Transforming growth factor-β and Th17 responses in resistance to primary murine schistosomiasis mansoni

Discovery of the T-helper (Th) 17 cell lineage and functions in immune responses of mouse and man prompted us to investigate the role of transforming growth factor-beta (TGF-β) and interleukin (IL)-17 in innate resistance to murine schistosomiasis mansoni. Schistosoma mansoni-infected BALB/c and C57BL/6 mice were administered with recombinant TGF-β or mouse monoclonal antibody to TGF-β to evaluate the impact of this cytokine on host immune responses against lung-stage schistosomula, and subsequent effects on adult worm parameters. Developing schistosomula elicited increase in peripheral blood mononuclear cells (PBMC) mRNA expression and/or plasma levels of IL-4, IL-17, and interferon-gamma (IFN-γ), cytokines known to antagonize each other, resulting in impaired Th1/Th2, and Th17 immune responses and parasite evasion. Mice treated with TGF-β showed elevated PBMC mRNA expression of IL-6, IL-17, TGF-β, and TNF-α mRNA and increased IL-23 and IL-17 or TGF-β plasma levels, associated with significantly (P < 0.02–<0.0001) lower S. mansoni adult worm burden compared to controls in both mouse strains, thus suggesting that TGF-β led to heightened Th17 responses that mediated resistance to the infection. Mice treated with antibody to TGF-β showed increase in PBMC mRNA expression and plasma levels of IL-4, IL-12p70, and IFN-γ, and significantly (P < 0.02 and <0.0001) reduced worm burden and liver worm egg counts than untreated mice, indicating that Th1/Th2 immune responses were potentiated, resulting in significant innate resistance to schistosomiasis. The implications of these observations for schistosome immune evasion and vaccination were discussed.     

The differential effect of Eriobotrya japonica hydrophilic leaf extract on cytokines production and modulation

Stimulating or modulating the release of cytokines by immunomodulators or immunostimulating agents is an attractive mode for treating several diseases such as viral infections. For instance, patients with viral infections may be in need of increasing or inducing T helper 1 (Th1) or proinflammatory cytokines, which ultimately activate T cytotoxic and Natural killer lymphocytes to kill virally infected cells. Of these agents, we found that Eriobotrya japonica hydrophilic leaf extract (EJHE) can induce and modulate cytokines in dose-dependent manner. Twenty-four hour exposure of increasing concentrations of EJHE increased significantly (p < 0.001) the production of IFN-γ and TNF-α, from PHA+LPS-stimulated whole blood. However, the production of IFN-γ and TNF-α plateaued at high EJHE concentrations (10–100 μg/ml). No significant changes in the production of IL-10 were seen. In addition, EJHE at 1 and 10 μg/ml reversed significantly (p < 0.01) the inhibitory effect of hydrocortisone on the IL-12 p70, IFN-γ and TNF-α production from PHAS+LPS stimulated whole blood. Without PHA and LPS, EJHE was found to induce significantly (p < 0.001) IFN-γ, IL-12 p70, TNF-α, and IL-10 from whole blood culture in concentration dependent manner. The maximum induction of IFN-γ, IL-12 p70, and TNF-α by EJHE was at 1 and 10 μg/ml. On the other hand, IL-10 induction kept increasing even at the highest concentration used (100 μg/ml) of EJHE. Furthermore, intra-peritoneal injection of EJHE into mice increased significantly serum cytokines level mainly at 10 and 100 μg/ml. Two-hour post i.p. injection, EJHE increased serum IFN-γ, TNF-α, and IL-10 to 750, 1000, and 250 pg/ml, respectively. However, 24 h post i.p. injection, the levels of TNF-α, and IL-10 were similar to basal levels but IFN-γ levels were 200 pg/ml. These results indicate that EJHE induces proinflammatory and Th1 cytokines in concentration dependent manner and the effect of this induction should be studied further in viral models to check the efficacy of such cytokine induction.     

Interleukin-10 modulates pro-apoptotic effects of TNF-alpha in human articular chondrocytes in vitro

The aim of this study is to determine if there is an antagonistic effect between tumour necrosis factor (TNF)-α and the immunoregulatory interleukin (IL)-10 on chondrocytes survival. Serum-starved primary human articular chondrocytes were stimulated with either 10 ng/ml recombinant TNF-α, IL-10 or a combination of both (at 10 ng/ml each). Chondrocyte apoptosis was determined by measuring caspase-3/7, -8 and -9 activities using caspase assays. Mitochondrial apoptotic inducer bax, and the suppressor bcl-2 were evaluated using western blotting at 48 h. Results indicated that TNF-α increased caspase activities and resulted in a significant (p = 0.001) increase in bax/bcl-2 ratio. Stimulation with IL-10 did not alter caspase activities, while co-treatment with IL-10 and TNF-α inhibited TNF-α induced caspase activities and significantly (p > 0.004) impaired bax/bcl-2 ratio. At 24 h, mRNA levels for collagen type II, TNF-α and IL-10 were determined using real-time RT-PCR. Stimulation with TNF-α or TNF-α and IL-10 significantly inhibited collagen type II and increased IL-10 and TNF-α mRNA expression. IL-10 modulated the pro-apoptotic capacity of TNF-α in chondrocytes as shown by the decrease in caspase activities and bax/bcl-2 ratio compared to TNF-α stimulated chondrocytes, suggesting a mostly antagonistic interplay of IL-10 and TNF-α on mitochondrial apoptotic pathways.     

N-Acetylhexosamine triad in one molecule: Chemoenzymatic introduction of 2-acetamido-2-deoxy-β-d-galactopyranosyluronic acid residue into a complex oligosaccharide

A complex trisaccharide β-d-GalpNAcA-(1 → 4)-β-d-GlcpNAc-(1 → 4)-d-ManpNAc (3) was prepared in a good yield (35%) in a transglycosylation reaction catalyzed by β-N-acetylhexosaminidase from Talaromyces flavus using p-nitrophenyl 2-acetamido-2-deoxy-β-d-galacto-hexodialdo-1,5-pyranoside (1) as a donor followed by the in situ oxidation of the aldehyde functionality by NaClO₂. The disaccharide β-d-GlcpNAc-(1 → 4)-d-ManpNAc (2) was used as galactosyl acceptor. A disaccharide β-d-GalpNAcA-(1 → 4)-d-GlcpNAc (4; 39%) originated as a by-product in the reaction. Oligosaccharides comprising a carboxy moiety at C-6 are shown to be very efficient ligands to natural killer cell activation receptors, particularly to human receptor CD69. Thus, oxidized trisaccharide 3 is the best-known oligosaccharidic ligand to this receptor, with IC₅₀ = 2.5 × 10⁻⁹ M. The presented method of introducing a β-d-GalpNAcA moiety into carbohydrate structures is versatile and can be applied in the synthesis of other complex oligosaccharides.     

Optimized methodology for extraction of (1 → 3)(1 → 6)-β-d-glucan from Saccharomyces cerevisiae and in vitro evaluation of the cytotoxicity and genotoxicity of the corresponding carboxymethyl derivative

The cell wall of Saccharomyces cerevisiae is an important source of β-d-glucan, a glucose homopolymer with immunostimulant properties. The standard methodologies described for its extraction involve acid and alkaline washings, which degrade part of its glucose chains and reduce the final yield. In the present study, an optimized methodology for extraction of β-d-glucan from S. cerevisiae cells, involving sonication and enzyme treatment, with a yield of 11.08 ± 0.19%, was developed. The high-purity (1 → 3)(1 → 6)-β-d-glucan was derivatized to carboxymethyl-glucan (CM-G). In vitro tests with CM-G in Chinese hamster epithelial cells (CHO-k1) did not reveal any cytotoxic or genotoxic effects or influences of this molecule on cell viability. The method described here is a convenient alternative for the extraction of (1 → 3)(1 → 6)-β-d-glucan under mild conditions without the generation of wastes that could be potentially harmful to the environment.     

The in vivo cytokine release profile following implantation

There has been no comprehensive study that maps the production of the range of inflammatory cytokines following implantation of a material. There is an urgent requirement for specific data on the real time production of biological markers in order to study their effects in vitro and more accurately predict the in vivo response. This study determined the production of IL-1β, IL-2, IL-4, IL-6, IL-10, IFNγ and TNF-α in response to a synthetic material implanted in a rat soft tissue model for up to 90 days. IL-1 β and TNF-α were elevated over the total experimental time course with values in the order of 500 pg/ml for IL-1β and 40 pg/ml for TNF-α. The cytokines IL-2, IL-4, IL-6 and IL-10 were also detected and their production reduced with increasing time.     

SERUM CONCENTRATIONS OF sIL-2R, IL-6, TGF-β1, NEOPTERIN, AND ZINC IN CHRONIC HEPATITIS C PATIENTS TREATED WITH INTERFERON-ALPHA

T lymphocytes and immunoregulatory cytokines play an important role in the host response to hepatitis C virus (HCV) infection. Zinc is required for a wide spectrum of immune functions, including T-cell activity. To determine the clinical significance of the cytokines sIL-2R, IL-6, TGF-β1, neopterin, and of zinc in chronic heptatitis C virus (HCV) infection, we investigated their concentrations in the serum of 16 patients with chronic HCV infection before, during and at the end of therapy with interferon (IFN) α (Roferon A), and after 6 months follow-up. Elevated concentrations of sIL-2R, IL-6, TGF-β1, and neopterin were found in the serum of all patients prior to therapy, as compared to healthy controls. sIL-2R patterns differed in responders and non-responders. While the mean concentration of sIL-2R (335.75 pg/ml) before therapy was about 40% higher in complete responders (n=4) than in controls (272.20 pg/ml), the mean concentration in non-responders (n=6) was 4-fold higher than in controls (1153.33 pg/ml). During therapy, sIL-2R levels in responders decreased by about 40%. Mean IL-6 concentrations in both complete and partial responders (n=6) decreased continuously during treatment, while mean concentrations in non-responders decreased for only a short time, and increased again after cessation of therapy. Mean levels of TGF-β1 behaved similarly to those of IL-6. Only negligible differences in mean neopterin levels were found between responders and non-responders over the entire observation time. The mean serum zinc concentrations slightly decreased in all 3 patient groups, the greatest reduction occurring in 3 of the 4 responders. The present findings underscore the importance of the immune system in the pathogenesis of chronic HCV infection. Serum sIL-2R levels may be used as a serological marker of outcome following IFN-α treatment.     

Enzymatic synthesis of N-acetylglucosaminobioses by reverse hydrolysis: characterisation and application of the library of fungal β-N-acetylhexosaminidases

The regioselectivity of 20 extracellular β-N-acetylhexosaminidases of fungal origin was screened in the reverse hydrolysis with 2-acetamido-2-deoxy- -glucopyranose. Most of the enzymes used yielded 2-acetamido-2-deoxy-β- -glucopyranosyl-(1→4)-2-acetamido-2-deoxy- -glucopyranose (3) and 2-acetamido-2-deoxy-β- -glucopyranosyl-(1→6)-2-acetamido-2-deoxy- -glucopyranose (4). So far unknown product of enzymatic condensation, 2-acetamido-2-deoxy-β- -glucopyranosyl-(1→3)-2-acetamido-2-deoxy- -glucopyranose (2) was synthesised using the β-N-acetylhexosaminidases from Penicillium funiculosum CCF 1994, P. funiculosum CCF 2325 and Aspergillus tamarii CCF 1665. Addition of salts ((NH₄)₂SO₄ or MgSO₄ (0.1–1.0 M)) to the reaction increased the yields and also enhanced the β-N-acetylhexosaminidase regioselectivity.     

Molecular screening of interleukin-6 gene promoter and influence of −174G/C polymorphism on breast cancer

Interleukin-6 (IL-6) is a cytokine involved in different physiologic and pathophysiologic processes including carcinogenesis. In 2003, a single nucleotide polymorphism (−174G/C) of the IL-6 gene promoter has been linked to breast cancer prognosis in node-positive (N+) breast cancer patients. Since, different studies have led to conflicting conclusions about its role as a prognostic and/or diagnostic marker. The primary aim of our study was to investigate the link between −174G/C polymorphism and breast cancer risk on the one hand, and −174G/C polymorphism and prognosis in different groups of patients: sporadic N+ breast cancers (n = 138), sporadic N− breast cancers (n = 95) and familial breast cancer (n = 60) on the other hand. The variables of interest were disease-free survival and overall survival. The secondary aim of the study was to screen IL-6 gene promoter using direct sequencing to identify new polymorphisms in our French Caucasian breast cancer population. No association or trend of association between −174G/C polymorphism of IL-6 gene promoter gene and breast cancer diagnosis or prognosis was shown, even in meta-analyses. Furthermore, we have identified four novel polymorphic sites in the IL-6 gene promoter region: −764G → A, −757C → T, −233T → A, 15C → A.     

Properties of a novel β-glucosidase from Fusarium proliferatum ECU2042 that converts ginsenoside Rg3 into Rh2

A novel β-glucosidase from Fusarium proliferatum ECU2042 (FPG) was successfully purified to homogeneity with a 506-fold increase in specific activity. The molecular mass of the native purified enzyme (FPG) was estimated to be approximately 78.7 kDa, with two homogeneous subunits of 39.1 kDa, and the pI of this enzyme was 4.4, as measured by two-dimensional electrophoresis. The optimal activities of FPG occurred at pH 5.0 and 50 °C, respectively. The enzyme was stable at pH 4.0–6.5 and temperatures below 60 °C, and the deactivation energy (E_d) for FPG was 88.6 kJ mo1⁻¹. Moreover, it was interesting to find that although the purified enzyme exhibited a very low activity towards p-nitrophenyl β-d-glucoside (pNPG), and almost no activity towards cellobiose, a relatively high activity was observed on ginsenoside Rg₃. The enzyme hydrolyzed the 3-C, β-(1 → 2)-glucoside of ginsenoside Rg₃ to produce ginsenoside Rh₂, but did not sequentially hydrolyze the β-d-glucosidic bond of Rh₂. The K_m and V_max values of FPG for ginsenoside Rg₃ were 2.37 mM and 0.568 μmol (h mg protein)⁻¹, respectively. In addition, this enzyme also exhibited significant activities towards various alkyl glucosides, aryl glucosides and several natural glycosides.     

Antibacterial and ulcer healing effects of organoselenium compounds in naproxen induced and Helicobacter pylori infected Wistar rat model

Aim of the present study was to evaluate in vitro toxicity and in vivo antibacterial, anti-inflammatory, antiulcer, and antioxidant activities of two organoselenium compounds, selenocystine (SeCys) and ebselen (Ebs). The study was conducted in experimentally induced ulcers in rodent model infected with Helicobacter pylori (H. pylori). In vitro toxicological studies on normal spleenic lymphocytes revealed that SeCys and Ebs were non-toxic to the cells even at 100 μM concentration. Antibacterial activity was observed at 500 μg/mL concentration of either of the compounds against H. pylori. In vivo studies after treatment with SeCys and Ebs (500 μg/kg/day) resulted in significant reduction in ROS production and inhibition of lipid peroxidation in gastric tissue. The antioxidant and anti-inflammatory activities of both the compounds were also confirmed by their ability to lower GSH reduction, to induce the expression of antioxidant genes such as GPx-4, and MnSOD and to suppress inflammatory genes namely COX-2, TNF-α and TGF-β. In addition, the immunomodulatory activity of both the compounds was evident by enhance of the CD4 levels and maintenance of the IgG, IL-6 and IL-10 levels. Persistent treatment (500 μg/kg, for 28 days) with both the compounds showed considerable (p < 0.05) ulcer healing property supporting its role in gastro protection. In conclusion, the results of our study suggest that both SeCys and Ebs possess broad spectrum of activities without any potential toxicity.     

Physiological and pathophysiological applications of sensitive ELISA methods for urinary deoxycorticosterone and corticosterone in rodents

Deoxycorticosterone (DOC: a weak mineralocorticoid) is the precursor to corticosterone (B: the major glucocorticoid in rodents) and aldosterone (the major mineralocorticoid). The genes Cyp11b1 and Cyp11b2 that encode the enzymes responsible for DOC to B (11β-hydroxylase) and DOC to aldosterone (aldosterone synthase) conversions are located on the same chromosome. The aim of this study was to develop sensitive and specific ELISA methods to quantify urinary DOC and B concentrations to assess the physiological and genetic control of the Cyp11b1/b2 locus. Antibodies raised in rabbits against DOC and B and horse radish peroxidase-goat anti-rabbit IgG enzyme tracer were used to develop the assays. Urine samples collected from mice held in metabolic cages were extracted with dichloromethane and reconstituted in assay buffer. The assays were validated for specificity, sensitivity, parallelism, accuracy and imprecision. Cross-reactivities with major interfering steroids were minimal: DOC assay (progesterone = 0.735% and corticosterone = 0.045%), and for B assay (aldosterone = 0.14%, 11-dehydro-B = 0.006%, cortisol = 0.016% and DOC = 0.04%) and minimum detection limit for DOC ELISA was 2.2 pg/mL (6.6 pmol/L), and for B ELISA was 6.2 pg/mL (17.9 pmol/L). The validity of urinary DOC and B ELISAs was confirmed by the excellent correlation between the results obtained before and after solvent extraction and HPLC (DOC ELISA: Y = 1.092X − 0.054, R² = 0.988; B ELISA: Y = 1.047X − 0.226, R² = 0.996). Accuracy studies, parallelism and imprecision data were determined and all found to be satisfactory. The methods were used in a series of metabolic cage studies which demonstrated that (i) females produce more DOC and corticosterone than males; (ii) DOC and corticosterone respond to ACTH treatment but not dietary sodium restriction; (iii) DOC:B ratios in Cyp11b1 null mice were >200-fold greater than wild type.     

Enzymatic synthesis of α- -fucosyl-N-acetyllactosamines and 3′-O-α- -fucosyllactose utilizing α- -fucosidases

An α- -fucosidase from porcine liver produced α- -Fuc-(1→2)-β- -Gal-(1→4)- -GlcNAc (2′-O-α- -fucosyl-N-acetyllactosamine, 1) together with its isomers α- -Fuc-(1→3)-β- -Gal-(1→4)- -GlcNAc (2) and α- -Fuc-(1→6)-β- -Gal-(1→4)- -GlcNAc (3) through a transglycosylation reaction from p-nitrophenyl α- -fucopyranoside and β- -Gal-(1→4)- -GlcNAc. The enzyme formed the trisaccharides 1–3 in 13% overall yield based on the donor, and in the ratio of 40:37:23. In contrast, transglycosylation by Alcaligenes sp. α- -fucosidase led to the regioselective synthesis of trisaccharides containing a (1→3)-linked α- -fucosyl residue. When β- -Gal-(1→4)- -GlcNAc and lactose were acceptors, the enzyme formed regioselectively compound 2 and α- -Fuc-(1→3)-β- -Gal-(1→4)- -Glc (3′-O-α- -fucosyllactose, 4), respectively, in 54 and 34% yields, based on the donor.     

Analysis of N-methylhistamine by gas chromatography–mass spectrometry

A gas chromatography–electron capture mass spectrometry assay has been developed for the histamine H₃ receptor agonist, N^α-methylhistamine (N^α-MH). The assay is linear from 50 pg–10 ng, with a limit of detection of 50 pg/ml for gastric juice and plasma, and 50 pg/sample for bacteria (10⁷–10⁸ CFU) and gastric tissue (5–10 mg wet weight). The limits of quantification are 100 pg/ml for gastric juice (%RSD=1.4) and plasma (%RSD=9.4), and 100 pg/sample for bacteria (%RSD=3.9) and tissue (%RSD=5.8). N^α-MH was not present in human plasma, but low levels (1.4 ng/ml and 0.4 ng/ml) were detected in two samples of human gastric juice obtained from patients infected with Helicobacter pylori.     

DNA sequence analysis of spontaneous and γ-radiation (anoxic)-induced lacI mutations in Escherichia coliumuC122::Tn5: differential requirement for umuC at G · C vs. A · T sites and for the production of transversions vs. transitions

Escherihica coliumC122::Tn5 cells were γ-radiated (¹³⁷Cs, 750 Gy, under N₂), and lac-constitutive mutants were produced at 36% of the wild-type level (the umC strain was not deficient in spontaneous mutagenesis, and the mutational spectrum determined by sequencing 263 spontaneous lacI^d mutations was very similar to that for the wild-type strain). The specific nature of the umC strain's partial radiation was determined by sequencing 325 radiation-induced lacI^d mutations. The yields of radiation-induced mutation classes in the umC strain (as a percentage of the wild-type yield) were: 80% for A · T → G · C transitions, 70% for multi-base additions, 60% for single-base deletions, 53% for A · T → C · G transversions, 36% for G · C → A · T transitions, 25% for multi-base deletions, 21% for A · T → T · A transversions, 11% for G · C → C · G transversions, 9% for G · C → T · A transversions and 0% for multiple mutations. Based on these deficiencies and other factors, it is concluded that the umC strain is near-normal for A · T → G · C transitions, single-base deletions and possibly A · T → C · G transversions; is generally deficient for mutagenesis at G · C sites fro transversions, and is grossly deficient in multiple mutations. Damage at G · C sites seems more difficult for translesion DNA synthesis to bypass than damage at A · T sites, and especially when trying to produced a transversion. The yield of G · C → A · T transitions in the umC strain *36% of the wild-type level) argues that a basic sites are involved in no more than 64% of γ-radiation-induced base substitutions in the wild-type strain. Altogether, these data suggest that the UmuC and UmuD′ proteins facilitate, rather than being absolutely required for, translesion DNA synthesis; with the degree of facilitation being dependent both on the nature of the noncoding DNA damage, i.e., at G · C vs A · T sites, and on the nature of the misincorporated base, i.e., whether it induces transversions or transitions.     

β-N-Acetylhexosaminidase-catalysed synthesis of non-reducing oligosaccharides

A large panel of fungal β-N-acetylhexosaminidases was tested for the regioselectivity of the β-GlcNAc transfer onto galacto-type acceptors ( -galactose, lactose, 2-acetamido-2-deoxy- -galactopyranose). A unique, non-reducing disaccharide β- -GlcpNAc-(1→1)-β- -Galp and trisaccharides β- -GlcpNAc-(1→4)-β- -GlcpNAc-(1→1)-β- -Galp, β- -Galp-(1→4)-β- -Glcp-(1→1)-β- -GlcpNAc and β- -Galp-(1→4)-α- -Glcp-(1→1)-β- -GlcpNAc were synthesised under the catalysis of the β-N-acetylhexosaminidase from the Aspergillus flavofurcatis CCF 3061 with -galactose and lactose as acceptors. The use of 2-acetamido-2-deoxy- -galactopyranose as an acceptor with the β-N-acetylhexosaminidases from A. flavofurcatis CCF 3061, A. oryzae CCF 1066 and A. tamarii CCF 1665 afforded only β- -GlcpNAc-(1→6)- -GalpNAc.     

Enhanced proinflammatory response of mononuclear cells to in vitro LPS-challenge in patients with ventricular fibrillation in the setting of acute myocardial infarction

Aims. Ventricular fibrillation (VF) in the setting of acute myocardial infarction (AMI) is the leading cause of sudden cardiac death. A potential role of intrinsic, subclinical inflammatory states in patients suffering from ischemia-related VF has not been investigated yet. The aim of the present study was (i) to examine serum levels of proinflammatory markers in VF survivors and (ii) to evaluate basal and lipopolysaccharide (LPS)-stimulated interleukin-8-mRNA (IL-8-mRNA) levels in patients with and without VF complicating AMI. Methods. Twenty-five patients with a history of VF during AMI and a control group of 25 AMI patients without VF were included. Blood samples were taken remote from AMI with a mean of 590 days. Circulating serum levels of IL-8, IL-6, soluble E-selectin (sE-selectin), tissue factor activity (TFA), tissue inhibitor of matrix-metalloproteinase-1 (TIMP-1) and matrix-metalloproteinase-9 (MMP-9) were measured. Mononuclear cells were isolated by density gradient centrifugation. The cells were stimulated with lipopolysaccharide (LPS) from Escherichia coli (700 ng/mL). IL-8-mRNA levels in mononuclear cells were determined by a colorimetric mRNA quantification assay. Results. Serum levels (median; range) of IL-8 (VF: 2.24 pg/mL; <0.10–19.3 pg/mL versus controls: 0.10 pg/mL; <0.10–7.7 pg/mL; p = 0.014), IL-6 (VF: 0.68 pg/mL; <0.05–2.9 pg/mL versus controls: 0.23 pg/mL; <0.05–1.8 pg/mL; p = 0.042) and TIMP-1 (VF: 229 ng/mL; 144–348 ng/mL versus controls: 186 ng/mL; 126–263 ng/mL; p = 0.014) were significantly higher among patients with VF as compared to controls. Baseline IL-8-mRNA concentrations of blood mononuclear cells were significantly higher among patients with VF (257 amol/mL; 52–2672 amol/mL) as compared to patients without VF (37 amol/mL, 3.2–770 amol/mL; p < 0.01). IL-8-mRNA levels after LPS-challenge were significantly higher among patients with VF (3503 amol/mL; 215–13,573 amol/mL) than in patients without VF (1003 amol/mL; 208–3386 amol/mL; p < 0.01). Conclusions. Circulating IL-8, IL-6, and TIMP-1 concentrations as well as IL-8-mRNA expression in mononuclear cells at baseline and after LPS-challenge are increased among patients with a history of VF in the setting of AMI as compared to patients without VF. These findings indicate an enhanced inflammatory response to a proinflammatory stimulus in VF survivors. The magnitude of this increased acute phase reactants may indicate a novel pathway of arrhythmogenesis in patients with AMI.     

Enzymatic syntheses of T antigen-containing glycolipid mimicry using the transglycosylation activity of endo-α-N-acetylgalactosaminidase

Thomsen–Friedenreich antigen (T antigen) disaccharide, β- -galactose-(1→3)-α-N-acetyl- -galactosamine (β- -Gal-(1→3)-α- -GalNAc), containing glycolipid mimicry was synthesized using the transglycosylation activity of endo-α-N-acetylgalactosaminidase from Bacillus sp. This enzyme could transfer the disaccharide from a p-nitrophenyl substrate to water-soluble 1-alkanols and other alcohols at a transfer ratio of 70% or more. Although the transfer ratios were lower for water-insoluble than water-soluble alcohols, they were shown to increase by adding sodium cholate to the reaction mixtures. The enzyme also transferred the disaccharide directly from asialofetuin to 1-alkanols. The anomeric bond between the disaccharide and 1-alkanols of the transglycosylation product is in the α configuration as determined by sequential digestion of jack bean β-galactosidase and Acremonium α-N-acetylgalactosaminidase. Since the transglycosylation product, β- -Gal-(1→3)-α- -GalNAc-(1→O)-hexyl, efficiently inhibits the binding of anti-T antigen monoclonal antibody to asialofetuin, it has potential as an agent for blocking T antigen-mediated cancer metastasis.     

New cell wall glycopolymers of the representatives of the genus Kribbella

Each of the cell walls of four representatives of the genus Kribbella (order Actinomycetales; suborder Propionibacterineae; family Nocardioidaceae) contains a neutral polysaccharide and an acidic polysaccharide with unusual structures. Common to all four strains studied is a mannan with the following repeating unit: In the cell wall of the strain VKM Ac-2541, a teichulosonic acid was identified with a monosaccharide component that has not hitherto been found in Gram-positive bacteria, viz., pseudaminic acid, and an unusual linkage type in the polymeric chain,

Full-size image (1K)

where R = Н (45%), α-d-Galp3OMe (37%) or α-d-Galp2,3OMe (18%).The anionic cell wall components of three other strains are represented by teichuronic acids with a rare constituent, viz., a diaminosugar, 2,3-diacetamido-2,3-dideoxyglucopyranose. The structures of their repeating units differ in the nature of the acidic components:→4)-β-d-Manp2,3NAcA-(1→6)-α-d-Glcp2,3NAc-(1→ (VKM Ас-2538 and VKM Ас-2540) and →4)-β-d-ManpNAcA-(1→6)-α-d-Glcp2,3NAc-(1→ (VKM Ас-2539).The structures of all the glycopolymers were established by chemical and NMR spectroscopic methods; they are identified in Gram-positive bacteria for the first time.