Molecular networks in neurological diseases are complex. Despite this fact, contemporary biomarkers are in most cases interpreted in isolation, leading to a significant loss of information and power. We present an analytical approach to scrutinize and combine information from biomarkers originating from multiple sources with the aim of discovering a condensed set of biomarkers that in combination could distinguish the progressive degenerative phenotype of multiple sclerosis (SPMS) from the relapsing-remitting phenotype (RRMS).Methods: Clinical and magnetic resonance imaging (MRI) data were integrated with data from protein and metabolite measurements of cerebrospinal fluid, and a method was developed to sift through all the variables to establish a small set of highly informative measurements. This prospective study included 16 SPMS patients, 30 RRMS patients and 10 controls. Protein concentrations were quantitated with multiplexed fluorescent bead-based immunoassays and ELISA. The metabolome was recorded using liquid chromatography-mass spectrometry. Clinical follow-up data of the SPMS patients were used to assess disease progression and development of disability.Results: Eleven variables were in combination able to distinguish SPMS from RRMS patients with high confidence superior to any single measurement. The identified variables consisted of three MRI variables: the size of the spinal cord and the third ventricle and the total number of T1 hypointense lesions; six proteins: galectin-9, monocyte chemoattractant protein-1 (MCP-1), transforming growth factor alpha (TGF-α), tumor necrosis factor alpha (TNF-α), soluble CD40L (sCD40L) and platelet-derived growth factor AA (PDGF-AA); and two metabolites: 20β-dihydrocortisol (20β-DHF) and indolepyruvate.The proteins myelin basic protein (MBP) and macrophage-derived chemokine (MDC), as well as the metabolites 20β-DHF and 5,6-dihydroxyprostaglandin F1a (5,6-DH-PGF1), were identified as potential biomarkers of disability progression.Conclusion: Our study demonstrates, in a limited but well-defined and data-rich cohort, the importance and value of combining multiple biomarkers to aid diagnostics and track disease progression.

To better understand the pathophysiological differences between secondary progressive multiple sclerosis (SPMS) and relapsing-remitting multiple sclerosis (RRMS), and to identify potential biomarkers of disease progression, we applied high-resolution mass spectrometry (HRMS) to investigate the metabolome of cerebrospinal fluid (CSF). The biochemical differences were determined using partial least squares discriminant analysis (PLS-DA) and connected to biochemical pathways as well as associated to clinical and radiological measures. Tryptophan metabolism was significantly altered, with perturbed levels of kynurenate, 5-hydroxytryptophan, 5-hydroxyindoleacetate, and N-acetylserotonin in SPMS patients compared with RRMS and controls. SPMS patients had altered kynurenine compared with RRMS patients, and altered indole-3-acetate compared with controls. Regarding the pyrimidine metabolism, SPMS patients had altered levels of uridine and deoxyuridine compared with RRMS and controls, and altered thymine and glutamine compared with RRMS patients. Metabolites from the pyrimidine metabolism were significantly associated with disability, disease activity and brain atrophy, making them of particular interest for understanding the disease mechanisms and as markers of disease progression. Overall, these findings are of importance for the characterization of the molecular pathogenesis of SPMS and support the hypothesis that the CSF metabolome may be used to explore changes that occur in the transition between the RRMS and SPMS pathologies.

The effect of a "normal" (n = 8) and "high" (n = 6) protein intake (1 and 2.5 g x kg(-1) x day(-1), respectively) and of exercise on plasma amino acid (AA) concentrations, insulin, and glucagon concentrations was followed throughout a continuous 24-h period in adult male subjects at energy balance after six days on a standardized diet and exercise program. Subjects were fasting from 2100 on day 6 to 1200 on day 7 and then fed 10 identical meals hourly until 2100. Physical exercise was performed (46% maximal oxygen uptake) between 0830 and 1000 (fasting) and in a fed state (1600-1730) on each day. The normal-protein group showed fasting plasma AA concentrations that were higher (P < 0.05) than those for the high-protein group, except for leucine, methionine, and tyrosine. Glutamine, glycine, alanine, taurine, and threonine concentrations were distinctly higher ( approximately 30% or greater) throughout the 24-h period in subjects consuming the normal- vs. the high-protein diets. Exercise appeared to increase, although not profoundly, the plasma concentrations of amino acids except for glutamate, histidine, ornithine, and tryptophan. The profound diet-related differences in plasma AA concentrations are only partially explained by differences in the renal clearance of the amino acids. We speculate on the possible metabolic basis for these findings.

Background Hydroxychloroquine (HCQ) is the standard of care in the treatment of systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), and other inflammatory rheumatic diseases and potentially for the treatment in COVID-19 patients. Determination of HCQ for therapeutic drug monitoring (TDM) can be performed in whole blood (WB), serum, and plasma. Direct comparisons of WB, serum, and plasma levels of HCQ in patients with SLE have not previously been reported. We describe a method for the determination of HCQ in human blood using liquid chromatography-high-resolution mass spectrometry (LC-HRMS) and compare the suitability of the three sample matrices. Methods A method for the determination of HCQ in human blood using LC-HRMS was developed, validated, and applied for the determination of HCQ levels in WB, serum, and plasma from 26 SLE patients. The reproducibility of the method, in the three matrices, was evaluated using quality control samples and repeated preparations and measurements of patient samples. The performance of the developed method for HCQ measurement in serum was further evaluated by comparison with two previously reported extraction methods. Results The performance of the presented method demonstrated high accuracy and precision. A large range of HCQ concentrations was observed for the SLE patients in all three matrices (WB, serum, and plasma). The mean levels in WB were approximately two-fold the levels in serum and plasma (813 ng/mL compared to 436 ng/mL and 362 ng/mL, respectively). Spiked quality controls showed high reproducibility for all matrices (coefficient of variation, CV, approx. 5%), whereas in patient samples, equally high-precision was only found using WB as the matrix (CV 3%). The CV for serum and plasma was 14% and 39%, respectively. Two alternative methods applied to serum samples did not demonstrate improved precision. Conclusions A LC-HRMS method for the measurement of HCQ in human blood was developed and validated. Whole blood was found to be the superior sample matrix in terms of sample reproducibility. Thus, whole blood samples should be used for HCQ analysis when patients are monitored for HCQ treatment effects. The assay is in clinical use to monitor levels of HCQ in patients.

Objectives:To determine whether treatment with autologous hematopoietic stem cell transplantation (HSCT) can induce sustained complete remission in patients with multiple sclerosis (MS). Material and methods:Case series of patients with relapsing-remitting MS (n = 10) treated at a single center between 2004 and 2007 and followed up for 10 years. The patients were treated with a BEAM/ATG conditioning regimen (n = 9) or a cyclophosphamide/ATG conditioning regimen (n = 1) followed by infusion of unmanipulated autologous hematopoietic stem cells. The primary endpoint was sustained complete remission. Sustained complete remission was defined as "no evidence of disease activity-4," sustained for a period of at least 5 years without any ongoing disease-modifying treatment. Furthermore, MS was considered as "resolved" if intrathecal IgG production and cerebrospinal fluid neurofilament light levels were normalized as well.Results: Five out of 10 patients were in sustained complete remission at the end of the study. In three of them, MS was resolved. Conclusions:Our data demonstrate that sustained complete remission after autologous HSCT for MS is possible. K E Y W O R D S cerebrospinal fluid, hematopoietic stem cell transplantation, magnetic resonance imaging, multiple sclerosis | 321 TOLF eT aL.

Estimation of kidney function is often part of daily clinical practice, mostly done by using the endoge-nous glomerular filtration rate (GFR)-markers creatinine or cystatin C. A recommendation to use both markers in parallel in 2010 has resulted in new knowledge concerning the pathophysiology of kidney disorders by the identification of a new set of kidney disorders, selective glomerular hypofiltration syndromes. These syndromes, connected to strong increases in mortality and morbidity, # Linnea Malmgren and Carl Öberg contributed equally as first authors.

The present work investigated serum vitamin D (25(OH)D) status in relation to bone and muscle qualities and functions in 19 female soccer players (13-16 years) resident at northern latitude with very low sun exposure (∼32-36 h/month) during winter season (late January to early March). Serum 25(OH)D, parathyroid hormone and bone turnover markers osteocalcin (OC) and beta carboxy-terminal collagen cross-links (β-Ctx), as well as body composition and muscle performance were examined. Hormones were tested using routine laboratory methods. Fat mass, lean mass, and bone mineral density in whole body, as well as femur and lumbar spine were evaluated with dual-energy X-ray absorptiometry. Muscle performance was assessed through isokinetic knee extension and flexion, countermovement jump, and sprint running. 25(OH)D was low (50.5 ±   12.8 nmol l), whereas the values of bone turnover markers were markedly high (OC: 59.4 ±   18.6 µg l; β-Ctx: 1075 ±   408 ng l). All bone and muscle measurements were normal or above normal. 25(OH)D was not significantly correlated with most of the parameters of bone and muscle quality or function, except the knee extension time to peak torque (r   =   -0.50, p =   .03). In conclusion, the level of vitamin D is markedly low in adolescent female soccer players during the winter in Sweden. However, vitamin D levels did not significantly correlate with measures of bone and muscle except a moderate correlation in time to peak torque in the knee extensors. The practical implication of low vitamin D levels in young growing female athletes remains unclear.

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