Proton NMR based serum metabolic profile correlates with the neurological recovery in treated acute spinal cord injury (ASCI) subjects: A pilot study

Highlights

• First study on serum metabolic profiling for differention using two treatment modalities in acute spinal cord injury (ASCI).

• ¹H NMR spectroscopy of serum samples correlated significantly with neurological status and recovery.

• The study provided a platform for biomarkers exploration in prognosis of ASCI subjects.

• This method is rapid and thus enables quick and effective patient management.

Abstract

Background

Acute Spinal Cord Injury (ASCI) is still having substantial morbidity and mortality despite of advanced therapeutics. Major obstacles are paucity of monitoring tools or biomarkers for severity determination, recovery and prognostication. A prospective case control pilot study with serum ¹H NMR spectroscopic metabolic profiling was carried out to evaluate metabolites perturbations and its relationship with recovery and to see role of stem cells in facilitating neurological recovery.

Methodology

Twenty subjects with ASCI were classified on the basis of therapeutic modality into surgical fixation alone (Group-1, n = 10), stem cell adjuvant (Group-2, n = 10) and healthy controls (Group-0, n = 10). Serum samples were collected at admission (baseline) and after six months (follow-up). NMR data of serum sample were quantified and subjected to Wilcoxon and ANOVA tests. Further validation was performed using supervised OSC-PCA and OPLS-DA by incorporating substantial control samples.

Result

Twenty-eight metabolites were identified; well resolved resonances of fifteen metabolites were quantified wherein seven were statistically significant. Predominantly amino acids and ketone bodies played vital role in the differentiation of groups.

Conclusions

Serum NMR spectroscopy reveals certain metabolites perturbations having clear correlation with pattern of recovery in treated ASCI subject. Stem cells treatment group had comparatively effective recovery.

Introduction

Spinal cord injury (SCI) is a devastating, severely debilitating traumatic disorder leading to complete or partial disability, affecting the physical and psycho-social wellbeing of the patient. According to WHO, the world wide incidence of SCI is estimated to be between 40 and 80 per million populations per year [[1], [2], [3], [4], [5], [6]]. Despite of best efforts, little has been achieved in terms of neurological recovery. Major successes have been achieved to rehabilitate the subjects to the wheel chair status, by not only modifications in the wheelchair dimensions but also by making the home and society paraplegic friendly. A surgical procedure to decompress the spinal cord and stabilize the fracture segment plays an important role in preventing any further damage. Recent advances in terms of functional electrical stimulation [7], retraining neural circuits to restore body functions and use of adaptive devices for communication are being studied. Ongoing spinal research is vast, in depth, rapidly expanding field involving multidisciplinary studies and working on every dimension possible. Spectacular key concepts are considered for future directions. Neuroprotection by drugs like steroid, antibiotics, erythropoietin and therapeutic hypothermia; axonal regeneration by anti-inflammatory drugs like Ibuprofen, Indomethacin and rolipram and monoclonal antibodies, anti-MAG (myelin associated glycoproteins) is also under study [8,9] Furthermore, experiments with variety of stem cell such as human oligodendrocyte progenitor cells, schwann cells, bone marrow stromal cells and nasal olfactory ensheathing cells etc. are being under extensive study [10].

In any biological organism there are various organ systems, the functions of which are based on a large number of physiochemical and biochemical reactions [11,12]. These reactions require participation of various small molecular species and a comprehensive study of these molecules is known as metabolomics. A metabolome is defined as a group or collection of low molecular weight compounds with molecular weight <1 kDa (Kilodalton). The diseases or traumatic events are defined by certain set of patho-physiological changes culminating into a milieu of various intermediate reactionary components or metabolites [13,14]. Earlier plasma ¹H-nuclear magnetic resonance (NMR) metabolomics study on rats with SCI has identified certain metabolites belonging to fatty acids and amino acids groups, which clearly differentiated between rats with severe SCI from those of healthy controls [15,16]. This study could be extrapolated to human subjects also. Determining the severity of injury by functional tests and clinical examination alone may not immediately give an accurate assessment of ongoing processes, and especially with respect to predicting future improvement or otherwise [17,18].

In ASCI primary injury occurs at the time of initial mechanical impact causing severe trauma to the cord in the form of contusion, laceration, abrasion, compression, stretch and haemorrhage. It is irreversible and incurable [19]. Secondary injury is a series of processes which occurs in tissues subsequently after primary injury, leading to further progression of inflammation, edema, ischemia, and micro vascular haemorrhage, along with onset of specific pathogenic processes like excitotoxicity due to presence of excess glutamate. Glutamate causes increase in the number of free radicals, ionic dysregulation and immune mediated damages. This phase aggravates the vicious cycle of cell injury and necrosis [20]. It is important to understand the pathophysiology of secondary injury, because it creates a window of therapeutic opportunities for planning interventions. Some studies suggest that surgery (internal fixation), over conservative treatment has a definite role in providing stability and minimizing neurological problems in acute spinal cord injuries.

Stem cells play a versatile role in an organism by maintaining the cellular stability, and indirectly maintaining different biochemical requirements of the body. Several studies have proved that stem cells play different roles including metabolite maintenance during an injury [21,22]. Among all the known sources of stem cells we have chosen bone marrow (BM) cells because of their ease of availability for autologous infusion and do not involve any ethical issues. The BM stem cell transplantation was safe for subjects in short and long-term assessments [[23], [24], [25]].

Curative management of SCI is still a distant dream but innovations of technologies and scientific devices have enabled patients to lead an active, productive and almost near normal life. In quest of new therapeutic modalities and its effect for prognostication and assessment of recovery status by NMR spectroscopy, therefore this pilot study was designed. This is a prospective case-control study where serum samples were analyzed for metabolic alterations and compared with those of healthy controls, to evaluate the status of recovery of subjects suffering from acute spinal cord injury. In addition, the use of stem cells as an adjuvant to conventional treatment has been performed to determine their role in neurological recovery based on metabolomic. The objective of this study was to correlate ¹H NMR based serum metabolic profile with the neurological recovery in acute spinal cord injury (ASCI) subjects.