Free Sialic Acid Concentrations Research Articles

Characterization of milk oligosaccharide and sialic acid content and their influence on brain sialic acid in a lean mouse model for gestational diabetes

Oligosaccharides and sialic acids (Sia) are bioactive components in milk that contribute to newborn development and health. Hyperglycemia in pregnancy (HIP) can have adverse effects on both mother and infant. HIP is associated with low-grade systemic inflammation. Inflammation influenced glycan composition, particularly of Sia-containing structures. We hypothesize that HIP and high-fat diet influence milk oligosaccharide composition, particularly sialylated oligosaccharides. Furthermore, we propose that milk Sia content influences pup brain Sia content. To test these hypotheses we (i) characterize mouse milk oligosaccharides and Sia concentrations in mouse milk of a GDM mouse model with dietary fat intake intervention; and (ii) determine Sia levels in offspring brains.The concentrations of oligosaccharides and Sia in mouse milk and offspring's brains were quantified using UPLC-FLD analysis. Analyses were performed on surplus samples from a previous study, where HIP was induced by combining high-fat diet (HF) feeding and low-dose streptozotocin injections in C57Bl/6NTac female mice. The previous study described the metabolic effects of HIP on dams and offspring.We detected 21 mouse milk oligosaccharides, including 9 neutral and 12 acidic structures using UPLC-MS. A total of 8 structures could be quantified using UPLC-FLD. Maternal HIP and HF diet during lactation influenced sialylated oligosaccharide concentrations in mouse milk and total and free sialic acid concentrations. Sia content in offspring brain was associated with total and free Neu5Gc in mouse milk of dams, but no correlations with HIP or maternal diet were observed.

Targeted versus untargeted omics - the CAFSA story.

In 2009, untargeted metabolomics led to the delineation of a new clinico-biological entity called cerebellar ataxia with elevated cerebrospinal free sialic acid, or CAFSA. In order to elucidate CAFSA, we applied sequentially targeted and untargeted omic approaches. First, we studied five of the six CAFSA patients initially described. Besides increased CSF free sialic acid concentrations, three patients presented with markedly decreased 5-methyltetrahydrofolate (5-MTHF) CSF concentrations. Exome sequencing identified a homozygous POLG mutation in two affected sisters, but failed to identify a causative gene in the three sporadic patients with high sialic acid but low 5-MTHF. Using targeted mass spectrometry, we confirmed that free sialic acid was increased in the CSF of a third known POLG-mutated patient. We then pursued pathophysiological analyses of CAFSA using mass spectrometry-based metabolomics on CSF from two sporadic CAFSA patients as well as 95 patients with an unexplained encephalopathy and 39 controls. This led to the identification of a common metabotype between the two initial CAFSA patients and three additional patients, including one patient with Kearns-Sayre syndrome. Metabolites of the CSF metabotype were positioned in a reconstruction of the human metabolic network, which highlighted the proximity of the metabotype with acetyl-CoA and carnitine, two key metabolites regulating mitochondrial energy homeostasis. Our genetic and metabolomics analyses suggest that CAFSA is a heterogeneous entity related to mitochondrial DNA alterations either through POLG mutations or a mechanism similar to what is observed in Kearns-Sayre syndrome.

Maltitol and Xylitol Sweetened Chewing-Gums Could Modulate Salivary Parameters Involved in Dental Caries Prevention

Objectives The effects of sugar-free chewing-gums (CG) sweetened with either 100% maltitol or 100% xylitol on salivary parameters were assessed in order to better understand the role polyols can play in dental caries prevention. Methods A double-blind, parallel, randomized, controlled study was conducted in China. Subjects (N = 258, age = 13 to 15 years old) were divided into 4 groups: 2 receiving polyols CG, containing respectively SweetPearl®-maltitol or XYLISORB®-xylitol, one placebo group receiving gum base and a negative control group not receiving any gum. CG were chewed for 30 days. Salivary parameters (flow rate, pH, glucan sucrase activity and free sialic acid concentration) were measured throughout the experimental period. Results All studied parameters were significantly modified with gum base compared with no-gum: salivary flow and pH increased. Glucan sucrase activity and saliva concentrations in free sialic acids decreased. Maltitol and xylitol CG showed even stronger effects on these parameters than gum base and led to a better resistance of the oral ecosystem to a sucrose challenge. This was tested in standardized in-vitro conditions. Conclusion Sugar-free CG sweetened with either maltitol or xylitol can similarly improve several parameters involved in global oral health and more specifically in dental caries prevention compared to gum base. The use of a gum base placebo allowed us to isolate the effects due to maltitol and xylitol themselves from those due to mastication, and to show that the effects were similar for both polyols.

Finding a Sugary Foothold: How Antibiotics Pave the Way for Enteric Pathogens

Antibiotic therapy predisposes the host to infections with human enteropathogens. In a recent study, Ng et al. (2013) demonstrate that antibiotic-mediated disruption of the microbial food web gives rise to free microbiota-liberated monosaccharides in the gut, which can promote growth of enteropathogenic bacteria.

Inhibitions of Several Antineoplastic Drugs on Serum Sialic Acid Levels in Mice Bearing Tumors

Six murine tumors, including ascetic tumors HepA, EC, P388 leukemia, S180 and solid tumor S180, and Lewis lung carcinoma, were employed in this work. The free sialic acid concentrations in both blood and ascites were measured in tumor-bearing mice. The results showed that the content of sialic acids in blood was increased in tumor growth and certain tumor types. Higher sialic acid content was observed in ascites than that present in blood. The influence of antineoplastic agents (vincristine, thiotepa, adriamycin, probimane, cisplatin, oxalysine, cortisone, nitrogen mustard, lycobetaine, Ara-C, harringtonine, and cyclophosphamide) on the content of sialic acids in mice blood bearing solid tumors of either S180 or Lewis lung carcinoma was observed. Different inhibitions of antineoplastic drugs on both tumor growth and serum sialic acid levels in mice bearing tumors were found. Among these antineoplastic drugs, probimane, cisplatin, nitrogen mustard, and lycobetaine were able to decrease the serum sialic acid levels in mice bearing tumors. Since these four antineoplastic drugs are all DNA chelating agents, it was proposed that the inhibition of tumor sialic acids by these drugs might be through the DNA template via two ways. Since we have found no effect of antineoplastic drugs on serum sialic acid levels in normal mice, this suggests that the inhibition of antineoplastic drugs on sialic acids is by tumor involvement.

In response to “Prenatal screening of sialic acid storage disease and confirmation in cultured fibroblasts by LC-MS/MS” by van den Bosch et al.

With great interest we have read the recent paper by Van den Bosch et al. (2011) who reported reference values of free sialic acid (FSA) in amniotic fluid supernatant (AFS). Reference values in AFS were lower (0–8.2 μmol/L) at 15–25 weeks of gestation compared to 26–38 weeks of gestation (3.2–12.0 μmol/L), indicating that FSA in AFS tends to increase with advancing gestation. Recently, we developed a method for quantification of sialic acid (SA) in cerebrospinal fluid (CSF) (van der Ham et al. 2010) since brain specific disorders of SA metabolism are recognized nowadays. When establishing reference values, we did not take into account the impact of gestational age (GA). The study of Van den Bosch et al. (2011) suggested SA metabolism to be dependent on GA. We therefore performed a study to evaluate CSF SA values in newborn infants (both preterm and full term born infants). This knowledge is essential for the interpretation of SA concentrations in CSF in these populations. The study included 27 CSF samples from preterm and 42 CSF samples from full term newborn infants. CSF was obtained for different clinical reasons. Prematurity was defined as a post menstrual age of less than 37 weeks at the time of CSF withdrawal. Post menstrual ages of preterm and full term infants were between 28 and 37 weeks and between 37 and 54 weeks, respectively. SA in CSF was quantified by HPLC-MS/MS (van der Ham et al. 2010). FSA concentrations were 72.2 ± 21.3 μmol/L and 49.3 ± 20.6 μmol/L in preterm and full term newborn infants, respectively (p < 0.0005). There was a significant negative correlation between FSA and post menstrual age (r = −0.664) with a 3.0 μmol/L decrease in FSA for each week of advancing post menstrual age (p < 0.0005). No difference was observed between conjugated sialic acid from preterm and full term newborn infants (80.0 ± 19.1 μmol/L versus 73.6 ± 28.0 μmol/L (p = 0.303)). In conclusion, with increasing gestational age, there was a considerable decrease in the amount of FSA in CSF. These results demonstrate that proper interpretation of metabolic investigations in preterm newborns depends on group-specific reference ranges, as concentrations of metabolites may differ significantly between full term and preterm born infants.

Comparative analysis of sialic acid levels in the colostrum and milk of ruminants: possible role in the passive immunity against neonatal infections

A study was conducted to determine the levels of free sialic acid in the colostrum and milk of apparently healthy indigenous Nigerian ruminants (Zebu cows, Yankassa sheep and Sokoto Red/Kano Brown goats). Haematological investigation indicates that the apparently healthy ruminants were also clinically healthy. The mean free sialic acid concentration in the colostrum of all the animal species examined was higher than those of milk and the difference in each case was statistically significant (P &lt; 0.05). The significance of the high sialic acid level in colostrum for the immunity of neonates against microbial infections is discussed.

Serum Free Sialic Acid as a Marker of Alcohol Abuse

Previous studies have shown that serum total sialic acid (TSA) concentration significantly increases during alcohol abuse. Chronic ethanol consumption impairs glycosylation of many proteins. The increased desialylation rate of serum glycoproteins is one of the effects of alcohol abuse. The aim of this study was to investigate the diagnostic value of free sialic acid (FSA) as a marker of alcohol abuse. We determined serum FSA concentrations in the group of 156 alcoholic subjects and 35 healthy control subjects by means of a modification of the thiobarbituric acid method. The alcoholic group was divided into subgroups according to their history of abuse. The FSA concentration was significantly higher in alcoholic subjects than in healthy controls. The subjects who consumed alcohol for longer than a week showed significantly higher FSA level than those who consumed alcohol for a shorter period. The serum FSA concentration was significantly higher in alcoholic subjects with elevated markers of liver dysfunction. The diagnostic accuracy of FSA was high, although it did not differ from TSA, and was limited by its low sensitivity. This study shows that FSA concentration in the sera of alcoholic subjects is increased. The low diagnostic sensitivity is accompanied by high specificity, however the accuracy is high and similar to the accuracy of TSA. Free sialic acid does not seem to be a better marker of alcohol abuse than TSA and current markers.

RED BLOOD CELL DESIALYLATION, AS OBSERVED IN SEPSIS ALTERS THEIR SHAPE AND BIOCHEMISTRY

Introduction: Red blood cell (RBC) metabolism is totally dependent on glycolysis to supply the ATP required for the maintenance of ionic homeostasis and structural integrity (1). Several studies have observed interactions between glycolytic enzymes and the RBC membrane (2,3). We have recently observed that RBCs from septic patients show a more spherical shape with decreased membrane sialic acid (SA) content (4). Moreover, RBC sphericity was inversely correlated with membrane SA content (4). We therefore hypothesized that desialylation as observed in septic patients could alter RBC glycolysis. Materials and Methods: Washed RBCs from 9 volunteers (adjusted at 20% hematocrit) were incubated with 0.125 U/mL of neuraminidase from Clostridium perfringens or RPMI as controls. RBC shape was estimated by flow cytometry after 10 hours of incubation. Normal RBCs show a bimodal distribution related to the biconcave form (4). On this histogram, we calculated the second Pearson Coefficient of Dissymmetry (= 3 × (mean-median)/SD) (PCD). The normal PCD value is around - 0.8 and a null value represents a perfect spherical shape. To confirm the release of SA from the RBC membrane by neuraminidase, free SA concentrations in the supernatant were measured at baseline and after 10 hours of incubation by an enzymatic colorimetric assay. 2,3 DPG and RBC lactate concentrations were measured after 10 hours of incubation with neuraminidase. Data are presented as mean±SD and were compared by a Student's test; correlations were evaluated by a Spearman's test. Results: Neuraminidase altered RBC shape and glycolysis. PCD values were correlated with 2,3 DPG concentrations (r2 = 0.37; P = 0.008).TableConclusions: Desialylation induces RBC sphericity and an increase in RBC lactate and 2,3 DPG concentrations, already present 10 hours after neuraminidase treatment. These changes may play a role in the alterations in RBC rheology observed in sepsis, and could be the target of new interventions to improve the management of septic patients. Funds: Erasme Foundation

Prenatal diagnosis of free sialic acid storage disorders (SASD)

Free sialic acid storage disorders, Salla disease (SD) and Infantile sialic acid storage disease (ISSD), are lysosomal storage diseases due to impaired function of a sialic acid transporter, sialin, at the lysosomal membrane. Several mutations of the sialin gene, SLC17A5, are known, leading either to the severe neonatal/infantile disease or to the milder, adult-type developmental disorder, Salla disease. Free sialic acid accumulation in lysosomes causes increased tissue concentration and consequently elevated urinary excretion. Prenatal diagnosis of SASD is possible either by determination of free sialic acid concentration or by mutation analysis of the SLC17A5 gene in fetal specimen, in chorionic villus biopsy particularly. Both techniques have been successfully applied in several cases, sialic acid assay more often in ISSD cases but mutation analysis preferentially in SD. Sialic acid assay of amniotic fluid supernatant or cultured amniotic fluid cells may give erroneous results and should not be used for prenatal diagnosis of these disorders. The present comments are mainly based on our experience of prenatal diagnosis of SD in Finnish families. A founder mutation in SLC17A5 gene, 115C-> T, represents 95% of the disease alleles in the Finnish SD patients, which provides a unique possibility to apply mutation analysis. Therefore, molecular studies have successfully been used in 17 families since the identification of the gene and the characterization of the SD mutations. Earlier, eight prenatal studies were performed by measuring the free sialic acid concentration in chorionic villus samples.

RAPID ALTERATIONS IN TRANSFERRIN SIALYLATION DURING SEPSIS

The inflammatory process is associated with alterations in iron metabolism. Transferrin, an acute-phase N-glycosylated glycoprotein, plays an important role in iron transport. Human serum transferrin contains two biantennary glycans, each consisting of 0 to 4 molecules of sialic acid (SA); its SA content is heterogeneous with high concentration of tetrasialotransferrin (4SA) and low amounts of disialo-, trisialo-, penta-, and hexasialotransferrin. The hepatic uptake of iron is greater for desialylated transferrin isoforms (disialotransferrin) than for the other forms. We hypothesized that serum levels of carbohydrate-deficient transferrin (CDT, disialotransferrin) may increase rapidly in septic patients. Blood samples were obtained from critically ill patients with (n = 15) and without (n = 14) documented sepsis and compared with healthy volunteers. The different forms of transferrin were studied by capillary zone electrophoresis; SA concentrations were measured by enzymatic colorimetric assay. There was a significant increase in the proportion of CDT in septic compared with nonseptic patients and volunteers (18.3% [1.3-30.5] vs. 0.7% [0.5-0.9]; P < 0.01 and 0.9% [0.5-1.1]; P < 0.05). Conversely, tri- and tetrasialotransferrin levels were lower in septic patients. Total and free SA concentrations were significantly higher in septic patients than in healthy volunteers. In a sheep model of septic shock secondary to peritonitis, serum free SA was already increased after 15 h. Sepsis is associated with decreased SA content on circulating transferrin and with an increase in blood free SA concentrations. In view of these rapid modifications and the long half-life of transferrin, the most likely explanation is degradation of transferrin by neuraminidase. Further studies including measurement of blood neuraminidase concentration and activity are needed to understand the process and exact role of SA decrease in septic patients.

English

Free and total sialic acid concentrations were determined in the midgut extract of&nbsp;Culex pipiense pipiense&nbsp;mosquitoes infected with&nbsp;Trypanosoma congolense.&nbsp; The mean total sialic acid concentrations were found to be 1.5 to 2 fold higher than the mean free sialic acid concentrations in the midgut extracts of all the groups of the&nbsp;T. congolense&nbsp;infectedC.p. pipiense. Infusion of 10 mg/ml galactose and 10 mg/ml lactose did not change the pattern of this difference but resulted to 1.3 to 1.4 fold decrease in the total sialic acid concentration. The relevance of these findings to the role of sialic acids in the midgut ofT. congolense&nbsp;infected&nbsp;C.p. pipiense&nbsp;mosquitoes is discussed in this paper. &nbsp; Key words:&nbsp;Trypanosoma congolense,&nbsp;Culex pipiense pipiense, sialic acid, midgut.

A Japanese case of infantile sialic acid storage disease

We report a 4-year-old Japanese girl with infantile sialic acid storage disease. She presented with failure to thrive, coarse facial features, hepatosplenomegaly, severe mental retardation and spastic quadriplegia. Electron microscopic examination of cultured skin fibroblasts revealed multiple vacuoles and inclusion material representing distended lysosomes, thus suggesting a lysosomal storage disorder. A high concentration of free sialic acid was present in the urine and cultured fibroblasts, but bound sialic acid was not increased. The activity of a variety of lysosomal enzymes was not diminished. The MR1 findings included brain atrophy and a diffuse high signal in the cerebral white matter and low signal in the basal ganglia on T2-weighted images. To our knowledge, this is the first case of infantile sialic acid storage disease described in a non-Caucasian family.

Determination of serum total lipid and free N-acetylneuraminic acid in genitourinary malignancies by fluorimetric high performance liquid chromatography. Relevance of free N-acetylneuraminic acid as tumour marker

The reliability of total sialic acid (TSA), lipid sialic acid (LSA) and free sialic acid (FSA) as markers in genitourinary malignancies was evaluated in 20 normal subjects, 21 patients with prostatic cancer, 22 patients with urinary bladder cancer and 14 patients with renal cell carcinoma. We introduce the new concept of ‘corrected’ lipid sialic acid (CLSA), which expresses the actual concentration of sialic acid bound to glycolipids by subtracting the concentration of FSA determined by a novel ultrafiltration method. TSA did not show significant differences with respect to normal controls, except for renal cell carcinoma, whose mean value (879 ± 145 μg/ml) showed a P value < 0.001. Instead, CLSA showed only significant differences ( P = 0.001), with respect to normal controls in stage I and in all grades of renal cell carcinoma. While all data indicated significant increases ( P = 0.001) in the FSA values, (means ± S.D.) of 0.621 ± 0.272 μg/ml were found in patients with prostatic cancer, 0.796 ± 0.443 μg/ml in patients with urinary bladder cancer and 0.667 ± 0.146 μg/ml in patients with renal cell carcinoma. Separate TSA and CLSA measurements appeared to be of limited value in the detection of genitourinary malignancies. However, results show that FSA was the most sensitive of the three markers tested for detecting malignancies.

Age-related reference values for urinary excretion of sialic acid and deoxysialic acid: application to diagnosis of storage disorders of free sialic acid

We have established by HPLC age-related reference intervals for sialic acid urinary excretion in 364 control individuals to assist in evaluating the clinical significance of the free sialic acid concentration in urine. In addition, an HPLC method for quantitative analysis of free deoxysialic acid was developed, and age-related reference intervals for excretion of this compound in urine were established. In patients with storage disorders of free sialic acid (n = 11) the sialic acid excretion was increased 2- to 35-fold, compared with the mean value of the control subjects in the corresponding age group, and exceeded the interval in each case. The excretion of deoxysialic acid was within the reference interval in all of the patients, indicating that its metabolism was not affected in the disorders. The age-related reference values assist in evaluating the excretion of free sialic acid in the diagnosis of storage disorders of free sialic acid, especially in young children.

Evaluation of serum sialic acid fractions as markers for malignant melanoma

Increased total serum sialic acid levels have proved to be a useful non-specific marker of various neoplastic diseases including melanoma. In some malignancies, specific fractions of sialic acid were reported to be more reliable for monitoring disease progression. In this study various fractions of sialic acid were measured in the sera of Bomirski melanoma-bearing Syrian hamsters using the modified thiobarbituric acid method of Skoza and Mohos. Significantly higher serum levels of total sialic acid, 601.0 ± 49.1 μg/ml, were demonstrated in animals with melanoma in comparison with control hamsters, 463.1 ± 33.0 μg/ml ( P < 0.01). Free sialic acid concentration was found to be negligible compared with that found in the sera of melanoma patients. Lipid bound sialic acid measurements carried out according to Bhatavdekar et al. were shown to be influenced by interfering substances. Hence, only the estimation of total sialic acid in sera can be recommended for monitoring melanoma progression. Bomirski melanoma proved to be a good model of human melanoma.

Sialic acid storage disease.

A baby girl with coarse facial features, hepatosplenomegaly, and developmental delay had raised free sialic acid concentrations in her urine and cultured fibroblasts. She died aged 13 months. Sialic acid is an important constituent of many glycoproteins and glycolipids; impaired release from the lysosome may be the underlying biochemical defect.

Overproduction of N-acetylneuraminic acid (sialic acid) by sialuria fibroblasts.

Fibroblasts from the original sialuria patient are shown to contain high levels of free sialic acid, i.e., an average of 87 versus a normal average of 2 nmol/mg of protein. Gas liquid chromatography-mass spectrometry analysis confirmed that the accumulated material is N-acetylneuraminic acid. Addition of D(+) glucosamine (0-5 mM) to the media of these cells increased the intracellular free sialic acid concentrations from 74 to 137 nmol/mg protein. In contrast, normal cells treated in an identical manner maintained their normal free sialic acid level of less than 4 nmol/mg protein. Addition of 20 mM N-acetylmannosamine in place of glucosamine resulted in a marked increase in free sialic acid in both the patient and the control, i.e., increases of 157 (from 95 to 252) and 120 (from 3 to 123) nmol/mg protein, respectively. Finally, while normal cells in the presence of glucosamine accumulated high levels of uridine diphosphate N-acetylhexosamine, the patient's cells accumulated much lower amounts of this compound. It is concluded that the elevated sialic acid level in sialuria is due, at least in part, to overproduction of free N-acetylneuraminic acid. Indirect evidence is presented that this may result from either hyperactivity or increased levels of the enzyme (uridine diphosphate N-acetylglucosamine 2-epimerase) that has been shown in other tissues to convert uridine diphosphate N-acetylglucosamine to N-acetylmannosamine.

Effects of thyrotropin and thiouracil on the sialic acid concentration in the thyroid gland

An investigation was made of sialic acid in the thyroid gland. A preparation of calf thyroglobulin which was homogeneous in the ultracentrifuge contained 1.2% sialic acid. Stimulation of the thyroid gland of the rat to cause resorption of colloid was accompanied by a decrease in the thyroidal sialic acid concentration. After feeding thiouracil for 2 days, the time when the rate of decrease of the sialic acid concentration was maximal, there was a doubling of the concentration of free sialic acid in the thyroid. The sialic acid was identified to be N-acetylneuraminic acid.