Screening for active tuberculosis in high-risk groups.

OBJECTIVE: To evaluate an active case-finding strategy among drug users (DUs), economically disadvantaged individuals and recent immigrants from hyperendemic countries, a population at high risk of developing tuberculosis (TB). METHODS: Retrospective, observational study carried out by the Tuberculosis Unit of the City of Barcelona from September 2009 to December 2012. All participants underwent chest X-ray and were screened for symptoms. RESULTS: Of 5982 participants screened, 30 TB cases were detected (total prevalence 0.5%): 26 were pulmonary, 8 were smear-positive and 2 were resistant to multiple drugs. Directly observed treatment was advised for 19 patients (63%). TB prevalence in the recent immigrants group was significantly greater (1.77%) than in all other groups studied: economically disadvantaged individuals 0.30% (RR 5.9, 95%CI 2.30-15.14); DUs 0.62% (RR 2.05, 95%CI 0.91-4.64), non-recent immigrants 0.41% (RR 4.31, 95%CI 1.68-11.4); and all native-born individuals 0.41% (RR 4.33, 95%CI 1.71-10.92). The rate was much greater than the estimated prevalence for the general population of the city (∼20 cases/100,000 population). CONCLUSIONS: In high-risk groups, active case finding can be used as a public health intervention to detect a large number of TB cases.

Rifampicin plus isoniazid for the prevention of tuberculosis in an immigrant population.

OBJECTIVES: To compare the tolerance, adherence and effectiveness of two approaches for the treatment of latent tuberculosis infection (LTBI): 6 months of isoniazid (6H) vs. 3 months of isoniazid plus rifampicin (3RH). POPULATION: Immigrants with LTBI. METHODS: Participants were enrolled in a controlled, randomised clinical trial in Barcelona, Spain, from April 2001 to April 2005. Monthly follow-up was done to assess tolerance, side effects and adherence. Effectiveness was evaluated at 5 years. RESULTS: In the 590 subjects enrolled, the rate of adherence was greater in the 3RH than in the 6H arm (72% vs. 52.4%, P = 0.001). No differences between study arms were observed with respect to hepatotoxicity or side effects. Variables associated with non-adherence were diagnosis by screening (OR 1.88, 95%CI 1.26-2.82, P = 0.001), illegal immigration status (OR 1.48, 95%CI 1.01-2.15, P = 0.03), unemployment (OR 1.91, 95%CI 1.28-2.85, P = 0.0008), illiteracy (OR 1.73, 95%CI 1.04-2.88, P = 0.02), lack of family support (OR 3.7, 95%CI 2.54-5.4, P = 0.001) and the 6-month treatment regimen (OR 2.45, 95%CI 1.68-3.57, P = 0.0001). None of the patients who completed either treatment developed tuberculosis. CONCLUSIONS: The 3RH regimen facilitates adherence to LTBI treatment and offers a safe, well-tolerated and effective alternative.

[Information for patients about hospital infections in psychiatry: An assessment of healthcare professionals' knowledge, opinion and attitude]. / Connaissances, opinions et pratiques des professionnels concernant l'information des patients sur les infections nosocomiales : évaluation en milieu psychiatrique.

INTRODUCTION: French legislation makes mandatory for healthcare providers the disclosure of hospital infection (HI) risk and actual occurrence to the patient. Given the specific diseases encountered in psychiatry, some difficulties may be expected in practical application of this regulation. OBJECTIVES: The aim of our study was to describe the knowledge, declared practices and opinions of healthcare workers (HCW) in psychiatry concerning information for patients about HI. METHODS: We randomly selected doctors, nurses and head nurses from four hospitals with psychiatric activity in Normandy. The HCW were asked to self-complete an anonymous questionnaire, including data describing the responding HCW and questions aiming at describing his/her knowledge, attitude in routine daily practice and opinion about information to patients about HI. RESULTS: One hundred and forty-one HCW were initially selected, of which 114 (80.9%) eventually agreed to complete the questionnaire. Only eight HCW (7.0%) were considered to have a correct overall knowledge of legal obligations. Main errors concerned the obligation to inform the patient of the HI risk according to the medical procedures that are to be performed (43.9% of correct answers) and the obligation to inform the patient of the HI risk according to his/her medical condition (46.5%). The obligation to inform the patient of the occurrence of a HI was largely known (84.2%). HCW usually giving information about the risk of HI to patients without HI accounted for 5.3%. Main reasons advocated for not informing patients were a low level risk of HI in psychiatry (80.4%) and the lack of patients' demand (59.8%). In the case of HI occurrence, the percentage of HCW routinely informing patients was 13.2%. HCW systematically informing the patient's family about the occurrence of HI accounted for 9.6%. A large proportion of HCW supported delivering information to patients about HI (86.0%). HCW expected from information better approval of prevention programs by the patients (87.7%) but feared an increased anxiety in patients (75.4%) and a higher rate of care refusal (48.2%). CONCLUSION: Whereas a very large proportion of HCW in psychiatry support delivering information to patients about HI, our study shows HCW's lack of awareness of regulations and lack of declared practices. Among factors explaining this contrast, a lower perceived HI risk and severity level are to be mentioned. Training programs focusing on risk and mechanisms of HI could be offered to professionals in psychiatry. The issue of specific communication difficulties with psychiatric patients should be addressed as well. In order to develop information on HI, specific methods suited to those patients should be developed.

Protease-activated receptor-4 (PAR 4): a role as inhibitor of visceral pain and hypersensitivity.

Protease-activated receptor-4 (PAR(4)) belongs to the family of receptors activated by the proteolytic cleavage of their extracellular N-terminal domain and the subsequent binding of the newly released N-terminus. While largely expressed in the colon, the role of PAR(4) in gut functions has not been defined. We have investigated the effects of PAR(4) agonist on colonic sensations and sensory neuron signalling, and its role in visceral pain. We observed that a single administration of the PAR(4) agonist peptide (AYPGKF-NH(2)), but not the control peptide (YAPGKF-NH(2)) into the colon lumen of mice significantly reduced the visceromotor response to colorectal distension at different pressures of distension. Further, intracolonic administration of the PAR(4) agonist, but not the control peptide, was able to significantly inhibit PAR(2) agonist- and transcient receptor potential vanilloid-4 (TRPV4) agonist-induced allodynia and hyperalgesia in response to colorectal distension. Protease-activated receptor-4 was detected in sensory neurons projecting from the colon, and isolated from the dorsal root ganglia, where it co-expressed with PAR(2) and TRPV4. In total sensory neurons, PAR(4) agonist exposure inhibited free intracellular calcium mobilization induced by the pro-nociceptive agonists of PAR(2) and TRPV4. Finally, PAR(4)-deficient mice experienced increased pain behaviour in response to intracolonic administration of mustard oil, compared with wild-type littermates. These results show that PAR(4) agonists modulate colonic nociceptive response, inhibit colonic hypersensitivity and primary afferent responses to pro-nociceptive mediators. Endogenous activation of PAR(4) also plays a major role in controlling visceral pain. These results identify PAR(4) as a previously unknown modulator of visceral nociception.

[Active tuberculosis case finding among immigrants in Barcelona]. / Búsqueda activa de tuberculosis en inmigrantes en Barcelona.

OBJECTIVE: To assess the prevalence of tuberculous infection and disease in recent economic immigrants in Barcelona. SUBJECTS AND METHOD: Examination and testing of immigrants. Tuberculin tests (TTs) were given and the presence of scars from tuberculosis vaccinations were noted. Thresholds of 5 and 15 mm were established for positivity in unvaccinated and vaccinated individuals, respectively. RESULTS: A total of 3651 persons were examined, but only 3151 completed the study. Eighteen were diagnosed with tuberculosis (571.2 per 100,000) and 50.6% were classified as positive TT reactors, 34.4% because of infection and 16.3% possibly because of tuberculosis vaccination. The percentage of reactors was significantly higher in the sample of economic immigrants than in the local population. Age, male sex, place of origin, greater poverty, and higher prevalence of disease in the country of origin were associated with tuberculous infection in the sample. DISCUSSION: Active case finding proved efficient. Interference from tuberculosis vaccination greatly affects the findings, depending on the positivity threshold that is established. We recommend that chest radiographs be used in addition to TTs. Immigration will change the nature of endemic tuberculosis in Spain, and strategies should be specifically designed to deal with the new challenges that will appear.

Estudio de la infección tuberculosa en adultos / A study of the tuberculous infection in adults

La infección tuberculosa en adultos no es muy conocida en nuestro país a pesar del interés epidemiológico que tiene. Se diseñó un estudio observacional en un grupo homogéneo de adultos funcionarios de Cataluña (n = 8.202) de 20 a 54 años de edad que se sometieron a un examen de salud, en el que se incluyó una prueba de tuberculina, con objeto de estudiar la infección tuberculosa en ellos y evaluar los factores relacionados con la misma. Resultados. La prevalencia global a la reactividad de la prueba de la tuberculina es del 22,36 por ciento y la prevalencia de la infección tuberculosa del 14,76 por ciento. Los factores relacionados con la infección tuberculosa son: edad, sexo masculino, antecedentes de exposición a fuentes de contagio y vacunación antituberculosa (BCG) previa. Conclusiones. La prevalencia de la infección tuberculosa en adultos ha descendido, siendo mayor en hombres que en mujeres, entre los que reconocen haber tenido contacto con un enfermo tuberculoso y entre los que fueron vacunados con BCG (AU)

[A study of the tuberculous infection in adults]. / Estudio de la infección tuberculosa en adultos.

UNLABELLED: Tuberculous infection in adults is not a well known entity in our country, despite its epidemiological importance. We have designed an observational study in a homogeneous group of adult civil servants of Catalonia (n = 8,202) from 20 to 54 years old that were submitted to a health examination which included a tuberculin test, in order to study the tuberculous infection in these people and to evaluate the factors associated with this infection. RESULTS: The global prevalence of reactivity in tuberculin test was 22.36% and the prevalence of the tuberculous infection was 14.76%. The factors related to the tuberculous infection were the following: age, male sex, background of exposure to sources of contagion, and previous BCG vaccination. CONCLUSIONS: The prevalence of tuberculous infection in adults has declined, and is currently greater in men than women, among patients who recognize previous contact with a tuberculous patient, and among patients with previous BCG vaccination.

[Pulmonary symptomatic tuberculosis' diagnostic delay study]. / Estudio del retraso diagnóstico de la tuberculosis pulmonar sintomática.

OBJECTIVE: To study symptomatic pulmonary tuberculosis (PTB) diagnostic delay. PATIENTS AND METHODS: Prospective study of new symptomatic PTB cases (aged > or = 15 years) by structured interview with the patients and their families. The main variables analyzed were patient's delay (PD), doctor's delay (DD), diagnostic process delay (DPD), health care system delay (HCSD) and total delay between the onset of symptoms and start of treatment (TD). Univariate and multivariate statistical analyses were performed for each component of delay. RESULTS: Two hundred eighty-seven patients were studied. The mean delays in days standard deviations were TD 81.8 77.3; PD 43.3 55.7; DD 28.4 59.6; DPD 10.0 17.7, and HCSD 38.5 62.5. CONCLUSIONS: Patients are responsible for 50% of excess delay in diagnosing symptomatic PTB. Patients in the health care system experienced diagnostic delays over 60 days in 18.5% of cases, doctors being responsible for 75% of the diagnostic delay attributable to the system.

Estudio del retraso diagnóstico de la tuberculosis pulmonar sintomática / Pulmonary symptomatic tuberculosis' diagnostic delay study

OBJETIVO: Estudiar el retraso diagnóstico de la tuberculosis pulmonar (TBP) sintomática. PACIENTES Y MÉTODOS: Estudio prospectivo de casos nuevos sintomáticos de TBP (edad 15 años) mediante entrevista estructurada al paciente y su familia. Las variables fundamentales analizadas fueron: retraso del enfermo (RE), retraso atribuible al médico (RM), retraso durante el proceso diagnóstico (RPD), retraso en el sistema sanitario (RSS) y retraso total (RT), esto es, el tiempo transcurrido desde el comienzo de los síntomas hasta el inicio del tratamiento de la TBP. Se realizó un análisis estadístico univariante, así como análisis multivariante para cada uno de los componentes del retraso diagnóstico. RESULTADOS: Se estudió a 287 enfermos. La media en días ñ desviación estándar (DE) fue para el RT y sus distintos componentes de 81,8 ñ 77,3 en el RT, 43,3 ñ 55,7 días en el RE, de 28,4 ñ 59,6 días en el RM; para el RD, de 10,0 ñ 17,7 días y en el RSS de 38,5 ñ 62,5 días. CONCLUSIONES: Dentro del elevado retraso diagnóstico de la TBP los enfermos son responsables del 50 por ciento. En el sistema sanitario el 18,5 por ciento de los enfermos sufrió un retraso diagnóstico mayor de 60 días, siendo los médicos responsables del 75 por ciento de la demora atribuible al sistema (AU)

Human immunodeficiency virus-infected patients receiving highly active antiretroviral therapy maintain activated CD8+ T cell subsets as a strong adaptive immune response to cytomegalovirus.

CD8(+) T lymphocyte function specific for human cytomegalovirus (CMV) was evaluated in 14 patients infected with human immunodeficiency virus (HIV) receiving highly active antiretroviral therapy (HAART) and 26 CMV-seropositive donors without HIV infection. Fifty-seven percent of the HIV-infected group had CMV-specific cytolytic activity in freshly isolated peripheral blood mononuclear cells (PBMC) against targets expressing CMV pp65. Both interferon (IFN)-gamma secretion by CD8(+) T cells and the frequency of human leukocyte antigen (HLA)-tetramer-positive T cells in HLA-A*0201-positive HIV-infected subjects correlated with CMV-specific cytolysis. In contrast, PBMC from healthy CMV-seropositive donors did not have either measurable CMV-specific cytolysis or secretion of IFN-gamma without in vitro stimulation. The T helper response to CMV antigens was vigorous in healthy CMV-seropositive donors but low in the cohort of HIV-infected patients. Potent CD8(+) cytotoxic T lymphocyte responses to CMV in HIV-infected patients receiving HAART is the converse of what is found in healthy CMV-seropositive subjects and may be the predominant adaptive immune response against CMV in HIV-infected patients.

Exploring the acceptor substrate recognition of the human beta-galactoside alpha 2,6-sialyltransferase.

Human beta1,4-galactoside alpha2,6-sialyltransferase I (ST6GalI) recognition of glycoprotein acceptors has been investigated using various soluble forms of the enzyme deleted to a variable extent in the N-terminal half of the polypeptide. Full-length and truncated forms of the enzyme have been investigated with respect to their specificity for a variety of desialylated glycoproteins of known complex glycans as well as related proteins with different carbohydrate chains. Differences in transfer efficiency have been observed between membrane and soluble enzymatic forms, indicating that deletion of the transmembrane fragment induces loss of acceptor preference. No difference in substrate recognition could be observed when soluble enzymes of similar peptide sequence were produced in yeast or mammalian cells, confirming that removal of the membrane anchor and heterologous expression do not alter enzyme folding and activity. When tested on free oligosaccharides, soluble ST6GalI displayed full ability to sialylate free N-glycans as well as various N-acetyllactosaminyl substrates. Progressive truncation of the N terminus demonstrated that the catalytic domain can proceed with sialic acid transfer with increased efficiency until 80 amino acids are deleted. Fusion of the ST6GalI catalytic domain to the N-terminal half of an unrelated transferase (core 2 beta1,6-N-acetylglucosaminyltransferase) further showed that a chimeric form of broad acceptor specificity and high activity could also be engineered in vivo. These findings therefore delineate a peptide region of approximately 50 amino acids within the ST6GalI stem region that governs both the preference for glycoprotein acceptors and catalytic activity, thereby suggesting that it may exert a steric control on the catalytic domain.

Cloning, expression and gene organization of a human Neu5Ac alpha 2-3Gal beta 1-3GalNAc alpha 2,6-sialyltransferase: hST6GalNAcIV.

On the basis of the detection of expressed sequence tag ('EST') similar to the rat N-acetylgalactosamine alpha2,6-sialyltransferase (ST6GalNAc) III cDNA, we have identified a novel member of the human ST6GalNAc family. We have isolated a cDNA clone containing an open reading frame that codes for a type II membrane protein of 302 amino acids with a seven-amino-acid cytoplasmic domain, an 18-amino-acid transmembrane domain and the smallest described catalytic domain of 277 amino acids. This predicted sialyltransferase sequence is similar to the rat ST6GalNAc III (46.6%), but was found to be even more similar to the recently reported mouse ST6GalNAc IV (88.1%) on the basis of amino acid sequence identity. Northern-blot analysis showed that the newly identified gene is expressed constitutively in various adult human tissues as a 2.2kb transcript, but was also found to be expressed at lower levels in brain, heart and skeletal muscle as a 2.5kb transcript. Expression of the hST6GalNAc IV gene was investigated by reverse transcription PCR in various human cancer cells, and was found to be present in the majority of cell types with the exception of the carcinoma cell line T47D and pro-monocyte THP cells. The transient expression in COS-7 cells of the full-length cDNA led to the production of an active enzyme sharing the acceptor specificity of the ST6GalNAc family towards Neu5Ac alpha 2-3Gal beta 1-3GalNAc alpha-O-R (where 'R' denotes H, benzyl, or a peptidic chain). Detailed analysis in vitro of substrate specificity revealed that the enzyme required the trisaccharide Neu5Ac alpha 2-3Gal beta1-3GalNAc found on O-glycans and arylglycosides. In addition, we have clarified the genomic organization of ST6GalNAc IV gene.

The acceptor and site specificity of alpha 3-fucosyltransferase V. High reactivity of the proximal and low of the distal galbeta 1-4GlcNAc unit in i-type polylactosamines.

We report here on in vitro acceptor and site specificity of recombinant alpha3-fucosyltransferase V (Fuc-TV) with 40 oligosaccharide acceptors. Galbeta1-4GlcNAc (LN) and GalNAcbeta1-4GlcNAc (LDN) reacted rapidly; Galbeta1-3GlcNAc (LNB) reacted moderately, and GlcNAcbeta1-4GlcNAc (N, N'-diacetyl-chitobiose) reacted slowly yet distinctly. In neutral and terminally alpha3-sialylated polylactosamines of i-type, the reducing end LN unit reacted rapidly and the distal (sialyl)LN group very slowly; the midchain LNs revealed intermediate reactivities. The data suggest that a distal LN neighbor enhances but a proximal LN neighbor reduces the reactivity of the midchain LNs. This implies that Fuc-TV may bind preferably the tetrasaccharide sequence Galbeta1-4GlcNAcbeta1-3Galbeta1-4GlcNAc for transfer at the underlined monosaccharide. Terminal alpha3-sialylation of i-type polylactosamines almost doubled the reactivities of the LN units at all positions of the chains. We conclude that, in comparison with human Fuc-TIV and Fuc-TIX, Fuc-TV reacted with a highly distinct site specificity with i-type polylactosamines. The Fuc-TV reactivity of free LNB resembled that of LNBbeta1-3'R of a polylactosamine, contrasting strongly with the dissimilarity of the reactivities of the analogous pair of LN and LNbeta1-3'R. This observation supports the notion that LN and LNB may be functionally bound at distinct sites on Fuc-TV surface. Our data show that Fuc-TV worked well with a very wide range of LN-glycans, showing weak reactivity only with distal (sialyl)LN units of i-type polylactosamines, biantennary N-glycans, and I branches of polylactosamines.

Induction of CTL response by a minimal epitope vaccine in HLA A*0201/DR1 transgenic mice: dependence on HLA class II restricted T(H) response.

CTL play a pivotal role in the immune response during viral infections. In this study, the HLA class II restricted T(H) requirement for optimal in vivo induction of HLA class I restricted CTL responses has been investigated. Towards this goal, transgenic mice expressing both HLA class I (A*0201 or A2.1) and class II (DRB1*0101 or DR1) molecules have been derived. Immunization of these mice with an HLA A*0201-restricted and CMV-specific CTL epitope (pp65(495-503)), and either of three different tetanus toxin-derived MHC class II-binding T(H) epitopes, resulted in a vigorous CTL response. CTL specific for the pp65(495-503) epitope were dramatically enhanced in mice expressing both the HLA-DR1 and HLA-A*0201 transgenes. Notably, preinjection of three TT peptides (TT(639-652), TT(830-843), and TT(947-967)) increased the capability of HLA A*0201/DR1 Tg mice to respond to subsequent immunization with the T(H) + CTL peptide mixture. These results indicate that the use of HLA A*0201/DR1 Tg mice constitute a versatile model system (in lieu of immunizing humans) for the study of both HLA class I and class II restricted T-cell responses. These studies provide a rational model for the design and assessment of new minimal-epitope vaccines based on their in vivo induction of a pathogen-specific CTL response.

[Oligosaccharide sulfate inhibitors of selectin-sugar interactions in inflammatory processes]. / Oligosaccharides sulfatés inhibiteurs des interactions sélectines-sucres intervenant dans les processus inflammatoires.

Leucocyte migration into lymphatic tissues or inflammatory sites depends upon the expression of adhesion molecules. Among these molecules, the selectins expressed on endothelial cells (E- and P-selectins) and leucocytes (L-selectin) recognize carbohydrate ligands such as sialyl Lewis A or sialyl Lewis X oligosaccharides due to the same positioning of NeuAc, Gal and Fuc residues in both isomeric structures. We have shown that the sialic acid residue could be replaced by a sulfate group such as in the sulfated Lewis A pentasaccharide, one of the most potent monovalent ligand for human E-selectin, which was shown to be very active in the prevention of ischemia reperfusion lung injury. In the same way, we have prepared through chemoenzymatic syntheses, two disulfated Lewis X pentasaccharides, the sulfated analogs of carbohydrate ligands found on GLYCAM 1, the natural receptor of L-selectin. Finally, based on the double recognition of L-selectin with Lewis type and glycosaminoglycan structures, we tentatively introduced a possible link between the selectin- and the integrin-mediated lymphocyte adhesion systems.

Porcine liver (2-->3)-alpha-sialyltransferase: substrate specificity studies and application of the immobilized enzyme to the synthesis of various sialylated oligosaccharide sequences.

In search of substrate analogues for the porcine liver beta-D-Gal p-(1-->3)-D-Gal p-NAc: CMP-Neu5Ac-(2-->3')-alpha-sialyltransferase, three disaccharides beta-D-Gal p-(1-->3)-beta-D-Gal p-O-CH3 (5), beta-D-Gal p-(1-->3)-beta-D-(2-OAc)-Gal p-O-CH3 (7) and beta-D-Gal p-(1-->3)-beta-D-(2-OAc)-Gal p-O-Bn (11) were synthesized and tested with the enzyme. Disaccharide 7 turned out to be a very good substrate allowing a rapid access to the trisaccharide alpha-Neu5Ac-(2-->3)-beta-D-Gal p-(1-->3)-beta-D-(2-OAc)-Gal p-O-CH3 (13) on a preparative scale using the crude enzyme immobilized on cationic exchanger. Trisaccharide 13 was further exploited, first as a sialyl donor in Trypanosoma cruzi trans-sialidase catalyzed reaction and second through acetolysis reaction as a source for the synthon alpha-Neu5Ac-(2-->3)-D-Gal (16).

Chemoenzymatic synthesis of a 3IV,6III-disulfated Lewis(x) pentasaccharide, a candidate ligand for human L-selectin.

The disulfated pentasaccharide 3-O-SO3(-)-beta-D-Galp-(1-->4)-[alpha-L-Fucp-(1-->3)]-6-O-SO3(-)- beta-D-GlcpNAc-(1-->3)-beta-D-Galp-(1-->4)-D-Glcp was prepared according to a chemoenzymatic approach, starting from 4-methoxybenzyl O-(4-O-acetyl-2,6-di-O-benzyl-beta- D-galactopyranosyl)-(1-->4)-O-2,3,6-tri-O-benzyl-beta-D-glucopyranoside, obtained in six steps from hepta-O-acetyl lactosyl bromide. Coupling of this lactose derivative with O-(3,4,6-tri-O-acetyl-2-deoxy-2-phthalimido-beta-D-glucopyranosyl) trichloracetimidate afforded, after dephthaloylation and re-N-acetylation, 4-methoxybenzyl O-(2-acetamido-2-deoxy-beta-D- glucopyranosyl)-(1-->3)-O-(2,6-di-O-benzyl-beta-D-galactopyranosyl)-(1-- >4)- O-2,3,6-tri-O-benzyl-beta-D-glucopyranoside. Regioselective sulfation at the primary position of the glucosamine residue was then successfully achieved and the benzyl groups were removed. Enzymatic galactosylation of 4-methoxybenzyl O-(2-acetamido-2-deoxy-6-O-sulfo-beta-D- glucopyranosyl)-(1-->3)-O-beta-D-galactopyranosyl-(1-->4)-O-beta-D- glucopyranoside sodium salt, and subsequent regioselective sulfation at position 3 of the outer galactose residue through the stannylene procedure, led then to 4-methoxybenzyl O-(3-sulfo-beta-D- galactopyranosyl)-(1-->4)-O-(2-acetamido-2-deoxy-6-sulfo-beta-D- glucopyranosyl)-(1-->3)-O-beta-D-galactopyranosyl)-(1-->4)-O-beta-D- glucopyranoside disodium salt, which was finally fucosylated using human milk alpha-(1-->3/4)-fucosyltransferase affording, after anomeric deprotection, the target pentasaccharide.

Ergot alkaloid glycosides with immunomodulatory activities.

New glycosides derived from ergot alkaloids elymoclavine and DH-lysergol were synthesized by chemoenzymatic methods. beta-Glucosides were obtained either by chemical method or by transglycosylation (glycosidase from Aspergillus oryzae), lactosides were prepared by further extension of carbohydrate chain using beta-1,4-galactosyltransferase (bovine milk) and alpha-5-N-acetylneuraminyl-(2-->6)-beta-D-galactopyranosyl-(l-->4)-2- acetamido-2-deoxy-beta-D-glucopyranosyl-(1-->O)-elymoclavine was prepared using alpha-2,6-sialyltransferase (rat liver). Immunomodulatory activity of elymoclavine and 9,10-dihydrolysergol and their glycosylated derivatives on natural killer (NK) cell-mediated cytotoxicity of human resting and activated human peripheral blood mononuclear cells (PBMC) was investigated. Addition of ergot alkaloid glycosides to the mixtures of effector and target cells potentiated the PBMC cytotoxicity against both NK-sensitive and -resistant target cells. The glycoconjugates of elymoclavine enhanced cytotoxicity of PBMC against NK-resistant target cells. The glycoconjugates of DH-lysergol potentiated NK cytotoxicity of PBMC against NK-sensitive target cells.

Combined chemical and enzymatic synthesis of the sialylated non reducing terminal sequence of GM1b glycolylated ganglioside, a potential human tumor marker.

N-Glycolylglucosamine 8 was synthesized in 4 steps from anisal glucosamine, via the new crystalline monochloracetyl derivatives 3, 4 and 7. N-Glycolylneuraminic acid 10 was prepared in 59% yield starting from pyruvate and a mixture of 8 and its manno epimer 9 in a 2:3 ratio, with immobilized sialic acid aldolase. Neu5Gc 10 was converted into CMP-NeuGc 11 in the presence of immobilized calf brain CMP-sialate synthetase. Finally 11 was used as a donor in the transfer to the acceptor beta-D-Gal-(1-3)-beta-D-GalNAc-OBn 12 catalyzed by a preparation of porcine liver (2-3)-alpha-sialyltransferase, roughly purified by a chromatography on Cibacron Blue-agarose. alpha-Neu5Gc-(2-3)-beta-D-Gal-(1-3)-beta-D-GalNac-OBn 13 isolated in 56% yield was deprotected to give the non-reducing terminal sequence of GM1b glycolylated ganglioside, which might be expressed in human tumors.