulin resistance (IR) and type 2 diabetes (T2D) remain unreported. We determined the mRNA levels of enzymes involved in glycerolipid synthesis in specific regions of the mouse brain and their changes in two models of severe IR, the lipodystrophic Agpat2−/− and the obese Leprdb/db mice. Methods Cerebral cortex, hypothalamus, hippocampus and cerebellum were dissected from adult Agpat2−/− mice, Leprdb/db mice and their respective wild type littermates. Total RNA was isolated and the relative mRNA abundance of enzymes was determined by RT-qPCR. Results GPAT1, AGPAT1-4, LIPIN1/2, DGAT1/2 and MOGAT1 mRNAs were detected in all studied brain regions, whereas GPAT2, LIPIN3 and MOGAT2 were undetectable. Abundance of GPAT1, AGPAT1, AGPAT2, AGPAT4, LIPIN1, and MOGAT1, was higher in the hypothalamus. AGPAT3 and DGAT1 were higher in cortex and cerebellum, and LIPIN2 and DGAT2 were higher in cortex and hippocampus. In Agpat2−/− mice, LIPIN1 levels were increased in all the brain regions. By contrast, GPAT1 and AGPAT4 in hypothalamus, AGPAT3 in hippocampus and hypothalamus, and MOGAT1 in cortex, hypothalamus and cerebellum were lower in Agpat2−/− mice. Leprdb/db mice showed fewer and milder changes, with increased levels of GPAT1 and LIPIN1 in cerebellum, and AGPAT3 in hypothalamus. Conclusions Enzymes involved in glycerolipids synthesis are differentially expressed across regions of the mouse brain and IR and T2D determine altered gene expression of these enzymes in the mouse brain.

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Messenger RNA levels of enzymes involved in glycerolipid synthesis in the brain of the mouse and its alterations in Agpat2-/- and db/db mice

10.1101/2020.04.02.021808 ◽

2020 ◽

Author(s):

Lila González-Hódar ◽

Anil K. Agarwal ◽

Víctor Cortés

Keyword(s):

Mouse Brain ◽

Messenger Rna ◽

Brain Regions ◽

Mrna Levels ◽

Expression Of Genes ◽

Genes Encoding ◽

Glycerolipid Synthesis ◽

Altered Gene ◽

The Brain

AbstractAimsExpression of genes encoding enzymes involved in glycerolipid and monoacylglycerol pathways in specific brain regions is poorly known and its impact in insulin resistance (IR) and type 2 diabetes (T2D) in the brain remains unreported. We determined mRNA levels of enzymes involved in glycerolipid synthesis in different regions of the mouse brain and evaluated their changes in two models of IR and T2D, the Agpat2-/- and Leprdb/db mice.MethodsCerebral cortex, hypothalamus, hippocampus and cerebellum were dissected from adult Agpat2-/- mice, Leprdb/db mice and their respective wild type littermates. Total RNA was isolated and mRNA abundance was measured by RT-qPCR.Key findingsGPAT1, AGPAT1-4, LIPIN1/2, DGAT1/2 and MOGAT1 mRNAs were detected in all studied brain regions, whereas GPAT2, LIPIN3 and MOGAT2 were undetectable. Abundance of AGPATs, LIPIN1 and DGAT1, was higher in cerebellum and hypothalamus. LIPIN2 and MOGAT1 levels were higher in hypothalamus, and DGAT2 was higher in cortex and hypothalamus. In Agpat2-/- mice, LIPIN1 levels were increased in all the brain regions. By contrast, GPAT1 in cortex and hypothalamus, AGPAT3 in hippocampus and hypothalamus, AGPAT4 in hypothalamus, and MOGAT1 in cortex, hypothalamus and cerebellum were lower in Agpat2-/- mice. Leprdb/db mice showed fewer and milder changes, with increased levels of GPAT1 and LIPIN1 in cerebellum, and AGPAT3 in hypothalamus.Conclusions and SignificanceEnzymes of glycerolipids synthesis are differentially expressed across regions of the mouse brain. Two mouse models of IR and T2D have altered gene expression of glycerolipid enzymes in the brain.

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β-Patchoulene, isolated from patchouli oil, suppresses inflammatory mediators in LPS-stimulated RAW264.7 macrophages

European Journal of Inflammation ◽

10.1177/1721727x17714694 ◽

2017 ◽

Vol 15 (2) ◽

pp. 136-141 ◽

Cited By ~ 3

Author(s):

Wen-Hao Yang ◽

Yu-Hong Liu ◽

Jia-Li Liang ◽

Zhi-Xiu Lin ◽

Qiu-Lin Kong ◽

...

Keyword(s):

Nitric Oxide ◽

Inflammatory Cytokines ◽

Messenger Rna ◽

Inflammatory Activity ◽

Prostaglandin E ◽

Mrna Levels ◽

Dependent Manner ◽

Raw264.7 Macrophages ◽

Cox 2 ◽

Anti Inflammatory

β-Patchoulene (β-PAE) is a tricyclic sesquiterpene isolated from patchouli oil. According to our previous study, β-PAE has anti-inflammatory activity in vivo; however, its anti-inflammatory response still remains unconfirmed in vitro. Therefore, this study is committed to demonstrate the anti-inflammatory effect of β-PAE on lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. According to our results, pre-treatment with β-PAE significantly decreased the protein and messenger RNA (mRNA) levels of pro-inflammatory cytokines including tumor necrosis factor-α (TNF-α), interleukin (IL)-6, and IL-1β while increased the expressions of anti-inflammatory cytokines like IL-10 in a dose-dependent manner. In addition, real-time polymerase chain reaction (PCR) also revealed that β-PAE could interrupt the mRNA expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and thus decreased the levels of nitric oxide (NO) and prostaglandin E2 (PGE2) in LPS-stimulated RAW264.7 macrophages. In conclusion, these results indicated that β-PAE exerted potent anti-inflammatory activity by maintaining the balance between pro- and anti-inflammatory cytokines as well as suppressing iNOS and COX-2 signaling pathways.

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Differential Effects of Gonadotropin-Releasing Hormone (GnRH) Pulse Frequency on Gonadotropin Subunit and GnRH Receptor Messenger Ribonucleic Acid Levels in Vitro*

Endocrinology ◽

10.1210/endo.138.3.4968 ◽

1997 ◽

Vol 138 (3) ◽

pp. 1224-1231 ◽

Cited By ~ 112

Author(s):

Ursula B. Kaiser ◽

Andrzej Jakubowiak ◽

Anna Steinberger ◽

William W. Chin

Keyword(s):

Gene Expression ◽

Messenger Rna ◽

Pulse Frequency ◽

Gnrh Receptor ◽

Pituitary Cells ◽

Mrna Levels ◽

Subunit Gene ◽

Differential Effects ◽

Messenger Ribonucleic Acid

Abstract The hypothalamic hormone, GnRH, is released and transported to the anterior pituitary in a pulsatile manner, where it binds to specific high-affinity receptors and regulates gonadotropin biosynthesis and secretion. The frequency of GnRH pulses changes under various physiological conditions, and varying GnRH pulse frequencies have been shown to regulate differentially the secretion of LH and FSH and the expression of the gonadotropin α, LHβ, and FSHβ subunit genes in vivo. We demonstrate differential effects of varying GnRH pulse frequency in vitro in superfused primary monolayer cultures of rat pituitary cells. Cells were treated with 10 nm GnRH pulses for 24 h at a frequency of every 0.5, 1, 2, or 4 h. α, LHβ, and FSHβ messenger RNA (mRNA) levels were increased by GnRH at all pulse frequencies. α and LHβ mRNA levels and LH secretion were stimulated to the greatest extent at a GnRH pulse frequency of every 30 min, whereas FSHβ mRNA levels and FSH secretion were stimulated maximally at a lower GnRH pulse frequency, every 2 h. GnRH receptor (GnRHR) mRNA levels also were increased by GnRH at all pulse frequencies and were stimulated maximally at a GnRH pulse frequency of every 30 min. Similar results were obtained when the dose of each pulse of GnRH was adjusted to maintain a constant total cumulative dose of GnRH over 24 h. These data show that gonadotropin subunit gene expression is regulated differentially by varying GnRH pulse frequencies in vitro, suggesting that the differential effects of varying GnRH pulse frequencies on gonadotropin subunit gene expression occur directly at the level of the pituitary. The pattern of regulation of GnRHR mRNA levels correlated with that of α and LHβ but was different from that of FSHβ. This suggests that α and LHβ mRNA levels are maximally stimulated when GnRHR levels are relatively high, whereas FSHβ mRNA levels are maximally stimulated at lower levels of GnRHR expression, and that the mechanism for differential regulation of the gonadotropins by varying pulse frequencies of GnRH may involve levels of GnRHR. Furthermore, these data suggest that the mechanisms whereby varying GnRH pulse frequencies stimulate α, LHβ, and GnRHR gene expression are similar, whereas the stimulation of FSHβ mRNA levels may be different.

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The Poor Osteoinductive Capability of Human Acellular Bone Matrix

The International Journal of Artificial Organs ◽

10.1177/039139881203501204 ◽

2012 ◽

Vol 35 (12) ◽

pp. 1061-1069 ◽

Cited By ~ 9

Author(s):

Yi-Zhou Huang ◽

Jia-Qin Cai ◽

Jing Xue ◽

Xiao-He Chen ◽

Chao-Liang Zhang ◽

...

Keyword(s):

Bone Regeneration ◽

Demineralized Bone Matrix ◽

Bone Matrix ◽

Structural Features ◽

Nodule Formation ◽

Porous Network ◽

Native Bone ◽

Genes Encoding ◽

Osteoid Formation ◽

Stem Cell Niches

Demineralized bone matrix (DBM) has extensive clinical use for bone regeneration because of its osteoinductive and osteoconductive aptitude. It is suggested that the demineralization process in bone matrix preparation is influential in maintaining osteoinductivity; however, relevant investigations, especially into the osteoinductivity of acellular bone matrix, are not often performed. This study addressed the osteoinductive capability of human acellular cancellous bone matrix (ACBM) after subcutaneous implantation in a rat model. The growth and osteogenic differentiation of rat bone marrow-derived mesenchymal stem cells (rBM-MSCs) seeded in this material were also studied. Without the demineralization process, the ACBM we obtained had an interconnected porous network and the micropores in the surface were clearly exposed. After the ACBM was subcutaneously implanted for 4 months, new osteoid formation was noted but not typical mature bone formation. rBM-MSCs grew well in the ACBM and kept a steady morphology after continuous culture for 28 days. However, no mineralized nodule formation was detected and the expression levels of genes encoding osteogenic markers were significantly decreased. These results demonstrated that human ACBM possess the structural features of native bone and poor osteoinductivity; nonetheless this material helped to preserve the undifferentiated phenotype of rBM-MSCs. Such insights may further broaden our understanding of the application of ACBM for bone regeneration and the creation of stem cell niches.

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Effect of Growth State on Transcription Levels of Genes Encoding Major Secreted Antigens of Mycobacterium tuberculosis in the Mouse Lung

Infection and Immunity ◽

10.1128/iai.72.4.2420-2424.2004 ◽

2004 ◽

Vol 72 (4) ◽

pp. 2420-2424 ◽

Cited By ~ 60

Author(s):

Lanbo Shi ◽

Robert North ◽

Maria Laura Gennaro

Keyword(s):

Mycobacterium Tuberculosis ◽

Adaptive Immunity ◽

Mouse Lung ◽

Mrna Levels ◽

Growth State ◽

Genes Encoding ◽

Secreted Antigens

ABSTRACT Arrest of the multiplication of Mycobacterium tuberculosis caused by expression of adaptive immunity in mouse lung was accompanied by a 10- to 20-fold decrease in levels of mRNAs encoding the secreted Ag85 complex and 38-kDa lipoprotein. esat-6 mRNA levels were high throughout infection. The data imply that multiplying and nonreplicating tubercle bacilli have different antigen compositions.

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Complement Factor C3 Methylation and mRNA Expression Is Associated to BMI and Insulin Resistance in Obesity

Genes ◽

10.3390/genes9080410 ◽

2018 ◽

Vol 9 (8) ◽

pp. 410 ◽

Cited By ~ 7

Author(s):

Daniel Castellano-Castillo ◽

Isabel Moreno-Indias ◽

Jose Carlos Fernandez-Garcia ◽

Mercedes Clemente-Postigo ◽

Manuel Castro-Cabezas ◽

...

Keyword(s):

Insulin Resistance ◽

Dna Methylation ◽

Adipose Tissue ◽

Messenger Rna ◽

Density Lipoprotein ◽

Complement Factor ◽

Mrna Levels ◽

Model Assessment ◽

Tissue Samples ◽

Obese Class

Epigenetic marks, and especially DNA methylation, are becoming an important factor in obesity, which could help to explain its etiology and associated comorbidities. Adipose tissue, now considered as an important endocrine organ, produces complement system factors. Complement component 3 (C3) turns out to be an important protein in metabolic disorders, via either inflammation or the C3 subproduct acylation stimulating protein (ASP) which directly stimulates lipid storage. In this study, we analyze C3 DNA methylation in adipose tissue from subjects with a different grade of obesity. Adipose tissue samples were collected from subjects with a different degree of obesity determined by their body mass index (BMI) as: Overweight subjects (BMI ≥ 25 and <30), obese class 1/2 subjects (BMI ≥ 30 and <40) and obese class 3 subjects (BMI ≥ 40). C3 DNA methylation was measured for 7 CpGs by pyrosequencition using the Pyromark technology (Qiagen, Madrid Spain). C3 messenger RNA (mRNA) levels were analyzed by pre-designed Taqman assays (Applied biosystems, Foster City, CA, USA) and ASP/C3a was measured using a ELISA kit. The data were analyzed using the statistic package SPSS. C3 DNA methylation levels were lower in the morbid obese group. Accordingly, C3 methylation correlated negatively with BMI and leptin. However, C3 mRNA levels were more associated with insulin resistance, and positive correlations with insulin, glucose and homeostasis model assessment-estimated insulin resistance (HOMA-IR) existed. ASP correlated negatively with high density lipoprotein (HDL) cholesterol. C3 methylation levels were associated to adiposity variables, such as BMI and leptin, while the C3 mRNA levels were associated to glucose metabolism.

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Oleifolioside A, a New Active Compound, Attenuates LPS-Stimulated iNOS and COX-2 Expression through the Downregulation of NF-κB and MAPK Activities in RAW 264.7 Macrophages

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2012/637512 ◽

2012 ◽

Vol 2012 ◽

pp. 1-8 ◽

Cited By ~ 15

Author(s):

Hai Yang Yu ◽

Kyoung-Sook Kim ◽

Young-Choon Lee ◽

Hyung-In Moon ◽

Jai-Heon Lee

Keyword(s):

Nitric Oxide ◽

Mapk Signaling ◽

Binding Activity ◽

Prostaglandin E ◽

Raw 264.7 ◽

Mrna Levels ◽

Dependent Manner ◽

Raw 264.7 Macrophages ◽

Dendropanax Morbifera ◽

Cox 2

Oleifolioside A, a new triterpenoid compound isolated fromDendropanax morbiferaLeveille (D. morbifera), was shown in this study to have potent inhibitory effects on lipopolysaccharide (LPS-)stimulated nitric oxide (NO) and prostaglandin E2(PGE2) production in RAW 264.7 macrophages. Consistent with these findings, oleifolioside A was further shown to suppress the expression of LPS-stimulated inducible nitric oxide synthase (iNOS) and cyclooxigenase-2 (COX-2) in a dose-dependent manner at both the protein and mRNA levels and to significantly inhibit the DNA-binding activity and transcriptional activity of NF-κB in response to LPS. These results were found to be associated with the inhibition of the degradation and phosphorylation of IκB-αand subsequent translocation of the NF-κB p65 subunit to the nucleus. Inhibition of NF-κB activation by oleifolioside A was also shown to be mediated through the prevention of p38 MAPK and ERK1/2 phosphorylation. Taken together, our results suggest that oleifolioside A has the potential to be a novel anti-inflammatory agent capable of targeting both the NF-κB and MAPK signaling pathways.

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