Effect of the organic component of Microcrystalline Hydroxyapatite Calcium (MCH-CALTM) supplement on musculoskeletal cell behaviour

Abstract

There are a vast number of diseases and injuries that occur in the musculoskeletal system including osteoporosis, non-union fractures, tendinopathy and tendon tears. The treatment options currently available for these diseases and injuries are limited and achieve poor healing. Waitaki MCH-CAL™ is a bovine bone-derived product that is marketed worldwide as a calcium supplement to treat degenerative diseases. The aim of this project is to investigate whether the organic component of Waitaki MCH-CAL™ has the potential to be used as a tool in musculoskeletal regenerative medicine. The protein levels in the protein digest of Waitaki MCH-CAL™ was measured via ELISA. The effect of Waitaki MCH-CAL™ (organic component) on cell number and differentiation was examined on MC3T3-EI pre-osteoblast-like cells and on rat tenocytes, by alamarBlue™ incorporation and real time PCR respectively. These effects were measured in 2D (plastic culture plates), collagen coated plates and 3D (collagen gels), with PDGF as a positive control. Detectable protein levels were higher in freeze-dried Waitaki MCH-CAL™ (537.5μg/ml) compared to vacuum-dried Waitaki MCH-CAL™ (323μg/ml) and the finished product extract of Waitaki MCH-CAL™ (379.7μg/ml). However, h-BMP2/7 levels were below detectable levels (<62.5pg/ml). The non-concentrated protein digest of Waitaki MCH-CAL™ (freeze dried) caused a dose dependent decrease in cell number in both the cell types, in all the culture systems, while PDGF increased cell number in both the cells types, as expected. In general, the gene expression of tenocyte and non-tenocyte markers decreased/were absent in tenocytes in response to Waitaki MCH-CAL™ protein digest. However, these results were not statistically significant and therefore inconclusive. In MC3T3-E1 cells, osteoblast marker gene expression appeared to increase, in response to Waitaki MCH-CAL™ protein digest. Gene expression in the PDGF treated osteoblasts, seeded on collagen coated plates, increased, whereas in the tenocytes gene expression decreased. These results suggest that the Waitaki MCH-CAL™ protein digest may have the potential to stimulate osteoblast differentiation, without causing an adverse effect in tendons, and therefore may be used as a tool for regenerative medicine. However, these studies are preliminary – further experiments need to be conducted in vitro, in animal models and finally in a clinical setting before any conclusive statements can be made about Waitaki MCH-CAL™ (organic component).