BackgroundPotential biomarkers to aid diagnosis and therapy need to be identified for Amyotrophic Lateral Sclerosis, a progressive motor neuronal degenerative disorder. The present study was designed to identify the factor(s) which are differentially expressed in the cerebrospinal fluid (CSF) of patients with sporadic amyotrophic lateral sclerosis (SALS; ALS-CSF), and could be associated with the pathogenesis of this disease.ResultsQuantitative mass spectrometry of ALS-CSF and control-CSF (from orthopaedic surgical patients undergoing spinal anaesthesia) samples showed upregulation of 31 proteins in the ALS-CSF, amongst which a ten-fold increase in the levels of chitotriosidase-1 (CHIT-1) was seen compared to the controls. A seventeen-fold increase in the CHIT-1 levels was detected by ELISA, while a ten-fold elevated enzyme activity was also observed. Both these results confirmed the finding of LC-MS/MS. CHIT-1 was found to be expressed by the Iba-1 immunopositive microglia.ConclusionElevated CHIT-1 levels in the ALS-CSF suggest a definitive role for the enzyme in the disease pathogenesis. Its synthesis and release from microglia into the CSF may be an aligned event of neurodegeneration. Thus, high levels of CHIT-1 signify enhanced microglial activity which may exacerbate the process of neurodegeneration. In view of the multifold increase observed in ALS-CSF, it can serve as a potential CSF biomarker for the diagnosis of SALS.

Background The incidence of invasive meningococcal disease in the UK decreased by approximately four times from 1999 to 2014, with reductions in serogroup C and serogroup B disease. Lower serogroup C invasive meningococcal disease incidence was attributable to implementation of the meningococcal serogroup C conjugate vaccine in 1999, through direct and indirect protection, but no vaccine was implemented against serogroup B disease. UK Meningococcal Carriage surveys 1-3 (UKMenCar1-3), conducted in 1999, 2000, and 2001, were essential for understanding the impact of vaccination. To investigate the decline in invasive meningococcal disease incidence, we did a large oropharyngeal carriage survey in 2014-15, immediately before the changes to meningococcal vaccines in the UK national immunisation schedule. Methods UKMenCar4 was a cross-sectional survey in adolescents aged 15-19 years who were enrolled from schools and colleges geographically local to one of 11 UK sampling centres between Sept 1, 2014, and March 30, 2015. Participants provided an oropharyngeal swab sample and completed a questionnaire on risk factors for carriage, including social behaviours. Samples were cultured for putative Neisseria spp, which were characterised with serogrouping and wholegenome sequencing. Data from this study were compared with the results from the UKMenCar1-3 surveys (1999-2001).Findings From the 19 641 participants (11 332 female, 8242 male, 67 not stated) in UKMenCar4 with culturable swabs and completed risk-factor questionnaires, 1420 meningococci were isolated, with a carriage prevalence of 7•23% (95% CI 6•88-7•60). Carriage prevalence was substantially lower in UKMenCar4 than in the previous surveys: carriage prevalence

Background: The survival of motor neurons is dependent upon neurotrophic factors both during childhood and adolescence and during adult life. In disease conditions, such as in patients with amyotrophic lateral sclerosis (ALS), the mRNA levels of trophic factors like brain-derived neurotrophic factor (BDNF), insulin-like growth factor-1 (IGF-1), fibroblast growth factor-2 (FGF-2), and vascular endothelial growth factor are downregulated. This was replicated in our in vivo experimental system following the injection of cerebral spinal fluid (CSF) of sporadic ALS (ALS-CSF) patients. Objective: To evaluate the protective role of BDNF in a model of sporadic ALS patients. Methods: The expressions of endogenous BDNF, its receptor TrkB, the enzyme choline acetyl transferase (ChAT), and phosphorylated neurofilaments were studied in NSC-34 cells. The calcium-buffering and proapoptotic effects were assessed by calbindin-D28K and caspase-3 expression, respectively. Results: ALS-CSF considerably depleted the endogenous BDNF protein, while its effect on IGF-1 and FGF-2 was inconsequential; this indirectly indicates a key role for BDNF in supporting motor neuronal survival. The exogenous supplementation of BDNF reversed autocrine expression; however, it may not be completely receptor mediated, as the TrkB levels were not restored. BDNF completely revived ChAT expression. It may inhibit apoptosis by restoring Ca²⁺ homeostasis, since caspase-3 and calbindin-D28K expression was back to normal. The organellar ultrastructural changes were only partially reversed. Conclusion: Our study provides evidence that BDNF supplementation ameliorates most but not all degenerative changes. The incomplete revival at the ultrastructural level signifies the requirement of factors other than BDNF for near-total protection of motor neurons, and, to an extent, it explains why only a partial success is achieved in clinical trials with BDNF in ALS patients.

Vascular endothelial growth factor (VEGF), the well-known angiogenic factor is both neurotrophic and neuroprotective. Altered VEGF signalling is implicated in the pathogenesis of amyotrophic lateral sclerosis (ALS), a fatal degenerative disease of motor neurons. We have shown earlier that VEGF protects NSC-34 motor neuronal cell line, when exposed to cerebrospinal fluid (CSF) from sporadic ALS patients (ALS-CSF). Here, we have investigated the consequences of ALS-CSF and VEGF supplementation on the VEGFR2 receptor and endogenous VEGF expression. ALS-CSF caused significant down-regulation of VEGFR2 as well as the Calbindin-D28K levels, but not endogenous VEGF. Exogenous supplementation restored the depletion of VEGFR2 and Calbindin-D28K with a concomitant up-regulation of endogenous VEGF. The up-regulated caspase 3 in the ALS-CSF group was reinstated to basal levels along with a significant reduction in the number of TUNEL-positive cells. Electron photomicrographs of ALS-CSF-exposed cells divulged presence of cytoplasmic vacuoles alongside severe damage to organelles like mitochondria, endoplasmic reticulum, etc. Substantial recovery of most of the damaged organelles was noted in response to VEGF supplementation. While the enhancement in endogenous VEGF levels highlights the autocrine functions, the up-regulation of VEGFR2 receptor emphasizes the paracrine functions of VEGF in modulating its neuroprotective effect against ALS-CSF. The revival of cellular organellar structure, increased calbindin expression and enhanced survival in response to VEGF supplementation consolidates the opinion that VEGF indeed has a therapeutic potential in sporadic ALS.

Background: Cerebrospinal fluid from amyotrophic lateral sclerosis patients (ALS-CSF) induces neurodegenerative changes in motor neurons and gliosis in sporadic ALS models. Search for identification of toxic factor(s) in CSF revealed an enhancement in the level and enzyme activity of chitotriosidase (CHIT-1). Here, we have investigated its upregulation in a large cohort of samples and more importantly its role in ALS pathogenesis in a rat model. Methods: CHIT-1 level in CSF samples from ALS (n = 158), non-ALS (n = 12) and normal (n = 48) subjects were measured using ELISA. Enzyme activity was also assessed (ALS, n = 56; non-ALS, n = 10 and normal-CSF, n = 45). Recombinant CHIT-1 was intrathecally injected into Wistar rat neonates. Lumbar spinal cord sections were stained for Iba1, glial fibrillary acidic protein and choline acetyl transferase to identify microglia, astrocytes and motor neurons respectively after 48 h of injection. Levels of tumour necrosis factor-α and interleukin-6 were measured by ELISA. Findings: CHIT-1 level in ALS-CSF samples was increased by 20-fold and it can distinguish ALS patients with a sensitivity of 87% and specificity of 83.3% at a cut off level of 1405.43 pg/ml. Enzyme activity of CHIT-1 was also 15-fold higher in ALS-CSF and has a sensitivity of 80.4% and specificity of 80% at cut off value of 0.1077989 μmol/μl/min. Combining CHIT-1 level and activity together gave a positive predictive value of 97.78% and negative predictive value of 100%. Administration of CHIT-1 increased microglial numbers and astrogliosis in the ventral horn with a concomitant increase in the levels of pro-inflammatory cytokines. Amoeboid-shaped microglial and astroglial cells were also present around the central canal. CHIT-1 administration also resulted in the reduction of motor neurons. Conclusions: CHIT-1, an early diagnostic biomarker of sporadic ALS, activates glia priming them to attain a toxic phenotype resulting in neuroinflammation leading to motor neuronal death.