Nerve growth factor promotes expression of novel genes in intervertebral disc cells that regulate tissue degradation

Kao, Tung‐Wei; Peng, Yi‐Jen; Tsou, Hsi‐Kai; Salter, Donald; Lee, Herng‐Sheng

doi:10.3171/2014.6.spine13756

Cited by 17 publications

(18 citation statements)

References 46 publications

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“…In this study, we have observed that NGF, which is likely to play a role in nerve ingrowth into the degenerated IVD [5,18], may also act as a catabolic factor promoting disc degeneration [15]. The observation that NGF induced overexpression of lipocalin 2 to enhance MMP9 activity in AF cells is previously unreported.…”

Section: Discussionmentioning

confidence: 74%

“…The detailed microarray methods and results were provided in our previous study [15]. Briefly, gene expression analysis of rat-tail disc cells with or without NGF stimulation for 5 days was undertaken using the Rat Whole Genome OneArray TM (Phalanx Biotech Group, Hsinchu, Taiwan), signals being scanned by an Axon 4000 scanner (Molecular Devices, Sunnyvale, CA, USA).…”

Section: Microarray Assaymentioning

confidence: 99%

“…Using purified RNA samples from the AF cells, we found that 97 of the 24,358 transcripts (\0.4 %) covered by Rat OneArray showed [twofold change after NGF stimulation [15]. The top 30 upregulated genes by a log2 ratio of more than 1.5 are shown as supplementary data.…”

Section: Effect Of Ngf On Annulus Fibrosus Cell Gene Expressionmentioning

confidence: 99%

“…As such NGF appears to be a prime target for intervention in the course of IVD degeneration with the potential to both ameliorate pain and decrease or reverse the rate of disease progression. NGF influences expression of several catabolic genes in animal and human IVD cells including ch3l1, lipocalin 2 (lcn2), mmp3 and mmp13 according to our previous microarray-based study [15]. Since Lcn2 protein has been reported as forming a complex with MMP9 to protect it from degradation and thereby enhancing its catabolic function in osteoarthritic cartilage [16], the current study was undertaken to test the hypothesis that NGF regulates expression of lipocalin 2 in AF cells resulting in an increase in MMP9 activity which would potentially contribute to IVD degeneration.…”

Section: Introductionmentioning

confidence: 99%

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Nerve growth factor increases MMP9 activity in annulus fibrosus cells by upregulating lipocalin 2 expression

et al. 2014

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Purpose Nerve growth factor (NGF) expression and activity is important in chronic lower back pain but may also act as a pro-catabolic factor in the pathogenesis of intervertebral disc (IVD) degeneration. Lipocalin 2 (Lcn2) expression in IVD was upregulated by NGF stimulation in our previous study. The current study was undertaken to identify potential mechanisms of the latter effect including potential interactions between Lcn2 and matrix metalloproteinase 9 (MMP9). Methods Rat annulus fibrosus (AF) cells were stimulated by NGF and subjected to microarray analysis, subsequent real-time PCR, western immunoblotting, and immunofluorescence. Cells were treated with NGF in the absence or presence of the NGF inhibitor Ro 08-2750. Zymography and functional MMP9 assays were used to determine MMP9 activity, whilst the dimethyl-methylene blue assay was used to quantify the release of glycosaminoglycans (GAGs) reflecting catabolic effects following NGF treatment. Immunoprecipitation with immunoblotting was used to identify interactions between MMP9 and Lcn2. Results Increased expression of Lcn2 gene and protein following NGF stimulation was confirmed by microarray analysis, real-time PCR, western blot and immunofluorescence. Zymography showed that NGF enhanced 125-kDa gelatinase activity, identified as a Lcn2/MMP9 complex by immunoprecipitation and immunoblotting. Functional assays showed increased MMP9 activity and GAG release in the presence of NGF. The effects of NGF were neutralized by the presence of Ro 08-2750. Conclusions NGF upregulates Lcn2 expression and increases MMP9 activity in AF cells; processes which are likely to potentiate degeneration of AF tissue in vivo. Anti-NGF treatment may have benefit for management of pain Electronic supplementary material The online version of this article

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Section: Discussionmentioning

confidence: 74%

Section: Microarray Assaymentioning

confidence: 99%

Section: Effect Of Ngf On Annulus Fibrosus Cell Gene Expressionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Nerve growth factor increases MMP9 activity in annulus fibrosus cells by upregulating lipocalin 2 expression

et al. 2014

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“…Increased neurotrophin activity is one potential cause of IDD. Nerve growth factor (NGF) stimulation has been shown to upregulate MMP-3 expression and increase the ratio of MMP-3 to tissue inhibitor of metalloproteinase (TIMP)-3 in rat and human IVD cells and tissue, which is abolished by Ro 08-2750, a specific inhibitor of NGF [65]. A later study by the same authors has demonstrated that NGF also augments the expression and activity of MMP-9 in rat AF cells by upregulating lipocalin 2 expression [66].…”

Section: Mmps and Iddmentioning

confidence: 99%

MMPs and ADAMTSs in intervertebral disc degeneration

Wang

et al. 2015

Clinica Chimica Acta

164

124

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Bicarbonate Recycling by HIF-1–Dependent Carbonic Anhydrase Isoforms 9 and 12 Is Critical in Maintaining Intracellular pH and Viability of Nucleus Pulposus Cells

Silagi

Schoepflin

Seifert

et al. 2017

Journal of Bone and Mineral Research

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Intervertebral disc degeneration is a ubiquitous condition closely linked to chronic low-back pain. The health of the avascular nucleus pulposus (NP) plays a crucial role in the development of this pathology. We tested the hypothesis that a network comprising HIF-1α, carbonic anhydrase (CA) 9 and 12 isoforms, and sodium-coupled bicarbonate cotransporters (NBCs) buffer intracellular pH through coordinated bicarbonate recycling. Contrary to the current understanding of NP cell metabolism, analysis of metabolic-flux data from Seahorse XF analyzer showed that CO hydration contributes a significant source of extracellular proton production in NP cells, with a smaller input from glycolysis. Because enzymatic hydration of CO is catalyzed by plasma membrane-associated CAs we measured their expression and function in NP tissue. NP cells robustly expressed isoforms CA9/12, which were hypoxia-inducible. In addition to increased mRNA stability under hypoxia, we observed binding of HIF-1α to select hypoxia-responsive elements on CA9/12 promoters using genomic chromatin immunoprecipitation. Importantly, in vitro loss of function studies and analysis of discs from NP-specific HIF-1α null mice confirmed the dependency of CA9/12 expression on HIF-1α. As expected, inhibition of CA activity decreased extracellular acidification rate independent of changes in HIF activity or lactate/H efflux. Surprisingly, CA inhibition resulted in a concomitant decrease in intracellular pH that was mirrored by inhibition of sodium-bicarbonate importers. These results suggested that extracellular bicarbonate generated by CA9/12 is recycled to buffer cytosolic pH fluctuations. Importantly, long-term intracellular acidification from CA inhibition lead to compromised cell viability, suggesting that plasma-membrane proton extrusion pathways alone are not sufficient to maintain homeostatic pH in NP cells. Taken together, our studies show for the first time that bicarbonate buffering through the HIF-1α-CA axis is critical for NP cell survival in the hypoxic niche of the intervertebral disc. © 2017 American Society for Bone and Mineral Research.

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Nerve growth factor promotes expression of novel genes in intervertebral disc cells that regulate tissue degradation

Cited by 17 publications

References 46 publications

Nerve growth factor increases MMP9 activity in annulus fibrosus cells by upregulating lipocalin 2 expression

Nerve growth factor increases MMP9 activity in annulus fibrosus cells by upregulating lipocalin 2 expression

MMPs and ADAMTSs in intervertebral disc degeneration

Bicarbonate Recycling by HIF-1–Dependent Carbonic Anhydrase Isoforms 9 and 12 Is Critical in Maintaining Intracellular pH and Viability of Nucleus Pulposus Cells

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