Mesenchymal stem cell (MSC) may have the potential to improve recovery and promote brain repair after stroke. However, therapeutic effect monitoring of biochemical compounds following transplantation of human MSC (hMSC) into stroke animal model have not been reported. To examine how organic compounds change in brain after transplantation of hMSC in rat that had undergone transient middle cerebral artery occlusion (MCAo), we investigated the content of organic acids (OAs), which are important biochemical indicators in pathological conditions.
An efficient method for the simultaneous profiling analysis of organic acids (OAs) including keto acids (KAs) as their methoximation/tert.-butyldimethylsilylation (MO/TBDMS) derivatives was developed by gas chromatography-mass spectrometry (GC-MS) in selected ion monitoring (SIM) mode. In this study, the keto groups of KAs were reacted with methoxyamine hydrochloride for the simultaneous analysis of 21 OAs including 3-hydroxybutyric acid. Then the aqueous solution was acidified with concentrated sulfuric acid (pH ≤ 2), followed by solvent extraction with diethyl ether and ethyl acetate in sequence. The combined extracts were evaporated to dryness under a gentle stream of nitrogen. The remaining active hydrogen atoms were converted to TBDMS derivatives for the direct analysis by GC-SIM-MS. Under the optimal conditions, linear response in the range of 0.01-5.0 μg/mL was achieved with good correlation coefficient (r ≥ 0.995). The precision showed low relative standard deviation less than 10 %, and the accuracy (% RE) varied from -8.0 to 6.7 for 21 OAs studied.
When applied to rat brain tissues from normal control group, MCAo group and MCAo group following intravenous hMSC transplantation, the levels of some OAs of the MCAo and MCAo + hMSC groups were significantly different from those of the control group. In addition, normalized OA levels of MCAo and MCAo + hMSC groups were transformed into distorted star patterns which were different from control mean. The present method has provided insights into understanding the complexity of biochemical and physiological events that occur in ischemic brain injury and the transplantation effects of MSCs in stroke.
