Fraser Collins - Academia.edu (original) (raw)
Papers by Fraser Collins
Annals of the Rheumatic Diseases
ObjectivesFibroblasts in synovium include fibroblast-like synoviocytes (FLS) in the lining andThy... more ObjectivesFibroblasts in synovium include fibroblast-like synoviocytes (FLS) in the lining andThy1+ connective-tissue fibroblasts in the sublining. We aimed to investigate their developmental origin and relationship with adult progenitors.MethodsTo discriminate betweenGdf5-lineage cells deriving from the embryonic joint interzone and otherPdgfrα-expressing fibroblasts and progenitors, adultGdf5-Cre;Tom;Pdgfrα-H2BGFPmice were used and cartilage injury was induced to activate progenitors. Cells were isolated from knees, fibroblasts and progenitors were sorted by fluorescence-activated cell-sorting based on developmental origin, and analysed by single-cell RNA-sequencing. Flow cytometry and immunohistochemistry were used for validation. Clonal-lineage mapping was performed usingGdf5-Cre;Confettimice.ResultsIn steady state,Thy1+ sublining fibroblasts were of mixed ontogeny. In contrast,Thy1-Prg4+ lining fibroblasts predominantly derived from the embryonic joint interzone and includedPrg...
![Research paper thumbnail of Death Receptor 3 orchestrates erosive pathology in inflammatory arthritis [Poster Abstract]](https://attachments.academia-assets.com/99955894/thumbnails/1.jpg)
](Immunology, Dec 1, 2010
IL-10 plays a central, non-redundant role in limiting inflammation in vivo. However the mechanism... more IL-10 plays a central, non-redundant role in limiting inflammation in vivo. However the mechanisms involved remain to be resolved. Here using primary human macrophages, we found that IL-10 inhibits selected inflammatory genes, primarily at a level of transcription. At the TNF gene this occurs not through an inhibition of RNA Polymerase II (Pol II) recruitment and transcription initiation, but through a mechanism targeting the stimulation of transcription elongation by cyclin dependent kinase (CDK)-9. We demonstrated an unanticipated requirement for a region downstream of the TNF 3¢ untranslated region (UTR) that contains the NF-jB binding motif (jB4) both for induction of transcription by LPS and its inhibition by IL-10. IL-10 not only inhibits the recruitment of RelA to regions containing jB sites at the TNF gene, but also to those found at other LPS induced genes. We show that while IL-10 elicits a general block in RelA recruitment to its genomic targets, the gene specific nature of IL-10's actions are defined through the differential recruitment of CDK9 and the control of transcription elongation. At TNF, but not NFKBIA, the consequence of RelA recruitment inhibition is a loss of CDK9 recruitment, preventing the stimulation of transcription elongation.
![Research paper thumbnail of Control of early cartilage destruction in inflammatory arthritis by death receptor 3 [Abstract]](https://attachments.academia-assets.com/99955893/thumbnails/1.jpg)
](Immunology, Dec 1, 2011
Journal of cells, molecules, systems and technologies Disclaimer: This abstract book has been pro... more Journal of cells, molecules, systems and technologies Disclaimer: This abstract book has been produced using author-supplied copy. Editing has been restricted to some corrections of spelling and style where appropriate. No responsibility is assumed for any claims, instructions, methods or drug dosages contained in the abstracts; it is recommended that these are verified independently.
Osteoarthritis and Cartilage
Analysis of antibody for the intracellular detection of Wnt10b in bone marrow immune cells by flo... more Analysis of antibody for the intracellular detection of Wnt10b in bone marrow immune cells by flow cytometry. <b></b>
Annals of the Rheumatic Diseases, 2021
ObjectiveWe aimed to understand the role of the transcriptional co-factor Yes-associated protein ... more ObjectiveWe aimed to understand the role of the transcriptional co-factor Yes-associated protein (Yap) in the molecular pathway underpinning the pathogenic transformation of synovial fibroblasts (SF) in rheumatoid arthritis (RA) to become invasive and cause joint destruction.MethodsSynovium from patients with RA and mice with antigen-induced arthritis (AIA) was analysed by immunostaining and qRT-PCR. SF were targeted using Pdgfrα-CreER and Gdf5-Cre mice, crossed with fluorescent reporters for cell tracing and Yap-flox mice for conditional Yap ablation. Fibroblast phenotypes were analysed by flow cytometry, and arthritis severity was assessed by histology. Yap activation was detected using Yap–Tead reporter cells and Yap–Snail interaction by proximity ligation assay. SF invasiveness was analysed using matrigel-coated transwells.ResultsYap, its binding partner Snail and downstream target connective tissue growth factor were upregulated in hyperplastic human RA and in mouse AIA synoviu...
Annals of the Rheumatic Diseases, 2021
In rheumatoid arthritis (RA), the fibroblast-like synoviocytes (FLS) in synovial lining become in... more In rheumatoid arthritis (RA), the fibroblast-like synoviocytes (FLS) in synovial lining become invasive and cause joint destruction. The molecular mechanisms underpinning this pathogenic FLS phenotype are incompletely understood. The FLS descend from Growth differentiation factor 5 (Gdf5)-expressing joint interzone cells in the embryo, and we showed that conditional ablation of the transcriptional co-activator Yes associated protein (Yap) in Gdf5-lineage cells prevents synovial lining hyperplasia after traumatic cartilage injury in mice [1].Here, we investigated a potential role for Yap in pathogenic FLS in immune-mediated inflammatory arthritis.Immunohistochemistry was used to detect Yap in human RA synovium and Yap, Snail and Ctgf in mouse synovium following antigen-induced arthritis (AIA). To determine the effect of Yap knockout (KO) in synovial stromal cells, AIA was induced in Gdf5-Cre;tdTomato;Yapfl/fl (Yap cKO) and Gdf5-Cre;tdTomato;Yapwt/wt (control) mice, or in Pdgfrα-CreER...
Advances in Experimental Medicine and Biology, 2017
In recent years a link between the gastro-intestinal tract and bone health has started to gain si... more In recent years a link between the gastro-intestinal tract and bone health has started to gain significant attention. Dysbiosis of the intestinal microbiota has been linked to the pathology of a number of diseases which are associated with bone loss. In addition modulation of the intestinal microbiota with probiotic bacteria has revealed to have both beneficial local and systemic effects. In the present chapter we discuss the intestinal and bone immune systems, explore how intestinal disease affects the immune system and examine how these pathologic changes could adversely impact bone health.
Molecular and Integrative Toxicology, 2017
In recent years, the intestinal microbiota has emerged as a crucial regulator of health with dysb... more In recent years, the intestinal microbiota has emerged as a crucial regulator of health with dysbiosis linked to a number of pathological states such as inflammatory bowel disease. In addition to a local intestinal effect, emerging evidence has demonstrated the potential for the microbiota to modulate systemic bone health via a gut-bone axis. In the present chapter, we discuss how diet can affect the composition of the intestinal microbiota, through the intake of prebiotics, and how these are utilized by the bacteria to influence the immune system and bone. In addition, we detail the recent murine studies that investigate how probiotic supplementation can increase bone mineral density in “healthy” individuals and protect against the pathological bone loss associated with menopausal estrogen deficiency. Finally, we highlight the advances made in unearthing the mechanisms that potentially lead to these observed beneficial effects.
Advances in Experimental Medicine and Biology, 2017
The intestinal epithelial barrier plays an essential role in maintaining host homeostasis. The ba... more The intestinal epithelial barrier plays an essential role in maintaining host homeostasis. The barrier regulates nutrient absorption as well as prevents the invasion of pathogenic bacteria in the host. It is composed of epithelial cells, tight junctions, and a mucus layer. Several factors, such as cytokines, diet, and diseases can affect this barrier. These factors have been shown to increase intestinal permeability, inflammation, and translocation of pathogenic bacteria. In addition, dysregulation of the epithelial barrier can result in inflammatory diseases such as inflammatory bowel disease. Our lab and others have also shown that barrier disruption can have systemic effects including bone loss. In this chapter, we will discuss the current literature to understand the link between intestinal barrier and bone. We will discuss how inflammation, aging, dysbiosis and metabolic diseases can affect intestinal barrier-bone link. In addition, we will highlight the current suggested mechanism between intestinal barrier and bone.
Annals of the Rheumatic Diseases, 2020
ObjectivesOsteophytes are highly prevalent in osteoarthritis (OA) and are associated with pain an... more ObjectivesOsteophytes are highly prevalent in osteoarthritis (OA) and are associated with pain and functional disability. These pathological outgrowths of cartilage and bone typically form at the junction of articular cartilage, periosteum and synovium. The aim of this study was to identify the cells forming osteophytes in OA.MethodsFluorescent genetic cell-labelling and tracing mouse models were induced with tamoxifen to switch on reporter expression, as appropriate, followed by surgery to induce destabilisation of the medial meniscus. Contributions of fluorescently labelled cells to osteophytes after 2 or 8 weeks, and their molecular identity, were analysed by histology, immunofluorescence staining and RNA in situ hybridisation. Pdgfrα-H2BGFP mice and Pdgfrα-CreER mice crossed with multicolour Confetti reporter mice were used for identification and clonal tracing of mesenchymal progenitors. Mice carrying Col2-CreER, Nes-CreER, LepR-Cre, Grem1-CreER, Gdf5-Cre, Sox9-CreER or Prg4-Cr...
Journal of Bone and Mineral Research, 2020
Glucocorticoids (GCs) are potent immune‐modulating drugs with significant side effects, including... more Glucocorticoids (GCs) are potent immune‐modulating drugs with significant side effects, including glucocorticoid‐induced osteoporosis (GIO). GCs directly induce osteoblast and osteocyte apoptosis but also alter intestinal microbiota composition. Although the gut microbiota is known to contribute to the regulation of bone density, its role in GIO has never been examined. To test this, male C57/Bl6J mice were treated for 8 weeks with GC (prednisolone, GC‐Tx) in the presence or absence of broad‐spectrum antibiotic treatment (ABX) to deplete the microbiota. Long‐term ABX prevented GC‐Tx‐induced trabecular bone loss, showing the requirement of gut microbiota for GIO. Treatment of GC‐Tx mice with a probiotic (Lactobacillus reuteri [LR]) prevented trabecular bone loss. Microbiota analyses indicated that GC‐Tx changed the abundance of Verrucomicobiales and Bacteriodales phyla and random forest analyses indicated significant differences in abundance of Porphyromonadaceae and Clostridiales operational taxonomic units (OTUs) between groups. Furthermore, transplantation of GC‐Tx mouse fecal material into recipient naïve, untreated WT mice caused bone loss, supporting a functional role for microbiota in GIO. We also report that GC caused intestinal barrier breaks, as evidenced by increased serum endotoxin level (2.4‐fold), that were prevented by LR and ABX treatments. Enhancement of barrier function with a mucus supplement prevented both GC‐Tx–induced barrier leakage and trabecular GIO. In bone, treatment with ABX, LR or a mucus supplement reduced GC‐Tx–induced osteoblast and osteocyte apoptosis. GC‐Tx suppression of Wnt10b in bone was restored by the LR and high‐molecular‐weight polymer (MDY) treatments as well as microbiota depletion. Finally, we identified that bone‐specific Wnt10b overexpression prevented GIO. Taken together, our data highlight the previously unappreciated involvement of the gut microbiota and intestinal barrier function in trabecular GIO pathogenesis (including Wnt10b suppression and osteoblast and osteocyte apoptosis) and identify the gut as a novel therapeutic target for preventing GIO. © 2019 American Society for Bone and Mineral Research.
Scientific Reports, 2019
Oral treatment with probiotic bacteria has been shown to prevent bone loss in multiple models of ... more Oral treatment with probiotic bacteria has been shown to prevent bone loss in multiple models of osteoporosis. In previous studies we demonstrated that oral administration of Lactobacillus reuteri in healthy male mice increases bone density. The host and bacterial mechanisms of these effects however are not well understood. The objective of this study was to understand the role of lymphocytes in mediating the beneficial effects of L. reuteri on bone health in male mice. We administered L. reuteri in drinking water for 4 weeks to wild type or Rag knockout (lack mature T and B lymphocytes) male mice. While L. reuteri treatment increased bone density in wild type, no significant increases were seen in Rag knockout mice, suggesting that lymphocytes are critical for mediating the beneficial effects of L. reuteri on bone density. To understand the effect of L. reuteri on lymphocytes in the intestinal tissues, we isolated mesenteric lymph node (MLN) from naïve wild type mice. In ex vivo st...
BMC Musculoskeletal Disorders, 2019
Background: Oestrogen-deficiency induced by menopause is associated with reduced bone density and... more Background: Oestrogen-deficiency induced by menopause is associated with reduced bone density and primary osteoporosis, resulting in an increased risk of fracture. While the exact etiology of menopause-induced primary osteoporotic bone loss is not fully known, members of the tumour necrosis factor super family (TNFSF) are known to play a role. Recent studies have revealed that the TNFSF members death receptor 3 (DR3) and one of its ligands, TNF-like protein 1A (TL1A) have a key role in secondary osteoporosis; enhancing CD14 + peripheral blood mononuclear cell (PBMC) osteoclast formation and bone resorption. Whether DR3 and TL1A contribute towards bone loss in menopause-induced primary osteoporosis however, remains unknown. Methods: To investigate this we performed flow cytometry analysis of DR3 expression on CD14 + PBMCs isolated from pre-and early post-menopausal females and late post-menopausal osteoporotic patients. Serum levels of TL1A, CCL3 and total MMP-9 were measured by ELISA. In vitro osteoclast differentiation assays were performed to determine CD14 + monocyte osteoclastogenic potential. In addition, splenic CD4 + T cell DR3 expression was investigated 1 week and 8 weeks post-surgery, using the murine ovariectomy model. Results: In contrast to pre-menopausal females, CD14 + monocytes isolated from post-menopausal females were unable to induce DR3 expression. Serum TL1A levels were decreased approx. 2-fold in early post-menopausal females compared to pre-menopausal controls and post-menopausal osteoporotic females; no difference was observed between pre-menopausal and late post-menopausal osteoporotic females. Analysis of in vitro CD14 + monocyte osteoclastogenic potential revealed no significant difference between the post-menopausal and postmenopausal osteoporotic cohorts. Interestingly, in the murine ovariectomy model splenic CD4 + T cell DR3 expression was significantly increased at 1 week but not 8 weeks post-surgery when compared to the sham control. Conclusion: Our results reveals for the first time that loss of oestrogen has a significant effect on DR3; decreasing expression on CD14 + monocytes and increasing expression on CD4 + T cells. These data suggest that while oestrogen-deficiency induced changes in DR3 expression do not affect late post-menopausal bone loss they could potentially have an indirect role in early menopausal bone loss through the modulation of T cell activity.
Journal of Bone and Mineral Research, 2019
Antibiotic treatment, commonly prescribed for bacterial infections, depletes and subsequently cau... more Antibiotic treatment, commonly prescribed for bacterial infections, depletes and subsequently causes long-term alterations in intestinal microbiota composition. Knowing the importance of the microbiome in the regulation of bone density, we investigated the effect of postantibiotic treatment on gut and bone health. Intestinal microbiome repopulation at 4-weeks postantibiotic treatment resulted in an increase in the Firmicutes:Bacteroidetes ratio, increased intestinal permeability, and notably reduced femoral trabecular bone volume (approximately 30%, p < 0.01). Treatment with a mucus supplement (a high-molecular-weight polymer, MDY-1001 [MDY]) prevented the postantibiotic-induced barrier break as well as bone loss, indicating a mechanistic link between increased intestinal permeability and bone loss. A link between the microbiome composition and bone density was demonstrated by supplementing the mice with probiotic bacteria. Specifically, Lactobacillus reuteri, but not Lactobacillus rhamnosus GG or nonpathogenic Escherichia coli, reduced the postantibiotic elevation of the Firmicutes:Bacteroidetes ratio and prevented femoral and vertebral trabecular bone loss. Consistent with causing bone loss, postantibiotic-induced dysbiosis decreased osteoblast and increased osteoclast activities, changes that were prevented by both L. reuteri and MDY. These data underscore the importance of microbial dysbiosis in the regulation of intestinal permeability and bone health, as well as identify L. reuteri and MDY as novel therapies for preventing these adverse effects.
mSphere
Annually, an estimated 2 million osteoporotic fractures occur in the United States alone. Osteopo... more Annually, an estimated 2 million osteoporotic fractures occur in the United States alone. Osteoporosis imparts a great burden on the health care system. The identification of novel regulators of bone health is critical for developing more effective therapeutics. A previous study on the colonization of germ-free (GF) mice with a microbial community has demonstrated that bacterial colonization dramatically increases bone loss. We therefore investigated the impact of multiple microbial communities in different mice to understand how generalizable the impact of bacterial colonization is on bone health. To investigate the impact of different microbial communities on bone health in outbred and inbred mouse strains, gavage was performed on GF Swiss Webster and GF C57BL/6 mice to introduce distinct microbiotas that originated from either humans or mice. GF mice displayed a high degree of colonization, as indicated by more than 90% of the operational taxonomic units present in the starting i...
PLOS ONE, 2017
Background & aims Wnt10b is a crucial regulator of bone density through its ability to promote os... more Background & aims Wnt10b is a crucial regulator of bone density through its ability to promote osteoblastogenesis. Parathyroid hormone has been shown to regulate Wnt10b expression in CD8 + T cells. However, the relative expression and other source(s) of Wnt10b in the bone marrow immune cells (BMICs) is unknown. Sex hormones and cytokines such as, estrogen and TNFα are critical regulators of bone physiology but whether they regulate BMIC Wnt10b expression is unclear. To determine the potential regulation of Wnt10b by estrogen and TNFα, we assessed Wnt10b expression by flow cytometry under estrogen-and TNFα-deficient conditions. Methods Effects of TNFα was determined in male and female C57BL/6 wildtype and TNFα knockout mice. Effect of estrogen was investigated 4, 6 and 8 weeks post-surgery in ovariectomized Balb/c mice. Intracellular Wnt10b was detected using goat anti-mouse Wnt10b and a conjugated secondary antibody and analyzed by flow cytometry. Results Wnt10b expression was sex-and lineage-specific. Females had 1.8-fold higher Wnt10b signal compared to males. Percent of Wnt10b + myeloid cells was higher in females than males (8.9% Vs 5.4%) but Wnt10b + lymphoid cells was higher in males than females (6.3% Vs 2.5%). TNFα ablation in males increased total BM Wnt10b expression 1.5-fold but significantly reduced the percentage of BM Wnt10b + CD4 + T cells (65%), CD8 + T cells (59%), dendritic cells (59%), macrophages (56%) and granulocytes (52%). These effects of TNFα on Wnt10b were observed only in males. In contrast to TNFα, estrogen-deficiency had indirect effects on BMIC Wnt10b levels; reducing the average percentage of BM Wnt10b + CD8 + T cells (25%) and granulocytes (26%) across an 8-week time course.
Annals of the Rheumatic Diseases
ObjectivesFibroblasts in synovium include fibroblast-like synoviocytes (FLS) in the lining andThy... more ObjectivesFibroblasts in synovium include fibroblast-like synoviocytes (FLS) in the lining andThy1+ connective-tissue fibroblasts in the sublining. We aimed to investigate their developmental origin and relationship with adult progenitors.MethodsTo discriminate betweenGdf5-lineage cells deriving from the embryonic joint interzone and otherPdgfrα-expressing fibroblasts and progenitors, adultGdf5-Cre;Tom;Pdgfrα-H2BGFPmice were used and cartilage injury was induced to activate progenitors. Cells were isolated from knees, fibroblasts and progenitors were sorted by fluorescence-activated cell-sorting based on developmental origin, and analysed by single-cell RNA-sequencing. Flow cytometry and immunohistochemistry were used for validation. Clonal-lineage mapping was performed usingGdf5-Cre;Confettimice.ResultsIn steady state,Thy1+ sublining fibroblasts were of mixed ontogeny. In contrast,Thy1-Prg4+ lining fibroblasts predominantly derived from the embryonic joint interzone and includedPrg...
Immunology, Dec 1, 2010
IL-10 plays a central, non-redundant role in limiting inflammation in vivo. However the mechanism... more IL-10 plays a central, non-redundant role in limiting inflammation in vivo. However the mechanisms involved remain to be resolved. Here using primary human macrophages, we found that IL-10 inhibits selected inflammatory genes, primarily at a level of transcription. At the TNF gene this occurs not through an inhibition of RNA Polymerase II (Pol II) recruitment and transcription initiation, but through a mechanism targeting the stimulation of transcription elongation by cyclin dependent kinase (CDK)-9. We demonstrated an unanticipated requirement for a region downstream of the TNF 3¢ untranslated region (UTR) that contains the NF-jB binding motif (jB4) both for induction of transcription by LPS and its inhibition by IL-10. IL-10 not only inhibits the recruitment of RelA to regions containing jB sites at the TNF gene, but also to those found at other LPS induced genes. We show that while IL-10 elicits a general block in RelA recruitment to its genomic targets, the gene specific nature of IL-10's actions are defined through the differential recruitment of CDK9 and the control of transcription elongation. At TNF, but not NFKBIA, the consequence of RelA recruitment inhibition is a loss of CDK9 recruitment, preventing the stimulation of transcription elongation.
Immunology, Dec 1, 2011
Journal of cells, molecules, systems and technologies Disclaimer: This abstract book has been pro... more Journal of cells, molecules, systems and technologies Disclaimer: This abstract book has been produced using author-supplied copy. Editing has been restricted to some corrections of spelling and style where appropriate. No responsibility is assumed for any claims, instructions, methods or drug dosages contained in the abstracts; it is recommended that these are verified independently.
Osteoarthritis and Cartilage
Analysis of antibody for the intracellular detection of Wnt10b in bone marrow immune cells by flo... more Analysis of antibody for the intracellular detection of Wnt10b in bone marrow immune cells by flow cytometry. <b></b>
Annals of the Rheumatic Diseases, 2021
ObjectiveWe aimed to understand the role of the transcriptional co-factor Yes-associated protein ... more ObjectiveWe aimed to understand the role of the transcriptional co-factor Yes-associated protein (Yap) in the molecular pathway underpinning the pathogenic transformation of synovial fibroblasts (SF) in rheumatoid arthritis (RA) to become invasive and cause joint destruction.MethodsSynovium from patients with RA and mice with antigen-induced arthritis (AIA) was analysed by immunostaining and qRT-PCR. SF were targeted using Pdgfrα-CreER and Gdf5-Cre mice, crossed with fluorescent reporters for cell tracing and Yap-flox mice for conditional Yap ablation. Fibroblast phenotypes were analysed by flow cytometry, and arthritis severity was assessed by histology. Yap activation was detected using Yap–Tead reporter cells and Yap–Snail interaction by proximity ligation assay. SF invasiveness was analysed using matrigel-coated transwells.ResultsYap, its binding partner Snail and downstream target connective tissue growth factor were upregulated in hyperplastic human RA and in mouse AIA synoviu...
Annals of the Rheumatic Diseases, 2021
In rheumatoid arthritis (RA), the fibroblast-like synoviocytes (FLS) in synovial lining become in... more In rheumatoid arthritis (RA), the fibroblast-like synoviocytes (FLS) in synovial lining become invasive and cause joint destruction. The molecular mechanisms underpinning this pathogenic FLS phenotype are incompletely understood. The FLS descend from Growth differentiation factor 5 (Gdf5)-expressing joint interzone cells in the embryo, and we showed that conditional ablation of the transcriptional co-activator Yes associated protein (Yap) in Gdf5-lineage cells prevents synovial lining hyperplasia after traumatic cartilage injury in mice [1].Here, we investigated a potential role for Yap in pathogenic FLS in immune-mediated inflammatory arthritis.Immunohistochemistry was used to detect Yap in human RA synovium and Yap, Snail and Ctgf in mouse synovium following antigen-induced arthritis (AIA). To determine the effect of Yap knockout (KO) in synovial stromal cells, AIA was induced in Gdf5-Cre;tdTomato;Yapfl/fl (Yap cKO) and Gdf5-Cre;tdTomato;Yapwt/wt (control) mice, or in Pdgfrα-CreER...
Advances in Experimental Medicine and Biology, 2017
In recent years a link between the gastro-intestinal tract and bone health has started to gain si... more In recent years a link between the gastro-intestinal tract and bone health has started to gain significant attention. Dysbiosis of the intestinal microbiota has been linked to the pathology of a number of diseases which are associated with bone loss. In addition modulation of the intestinal microbiota with probiotic bacteria has revealed to have both beneficial local and systemic effects. In the present chapter we discuss the intestinal and bone immune systems, explore how intestinal disease affects the immune system and examine how these pathologic changes could adversely impact bone health.
Molecular and Integrative Toxicology, 2017
In recent years, the intestinal microbiota has emerged as a crucial regulator of health with dysb... more In recent years, the intestinal microbiota has emerged as a crucial regulator of health with dysbiosis linked to a number of pathological states such as inflammatory bowel disease. In addition to a local intestinal effect, emerging evidence has demonstrated the potential for the microbiota to modulate systemic bone health via a gut-bone axis. In the present chapter, we discuss how diet can affect the composition of the intestinal microbiota, through the intake of prebiotics, and how these are utilized by the bacteria to influence the immune system and bone. In addition, we detail the recent murine studies that investigate how probiotic supplementation can increase bone mineral density in “healthy” individuals and protect against the pathological bone loss associated with menopausal estrogen deficiency. Finally, we highlight the advances made in unearthing the mechanisms that potentially lead to these observed beneficial effects.
Advances in Experimental Medicine and Biology, 2017
The intestinal epithelial barrier plays an essential role in maintaining host homeostasis. The ba... more The intestinal epithelial barrier plays an essential role in maintaining host homeostasis. The barrier regulates nutrient absorption as well as prevents the invasion of pathogenic bacteria in the host. It is composed of epithelial cells, tight junctions, and a mucus layer. Several factors, such as cytokines, diet, and diseases can affect this barrier. These factors have been shown to increase intestinal permeability, inflammation, and translocation of pathogenic bacteria. In addition, dysregulation of the epithelial barrier can result in inflammatory diseases such as inflammatory bowel disease. Our lab and others have also shown that barrier disruption can have systemic effects including bone loss. In this chapter, we will discuss the current literature to understand the link between intestinal barrier and bone. We will discuss how inflammation, aging, dysbiosis and metabolic diseases can affect intestinal barrier-bone link. In addition, we will highlight the current suggested mechanism between intestinal barrier and bone.
Annals of the Rheumatic Diseases, 2020
ObjectivesOsteophytes are highly prevalent in osteoarthritis (OA) and are associated with pain an... more ObjectivesOsteophytes are highly prevalent in osteoarthritis (OA) and are associated with pain and functional disability. These pathological outgrowths of cartilage and bone typically form at the junction of articular cartilage, periosteum and synovium. The aim of this study was to identify the cells forming osteophytes in OA.MethodsFluorescent genetic cell-labelling and tracing mouse models were induced with tamoxifen to switch on reporter expression, as appropriate, followed by surgery to induce destabilisation of the medial meniscus. Contributions of fluorescently labelled cells to osteophytes after 2 or 8 weeks, and their molecular identity, were analysed by histology, immunofluorescence staining and RNA in situ hybridisation. Pdgfrα-H2BGFP mice and Pdgfrα-CreER mice crossed with multicolour Confetti reporter mice were used for identification and clonal tracing of mesenchymal progenitors. Mice carrying Col2-CreER, Nes-CreER, LepR-Cre, Grem1-CreER, Gdf5-Cre, Sox9-CreER or Prg4-Cr...
Journal of Bone and Mineral Research, 2020
Glucocorticoids (GCs) are potent immune‐modulating drugs with significant side effects, including... more Glucocorticoids (GCs) are potent immune‐modulating drugs with significant side effects, including glucocorticoid‐induced osteoporosis (GIO). GCs directly induce osteoblast and osteocyte apoptosis but also alter intestinal microbiota composition. Although the gut microbiota is known to contribute to the regulation of bone density, its role in GIO has never been examined. To test this, male C57/Bl6J mice were treated for 8 weeks with GC (prednisolone, GC‐Tx) in the presence or absence of broad‐spectrum antibiotic treatment (ABX) to deplete the microbiota. Long‐term ABX prevented GC‐Tx‐induced trabecular bone loss, showing the requirement of gut microbiota for GIO. Treatment of GC‐Tx mice with a probiotic (Lactobacillus reuteri [LR]) prevented trabecular bone loss. Microbiota analyses indicated that GC‐Tx changed the abundance of Verrucomicobiales and Bacteriodales phyla and random forest analyses indicated significant differences in abundance of Porphyromonadaceae and Clostridiales operational taxonomic units (OTUs) between groups. Furthermore, transplantation of GC‐Tx mouse fecal material into recipient naïve, untreated WT mice caused bone loss, supporting a functional role for microbiota in GIO. We also report that GC caused intestinal barrier breaks, as evidenced by increased serum endotoxin level (2.4‐fold), that were prevented by LR and ABX treatments. Enhancement of barrier function with a mucus supplement prevented both GC‐Tx–induced barrier leakage and trabecular GIO. In bone, treatment with ABX, LR or a mucus supplement reduced GC‐Tx–induced osteoblast and osteocyte apoptosis. GC‐Tx suppression of Wnt10b in bone was restored by the LR and high‐molecular‐weight polymer (MDY) treatments as well as microbiota depletion. Finally, we identified that bone‐specific Wnt10b overexpression prevented GIO. Taken together, our data highlight the previously unappreciated involvement of the gut microbiota and intestinal barrier function in trabecular GIO pathogenesis (including Wnt10b suppression and osteoblast and osteocyte apoptosis) and identify the gut as a novel therapeutic target for preventing GIO. © 2019 American Society for Bone and Mineral Research.
Scientific Reports, 2019
Oral treatment with probiotic bacteria has been shown to prevent bone loss in multiple models of ... more Oral treatment with probiotic bacteria has been shown to prevent bone loss in multiple models of osteoporosis. In previous studies we demonstrated that oral administration of Lactobacillus reuteri in healthy male mice increases bone density. The host and bacterial mechanisms of these effects however are not well understood. The objective of this study was to understand the role of lymphocytes in mediating the beneficial effects of L. reuteri on bone health in male mice. We administered L. reuteri in drinking water for 4 weeks to wild type or Rag knockout (lack mature T and B lymphocytes) male mice. While L. reuteri treatment increased bone density in wild type, no significant increases were seen in Rag knockout mice, suggesting that lymphocytes are critical for mediating the beneficial effects of L. reuteri on bone density. To understand the effect of L. reuteri on lymphocytes in the intestinal tissues, we isolated mesenteric lymph node (MLN) from naïve wild type mice. In ex vivo st...
BMC Musculoskeletal Disorders, 2019
Background: Oestrogen-deficiency induced by menopause is associated with reduced bone density and... more Background: Oestrogen-deficiency induced by menopause is associated with reduced bone density and primary osteoporosis, resulting in an increased risk of fracture. While the exact etiology of menopause-induced primary osteoporotic bone loss is not fully known, members of the tumour necrosis factor super family (TNFSF) are known to play a role. Recent studies have revealed that the TNFSF members death receptor 3 (DR3) and one of its ligands, TNF-like protein 1A (TL1A) have a key role in secondary osteoporosis; enhancing CD14 + peripheral blood mononuclear cell (PBMC) osteoclast formation and bone resorption. Whether DR3 and TL1A contribute towards bone loss in menopause-induced primary osteoporosis however, remains unknown. Methods: To investigate this we performed flow cytometry analysis of DR3 expression on CD14 + PBMCs isolated from pre-and early post-menopausal females and late post-menopausal osteoporotic patients. Serum levels of TL1A, CCL3 and total MMP-9 were measured by ELISA. In vitro osteoclast differentiation assays were performed to determine CD14 + monocyte osteoclastogenic potential. In addition, splenic CD4 + T cell DR3 expression was investigated 1 week and 8 weeks post-surgery, using the murine ovariectomy model. Results: In contrast to pre-menopausal females, CD14 + monocytes isolated from post-menopausal females were unable to induce DR3 expression. Serum TL1A levels were decreased approx. 2-fold in early post-menopausal females compared to pre-menopausal controls and post-menopausal osteoporotic females; no difference was observed between pre-menopausal and late post-menopausal osteoporotic females. Analysis of in vitro CD14 + monocyte osteoclastogenic potential revealed no significant difference between the post-menopausal and postmenopausal osteoporotic cohorts. Interestingly, in the murine ovariectomy model splenic CD4 + T cell DR3 expression was significantly increased at 1 week but not 8 weeks post-surgery when compared to the sham control. Conclusion: Our results reveals for the first time that loss of oestrogen has a significant effect on DR3; decreasing expression on CD14 + monocytes and increasing expression on CD4 + T cells. These data suggest that while oestrogen-deficiency induced changes in DR3 expression do not affect late post-menopausal bone loss they could potentially have an indirect role in early menopausal bone loss through the modulation of T cell activity.
Journal of Bone and Mineral Research, 2019
Antibiotic treatment, commonly prescribed for bacterial infections, depletes and subsequently cau... more Antibiotic treatment, commonly prescribed for bacterial infections, depletes and subsequently causes long-term alterations in intestinal microbiota composition. Knowing the importance of the microbiome in the regulation of bone density, we investigated the effect of postantibiotic treatment on gut and bone health. Intestinal microbiome repopulation at 4-weeks postantibiotic treatment resulted in an increase in the Firmicutes:Bacteroidetes ratio, increased intestinal permeability, and notably reduced femoral trabecular bone volume (approximately 30%, p < 0.01). Treatment with a mucus supplement (a high-molecular-weight polymer, MDY-1001 [MDY]) prevented the postantibiotic-induced barrier break as well as bone loss, indicating a mechanistic link between increased intestinal permeability and bone loss. A link between the microbiome composition and bone density was demonstrated by supplementing the mice with probiotic bacteria. Specifically, Lactobacillus reuteri, but not Lactobacillus rhamnosus GG or nonpathogenic Escherichia coli, reduced the postantibiotic elevation of the Firmicutes:Bacteroidetes ratio and prevented femoral and vertebral trabecular bone loss. Consistent with causing bone loss, postantibiotic-induced dysbiosis decreased osteoblast and increased osteoclast activities, changes that were prevented by both L. reuteri and MDY. These data underscore the importance of microbial dysbiosis in the regulation of intestinal permeability and bone health, as well as identify L. reuteri and MDY as novel therapies for preventing these adverse effects.
mSphere
Annually, an estimated 2 million osteoporotic fractures occur in the United States alone. Osteopo... more Annually, an estimated 2 million osteoporotic fractures occur in the United States alone. Osteoporosis imparts a great burden on the health care system. The identification of novel regulators of bone health is critical for developing more effective therapeutics. A previous study on the colonization of germ-free (GF) mice with a microbial community has demonstrated that bacterial colonization dramatically increases bone loss. We therefore investigated the impact of multiple microbial communities in different mice to understand how generalizable the impact of bacterial colonization is on bone health. To investigate the impact of different microbial communities on bone health in outbred and inbred mouse strains, gavage was performed on GF Swiss Webster and GF C57BL/6 mice to introduce distinct microbiotas that originated from either humans or mice. GF mice displayed a high degree of colonization, as indicated by more than 90% of the operational taxonomic units present in the starting i...
PLOS ONE, 2017
Background & aims Wnt10b is a crucial regulator of bone density through its ability to promote os... more Background & aims Wnt10b is a crucial regulator of bone density through its ability to promote osteoblastogenesis. Parathyroid hormone has been shown to regulate Wnt10b expression in CD8 + T cells. However, the relative expression and other source(s) of Wnt10b in the bone marrow immune cells (BMICs) is unknown. Sex hormones and cytokines such as, estrogen and TNFα are critical regulators of bone physiology but whether they regulate BMIC Wnt10b expression is unclear. To determine the potential regulation of Wnt10b by estrogen and TNFα, we assessed Wnt10b expression by flow cytometry under estrogen-and TNFα-deficient conditions. Methods Effects of TNFα was determined in male and female C57BL/6 wildtype and TNFα knockout mice. Effect of estrogen was investigated 4, 6 and 8 weeks post-surgery in ovariectomized Balb/c mice. Intracellular Wnt10b was detected using goat anti-mouse Wnt10b and a conjugated secondary antibody and analyzed by flow cytometry. Results Wnt10b expression was sex-and lineage-specific. Females had 1.8-fold higher Wnt10b signal compared to males. Percent of Wnt10b + myeloid cells was higher in females than males (8.9% Vs 5.4%) but Wnt10b + lymphoid cells was higher in males than females (6.3% Vs 2.5%). TNFα ablation in males increased total BM Wnt10b expression 1.5-fold but significantly reduced the percentage of BM Wnt10b + CD4 + T cells (65%), CD8 + T cells (59%), dendritic cells (59%), macrophages (56%) and granulocytes (52%). These effects of TNFα on Wnt10b were observed only in males. In contrast to TNFα, estrogen-deficiency had indirect effects on BMIC Wnt10b levels; reducing the average percentage of BM Wnt10b + CD8 + T cells (25%) and granulocytes (26%) across an 8-week time course.