Smaller hippocampal volume in APOE ε4 carriers independent of amyloid-ß (Aß) burden.

OBJECTIVE: To investigate the association of the APOE ε4 genotype with hippocampal volume, independent of Aß burden. METHOD: This cross-sectional study included 71 participants with mild cognitive impairment or mild AD. All participants were divided into carriers or non-carriers of the ε4 allele. The main outcome was hippocampal volume measured using structural magnetic resonance imaging; 18F-florbetaben positron emission tomography was additionally performed to investigate the association of APOE ε4 genotype with hippocampal volumes, independently of Aß burden. Analysis of covariance was conducted to compare the differences in hippocampal volumes between carriers and non-carriers of the ε4 allele after controlling for global Aß burden or local hippocampal Aß burden. RESULTS: The APOE ε4 genotype was associated with a smaller right and total hippocampal volume (right: 3160.16 ± 365.71 vs. 3365.24 ± 434.88, p < 0.05; total: 6257.48 ± 790.60 vs. 6599.52 ± 840.58, p < 0.05), independent of Aß burden. CONCLUSION: Our findings on the association of APOEε4 genotype with hippocampal volume independent of Aß burden suggest that the APOEε4 genotype may contribute to hippocampal neurodegeneration through an Aß-independent mechanism.

Development of a Phosphor Screen with Mirror Coating to Improve Optical Properties in Radiography.

Phosphor screens have attracted increasing global interest because they can aid the acquisition of high-quality images while simultaneously reducing exposure. However, although increasing the thickness of the phosphor screen increases exposure efficiency due to scattered light, it also leads to a broader light spread, which results in poorer resolution. Hence, in this study, we implemented a reflector using a mirror-coating technique on the surface of a phosphor screen and analyzed its characteristics in terms of luminescence intensity and resolution. We present the fabrication and measurement of the phosphor screen based on a reflector containing Gd2O2S:Tb. The phosphor screen containing the reflector can be fabricated via the screen-printing method and roll-to-roll sputtering method. In particular, when compared to the condition without a reflector, Al and Cu reflectors showed improvements in luminescence intensity of 15.7% and 12.2%, respectively, as well as lower full widths at half maximum of 11.45 and 9.08, respectively. This quantitatively demonstrates that Cu and Al are suitable reflector materials to improve exposure efficiency while maintaining resolution in radiography systems. Moreover, because we did not utilize an optical assembly, which improves the transmission efficiency by matching the different refractive indices of the phosphor screen and photo-detector, we believe that the quantity of photons could be further improved if the reflectors were applied to a commercial product. Thus, future studies using single-layer anti-reflector coating techniques with optical assemblies are warranted.

Comparison Analysis between the Medication Efficacy of the Milnacipran and Functional Connectivity of Neural Networks in Fibromyalgia Patients.

Milnacipran is a reuptake inhibitor of both serotonin and noradrenaline, used in the treatment of fibromyalgia with severe depression. However, few studies have been conducted on the efficacies of milnacipran drug on the functional connectivity of the neural network. The authors aimed to find the correlation between the drug efficacy and the changes in neural network in fibromyalgia patients. Resting-state-functional magnetic resonance imaging (rs-fMRI) were obtained before and after milnacipran drug administration. Graph theory indexes and small-worldness were calculated using preprocessed blood-oxygen-level-dependent signals from the rs-fMRI scans of 14 brain regions-of-interest. Statistical analyses were conducted to compare the topological network parameters. Significant changes in the neural network indexes appeared in three of the 14 brain regions-of-interest. In the pain network, the average path length on the left side of Brodmann area 32 was shortened. In the default mode network, functional connectivity changes were observed in the left lateral parietal cortex and medial prefrontal cortex. In the left lateral parietal cortex, the degree and betweenness centrality increased, whereas the clustering coefficient decreased. In the medial prefrontal cortex, local efficiency decreased. The small-worldness declined after milnacipran medication. The present results demonstrate that functional connectivity indexes in the brains of female fibromyalgia patients obtained from rs-fMRI data can be used as potential prognosis markers of milnacipran drug treatment.

Quantitative Validation of a Visual Rating Scale for Defining High-Iron Putamen in Patients With Multiple System Atrophy.

Objectives: To validate a visual rating scale reflecting sub-regional patterns of putaminal hypointensity in susceptibility-weighted imaging of patients with multiple system atrophy (MSA). Methods: Using a visual rating scale (from G0 to G3), 2 examiners independently rated putaminal hypointensities of 37 MSA patients and 21 control subjects. To investigate the correlation with the scales, R2* values and the volume of the entire putamen were measured. Results: MSA patients with parkinsonian variant had significantly higher scores than those with cerebellar variant. Visual rating scores in MSA were correlated with R2* values [General estimating equation (GEE), Wald chi-square = 25.89, corrected p < 0.001] and volume (Wald chi-square = 75.44, corrected p < 0.001). They correlated with UPDRS motor scores. Binary logistic regression analyses revealed that the visual rating scale was a significant predictor for discriminating MSA patients from controls [multivariate model adjusted for age and sex, odds ratio 52.722 (corrected p = 0.009)]. Pairwise comparison between areas under the curve (AUCs) revealed that the visual rating scale demonstrated higher accuracy than R2* values [difference between AUCs; univariate model = 0.247 (corrected p < 0.001); multivariate model = 0.186 (corrected p = 0.003)]. There were no significant differences in clinical characteristics between the high-iron group, defined as putamen with visual rating scale ≥ G2 and R2* values ≥ third quartile, and the remaining patients. Conclusion: The visual rating scale, which reflects quantitative iron content and atrophy of the putamen as well as motor severities, could be useful for the discrimination and evaluation of patients with MSA.

Speculating the timing of iron deposition in the putamen in multiple system atrophy.

BACKGROUND & OBJECTIVE: Although iron accumulation is thought to be associated with neurodegenerative processes, the timing of putaminal iron deposition during the disease course of multiple system atrophy (MSA) remains unclear. We sought to investigate the temporal pattern of iron deposition in the putamen of MSA patients by calculating the conditional probabilities (CPs) of multimodal MRI changes. METHODS: We simultaneously measured putaminal R2*, volume and MD values of 39 probable MSA patients and 22 control subjects. The presence of significant MRI changes was defined as higher R2* and MD values, or lower volumes than cut-off values derived from mean values in control putamen. The CPs of R2* changes without MD or volume changes were then compared with those of MD or volume changes without R2* changes. RESULTS: Regardless of the cut-off values, the CPs of R2* increments without MD or volume changes were significantly lower than those for MD or volume changes without R2* increments. The associations of R2* with volume and MD values appeared in non-linear exponential and quadratic patterns, respectively. CONCLUSIONS: Our findings suggest that putaminal iron accumulation would occur under volume atrophy or MD increments. Thus, iron deposition in the putamen of MSA patients is likely a secondary byproduct of neurodegeneration.

Feasibility Study of Phosphor Screen Containing Nano-Scale Anti-Reflection Layer for Improved Optical Properties in Indirect X-ray Detector.

With increasingly strict regulations regarding patient exposure, research on digital radiography technology has recently focused on indirect methods that can produce high-quality images for a low radiation dose. In particular, medical imaging systems based on indirect methods universally use rare-earth metal phosphors, because of their high atomic number and excellent luminescence efficiency. Thus, various studies aiming to improve the luminescence efficiency of phosphors have been conducted. Despite this research, however, the current luminescence efficiencies are insufficient. Here, we report a basic study aiming to develop a phosphor screen containing a three-quarter-wave optical-thickness layer to improve the light transmission efficiency. Specifically, the fabrication and measurement of a Gd2O2S:Tb phosphor screen containing a single three-quarter-wave optical-thickness layer is presented. The screen is fabricated via a screen-printing and spin-coating method. Based on histograms of the degree of luminescence and the pixel values, we demonstrate that the light transmission efficiency is improved by the three-quarter-wave optical-thickness layer. Note that analysis of the full width at half maximum of the pixel value distribution reveals the possibility of resolution loss when obtaining medical images. Overall, the results of this study confirm that the light transmission efficiency can be improved through use of a single-layer anti-reflection coating. However, because the emission spectrum of the Gd2O2S:Tb screen is in the 480-600-nm band, it is necessary to expand the areas exhibiting the lowest reflectance to the wavelengths at the edge of this band. Thus, further study should be conducted to optimize the optical thickness.

Spatial correlation and segregation of multimodal MRI abnormalities in multiple system atrophy.

OBJECTIVE: The variability of the severity and regional distribution of pathological process in basal ganglia (BG) and brainstem-cerebellar systems results in clinical heterogeneity and represents the motor subtype of multiple system atrophy (MSA). This study aimed to quantify spatial patterns of multimodal MRI abnormalities in BG and stem-CB regions and define structural MRI findings that correlate with clinical characteristics. METHODS: We simultaneously measured R2*, mean diffusivity (MD), and volume in the subcortical structures (BG, thalamus, brainstem-cerebellar regions) of 39 probable MSA and 22 control subjects. Principal component analysis (PCA) and structural equation modeling (SEM) were performed to show a model consisting of multiple inter-dependencies. RESULTS: Structural MRI alterations were found to be significantly interrelated within BG as well as brainstem-cerebellar regions in MSA patients. PCA extracted four factors: three factors reflected alterations in R2*, MD and volume of the BG region including the caudate nucleus, putamen, and pallidum, and the remaining one factor represented degenerative changes in MD and volume of stem-CB region. In SEM, a latent variable reflecting brainstem-cerebellar degeneration did not show a significant correlation with the other latent variables associated with BG degeneration. Putaminal MD values and a PCA-driven factor reflecting MD values in the BG showed a significant correlation with UPDRS and UMSARS scores. CONCLUSION: Multimodal structural MRI abnormalities in MSA appear to be segregated into BG and stem-CB-related factors that can be associated with the clinical phenotype and motor severity.

Decreased gray matter volume is associated with the subtypes of psychotic symptoms in patients with antipsychotic-naïve mild or moderate Alzheimer's disease: A voxel-based morphometry study.

The purpose of this study was to investigate the association between brain regional gray matter volume and two subtypes of psychotic symptoms, namely paranoid and misidentification subtypes, in antipsychotic-naïve mild or moderate Alzheimer's disease (AD) patients. Forty AD patients with psychotic symptoms and 25 AD patients without psychotic symptoms were assessed for cognitive and functional impairment. Presence and subtype of psychotic symptoms were assessed by using the delusion and hallucination subscale of the Korean Neuropsychiatric Inventory (K-NPI). Structural MRI images were acquired on a 3 T scanner, and were analyzed using voxel-based morphometry (VBM) for automated analysis. The misidentification subtype is associated with more severe gray matter atrophy, and paranoid subtype is associated with less severe gray matter atrophy compared to non-psychosis group. These results suggest that the misidentification, the paranoid subtype and the non-psychosis group have a distinct neural correlation.

Gray matter volumes and treatment response of psychotic symptoms to risperidone in antipsychotic-naïve Alzheimer's disease patients.

OBJECTIVE: The purpose of this study was to determine whether gray matter volumes are associated with treatment response of psychotic symptoms in Alzheimer's disease (AD) patients. METHOD: Risperidone, which is commonly used as an atypical antipsychotic drug, was administered in a clinical setting for 6 weeks from April 2012 to February 2013 to 25 antipsychotic-naïve AD patients with psychosis, diagnosed according to Jeste and Finkel's proposed diagnostic criteria for psychosis of Alzheimer's disease. Psychotic symptoms were rated with the Korean version of the Neuropsychiatric Inventory (K-NPI) at baseline and at 6 weeks, and treatment response was defined as the change in K-NPI score from baseline to 6 weeks. Gray matter volumes were measured with magnetic resonance imaging and voxel-based morphometry at baseline. Age, gender, years of education, total intracranial volume, apolipoprotein E genotype, dosage of risperidone, the baseline scores on the Korean version of the Mini-Mental State Examination, and the baseline psychotic and nonpsychotic symptoms scores on the K-NPI were measured as covariates of no interest. RESULTS: We found that treatment response of psychotic symptoms to risperidone in antipsychotic-naïve AD patients was positively associated with both left and right putamina, left parahippocampal gyrus, and left amygdala volume after controlling covariates of no interest (uncorrected P < .001, KE > 100 voxels). CONCLUSIONS: Therefore, we conclude that gray matter volumes of putamina, left parahippocampal gyrus, and left amygdala are associated with treatment response of psychotic symptoms after 6 weeks of treatment with risperidone in antipsychotic-naïve AD patients with psychosis. These results suggest that the volumes of specific gray matter regions probably contribute to treatment response of psychotic symptoms in AD patients. TRIAL REGISTRATION: ClinicalTrials.gov identifier: NCT01198093.

Noninvasive and repetitive measurement of cellular metabolite from human osteosarcoma cells (MG-63) using 3.0 tesla proton (1 H) MR spectroscopy.

PURPOSE: This study suggests a noninvasive and repetitive measurement method using 1 H magnetic resonance spectroscopy to monitor changes in cellular metabolites within a single sample. METHODS: Longitudinal acquisition of cellular metabolites from three-dimensional cultured human osteosarcoma (MG-63) cells was conducted using 3.0 Tesla 1 H MRS for 2 weeks at three time points: days 1, 7, and 14. During the MR spectroscopy (MRS) scan, cell specimen temperatures were kept constant at 37°C by a lab-developed magnetic resonance compatible thermostatic device. A DNA assay and live/dead staining of the cell specimens were carried out at each time point to verify the MRS measurements. RESULTS: Cell viability in the proposed device did not significantly differ from that of cells in a conventional incubator (P = 0.946). Cell proliferation and choline concentration increased during the first week, but remained constant during the second week. Lactate did not change during the first week, but increased during the second week. Likewise, cell viability remained constant until day 7, then decreased. CONCLUSION: The proposed MRS technique results in a survivable environment for longitudinal studies of cells and provides a new way to measure metabolomic changes over time in single specimens of cells. Magn Reson Med 76:1912-1918, 2016. © 2016 International Society for Magnetic Resonance in Medicine.

Impact of Apolipoprotein E4 Polymorphism on the Gray Matter Volume and the White Matter Integrity in Subjective Memory Impairment without White Matter Hyperintensities: Voxel-Based Morphometry and Tract-Based Spatial Statistics Study under 3-Tesla MRI.

OBJECTIVE: The aim of this study is to compare gray matter (GM) volume and white matter (WM) integrity in Apolipoprotein E4 (ApoE ε4) carriers with that of ApoE ε4 noncarriers using the voxel-based morphometry and diffusion tensor imaging (DTI) to investigate the effect of the ApoE ε4 on brain structures in subjective memory impairment (SMI) without white matter hyperintensities (WMH). METHODS: Altogether, 26 participants with SMI without WMH were finally recruited from the Memory impairment clinics of Pusan National University Hospital in Korea. All participants were ApoE genotyped (ApoE ε4 carriers: n = 13, matched ApoE ε4 noncarriers: n = 13) and underwent 3-tesla magnetic resonance imaging (MRI) including 3-dimensional volumetric images for GM volume and DTI for WM integrity. RESULTS: ApoE ε4 carriers compared with noncarriers in SMI without WMH showed the atrophy of GM in inferior temporal gyrus, inferior parietal lobule, anterior cingulum, middle frontal gyrus, and precentral gyrus and significantly lower fractional anisotropy WM values in the splenium of corpus callosum and anterior corona radiate. CONCLUSION: Our findings suggest that the ApoE ε4 is associated with both atrophy of GM volume and disruption of WM integrity in SMI without WMH.

Apolipoprotein E genotype modulates effects of vitamin B12 and homocysteine on grey matter volume in Alzheimer's disease.

AIMS: The aim of the present study was to investigate whether the effects of vitamin B12 and homocysteine on brain volume are influenced by apolipoprotein E (APOE) genotype. We examined the effects in each subgroup (APOE ε4 carriers and non-carriers) of Alzheimer's disease (AD) patients and healthy normal controls. METHODS: Forty participants with AD and 20 healthy normal controls were recruited from memory impairment clinics at Pusan National University Hospital in Korea. All participants were APOE genotyped and underwent magnetic resonance imaging, including 3-D volumetric images for grey matter (GM) volume. A multiple regression model integrated into statistical parametric mapping was used to see if there was any correlation between vitamin B12 or homocysteine and GM volume in each subgroup (APOE ε4 carriers and non-carriers) of AD patients and healthy normal controls. RESULTS: There was a significant positive correlation between serum concentrations of vitamin B12 and regional GM volume in APOE ε4 carriers with AD but not in non-carriers. We also found that there was a significant negative correlation between serum concentrations of homocysteine and regional GM volume in APOE ε4 non-carriers with AD but not in carriers (P < 0.001, uncorrected for multiple comparisons; extent threshold = 100 voxel). CONCLUSION: The present findings suggest that the effects of vitamin B12 and homocysteine on GM volume might be influenced by APOE genotype.

Distinguishing tendon and ligament fibroblasts based on 1H nuclear magnetic resonance spectroscopy.

Tendon and ligament (T/L) have been known to be obviously different from each other in tissue level. However, due to the overlapping gene markers, distinction in cellular level has not been clearly verified yet. Recently, the use of nuclear magnetic resonance (NMR) spectroscopy has shown the potential to detect biological markers in cellular level. Therefore, in this study we applied a non-invasive technique based on NMR spectroscopy to establish biomarkers to distinguish between T/L fibroblasts. In addition the cellular morphologies and gene expression patterns were also investigated for comparison through optical microscopy and real-time polymerase chain reaction (PCR). No difference was observed from morphology and real-time PCR results, either as expected. However, we found clear differences in their metabolomic spectra using 1H NMR spectroscopy. The calculated integral values of fatty acids (with chemical shifts at ~0.9, 1.26, 1.59, 2.05, 2.25, and 2.81 ppm), lactate (~1.33 ppm), and leucine (~2.72 ppm) were significantly different between the two types of fibroblasts. To be specific tendon group exhibited higher level of the metabolite than ligament group. In conclusion, in-cell metabolomic evaluation by NMR technique used in this study is believed to provide a promising tool in distinguishing cell types, especially T/L cells, which cannot be classified by conventional biological assays.

Cytotoxicity of TSP in 3D Agarose Gel Cultured Cell.

PURPOSE: A reference reagent, 3-(trimethylsilyl) propionic-2, 2, 3, 3-d4 acid sodium (TSP), has been used frequently in nuclear magnetic resonance (NMR) and magnetic resonance spectroscopy (MRS) as an internal reference to identify cell and tissue metabolites, and determine chemical and protein structures. This reference material has been exploited for the quantitative and dynamic analyses of metabolite spectra acquired from cells. The aim of this study was to evaluate the cytotoxicity of TSP on three-dimensionally, agarose gel, cultured cells. MATERIALS AND METHODS: A human osteosarcoma cell line (MG-63) was selected, and cells were three dimensionally cultured for two weeks in an agarose gel. The culture system contained a mixture of conventional culture medium and various concentrations (0, 1, 3, 5, 7, 10, 20 30 mM) of TSP. A DNA quantification assay was conducted to assess cell proliferation using Quant-iT PicoGreen dsDNA reagent and kit, and cell viability was determined using a LIVE/DEAD Viability/Cytotoxicity kit. Both examinations were performed simultaneously at 1, 3, 7 and 14 days from cell seeding. RESULTS: In this study, the cytotoxicity of TSP in the 3D culture of MG-63 cells was evaluated by quantifying DNA (cell proliferation) and cell viability. High concentrations of TSP (from 10 to 30 mM) reduced both cell proliferation and viability (to 30% of the control after one week of exposure), but no such effects were found using low concentrations of TSP (0-10 mM). CONCLUSIONS: This study shows that low concentrations of TSP in 3D cell culture medium can be used for quantitative NMR or MRS examinations for up to two weeks post exposure.

Progression of subcortical atrophy and iron deposition in multiple system atrophy: a comparison between clinical subtypes.

Magnetic resonance imaging (MRI) can be useful not only for the diagnosis of multiple system atrophy (MSA) itself, but also to distinguish between different clinical subtypes. This study aimed to investigate whether there are differences in the progression of subcortical atrophy and iron deposition between two variants of MSA. Two serial MRIs at baseline and follow-up were analyzed in eight patients with the parkinsonian variant MSA (MSA-P), nine patients with cerebellar variant MSA (MSA-C), and fifteen patients with Parkinson's disease (PD). The R2* values and volumes were calculated for the selected subcortical structures (caudate nucleus, putamen, globus pallidus, and thalamus) using an automated region-based analysis. In both volume and R2*, a higher rate of progression was identified in MSA-P patients. Volumetric analysis showed significantly more rapid progression of putamen and caudate nucleus in MSA-P than in MSA-C. With regard to R2* changes, a significant increase at follow-up and a higher rate of progression were identified in the putamen of MSA-P group compared to MSA-C and PD groups. This longitudinal study revealed different progression rates of MRI markers between MSA-P and MSA-C. Iron-related degeneration in the putamen may be more specific for MSA-P.

Automated Classification to Predict the Progression of Alzheimer's Disease Using Whole-Brain Volumetry and DTI.

OBJECTIVE: This study proposes an automated diagnostic method to classify patients with Alzheimer's disease (AD) of degenerative etiology using magnetic resonance imaging (MRI) markers. METHODS: Twenty-seven patients with subjective memory impairment (SMI), 18 patients with mild cognitive impairment (MCI), and 27 patients with AD participated. MRI protocols included three dimensional brain structural imaging and diffusion tensor imaging to assess the cortical thickness, subcortical volume and white matter integrity. Recursive feature elimination based on support vector machine (SVM) was conducted to determine the most relevant features for classifying abnormal regions and imaging parameters, and then a factor analysis for the top-ranked factors was performed. Subjects were classified using nonlinear SVM. RESULTS: Medial temporal regions in AD patients were dominantly detected with cortical thinning and volume atrophy compared with SMI and MCI patients. Damage to white matter integrity was also accredited with decreased fractional anisotropy and increased mean diffusivity (MD) across the three groups. The microscopic damage in the subcortical gray matter was reflected in increased MD. Classification accuracy between pairs of groups (SMI vs. MCI, MCI vs. AD, SMI vs. AD) and among all three groups were 84.4% (±13.8), 86.9% (±10.5), 96.3% (±4.6), and 70.5% (±11.5), respectively. CONCLUSION: This proposed method may be a potential tool to diagnose AD pathology with the current clinical criteria.

Cortical atrophy, reduced integrity of white matter and cognitive impairment in subcortical vascular dementia of Binswanger type.

AIMS: An association between white matter hyperintensities (WMH) and cognitive dysfunction has long been recognized. However, subjects with identically appearing WMH on magnetic resonance imaging present with a wide variance in cognitive function ranging from normal cognition to dementia. The aim of this study was to compare cortical atrophy and integrity of white matter of patients with subcortical vascular dementia of Binswanger type (SVaD-BT) with those of the normal cognition group with WMH (ncWMH). METHODS: Eleven patients with SVaD-BT and 11 age-, sex-, education- and grade of WMH-matched ncWMH underwent magnetic resonance imaging, including 3-D volumetric images for cortical atrophy and diffusion tensor imaging for integrity of white matter. RESULTS: Compared to ncWMH, SVaD-BT patients showed cortical atrophies in frontal (i.e. frontal pole, precentral gyrus and frontal medial cortex) and occipital areas (i.e. lingual gyrus) followed by atrophies in temporal (i.e. fusiform cortex and middle temporal gyrus) areas. Along with cortical atrophies, reduced integrity with low fractional anisotropy and high mean diffusivity values in genu and splenium of the corpus callosum were detected in SVaD-BT patients. CONCLUSIONS: Our findings suggest that cognitive decline from ncWMH to SVaD-BT may be associated with cortical atrophy and reduced integrity of white matter.

Evaluation of brain iron content in idiopathic REM sleep behavior disorder using quantitative magnetic resonance imaging.

BACKGROUND: Neuroimaging studies in patients with idiopathic rapid eye movement sleep behavior disorder (iRBD) show similar structural and functional changes to alpha-synucleinopathies, including PD. Until now, there have been few attempts to characterize brain iron deposition in iRBD. The aim of this study was to investigate brain iron content in patients with iRBD using quantitative magnetic resonance imaging (MRI). METHODS: 3-T MRI was performed in 15 patients with iRBD and 20 age-matched healthy control subjects. In order to evaluate the iron-related neurodegenerative changes, we assessed volume and transverse relaxation rate (R2*) simultaneously. We used both region-based and voxel-based analysis. RESULTS: No significant differences in R2* values were found between iRBD groups and healthy control subjects. There were no areas of significantly reduced or increased gray matter and white matter volume in the iRBD group. Instead, lateral ventricle volumes measured automatically by FreeSurfer were significantly larger in patients with iRBD than in healthy controls (P < 0.05). CONCLUSION: The present study suggests that iron-related R2* values may not be an imaging biomarker for neurodegeneration in iRBD.

Evaluation of metabolomic changes as a biomarker of chondrogenic differentiation in 3D-cultured human mesenchymal stem cells using proton (1H) nuclear magnetic resonance spectroscopy.

PURPOSE: The purpose of this study was to evaluate the metabolomic changes in 3D-cultured human mesenchymal stem cells (hMSCs) in alginate beads, so as to identify biomarkers during chondrogenesis using (1)H nuclear magnetic resonance (NMR) spectroscopy. MATERIALS AND METHODS: hMSCs (2×10(6) cells/mL) were seeded into alginate beads, and chondrogenesis was allowed to progress for 15 days. NMR spectra of the chondrogenic hMSCs were obtained at 4, 7, 11, and 15 days using a 14.1-T (600-MHz) NMR with the water suppression sequence, zgpr. Real-Time polymerase chain reaction (PCR) was performed to confirm that that the hMSCs differentiated into chondrocytes and to analyze the metabolomic changes indicated by the NMR spectra. RESULTS: During chondrogenesis, changes were detected in several metabolomes as hMSC chondrogenesis biomarkers, e.g., fatty acids, alanine, glutamate, and phosphocholine. The metabolomic changes were compared with the Real-Time PCR results, and significant differences were determined using statistical analysis. We found that changes in metabolomes were closely related to biological reactions that occurred during the chondrogenesis of hMSCs. CONCLUSIONS: In this study, we confirm that metabolomic changes detected by (1)H-NMR spectroscopy during chondrogenic differentiation of 3D-cultured hMSCs in alginate beads can be considered as biomarkers of stem cell differentiation.