Although many therapeutic options offer some short and long-term immune-modulating effects, these drugs do not exert clinical sustainable neuroprotective effects and thereby fail to improve the long-term disease course in MS [21]. Additionally, no pharmacological agent still exerts beneficial effects in MS-related cognitive impairment. Here, the true challenge with translational MS studies is in connecting the preclinical results to human biological processes, which are complex and can involve combinations of many key pathophysiological components, acting simultaneously and feeding back on one another [40, 41]. Consequently, the complexity of the resulting pathophysiology is such that therapeutic interventions that act on singular pathways may not provide adequate benefit. This led to hypothesize that alternative drugs, including nutritional supplements targeting multiple pathways, may be needed to improve the disease course. Our data show for the first time that magnesium and high-dose biotin was the most effective combination in enhancing cognitive scores and restoring myelination (OPG l) and inflammatory parameters (ILs, TNF-α, chemokine, NF-κB, and GFAP) without producing any side effects. Indeed, improved cognitive scores in the magnesium and high-dose biotin group were correlated with more significantly increased bioenergetic mechanisms, including the upregulation of critical biotin-related enzymes, such as ACC-1, ACC-2, PC, PCC, and MCC, which are involved in cognitive processes.
According to the previous effects of BDNF on synaptogenesis, cognition, and learning, we have also analyzed the expression of BDNF, GAP43, ICAM, and synaptic proteins (i.e., PSD 93, PSD 95, and synapsin). In line with hitherto studies, we observed that BDNF levels and markers for axonal regeneration and synaptic remodeling [42–44] were dose-dependently increased after the treatment with biotin and MgBiotin We next explored whether the pro-cognitive potential of BDNF and other neuro-regeneration markers might still be adequate to alter the neurobehavioral outcomes in the whole study groups. Not surprisingly, we have observed that biotin and magnesium biotinate combination dose-dependently improved cognitive scores, which is in line with the effects of BDNF on learning, memory, and depression [43, 45]. More interestingly, we have noticed that improved neurobehavioral outcomes were also associated with improved remyelination process after biotin and magnesium application. Again, this might suggest interactive processes between re-myelination, synaptogenesis, and (region-specific) energetic requirements involved in improved neurological status. For instance, the different structural expression patterns of the biotin-related enzyme and varying energy requirements of neuronal structures might be responsible for additive beneficial effects. A good example could be acetyl-CoA carboxylase1 and acetyl-CoA carboxylase2, which are expressed in oligodendrocytes and induce myelin repair by supporting the fatty acid synthesis, whereas other biotin-related enzymes are primarily expressed in neurons and protect axons by enhancing energy production [1, 46, 47].
Although there are inconclusive results regarding the routine clinical use of biotin in MS for safety reasons, our results have revealed that serum levels of magnesium and biotin were directly correlated with the brain concentrations of magnesium biotinate. Furthermore, we have observed no significant differences in body weight and biochemical parameters after applying biotin with magnesium. Since poor oral bioavailability can not only result in low efficacy but also lead to some unpredictable side effects, our present data might indicate that the combination of Mg and Biotin might exert stronger bioavailability than other formulations without producing any side effects. From the translational point of view, when our results were compared with data from recent MS clinical trial results, where high-dose biotin did not seem to be associated with a beneficial effect, the main results presented here indicate that low and high doses of magnesium-biotin exerted a significant beneficial impact on myelination, inflammation, regeneration, and neurobehavioral scores. Considering that cognitive impairment is the main factor influencing the quality of life in MS patients [48], our present data is highly relevant for understanding the mechanism of cognitive impairment in these specific groups of patients. From another point of view, biotin-based treatment options, which led to increased availability of alternative energy sources, have been increasingly recommended as a therapeutic tool in degenerative cognitive diseases[49]. Although we have finally noticed that magnesium and high dose biotinate was the most effective combination in reducing the injury markers and functional outcomes, it was interesting to observe that magnesium significantly improved almost all parameters when combined with low and high doses of biotin. For instance, biotin-related enzymes, PSD 95, GFAP, MMP, OPG, IL 17, which did not change even when applying higher doses of biotin, responded well to both magnesium and biotin combinations (MgB1 and MgB2), suggesting that combining biotin with magnesium instead of increasing its dosages might provide more beneficial effects on the process of demyelination and neurobehavioral outcomes. This is consistent with recent prospective MS trials revealing that applying high-dose-biotin as a single agent did not lead to a clear improvement in disability scores [50].
Taken together, we show for the first time that MgB2 was the most effective formulation to improve cognitive scores-neurobehavioral scores, which is associated with improved synaptogenesis, remyelination, bioenergetic mechanisms BDNF levels, and inflammation. To elucidate the underlying neuroprotective and pro-cognitive efficacy of biotin and magnesium in detail, further studies should provide more insights on dose-response relationships of magnesium and biotin. Because biotin and magnesium exert no toxic effects when combined, our data may have a strong translational therapeutic message in cognitive impairment both in progressive and relapsing-remitting MS forms. In conclusion, increasing the bioavailability of biotin could be a promising neuroprotective and pro-cognitive treatment approach in MS, while bioenergetic enzymes could have critical importance, of which researchers should be aware.