An &lt;em&gt;In Vitro&lt;/em&gt; Organ Culture Model of the Murine Intervertebral Disc

Liu, Jennifer W.; Lin, Kevin; Weber, C.; Bhalla, Sameer; Kelso, Sean; Wang, Kaixi; Tang, S. Y.

doi:10.3791/55437-v

Cited by 4 publications

(5 citation statements)

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“…According to previous research, the functional spine unit can only survive for 3‐5 days in the condition of a low concentration of foetal bovine serum medium for in vitro culture 24 . Whereas, survival time can be prolonged for up to at least 10 days if 10% foetal bovine serum is present in culture medium 25,26 . In our experiment, intervertebral discs survived up to 14 days in 10% foetal bovine serum medium.…”

Section: Discussionsupporting

confidence: 50%

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Bmi deficiency causes oxidative stress and intervertebral disc degeneration which can be alleviated by antioxidant treatment

Zhang

et al. 2020

J Cellular Molecular Medi

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The transcriptional repressor Bmi‐1 is involved in cell‐cycle regulation and cell senescence, the deficiency of which has been shown to cause oxidative stress. This study investigated whether Bmi‐1 deficiency plays a role in promoting disc degeneration and the effect of treatment with antioxidant N‐acetylcysteine (NAC) on intervertebral disc degeneration. Bmi‐1 −/− mice were treated with the antioxidant NAC, supplied in drinking water (Bmi‐1 −/− +NAC). For in vitro experiments, mouse intervertebral discs were cultured under low oxygen tension and serum‐limiting conditions in the presence of tumour necrosis factor α and interleukin 1β in order to mimic degenerative insult. Disc metabolism parameters in these in vitro and in vivo studies were evaluated by histopathological, immunohistochemical and molecular methods. Bmi‐1 −/− mice showed lower collagen Ⅱ and aggrecan levels and higher collagen Ⅹ levels than wild‐type and Bmi‐1 −/− +NAC mice. Bmi‐1 −/− mice showed significantly lower superoxide dismutase (SOD)‐1, SOD‐2, glutathione peroxidase (GPX)‐1 and GPX‐3 levels than their wild‐type littermates and Bmi‐1 −/− + NAC mice. Relative to Bmi‐1 −/− mice, the control and Bmi‐1 −/− +NAC mice showed significantly lower p16, p21, and p53 levels. These results demonstrate that Bmi‐1 plays an important role in attenuating intervertebral disc degeneration in mice by inhibiting oxidative stress and cell apoptosis.

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Section: Discussionsupporting

confidence: 50%

“…24 Whereas, survival time can be prolonged for up to at least 10 days if 10% foetal bovine serum is present in culture medium. 25,26 In our experiment, intervertebral discs survived up to 14 days in 10% foetal bovine serum medium. Previous studies have established degeneration models using enzyme digestion or acupuncture methods.…”

Section: Discussionmentioning

confidence: 68%

Bmi deficiency causes oxidative stress and intervertebral disc degeneration which can be alleviated by antioxidant treatment

Zhang

et al. 2020

J Cellular Molecular Medi

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“…FSUs were immediately placed in culture media in 24‐well plates and pre‐conditioned for 7 days prior to all experiments. Culture media consisted of 1:1 Dulbecco's modified Eagle's medium: Nutrient mixture F‐12 (DMEM:F12) supplemented with 20% fetal bovine serum and 1% penicillin–streptomycin 35 . The FSUs were cultured under the following conditions (37°C, 5% CO 2 , 100% humidity) and were assigned to one of three distinct experiments: transient activation of TRPV4 repeated activation of TRPV4 and sustained activation of TRPV4 (Figure 1).…”

Section: Methodsmentioning

confidence: 99%

“…Culture media consisted of 1:1 Dulbecco's modified Eagle's medium: Nutrient mixture F-12 (DMEM:F12) supplemented with 20% fetal bovine serum and 1% penicillin-streptomycin. 35 The FSUs were cultured under the following conditions (37°C, 5% CO 2 , 100% humidity) and were assigned to one of three distinct experiments: transient activation of TRPV4 repeated activation of TRPV4 and sustained activation of TRPV4 (Figure 1). Our prior work demonstrated that these culture conditions maintain the viability of all cells in the IVD, including those in the cartilaginous end-plate, but not cells in the bone marrow space or osteocytes.…”

Section: Animalsmentioning

confidence: 99%

Modulation of TRPV4 protects against degeneration induced by sustained loading and promotes matrix synthesis in the intervertebral disc

et al. 2022

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“…The FSUs were immediately placed in culture media in 24-well plates and preconditioned for 7 days before the administration of the experimental conditions. 26 The culture media consisted of a mixture of Dulbecco's modified Eagle's medium and nutrient mixture F-12 in a 1:1 ratio (DMEM:F12), supplemented with 20% fetal bovine serum and 1% penicillin-streptomycin. The FSUs were cultured under controlled conditions of 37 C, 5% CO 2 , and 100% humidity.…”

Section: Animalsmentioning

confidence: 99%

TRPV4 differentially controls inflammatory cytokine networks during static and dynamic compression of the intervertebral disc

Easson,

Savadipour,

Gonzalez

et al. 2023

JOR Spine

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BackgroundThe ion channel transient receptor potential vanilloid 4 (TRPV4) critically transduces mechanical forces in the IVD, and its inhibition can prevent IVD degeneration due to static overloading. However, it remains unknown whether different modes of loading signals through TRPV4 to regulate the expression of inflammatory cytokines. We hypothesized that TRPV4 signaling is essential during static and dynamic loading to mediate homeostasis and mechanotransduction.MethodsMouse functional spine units were isolated and either cyclically compressed for 5 days (1 Hz, 1 h, 10% strain) or statically compressed (24 h, 0.2 MPa). Conditioned media were monitored at 6 h, 24 h, 2 days, and 5 days, with and without TRPV4 inhibition. Effects of TRPV4 activation was also evaluated without loading. The media was analyzed for a panel of 44 cytokines using a microbead array and then a correlative network was constructed to explore the regulatory relationships during loading and TRPV4 inhibition. After the loading regimen, the IVDs were evaluated histologically for degeneration.ResultsActivation of TRPV4 led to an increase interleukin‐6 (IL‐6) family of cytokines (IL‐6, IL‐11, IL‐16, and leukemia inhibitory factor [LIF]) and decreased the T‐cell (CCL3, CCL4, CCL17, CCL20, CCL22, and CXCL10) and monocyte (CCL2 and CCL12) recruiting chemokines by the IVD. Dynamic and static loading each provoked unique chemokine correlation networks. The inhibition of TRPV4 during dynamic loading dysregulated the relationship between LIF and other cytokines, while the inhibition of TRPV4 during static loading disrupted the connectivity of IL‐16 and VEGFA.ConclusionsWe demonstrated that TRPV4 critically mediates the cytokine production following dynamic and static loading. The activation of TRPV4 upregulated a diverse set of cytokines that may suppress the chemotaxis of T‐cells and monocytes, implicating the role of TRPV4 in maintaining the immune privilege of healthy IVD.

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An <em>In Vitro</em> Organ Culture Model of the Murine Intervertebral Disc

Cited by 4 publications

References 0 publications

Bmi deficiency causes oxidative stress and intervertebral disc degeneration which can be alleviated by antioxidant treatment

Bmi deficiency causes oxidative stress and intervertebral disc degeneration which can be alleviated by antioxidant treatment

Modulation of TRPV4 protects against degeneration induced by sustained loading and promotes matrix synthesis in the intervertebral disc

TRPV4 differentially controls inflammatory cytokine networks during static and dynamic compression of the intervertebral disc

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