Discussion of study 1 part 4: endothelial dysfunction markers in relation to G-

3.3.4.1 Discussion of Thrombomodulin levels in relation to G-gap results

When examining the results, there was no significant difference in plasma

thrombomodulin levels between the two groups (negative or positive G-gap) with a p value >0.05(figure 3.2.1b, table 3.2.1d), thus whether the value of G-gaps is positive or negative is unlikely to alter the levels of plasma thrombomodulin. Considering thrombomodulin is an important marker for endothelial dysfunction (Takahashi et al., 1992) and the results of the assay suggest that there is no difference in endothelial

172

dysfunction occurrence with G-gap, this means G-gap may neither effect nor predict the occurrence of endothelial dysfunction.

As discussed in the introduction chapter, the rate of glycation is connected to the production of AGEs and ROS, both of which contribute to damage. Therefore, a positive G-gap means a higher level of intracellular glycated haemoglobin than the predicted level from fructosamine, which can be translated as greater chance of having endothelial dysfunction. Our findings are contrary to our prediction.

As thrombomodulin is only one marker of endothelial dysfunction, therefore it was recommended to measure more makers of endothelial damage which might show a different conclusion. In other words, it might show the expected relationship between incidence of endothelial dysfunction and G-gap values. Therefore we decided to measure other markers associated with endothelial damage such as E- selectin and PAI-1, although the result of this assay may have some power in the prediction of many cardiovascular events that are associated with diabetes, of which endothelial dysfunction is a precursor.

Identification of endothelial damage as an early precursor of several diabetic complications can help identify the predictive importance of G-gap on endothelial dysfunction and can also have important benefits for prediction of CVD

complications.

3.3.4.2 Discussion of E-selectin in relation to G-gap results

In the E-selectin immunoassay there was no significant difference in plasma E- Selectin levels between the two groups (negative or positive G-gap) with a p value >0.05 (figure 3.2.2b). This means G-gaps status is unlikely to affect the

173

As described in the introduction chapter, E-selectin is an important marker of endothelial dysfunction alongside thrombomodulin (Muniyappa & Sowers, 2013). Our findings showed a non-significant difference in E-selectin levels between the two glycation gap groups which means there will be no difference in the presence of endothelial dysfunction between those two groups. As we predicted that a positive G-gap means a higher level of intracellular glycated haemoglobin than the predicted level from fructosamine, which in turn can be translated as greater chance of having endothelial dysfunction, our findings are therefore contrary to our prediction.

According to Endemann & Schiffrin (2004), E-selectin is a well know marker of endothelial dysfunction and an increase in its concentration is well connected to the presence of endothelial dysfunction, E-selectin is expressed on the surface of the cell for cell adhesion, a small proportion of it also found in the circulation which can be measured via immunoassays and this make it an important candidate of

endothelial damage.

Despite our results showing no difference in Plasma E-selectin and thrombomodulin levels between the two glycation groups, many studies have reported contradictory results. For example, glycation gap status has been used in patients with diabetes for disease monitoring and prediction of diabetic

complications, i.e. When the value is positive it is associated with certain diabetes- related complications, positive G-gap associated-retinopathy, nephropathy and some CVD diseases (Hempe et al., 2002, McCarter et al., 2000 and Cohen et al., 2003).

In general, several studies have reported that the G-gap value is correlated with various diabetic complications. As an example, Cohen et al., (2008) studied a cohort for 4 years with random measurements of G-gaps, blood glucose levels and glycated haemoglobin and found that an increased risk of diabetic nephropathy was

174

associated with positive G-gap but not with negative G-gap. In regard to other diabetes-related complications such as nephropathy, both Cohen et al., (2003) and Rodriguez-Segade, et al., (2011) found that the risk of nephropathy is higher with positive G-gap rather negative G-gap (positive G-gap was defined as >1% whereas the negative was defined <-1%): both studies show that G-gap is a predictor of nephropathy in NIDDM patients, taking into consideration that nephropathy is an important link to microvascular diseases.

Despite the previous studies found that positive G-gap was associated with diabetic complication such as nephropathy and neuropathy, Nayak et al., (2013) found that the risk of mortality is increased in both positive and negative G-gaps, indicating a lack of correlation between mortality rate and the variation of G-gap values. This study describes a U shape quadratic relationship, proposing that neither positive nor negative G-gap status is linked with improving mortality rate. This result agrees with our findings that there is no link between difference in G-gap with

plasma levels of thrombomodulin and E-selectin.

As both thrombomodulin and E-selectin are well known established markers of endothelial damage with previous studies showed that individuals exhibiting positive G-gaps have high risk of diabetes related complication and considering that ED is a well-known precursor of these complications, it could be assumed that those two markers of endothelial dysfunction are increased with positive G-gap (Cohen, et

al. 2003, Nayak, et al., 2013). However, our findings did not support this assumption.

This finding suggests that G-gap is not associated with the early phases of

developing CVD complications. Considering larger cohort might demonstrate a link between ED and G-gap variation, or these two markers alone are insufficient to support this assumption.

175