Cerebrospinal fluid ß-glucocerebrosidase activity is reduced in parkinson's disease patients.

BACKGROUND: Reduced ß-glucocerebrosidase activity was observed in postmortem brains of both GBA1 mutation carrier and noncarrier Parkinson's disease patients, suggesting that lower ß-glucocerebrosidase activity is a key feature in the pathogenesis of PD. The objectives of this study were to confirm whether there is reduced ß-glucocerebrosidase activity in the CSF of GBA1 mutation carrier and noncarrier PD patients and verify if other lysosomal enzymes show altered activity in the CSF. METHODS: CSF ß-glucocerebrosidase, cathepsin D, and ß-hexosaminidase activities were measured in 79 PD and 61 healthy controls from the BioFIND cohort. The whole GBA1 gene was sequenced. RESULTS: Enzyme activities were normalized according to CSF protein content (specific activity). ß-glucocerebrosidase specific activity was significantly decreased in PD versus controls (-28%, P < 0.001). GBA1 mutations were found in 10 of 79 PD patients (12.7%) and 3 of 61 controls (4.9%). GBA1 mutation carrier PD patients showed significantly lower ß-glucocerebrosidase specific activity versus noncarriers. ß-glucocerebrosidase specific activity was also decreased in noncarrier PD patients versus controls (-25%, P < 0.001). Cathepsin D specific activity was lower in PD versus controls (-21%, P < 0.001). ß-Hexosaminidase showed a similar trend. ß-Glucocerebrosidase specific activity fairly discriminated PD from controls (area under the curve, 0.72; sensitivity, 0.67; specificity, 0.77). A combination of ß-glucocerebrosidase, cathepsin D, and ß-hexosaminidase improved diagnostic accuracy (area under the curve, 0.77; sensitivity, 0.71; specificity, 0.85). Lower ß-glucocerebrosidase and ß-hexosaminidase specific activities were associated with worse cognitive performance. CONCLUSIONS: CSF ß-glucocerebrosidase activity is reduced in PD patients independent of their GBA1 mutation carrier status. Cathepsin D and ß-hexosaminidase were also decreased. The possible link between altered CSF lysosomal enzyme activities and cognitive decline deserves further investigation. © 2017 International Parkinson and Movement Disorder Society.

Ym1, an eosinophilic chemotactic factor, participates in the brain inflammation induced by Angiostrongylus cantonensis in mice.

Angiostrongylus cantonensis is an emerging zoonotic pathogen that has caused hundreds of cases of human angiostrongyliasis worldwide. The larva in nonpermissive hosts cannot develop into an adult worm and can cause eosinophilic meningitis and ocular angiostrongyliasis. The mechanism of brain inflammation caused by the worm remains poorly defined. According to previous data of GeneChip, Ym1 in the brain of mice 21 days after infection with A. cantonensis was highly upregulated to over 7,300 times than the untreated mice. Ym1 is an eosinophilic chemotactic factor with the alternative names of chitinase-3-like protein 3, eosinophil chemotactic cytokine, and ECF-L. Ym1 displays chemotactic activity for T lymphocytes, bone marrow cells, and eosinophils and may favor inflammatory responses induced by parasitic infections and allergy. It has been reported that Ym1 is synthesized and secreted by activated macrophages during parasitic infection (Chang et al., J Biol Chem 276(20):17497-17506, 2001). In the brain, microglia are alternatively activated macrophage-derived cells which are the key immune cells in central nervous system inflammation. To explore the role of Ym1 in inflammation caused by A. cantonensis-infected mice, we examined the levels of Ym1 in the sera and cerebrospinal fluid (CSF) of the infected animals, followed by detection of the mRNA expression level of Ym1 in various organs including the brain, lung, liver, spleen, and kidney and of the cytokines IL-5 and IL-13 in the brain of the infected mice with or without intraperitoneal injection of minocycline (an inhibitor of microglial activation) by real-time reserve transcription PCR. Furthermore, immunolocalization of Ym1 in the brains of the infected mice was observed by using a fluorescence microscope. Our results showed that Ym1 was most highly expressed in the brains and CSF of the infected mice along with the process of inflammation. The antibody localized Ym1 to the microglia in the brain of the mice in both infection and minocycline + infection groups. And as in the brain, the mRNA level of Ym1 changed more obviously than IL-5 and IL-13. The study implies that Ym1 might serve as an alternative potential pathological marker which is detected not only in the sera and CSF but also in the brains of the infected mice and Ym1 secreted by microglia might be involved in eosinophilic meningitis and meningoencephalitis caused by A. cantonensis infection.

Cerebrospinal fluid beta-glucocerebrosidase activity is reduced in Dementia with Lewy Bodies.

The autophagy-lysosomal degradation pathway plays a role in the onset and progression of neurodegenerative diseases. Clinical and genetic studies indicate that mutations of beta-glucocerebrosidase represent genetic risk factors for synucleinopathies, including Parkinson's Disease (PD) and Dementia with Lewy Bodies (DLB). We recently found a decreased activity of lysosomal hydrolases, namely beta-glucocerebrosidase, in cerebrospinal fluid of PD patients. We have thus measured the activity of these enzymes - alpha-mannosidase (EC 3.2.1.24), beta-mannosidase (EC 3.2.1.25), beta-glucocerebrosidase (EC 3.2.1.45), beta-galactosidase (EC 3.2.1.23) and beta-hexosaminidase (EC 3.2.1.52) - in cerebrospinal fluid of patients suffering from DLB, Alzheimer's Disease (AD), Fronto-Temporal Dementia (FTD) and controls. Alpha-mannosidase activity showed a marked decrease across all the pathological groups as compared to controls. Conversely, beta-glucocerebrosidase activity was selectively reduced in DLB, further suggesting that this enzyme might specifically be impaired in synucleinopathies.

beta-N-acetylhexosaminidases in human cerebrospinal fluid and serum of patients with multiple sclerosis.

Beta-N-Acetylhexosaminidase activity and isoenzyme have been investigated in normal human cerebrospinal fluid and that of patients with multiple sclerosis. beta-N-acetylhexosaminidase activity in normal cerebrospinal fluids has been resolved into five components. The major component was in a form that eluted from DEAE cellulose at the same salt concentration as hexosaminidase As, the isoenzyme previously identified in human serum. Cerebrospinal fluid from patients exhibited a different isoenzyme profile, showing a remarkable increase in a form having a pI which was more acidic than that of As. These changes have a potential use in the diagnosis and further biochemical characterization of multiple sclerosis.

Laboratory diagnosis of canine GM2-gangliosidosis using blood and cerebrospinal fluid.

In the present study, laboratory techniques were used to diagnose canine GM2-gangliosidosis using blood and cerebrospinal fluid (CSF) that can be collected noninvasively from living individuals. Lysosomal acid beta-hexosaminidase (Hex) was measured spectrofluorometrically using 4-methylumbelliferyl N-acetyl-beta-D-glucosaminide and 4-methylumbelliferyl 7-(6-sulfo-2-acetamido-2-deoxy-beta-D-glucopyranoside) as substrates. Main isoenzymes A and B of Hex in leukocytes were also analyzed using cellulose acetate membrane electrophoresis. GM2-ganglioside in CSF was detected and determined quantitatively by using thin-layer chromatography/enzyme-immunostaining method with anti-GM2-ganglioside antibody. In normal dogs, Hex activities could be determined in leukocytes, serum, and CSF and the total activities were markedly reduced in all the enzyme sources in a dog with Sandhoff disease. Electrophoresis of a leukocyte lysate from a normal dog showed that the Hex A and Hex B were not separated distinctively with formation of a broad band, whereas there were no bands in electrophoresis of a lysate from a dog with Sandhoff disease, showing a deficiency in the total enzyme activity. GM2-ganglioside could be detected and determined quantitatively in as little as 100 microl of canine CSE GM2-ganglioside in CSF in a dog with Sandhoff disease increased to 46 times the normal level. In conclusion, the methods in the present study are useful for diagnosis of canine GM2-gangliosidosis. These techniques enable definitive and early diagnosis of canine GM2-gangliosidosis even if tissues and organs cannot be obtained.

Factors influencing the measurement of lysosomal enzymes activity in human cerebrospinal fluid.

Measurements of the activities of lysosomal enzymes in cerebrospinal fluid have recently been proposed as putative biomarkers for Parkinson's disease and other synucleinopathies. To define the operating procedures useful for ensuring the reliability of these measurements, we analyzed several pre-analytical factors that may influence the activity of ß-glucocerebrosidase, α-mannosidase, ß-mannosidase, ß-galactosidase, α-fucosidase, ß-hexosaminidase, cathepsin D and cathepsin E in cerebrospinal fluid. Lysosomal enzyme activities were measured by well-established fluorimetric assays in a consecutive series of patients (n = 28) with different neurological conditions, including Parkinson's disease. The precision, pre-storage and storage conditions, and freeze/thaw cycles were evaluated. All of the assays showed within- and between-run variabilities below 10%. At -20°C, only cathepsin D was stable up to 40 weeks. At -80°C, the cathepsin D, cathepsin E, and ß-mannosidase activities did not change significantly up to 40 weeks, while ß-glucocerebrosidase activity was stable up to 32 weeks. The ß-galactosidase and α-fucosidase activities significantly increased (+54.9±38.08% after 4 weeks and +88.94±36.19% after 16 weeks, respectively). Up to four freeze/thaw cycles did not significantly affect the activities of cathepsins D and E. The ß-glucocerebrosidase activity showed a slight decrease (-14.6%) after two freeze/thaw cycles. The measurement of lysosomal enzyme activities in cerebrospinal fluid is reliable and reproducible if pre-analytical factors are accurately taken into consideration. Therefore, the analytical recommendations that ensue from this study may contribute to the establishment of actual values for the activities of cerebrospinal fluid lysosomal enzymes as putative biomarkers for Parkinson's disease and other neurodegenerative disorders.

A thermodynamic study of beta-N-acetylhexosaminidase enzyme heterogeneity in cerebrospinal fluid from patients with multiple sclerosis.

BACKGROUND: ss-N-acetylhexosaminidase (Hex) is a lysosomal hydrolase, whose determination in the cerebrospinal fluid (CSF) of patients with multiple sclerosis (MS) has provided discordant results. METHODS: Total Hex and its isoenzymes Hex A and Hex B were determined using a thermodynamic procedure in the CSF of 27 patients with definitive MS, 8 with possible MS, 9 with meningitis, 14 with other neurological diseases, and in 10 controls without any neurological disease. RESULTS: In the group of patients with definitive MS, the total Hex and Hex A were significantly higher than in the control group (p<0.001), with a possible association of greater enzymatic activities with the presence of oligoclonal bands and recent relapse; however, an overlap was detected for the activities of total Hex and its isoenzymes between the groups of patients with different neurological diseases. A significant correlation was obtained for neuron-specific enolase (NSE) with total Hex and Hex A and Hex B isoenzymes (p<0.001); however, in the partial correlation statistical significance was only obtained between NSE and Hex A (p<0.001) which is the most abundant Hex isoenzyme in the brain. CONCLUSIONS: Although the inflammatory process in MS mainly takes place in the perivascular zone, with little activity in the cerebral parenchyma, the significant increase of NSE and Hex A isoenzyme in CSF reveals a neuronal damage. The disease status may have effect on the CSF Hex activity.

Increased activity of lysosomal acid hydrolases in the cell-free cerebrospinal fluid of bacterial meningitis.

Because inflammation could affect lysosomal enzyme trafficking, resulting in increased enzyme release from the cells, tissue necrosis, or altered blood- and the brain-cerebrospinal fluid (CSF) barrier, the activity of four lysosomal enzymes in the cell-free CSF of 34 patients with bacterial meningitis, 20 with aseptic meningitis, and 39 control subjects was measured. Activities are expressed in nanomoles of 4-methylumbelliferone mL/h. The median beta-hexosaminidase A activity in bacterial meningitis was 313, in aseptic meningitis it was 173, and in the control subjects it was 175, the median beta-hexosaminidase B activity was 417, 165, and 120; the median alpha-mannosidase activity was 171, 124, and 113; and the median beta-glucuronidase activity was 133.7, 14.3, and 10.0, respectively. The difference of the activities of the four enzymes measured between the bacteria meningitis and the controls is significant (p < 0.000). Also significant is the difference between bacterial and aseptic meningitis (p = 0.005 to < 0.000), but it is not significant between aseptic and control subjects. Both the sensitivity and specificity of the beta-glucuronidase activity between bacterial meningitis and control subjects were 100%, whereas the corresponding values between bacterial and aseptic meningitis were 100% and 90%, respectively. No significant correlation was observed between the activities of the enzymes measured and the number of the polymorphonuclear leukocytes or other laboratory characteristics of the CSF. The increased lysosomal enzyme activities in the CSF of patients with meningitis may result from diffusion across the blood-CSF or the brain-CSF barrier or from enzyme leakage through the cell membranes.

Sandhoff disease in a golden retriever dog.

A golden retriever dog is described with total hexosaminidase deficiency and raised GM2-ganglioside in CSF. The animal represents a model for human Sandhoff disease.