Schistosomulum-released products (SRP) have been shown to enhance both expression of rat and huma... more Schistosomulum-released products (SRP) have been shown to enhance both expression of rat and human eosinophil Fc receptors and IgG-dependent cytotoxicity. The present work provides additional evidence of the secretion of eosinophil-enhancing factors by schistosomula and other developmental stages of schistosomes, including adult worms. The heat lability, as well as the strong inhibition of the stimulating activity of SRP by the protease inhibitor Trasylol, suggest that thermolabile proteases secreted by the parasite are involved in this mechanism. The purification of the schistosome proteases by preparative isoelectric focusing and gel filtration demonstrated that neutral proteases able to hydrolyze the collagenase substrates Azocoll and Z-Gly-Pro-Leu-Gly-Pro are able to significantly enhance eosinophil effector functions. Purified Clostridium histolyticum collagenase was also able to mimic the enhancing effect of schistosome proteases, suggesting involvement of a collagenase-like a...
To know if alkaline phosphatase (AP) from schistosomes other than Schistosoma mansoni can be used... more To know if alkaline phosphatase (AP) from schistosomes other than Schistosoma mansoni can be used as diagnostic marker for schistosomiasis in alkaline phosphatase immunocapture assay (APIA), we comparatively tested n-butanol extracts of adult worm membranes from a Venezuelan (JL) strain of S. mansoni (Ven/AWBE/Sm); a Cameroonian (EDEN) strain of Schistosoma intercalatum (Cam/AWBE/Si) and a Yemeni strain of Schistosoma haematobium (Yem/AWBE/Sh). APIA was evaluated with sera of patients from Venezuela, Senegal, and Gabon infected with S. mansoni, from Gabon infected with S. intercalatum or S. haematobium, from Chine infected with Schistosoma japonicum and from Cambodian patients infected with Schistosoma mekongi. Results indicate that 92.5% (37/40) of Venezuela sera, 75% (15/20) of Senegal sera, 39.5% (17/43) of S. haematobium sera, and 19.2% (5/26) S. intercalatum sera were APIA-positive with the Ven/AWBE/Sm preparation. APIA with the Cam/AWBE/Si preparation showed that 53.8% of S. intercalatum-positive sera had anti-AP antibodies, and 51.2% S. haematobium-positive sera cross-immunocapturing the S. intercalatum AP. APIA performed with Yem/AWBE/Sh showed that 55.8% S. haematobium sera were positive. Only two out of nine S. japonicum sera were APIA-positive with the Ven/AWBE/Sm and Cam/AWBE/Si, and no reaction was observed with Cambodian S. mekongi-positive sera. AP activity was shown to be present in all the schistosome species/strains studied. The use of APIA as a tool to explore the APs antigenicity and the presence of Schistosoma sp. infections through the detection of anti-Schistosoma sp. AP antibodies in a host, allowed us to demonstrate the antigenicity of APs of S. mansoni, S. intercalatum, and S. haematobium.
Schistosoma mansoni cathepsin B (Sm31) is a major antigen from adult worms that circulates in the... more Schistosoma mansoni cathepsin B (Sm31) is a major antigen from adult worms that circulates in the blood of infected patients (Li et al., Parasitol Res 1996; 82: 14-18). An analysis of the Sm31 sequence (Klinkert et al., Mol Biochem Parasitol 1989; 33: 113-122) allowed the prediction of seven hydrophilic regions that were confirmed to be exposed on the surface of a 3D model of Sm31; the species specificity of these regions was checked using BLAST analysis. The corresponding peptides were chemically synthesized in polymerazible forms using the t-Boc technique. Rabbits developed a high humoral response against these peptides as tested by a multiple antigen blot assay; it recognized native Sm31 in crude S. mansoni extracts and as circulating antigen in sera of S. mansoni-infected patients by western blot. Relevant antigenic determinants were located at the N- and C-terminus sequences. Antibodies against these regions recognized the native enzyme in an ELISA-like assay called cysteine protease immuno assay in which the immunocaptured enzyme was revealed by the intrinsic cathepsin B hydrolytic activity of Sm31. The method successfully and specifically detected Sm31 in sera of infected individuals, most of them (83.3%) with light infections, offering a rationale for the development of parasite enzyme capture assays using anti-synthetic peptide antibodies for possible use in the diagnosis of schistoso,iasis.
1<p>Average n° of adult worms (males and females) recovered in the perfusion fluid of liver... more 1<p>Average n° of adult worms (males and females) recovered in the perfusion fluid of liver and small mesenteric veins from each experimental group (Mean ± SEM, <i>n</i> = 10).</p><p>The reduction of the parasitic load was calculated according to formula: % Protection = (1−B/A)×100 <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002254#pntd.0002254-Shuxian1" target="_blank">[31]</a> where A is the average n° of worms in the AWBE-immunized group and, B the average n° of worms in the adjuvant (ADJ) inoculated group (Control). AW: Adult Worms; AWBE: Adult Worm <i>n</i>-Butanol Extract; ADJ: adjuvant inoculated mice; B: mice injected with buffer. Significance by <i>t</i> student test (compared to group of ADJ-inoculated animals):</p>*<p>significant (<i>p</i>≤0.05).</p
... FROM DIFFERENT ANIMALS BY THE S. manSOm AGGLUTININ Erythrocyte source Titer NMRI/T/IVIC Mice ... more ... FROM DIFFERENT ANIMALS BY THE S. manSOm AGGLUTININ Erythrocyte source Titer NMRI/T/IVIC Mice A Mice AKR Mice BALB/c ... Acknowledgements The author wishes to thank Miss Maria Elena Garcia for the excellent technical assistance and Mr. Erick Fernandez and ...
The S. mansoni adult worm n-butanol extract (Sm-AWBE) has been previously shown to contain specif... more The S. mansoni adult worm n-butanol extract (Sm-AWBE) has been previously shown to contain specific S. mansoni antigens that have been used for immunodiagnosis of schistosomiasis in solid phase alkaline phosphatase immunoassay (APIA) and western blot (WB) analyses. Sm-AWBE was also used in immunoprotection studies against a fatal live-cercariae challenge in experimental mouse vaccination (~43% protection). The Sm-AWBE fraction was prepared by mixing adult worm membranous suspensions with aqueous-saturated n-butanol, centrifuging and recovering n-butanol-resistant proteins in the aqueous phase. Here we report a preliminary identification of Sm-AWBE protein components as revealed from a qualitative proteomic study after processing Sm-AWBE by 1D-gel electrophoresis, in-gel and in-solution tryptic digestions, and mass spectrometry analyses. We identified 33 proteins in Sm-AWBE, all previously known S. mansoni proteins and antigens; among them, immunomodulatory proteins and proteins most...
A study of the subcellular localization of the nicotinamide adenine dinucleotide (NADH)-3-(4, 3-d... more A study of the subcellular localization of the nicotinamide adenine dinucleotide (NADH)-3-(4, 3-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) oxido-reductase systems in Mycobacterium was presented. Evidence based on starch gel electrophoresis and different responses of the subcellular fractions to heat inactivation suggested the existence of more than one enzyme responsible for the NADH-MTT oxido-reductase activity. One type of activity was found in the membrane-mesosome fraction which contained labile and electrophoretically non-migrating enzymes. Another type of activity was also detected in the soluble fraction which on starch gel electrophoresis exhibited 4 bands of activity, two of which showed heat resistance.
1. Incubation of Schistosoma mansoni for 5 min in a phosphate-buffered medium, pH 7.4, released t... more 1. Incubation of Schistosoma mansoni for 5 min in a phosphate-buffered medium, pH 7.4, released tegumental material containing the following phosphohydrolase activities: alkaline phosphatase, 5′-nucleotidase, glycerol-2-phosphatase, glucose 6-phosphatase, phosphodiesterase and ATPase. 2. Maximum activity of these enzymes was measured at pH 9.5; however, the phosphodiesterase and ATPase activities were also appreciable at pH 7.0. 3. Solubilization of the released tegumental material in 1% Triton X-100 followed by gel filtration distinguished three peaks of enzyme activity: an ATPase (mol.wt. greater than 1000 000), a phosphodiesterase (mol.wt. 1 000 000) and an alkaline phosphomonoesterase with broad specificity (mol.wt. 232 000). 4. The ATPase activity was highly activated by 10 mM-Mg2+ or 1 mM-Ca2+ and was inhibited by chelating agents. Ouabain, Na+ and K+ had little effect on enzyme activity, whereas activity was increased by 50% in the presence of calmodulin. The phosphodiesteras...
Proceedings of the National Academy of Sciences, 1973
Amino-acid sequences of the variable regions of three lambda chains produced by plasmacytomas of ... more Amino-acid sequences of the variable regions of three lambda chains produced by plasmacytomas of BALB/c mice are compared. Two are almost certainly identical and one differs from these by three amino acids. These findings extend our earlier conclusion on the relative uniformity of sequences in this type of immunoglobulin light chain. With amino-acid sequence data on two additional lambda chains, eight mouse lambda chains studied to date are indistinguishable and four probably differ from these by one, two, or three amino acids.
An aminopeptidase activity capable of hydrolysing leucine 4-nitroanilide and alanine 4-nitroanili... more An aminopeptidase activity capable of hydrolysing leucine 4-nitroanilide and alanine 4-nitroanilide at pH 7.0 was detected in saponin-CaCl2 extracts and homogenates of adult Schistosoma mansoni. The extracts were also capable of acting on synthetic dipeptides at the same pH, preferentially hydrolysing peptide bonds following leucine, alanine, or proline N-terminal residues. Imide bonds were not hydrolysed. The hydrolysis of leucine 4-nitroanilide was apparently stimulated by thiols, strongly inhibited by 1 mM 4-chloromercuric benzene sulfonic acid, and partially inhibited by 1 mM 1,10-phenanthroline.
Control de la esquistosomiasis en Venezuela ha sido un tema de gran interés y controversia entre ... more Control de la esquistosomiasis en Venezuela ha sido un tema de gran interés y controversia entre las parasitosis metaxénicas. Una pequeña área de transmisión de aproximadamente 15.000 km2 fue pensado para ser erradicada hace unos años. Sin embargo, algunas ...
Methyl jasmonate (MJ), an oxylipid that induces defense-related mechanisms in plants, has been sh... more Methyl jasmonate (MJ), an oxylipid that induces defense-related mechanisms in plants, has been shown to be active against cancer cells bothin vitroandin vivo, without affecting normal cells. Here we review most of the described MJ activities in an attempt to get an integrated view and better understanding of its multifaceted modes of action. MJ (1) arrests cell cycle, inhibiting cell growth and proliferation, (2) causes cell death through the intrinsic/extrinsic proapoptotic, p53-independent apoptotic, and nonapoptotic (necrosis) pathways, (3) detaches hexokinase from the voltage-dependent anion channel, dissociating glycolytic and mitochondrial functions, decreasing the mitochondrial membrane potential, favoring cytochromecrelease and ATP depletion, activating pro-apoptotic, and inactivating antiapoptotic proteins, (4) induces reactive oxygen species mediated responses, (5) stimulates MAPK-stress signaling and redifferentiation in leukemia cells, (6) inhibits overexpressed proinfla...
Schistosomulum-released products (SRP) have been shown to enhance both expression of rat and huma... more Schistosomulum-released products (SRP) have been shown to enhance both expression of rat and human eosinophil Fc receptors and IgG-dependent cytotoxicity. The present work provides additional evidence of the secretion of eosinophil-enhancing factors by schistosomula and other developmental stages of schistosomes, including adult worms. The heat lability, as well as the strong inhibition of the stimulating activity of SRP by the protease inhibitor Trasylol, suggest that thermolabile proteases secreted by the parasite are involved in this mechanism. The purification of the schistosome proteases by preparative isoelectric focusing and gel filtration demonstrated that neutral proteases able to hydrolyze the collagenase substrates Azocoll and Z-Gly-Pro-Leu-Gly-Pro are able to significantly enhance eosinophil effector functions. Purified Clostridium histolyticum collagenase was also able to mimic the enhancing effect of schistosome proteases, suggesting involvement of a collagenase-like a...
To know if alkaline phosphatase (AP) from schistosomes other than Schistosoma mansoni can be used... more To know if alkaline phosphatase (AP) from schistosomes other than Schistosoma mansoni can be used as diagnostic marker for schistosomiasis in alkaline phosphatase immunocapture assay (APIA), we comparatively tested n-butanol extracts of adult worm membranes from a Venezuelan (JL) strain of S. mansoni (Ven/AWBE/Sm); a Cameroonian (EDEN) strain of Schistosoma intercalatum (Cam/AWBE/Si) and a Yemeni strain of Schistosoma haematobium (Yem/AWBE/Sh). APIA was evaluated with sera of patients from Venezuela, Senegal, and Gabon infected with S. mansoni, from Gabon infected with S. intercalatum or S. haematobium, from Chine infected with Schistosoma japonicum and from Cambodian patients infected with Schistosoma mekongi. Results indicate that 92.5% (37/40) of Venezuela sera, 75% (15/20) of Senegal sera, 39.5% (17/43) of S. haematobium sera, and 19.2% (5/26) S. intercalatum sera were APIA-positive with the Ven/AWBE/Sm preparation. APIA with the Cam/AWBE/Si preparation showed that 53.8% of S. intercalatum-positive sera had anti-AP antibodies, and 51.2% S. haematobium-positive sera cross-immunocapturing the S. intercalatum AP. APIA performed with Yem/AWBE/Sh showed that 55.8% S. haematobium sera were positive. Only two out of nine S. japonicum sera were APIA-positive with the Ven/AWBE/Sm and Cam/AWBE/Si, and no reaction was observed with Cambodian S. mekongi-positive sera. AP activity was shown to be present in all the schistosome species/strains studied. The use of APIA as a tool to explore the APs antigenicity and the presence of Schistosoma sp. infections through the detection of anti-Schistosoma sp. AP antibodies in a host, allowed us to demonstrate the antigenicity of APs of S. mansoni, S. intercalatum, and S. haematobium.
Schistosoma mansoni cathepsin B (Sm31) is a major antigen from adult worms that circulates in the... more Schistosoma mansoni cathepsin B (Sm31) is a major antigen from adult worms that circulates in the blood of infected patients (Li et al., Parasitol Res 1996; 82: 14-18). An analysis of the Sm31 sequence (Klinkert et al., Mol Biochem Parasitol 1989; 33: 113-122) allowed the prediction of seven hydrophilic regions that were confirmed to be exposed on the surface of a 3D model of Sm31; the species specificity of these regions was checked using BLAST analysis. The corresponding peptides were chemically synthesized in polymerazible forms using the t-Boc technique. Rabbits developed a high humoral response against these peptides as tested by a multiple antigen blot assay; it recognized native Sm31 in crude S. mansoni extracts and as circulating antigen in sera of S. mansoni-infected patients by western blot. Relevant antigenic determinants were located at the N- and C-terminus sequences. Antibodies against these regions recognized the native enzyme in an ELISA-like assay called cysteine protease immuno assay in which the immunocaptured enzyme was revealed by the intrinsic cathepsin B hydrolytic activity of Sm31. The method successfully and specifically detected Sm31 in sera of infected individuals, most of them (83.3%) with light infections, offering a rationale for the development of parasite enzyme capture assays using anti-synthetic peptide antibodies for possible use in the diagnosis of schistoso,iasis.
1<p>Average n° of adult worms (males and females) recovered in the perfusion fluid of liver... more 1<p>Average n° of adult worms (males and females) recovered in the perfusion fluid of liver and small mesenteric veins from each experimental group (Mean ± SEM, <i>n</i> = 10).</p><p>The reduction of the parasitic load was calculated according to formula: % Protection = (1−B/A)×100 <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0002254#pntd.0002254-Shuxian1" target="_blank">[31]</a> where A is the average n° of worms in the AWBE-immunized group and, B the average n° of worms in the adjuvant (ADJ) inoculated group (Control). AW: Adult Worms; AWBE: Adult Worm <i>n</i>-Butanol Extract; ADJ: adjuvant inoculated mice; B: mice injected with buffer. Significance by <i>t</i> student test (compared to group of ADJ-inoculated animals):</p>*<p>significant (<i>p</i>≤0.05).</p
... FROM DIFFERENT ANIMALS BY THE S. manSOm AGGLUTININ Erythrocyte source Titer NMRI/T/IVIC Mice ... more ... FROM DIFFERENT ANIMALS BY THE S. manSOm AGGLUTININ Erythrocyte source Titer NMRI/T/IVIC Mice A Mice AKR Mice BALB/c ... Acknowledgements The author wishes to thank Miss Maria Elena Garcia for the excellent technical assistance and Mr. Erick Fernandez and ...
The S. mansoni adult worm n-butanol extract (Sm-AWBE) has been previously shown to contain specif... more The S. mansoni adult worm n-butanol extract (Sm-AWBE) has been previously shown to contain specific S. mansoni antigens that have been used for immunodiagnosis of schistosomiasis in solid phase alkaline phosphatase immunoassay (APIA) and western blot (WB) analyses. Sm-AWBE was also used in immunoprotection studies against a fatal live-cercariae challenge in experimental mouse vaccination (~43% protection). The Sm-AWBE fraction was prepared by mixing adult worm membranous suspensions with aqueous-saturated n-butanol, centrifuging and recovering n-butanol-resistant proteins in the aqueous phase. Here we report a preliminary identification of Sm-AWBE protein components as revealed from a qualitative proteomic study after processing Sm-AWBE by 1D-gel electrophoresis, in-gel and in-solution tryptic digestions, and mass spectrometry analyses. We identified 33 proteins in Sm-AWBE, all previously known S. mansoni proteins and antigens; among them, immunomodulatory proteins and proteins most...
A study of the subcellular localization of the nicotinamide adenine dinucleotide (NADH)-3-(4, 3-d... more A study of the subcellular localization of the nicotinamide adenine dinucleotide (NADH)-3-(4, 3-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT) oxido-reductase systems in Mycobacterium was presented. Evidence based on starch gel electrophoresis and different responses of the subcellular fractions to heat inactivation suggested the existence of more than one enzyme responsible for the NADH-MTT oxido-reductase activity. One type of activity was found in the membrane-mesosome fraction which contained labile and electrophoretically non-migrating enzymes. Another type of activity was also detected in the soluble fraction which on starch gel electrophoresis exhibited 4 bands of activity, two of which showed heat resistance.
1. Incubation of Schistosoma mansoni for 5 min in a phosphate-buffered medium, pH 7.4, released t... more 1. Incubation of Schistosoma mansoni for 5 min in a phosphate-buffered medium, pH 7.4, released tegumental material containing the following phosphohydrolase activities: alkaline phosphatase, 5′-nucleotidase, glycerol-2-phosphatase, glucose 6-phosphatase, phosphodiesterase and ATPase. 2. Maximum activity of these enzymes was measured at pH 9.5; however, the phosphodiesterase and ATPase activities were also appreciable at pH 7.0. 3. Solubilization of the released tegumental material in 1% Triton X-100 followed by gel filtration distinguished three peaks of enzyme activity: an ATPase (mol.wt. greater than 1000 000), a phosphodiesterase (mol.wt. 1 000 000) and an alkaline phosphomonoesterase with broad specificity (mol.wt. 232 000). 4. The ATPase activity was highly activated by 10 mM-Mg2+ or 1 mM-Ca2+ and was inhibited by chelating agents. Ouabain, Na+ and K+ had little effect on enzyme activity, whereas activity was increased by 50% in the presence of calmodulin. The phosphodiesteras...
Proceedings of the National Academy of Sciences, 1973
Amino-acid sequences of the variable regions of three lambda chains produced by plasmacytomas of ... more Amino-acid sequences of the variable regions of three lambda chains produced by plasmacytomas of BALB/c mice are compared. Two are almost certainly identical and one differs from these by three amino acids. These findings extend our earlier conclusion on the relative uniformity of sequences in this type of immunoglobulin light chain. With amino-acid sequence data on two additional lambda chains, eight mouse lambda chains studied to date are indistinguishable and four probably differ from these by one, two, or three amino acids.
An aminopeptidase activity capable of hydrolysing leucine 4-nitroanilide and alanine 4-nitroanili... more An aminopeptidase activity capable of hydrolysing leucine 4-nitroanilide and alanine 4-nitroanilide at pH 7.0 was detected in saponin-CaCl2 extracts and homogenates of adult Schistosoma mansoni. The extracts were also capable of acting on synthetic dipeptides at the same pH, preferentially hydrolysing peptide bonds following leucine, alanine, or proline N-terminal residues. Imide bonds were not hydrolysed. The hydrolysis of leucine 4-nitroanilide was apparently stimulated by thiols, strongly inhibited by 1 mM 4-chloromercuric benzene sulfonic acid, and partially inhibited by 1 mM 1,10-phenanthroline.
Control de la esquistosomiasis en Venezuela ha sido un tema de gran interés y controversia entre ... more Control de la esquistosomiasis en Venezuela ha sido un tema de gran interés y controversia entre las parasitosis metaxénicas. Una pequeña área de transmisión de aproximadamente 15.000 km2 fue pensado para ser erradicada hace unos años. Sin embargo, algunas ...
Methyl jasmonate (MJ), an oxylipid that induces defense-related mechanisms in plants, has been sh... more Methyl jasmonate (MJ), an oxylipid that induces defense-related mechanisms in plants, has been shown to be active against cancer cells bothin vitroandin vivo, without affecting normal cells. Here we review most of the described MJ activities in an attempt to get an integrated view and better understanding of its multifaceted modes of action. MJ (1) arrests cell cycle, inhibiting cell growth and proliferation, (2) causes cell death through the intrinsic/extrinsic proapoptotic, p53-independent apoptotic, and nonapoptotic (necrosis) pathways, (3) detaches hexokinase from the voltage-dependent anion channel, dissociating glycolytic and mitochondrial functions, decreasing the mitochondrial membrane potential, favoring cytochromecrelease and ATP depletion, activating pro-apoptotic, and inactivating antiapoptotic proteins, (4) induces reactive oxygen species mediated responses, (5) stimulates MAPK-stress signaling and redifferentiation in leukemia cells, (6) inhibits overexpressed proinfla...
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