Limits...
Effects of Age and Estrogen on Skeletal Gene Expression in Humans as Assessed by RNA Sequencing.

Farr JN, Roforth MM, Fujita K, Nicks KM, Cunningham JM, Atkinson EJ, Therneau TM, McCready LK, Peterson JM, Drake MT, Monroe DG, Khosla S - PLoS ONE (2015)

Bottom Line: Interestingly, the LEF1 transcription factor, which is a classical downstream target of the Wnt/β-catenin signaling pathway, was significantly downregulated in the bones from the old versus young women; consistent with this, LEF1 binding sites were significantly enriched in the promoter regions of the differentially expressed genes in the old versus young women, suggesting that aging was associated with alterations in Wnt signaling in bone.In conclusion, our data demonstrate that aging alters a substantial portion of the skeletal transcriptome, whereas E therapy appears to have significant, albeit less wide-ranging effects.These data provide a valuable resource for the potential identification of novel biomarkers associated with age-related bone loss and also highlight potential pathways that could be targeted to treat osteoporosis.

View Article: PubMed Central - PubMed

Affiliation: Division of Endocrinology, Department of Medicine, Mayo Clinic College of Medicine, Rochester, MN, 55905, United States of America; Robert and Arlene Kogod Center on Aging, Mayo Clinic College of Medicine, Rochester, MN, 55905, United States of America.

ABSTRACT

Unlabelled: Precise delineation of the specific genes and pathways altered with aging and estrogen (E) therapy may lead to new skeletal biomarkers and the development of novel bone therapeutics. Previous human bone studies, however, have been limited by only examining pre-specified genes and pathways. High-throughput RNA sequencing (RNAseq), on the other hand, offers an unbiased approach to examine the entire transcriptome. Here we present an RNAseq analysis of human bone samples, obtained from iliac crest needle biopsies, to yield the first in vivo interrogation of all genes and pathways that may be altered in bone with aging and E therapy in humans. 58 healthy women were studied, including 19 young women (mean age ± SD, 30.3 ± 5.4 years), 19 old women (73.1 ± 6.6 years), and 20 old women treated with 3 weeks of E therapy (70.5 ± 5.2 years). Using generally accepted criteria (false discovery rate [q] < 0.10), aging altered a total of 678 genes and 12 pathways, including a subset known to regulate bone metabolism (e.g., Notch). Interestingly, the LEF1 transcription factor, which is a classical downstream target of the Wnt/β-catenin signaling pathway, was significantly downregulated in the bones from the old versus young women; consistent with this, LEF1 binding sites were significantly enriched in the promoter regions of the differentially expressed genes in the old versus young women, suggesting that aging was associated with alterations in Wnt signaling in bone. Further, of the 21 unique genes altered in bone by E therapy, the expression of INHBB (encoding for the inhibin, beta B polypeptide), which decreased with aging (by 0.6-fold), was restored to young adult levels in response to E therapy. In conclusion, our data demonstrate that aging alters a substantial portion of the skeletal transcriptome, whereas E therapy appears to have significant, albeit less wide-ranging effects. These data provide a valuable resource for the potential identification of novel biomarkers associated with age-related bone loss and also highlight potential pathways that could be targeted to treat osteoporosis.

Trial registration: ClinicalTrials.gov NCT02349113.

No MeSH data available.


Related in: MedlinePlus

(A) Notch pathway and (B) Wnt/β-catenin pathway genes are altered with aging in human bone biopsies.Genes in the Notch pathway and Wnt/β-catenin pathway are significantly altered in old relative to young women revealed by RNAseq based on pathway analysis using the Ingenuity Pathway Analysis software (see Statistical analyses). Values are presented as median fold changes (95% CIs) for old relative to young subjects. †p < 0.01; ‡p < 0.001.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4581624&req=5

pone.0138347.g002: (A) Notch pathway and (B) Wnt/β-catenin pathway genes are altered with aging in human bone biopsies.Genes in the Notch pathway and Wnt/β-catenin pathway are significantly altered in old relative to young women revealed by RNAseq based on pathway analysis using the Ingenuity Pathway Analysis software (see Statistical analyses). Values are presented as median fold changes (95% CIs) for old relative to young subjects. †p < 0.01; ‡p < 0.001.

Mentions: Particularly noteworthy, and consistent with our previous study using QPCR [6], was the significant alteration (q = 0.021) of the Notch signaling pathway with aging in the bone biopsies (Table 2). Indeed, expression levels of key regulatory genes from the Notch pathway including NOTCH3 and NOTCH4 (both single-pass transmembrane cell surface receptors), as well as JAG2 and DLL4 (two Notch receptor ligands that cause proteolytic cleavage of Notch receptors upon binding) and two critical direct downstream Notch pathway targets (HES1 and HEY1) were significantly up-regulated in bone biopsies of old versus young subjects (Fig 2A). It is also noteworthy that the Wnt/β-catenin signaling pathway was altered with aging, although as noted above, this pathway fell into the category of bone-related pathways with p < 0.05, but q > 0.10 (Table 3). In this pathway, transcriptional levels of the genes for SOX4, SFRP5, and LEF1 were decreased, whereas levels for SOX17, SOX7, FZD4, CDH3, SFRP1, SOX18, SOX13, and CCND1 were all increased in bone biopsies of old versus young women (Fig 2B). By contrast, however, there were no changes in any of the lipoprotein receptor-related proteins previously linked to bone (e.g., LRP4, LRP5, and LRP6) or the associated DKK family of Wnt antagonists with aging. Finally, as evident in Table 3, additional pathways with known roles in bone metabolism, including Inhibition of Matrix Metalloproteases, eNOS signaling, Nitric Oxide Signaling in the Cardiovascular System, PDGF Signaling, Gαi Signaling, and Oncostatin M Signaling were all altered in bone biopsies of old versus young subjects (at least at the p < 0.05 level, but only the first 2 at the q < 0.10 level).


Effects of Age and Estrogen on Skeletal Gene Expression in Humans as Assessed by RNA Sequencing.

Farr JN, Roforth MM, Fujita K, Nicks KM, Cunningham JM, Atkinson EJ, Therneau TM, McCready LK, Peterson JM, Drake MT, Monroe DG, Khosla S - PLoS ONE (2015)

(A) Notch pathway and (B) Wnt/β-catenin pathway genes are altered with aging in human bone biopsies.Genes in the Notch pathway and Wnt/β-catenin pathway are significantly altered in old relative to young women revealed by RNAseq based on pathway analysis using the Ingenuity Pathway Analysis software (see Statistical analyses). Values are presented as median fold changes (95% CIs) for old relative to young subjects. †p < 0.01; ‡p < 0.001.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4581624&req=5

pone.0138347.g002: (A) Notch pathway and (B) Wnt/β-catenin pathway genes are altered with aging in human bone biopsies.Genes in the Notch pathway and Wnt/β-catenin pathway are significantly altered in old relative to young women revealed by RNAseq based on pathway analysis using the Ingenuity Pathway Analysis software (see Statistical analyses). Values are presented as median fold changes (95% CIs) for old relative to young subjects. †p < 0.01; ‡p < 0.001.
Mentions: Particularly noteworthy, and consistent with our previous study using QPCR [6], was the significant alteration (q = 0.021) of the Notch signaling pathway with aging in the bone biopsies (Table 2). Indeed, expression levels of key regulatory genes from the Notch pathway including NOTCH3 and NOTCH4 (both single-pass transmembrane cell surface receptors), as well as JAG2 and DLL4 (two Notch receptor ligands that cause proteolytic cleavage of Notch receptors upon binding) and two critical direct downstream Notch pathway targets (HES1 and HEY1) were significantly up-regulated in bone biopsies of old versus young subjects (Fig 2A). It is also noteworthy that the Wnt/β-catenin signaling pathway was altered with aging, although as noted above, this pathway fell into the category of bone-related pathways with p < 0.05, but q > 0.10 (Table 3). In this pathway, transcriptional levels of the genes for SOX4, SFRP5, and LEF1 were decreased, whereas levels for SOX17, SOX7, FZD4, CDH3, SFRP1, SOX18, SOX13, and CCND1 were all increased in bone biopsies of old versus young women (Fig 2B). By contrast, however, there were no changes in any of the lipoprotein receptor-related proteins previously linked to bone (e.g., LRP4, LRP5, and LRP6) or the associated DKK family of Wnt antagonists with aging. Finally, as evident in Table 3, additional pathways with known roles in bone metabolism, including Inhibition of Matrix Metalloproteases, eNOS signaling, Nitric Oxide Signaling in the Cardiovascular System, PDGF Signaling, Gαi Signaling, and Oncostatin M Signaling were all altered in bone biopsies of old versus young subjects (at least at the p < 0.05 level, but only the first 2 at the q < 0.10 level).

Bottom Line: Interestingly, the LEF1 transcription factor, which is a classical downstream target of the Wnt/β-catenin signaling pathway, was significantly downregulated in the bones from the old versus young women; consistent with this, LEF1 binding sites were significantly enriched in the promoter regions of the differentially expressed genes in the old versus young women, suggesting that aging was associated with alterations in Wnt signaling in bone.In conclusion, our data demonstrate that aging alters a substantial portion of the skeletal transcriptome, whereas E therapy appears to have significant, albeit less wide-ranging effects.These data provide a valuable resource for the potential identification of novel biomarkers associated with age-related bone loss and also highlight potential pathways that could be targeted to treat osteoporosis.

View Article: PubMed Central - PubMed

Affiliation: Division of Endocrinology, Department of Medicine, Mayo Clinic College of Medicine, Rochester, MN, 55905, United States of America; Robert and Arlene Kogod Center on Aging, Mayo Clinic College of Medicine, Rochester, MN, 55905, United States of America.

ABSTRACT

Unlabelled: Precise delineation of the specific genes and pathways altered with aging and estrogen (E) therapy may lead to new skeletal biomarkers and the development of novel bone therapeutics. Previous human bone studies, however, have been limited by only examining pre-specified genes and pathways. High-throughput RNA sequencing (RNAseq), on the other hand, offers an unbiased approach to examine the entire transcriptome. Here we present an RNAseq analysis of human bone samples, obtained from iliac crest needle biopsies, to yield the first in vivo interrogation of all genes and pathways that may be altered in bone with aging and E therapy in humans. 58 healthy women were studied, including 19 young women (mean age ± SD, 30.3 ± 5.4 years), 19 old women (73.1 ± 6.6 years), and 20 old women treated with 3 weeks of E therapy (70.5 ± 5.2 years). Using generally accepted criteria (false discovery rate [q] < 0.10), aging altered a total of 678 genes and 12 pathways, including a subset known to regulate bone metabolism (e.g., Notch). Interestingly, the LEF1 transcription factor, which is a classical downstream target of the Wnt/β-catenin signaling pathway, was significantly downregulated in the bones from the old versus young women; consistent with this, LEF1 binding sites were significantly enriched in the promoter regions of the differentially expressed genes in the old versus young women, suggesting that aging was associated with alterations in Wnt signaling in bone. Further, of the 21 unique genes altered in bone by E therapy, the expression of INHBB (encoding for the inhibin, beta B polypeptide), which decreased with aging (by 0.6-fold), was restored to young adult levels in response to E therapy. In conclusion, our data demonstrate that aging alters a substantial portion of the skeletal transcriptome, whereas E therapy appears to have significant, albeit less wide-ranging effects. These data provide a valuable resource for the potential identification of novel biomarkers associated with age-related bone loss and also highlight potential pathways that could be targeted to treat osteoporosis.

Trial registration: ClinicalTrials.gov NCT02349113.

No MeSH data available.


Related in: MedlinePlus