Effects of low glucose concentrations on oxygen consumption rates of intervertebral disc cells

Abstract

Study design: Investigation of the effects of low glucose concentrations on the oxygen consumption rates of intervertebral disc cells.

Objectives: To determine the oxygen consumption rate of porcine anulus fibrosus (AF) cells at different glucose concentrations and to examine the differences in the oxygen consumption rate between AF and nucleus pulposus (NP) cells at different glucose levels.

Summary of background data: Poor nutrient supply has been suggested as a potential mechanism for degeneration of the intervertebral disc (IVD). Distribution of nutrients in the IVD is strongly dependent on transport properties of the tissue and cellular metabolic rates. Previous studies have shown dependence of oxygen consumption rate of IVD cells on oxygen tension, pH levels, and glucose levels outside the physiologic range. However, the oxygen consumption rate of AF cells at in vivo glucose levels has not been investigated.

Methods: IVD cells were isolated from the outer AF and NP of 4- to 5-month-old porcine lumbar discs. The changes in oxygen tension were recorded when cells were cultured in sealed metabolism chamber. The oxygen consumption rate of cells was determined by theoretical curve fitting using the Michaelis-Menten equation.

Results: The outer AF cells cultured in high glucose medium (25 mmol/L) exhibited the lowest oxygen consumption rate, whereas no significant differences in oxygen consumption rates were found among outer AF cells cultured at physiologic glucose levels (i.e., 1 mmol/L, 2.5 mmol/L, 5 mmol/L). The oxygen consumption rate of NP cells was significantly greater than that of outer AF cells.

Conclusion: Since the oxygen consumption rates determined in this study are comparable to the findings in the literature, this study has developed a new alternative method for determining oxygen consumption rate. The oxygen consumption rates of IVD cells reported in this study will be valuable for theoretically predicting local oxygen concentrations in IVD, which can provide a better understanding of transport of oxygen in the discs.

Figure 1

(a) Anatomy of the intervertebral disc. (b) Different regions of porcine intervertebral disc.

Figure 1

(a) Anatomy of the intervertebral disc. (b) Different regions of porcine intervertebral disc.

Figure 2

(a) Schematic of experimental setup. (b) Typical experimental data of oxygen concentration measurement for NP and outer AF cells along with theoretical curve fittings. Cells were cultured in the metabolism chamber containing DMEM medium and 1 mM glucose (pH=7.4).

Figure 2

(a) Schematic of experimental setup. (b) Typical experimental data of oxygen concentration measurement for NP and outer AF cells along with theoretical curve fittings. Cells were cultured in the metabolism chamber containing DMEM medium and 1 mM glucose (pH=7.4).

Figure 3

Comparison of (a) V_max and (b) K_m among the outer AF or NP cells cultured in the media with different glucose concentrations (n=5 for each group).

Figure 4

Comparison of the average of (a) V_max and (b) K_m between the outer AF or NP cells (n=20 for each group).

Figure 5

Comparison of oxygen consumption rate at 5% oxygen tension (a) between the outer AF and NP cells (n=20 for each group) and (b) among the outer AF or NP cells cultured in the media with different glucose concentrations (n=5 for each group). The oxygen consumption rate was calculated using Eq. 1 based on V_max and K_m which were determined from theoretical curve fitting.

Figure 6

Comparison of oxygen consumption rate within the range of oxygen tension from 0 to 20% between this study and those in the literature. The consumption rate of bovine NP cells at pH 7.4 was calculated based on the modified Michaelis-Menten equation proposed in the study of Bibby et al [22]. The oxygen consumption rate of canine NP cells was converted from the oxygen consumption of canine NP tissue based on its cell density reported in the study of Holm et al. [5].