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Investigating the human cartilage endplate in chronic low back pain: from mechanisms of degeneration to molecular, cell and tissue level characterization

Lakstins, Katherine S
http://rave.ohiolink.edu/etdc/view?acc_num=osu1584627459584403

Abstract Details

2020, Doctor of Philosophy, Ohio State University, Biomedical Engineering.
Degeneration/calcification of the cartilage endplate (CEP) has been linked to the onset and progression of intervertebral disc (IVD) degeneration yet the CEP remains under characterized and understudied. The aims of this dissertation were to evaluate the effect of in vitro cell culture conditions, including biochemical hypertrophic stimuli (10% fetal bovine serum (FBS) and Wnt agonist), oxygen tension (Aim 1), and substrate stiffness (Aim 2), on hypertrophic differentiation, a possible mechanism of degeneration/calcification, in human CEP cells and to characterize the human CEP on the molecular, cell and tissue level (Aim 3) and compare it to the CEP of animal models commonly used to study IVD degeneration, specifically bovine and canine species (Aim 4). For Aim 1, isolated human CEP cells were cultured as pellets for 21 days at either 5% or 20.7% O2 and treated with 10% FBS or Wnt agonist, two biochemical stimuli known to induce hypertrophy in articular chondrocytes. CEP cells did not exhibit a hypertrophic morphology in response to hypertrophic stimuli but did display other hallmarks of chondrocyte hypertrophy and degeneration including hypertrophic gene and protein expression, and a decrease in healthy proteoglycans (glycosaminoglycan (GAG)) and increase in fibrous collagen accumulation. For Aim 2, human CEP cells were cultured in either 2% (soft) or 4.5% (stiff) agarose gels at 5% O2 for 11 days. No significant differences in cell morphology, or protein expression of hypertrophic cell markers were observed between substrate stiffnesses. For Aims 3 and 4, isolated human, bovine and canine CEP, nucleus pulposus (NP), annulus fibrosus (AF) and articular cartilage (AC) tissue and cells were evaluated for cell morphology, matrix composition/structure, GAG content, and gene and protein expression. Significant differences in matrix and cell marker gene expression were observed between human CEP and NP or AF, with greatest differences between the human CEP and AC. We were able to distinguish human NP from CEP cells using collagen-X, highlighting collagen-X as a potential CEP marker. The bovine and canine CEPs were thinner with rounded cells and the canine CEP contained significantly more GAGs compared to human. The bovine CEP demonstrated increased expression of ACAN, COL1 and COL2 and decreased expression of T, FBLN1 and COLX and the canine CEP had increased COL2 and decreased COL1, KRT19, MKX, FBLN1, COLX expression compared to human. These findings demonstrate that CEP cells take on a degenerative phenotype in response to biochemical hypertrophic stimuli in vitro, but do not undergo classical changes in cell morphology associated with hypertrophic differentiation regardless of oxygen levels, highlighting potential differences in the response of CEP cells versus AC cells to the same stimuli. Furthermore, at the cell and tissue level, the human CEP demonstrates a unique phenotype that is different from the NP, AF and AC. Finally, fewer differences were observed between the bovine and human CEP, therefore the bovine CEP may be a more appropriate animal model for studying CEP degeneration. Overall, this work can help inform future mechanistic studies and regenerative strategies that target the CEP and its role in chronic LBP.
Devina Purmessur (Advisor)
Alan Litsky (Committee Member)
Sarah Moore (Committee Member)
Gregory Lafyatis (Committee Member)
190 p.
Biomedical Engineering
low back pain; cartilage endplate; intervertebral disc; intervertebral disc degeneration

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Citations

  • Lakstins, K. S. (2020). Investigating the human cartilage endplate in chronic low back pain: from mechanisms of degeneration to molecular, cell and tissue level characterization [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1584627459584403

    APA Style (7th edition)

  • Lakstins, Katherine. Investigating the human cartilage endplate in chronic low back pain: from mechanisms of degeneration to molecular, cell and tissue level characterization. 2020. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1584627459584403.

    MLA Style (8th edition)

  • Lakstins, Katherine. "Investigating the human cartilage endplate in chronic low back pain: from mechanisms of degeneration to molecular, cell and tissue level characterization." Doctoral dissertation, Ohio State University, 2020. http://rave.ohiolink.edu/etdc/view?acc_num=osu1584627459584403

    Chicago Manual of Style (17th edition)

osu1584627459584403
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