Biomedical subjects

J L Molinari

Publications and source records attributed to J L Molinari.

At least 19 recordsLinked to original sources

PubMed2001

A factor isolated from Taenia solium metacestodes stimulates T lymphocytes to proliferate and produce gamma interferon.

A metacestode factor (MF) isolated from live metacestodes of Taenia solium suppresses humoral and cellular responses, and inhibits the inflammatory reaction around metacestodes implanted subcutaneously in mice. When this MF is digested with RNase (dMF), it loses the suppressive capacity, but acquires T-cell stimulant ability. By filtering MF through a Bio-gel P6 column, two components were separated. The first (F1) was suppressive. while the second (F2) stimulated T cells to proliferate. In these experiments, F2 or dMF was used with mouse spleen cells in stimulation assays in vitro. Spleen cells from mice treated with F2 or dMF were also stimulated with concanavalin A (Con-A) ex vivo. Flow cytometry analyses were performed to estimate cell proliferation, intracellular cytokine production. and restoration of CD4 cells. Spleen lymphocytes from mice previously treated with F2 or dMF and then stimulated with Con-A ex vivo exhibited a significant increase in cell proliferation and gamma interferon production by CD4+ (P<0.05) and CD8+ cells. These effects were concentration-dependent and inversely correlated with the amount of dMF or F2. Similar results were observed in normal mouse spleen T cells incubated with F2 or dMF and Con-A in vitro. Finally, dMF induced a significant restoration of CD4-cells in mice depleted of these cells.

Animals↗

PubMed2000

Taenia solium: a cysteine protease secreted by metacestodes depletes human CD4 lymphocytes in vitro.

Excreted/secreted products from Taenia solium metacestodes cultured in vitro were analyzed for peptidase activity using peptide substrates Z-Phe-Arg-AFC, Arg-AFC, and Z-Gly-Gly-Arg-AFC and zymography studies. Specific inhibitor profiles revealed mainly cysteine and metalloprotease activities. Hydrolysis of substrate Z-Phe-Arg-AFC was augmented by the addition of L-cysteine and acid pH, consistent with cysteine protease activity. Cysteine protease activity was more prominent in supernatants from living metacestodes cultured in PBS than in either RPMI or RPMI plus fetal calf serum and was proportional to the number of metacestodes. Flow cytometry analysis showed depletion of human T lymphocytes cultured with living T. solium metacestodes. CD4(+) expression was significantly decreased when metacestode E/S products and L-cysteine were added to lymphocyte cultures (P = 0.027). This peptidase activity was inhibited by E-64 indicating that the depletion of CD4(+) cells was due to cysteine protease activity. Thus, T. solium metacestodes produce excretory/secretory proteases. These enzymes may cleave molecules critical for the host immune response allowing the parasites to survive in the host tissues.

Animals↗

PubMed1998

Depressed immunity to a Salmonella typhimurium vaccine in mice experimentally parasitized by Taenia crassiceps.

To assess the immunological status of mice parasitized with Taenia crassiceps metacestodes, 6-month old female BALB/c mice experimentally parasitized with T. crassiceps and immunized with Salmonella typhimurium antigens were infected with S. typhimurium virulent bacilli (1.6 x LD50). Both T. crassiceps-parasitized and immunized and parasitized mice showed a very high susceptibility to infection (**P < 0.01) with higher bacteremia than control and immunized-control animals and produced a reduced IgG response to S. typhimurium, antigens (* P < 0.05). This indicates that T. crassiceps is able to preclude development of immunity to S. typhimurium, because appropriate antibody production to a heterologous antigenic stimulus did not take place, and the bacteremia results suggest the parasitosis altered the mononuclear phagocyte system. It has been demonstrated that Taenia solium metacestodes produce a small RNA molecule in culture which suppresses humoral and cellular responses against homologous antigens in mice. We propose that T. crassiceps may be actively synthesizing such a factor, apart from other simultaneously acting immunomodulatory mechanisms, to induce an immunosuppressed state favorable to its development in the host.

Animals↗

PubMed1998

A Taenia solium metacestode factor nonspecifically inhibits cytokine production.

Studies of the immune response in chronic helminth infections suggest that parasites modulate the host's immune response. Taenia solium metacestodes, in particular, produce molecules that down-regulate cell-mediated immunity. We have described a small RNA peptide termed metacestode factor (MF) that depresses the murine immune response to Salmonella typhimurium antigens. MF inhibits mitogen-induced proliferation, humoral and cellular responses to metacestode antigens, and inflammation surrounding metacestodes implanted subcutaneously in mice. To assess the effects of MF on cytokine production we stimulated murine spleen cells in vitro with concanavalin A and measured cytokine concentrations in the culture supernatants by enzyme-linked immunosorbent assay. When cultured with MF, the cells showed significantly decreased production of interleukin 2 (IL-2), interferon-gamma (IFN-gamma), and IL-4 as compared with mitogen alone. Exogenous rIL-2 and rIL-4 largely restored the proliferative response (85% and 71% of control cells, respectively). MF also decreased production of tumor necrosis factor-alpha (TNF-alpha) by macrophages stimulated with lipopolysaccharide and IFN-gamma. The TNF-alpha concentration was inversely correlated with the MF concentration. Experiments using spleen cells from mice treated with MF also showed a significant reduction in IL-4 concentration. These results suggest that MF inhibits cytokine production without regard to cell type or cytokine. This may explain the function of this molecule as an inhibitor of the host inflammatory and immune responses.

Animals↗

PubMed1998

Impairment of the inflammatory reaction on implanted Taenia solium metacestodes in mice by a T. solium RNA-peptide: a scanning electron microscopy study.

Inhibition of inflammation by a Taenia solium RNA-peptide (metacestode factor, MF) was studied by scanning electron microscopy (SEM). Viable (96%) T. solium metacestodes obtained from a naturally infected pig were dissected and implanted in treated and control mice, removed at 6 and 12 days postimplantation (p.i.), and studied by SEM. At day 6, metacestodes in control mice showed vigorous inflammation, whereas in mice treated with MF they were apparently intact with exiguous inflammation. Mice immunized with T. solium metacestode antigens showed a moderate inflammation; those treated with both MF and T. solium antigens presented scanty inflammation. At day 12, metacestodes presented copious inflammation and severe damage to the sucker tissues in mice immunized with T. solium; in mice treated with either MF or MF and T. solium antigens there was only discrete inflammation. These observations illustrate the central role of MF in the inhibition of the early events leading to the parasite's destruction by means of an inflammatory response.

Animals↗

PubMed1997

Field trial for reducing porcine Taenia solium cysticercosis in Mexico by systematic vaccination of pigs.

It has previously been demonstrated that immunization of pigs with a crude extract of Taenia solium metacestodes can confer a high level of protection against an egg challenge. Furthermore, vaccination of infected animals also induces an immune response against the larvae, which are either destroyed or rendered non-infectious. To assess the efficacy of immunization as a strategy for reducing the prevalence of porcine cysticercosis, a field trial of this vaccine was performed in an endemic area in the northern region of the Guerrero State, Mexico, Random samples of pigs belonging to 17 villages were examined for metacestodes by inspection of their tongues. Each animal was immunized with a dose of 150 micrograms of protein (antigenic extract from Taenia solium metacestodes) by the intramuscular route. A prevalence of 2.4% of porcine cysticercosis on average was found in these villages at the beginning of the trial (62 cysticercotic pigs out of 2650 inspected). Six of these villages were selected for the periodic vaccination of new random samples of pigs. A statistically significant decline in the prevalence of porcine cysticercosis was observed at the end of the trial, decreasing from 2.4% at the beginning of vaccination to 0.45% at the end of the trial. A reduction of 82% was observed in spite of the poor living conditions in these villages. These results are consistent with previous data and suggest that it may be possible to turn a susceptible pig population into a protected one by systematic vaccination.

Animals↗

PubMed1996

Immunosuppression and inhibition of inflammation in mice induced by a small Taenia solium RNA-peptide to implanted T. solium metacestodes.

Subcutaneous implantation of Taenia solium metacestodes in mice induces an inflammatory reaction made up mainly of neutrophils and eosinophils after 12 days. Administration of a small RNA-peptide (metacestode factor, MF) purified from T. solium metacestodes significantly reduces the inflammatory site in both size and composition, yielding a very low number of eosinophils. The metacestodes implanted in control mice were completely destroyed and their remnants were surrounded by an intense inflammation predominantly made up of neutrophils and eosinophils. In contrast, metacestodes implanted in mice treated with MF showed apparently intact suckers, rostellum, hooks, and tegument. Inhibition of inflammation around the parasites was also observed in mice immunized with T. solium metacestode antigens and inoculated simultaneously with MF. Mice immunized only with T. solium metacestode antigens produced a granulomatous process around metacestodes that destroyed most of the large metacestode structures: suckers, rostellum, hooks, and tegument-wall tissues. Furthermore, treatment of mice with MF or implanted metacestodes decreased the antibody (P < 0.05) and cellular responses (P < 0.05) to metacestode antigens. The antibody responses was even lower when both of these treatments were given simultaneously. These findings support the idea that MF plays a key role in the down-regulation of the host immune response, contributing to the parasite's survival.

Animals↗

PubMed1995

Suppression of murine lymphocyte proliferation induced by a small RNA purified from the Taenia solium metacestode.

A substance from Taenia solium metacestodes that decreases lymphocyte proliferation induced by concanavalin A was isolated. The molecular weight of this substance was estimated to be slightly more than 1,450 Da. Crude metacestode factor was fractionated through a Bio-gel P-6 column. Peak 1 showed suppressive activity. After incubation with RNase the substance lost its activity. Incubation of this material with trypsin or papain increased its suppressive activity. It was stable at boiling temperature for 10 min. The incubation of this substance with murine macrophages had no effect on [3H]-thymidine uptake by cocultured fresh splenic lymphocytes stimulated with concanavalin A. Conversely, cocultures of lymphocytes pretreated with the substance and fresh splenic lymphocytes showed a decreased incorporation of [3H]-thymidine. These results suggest that this substance is a RNA-peptide molecule whose RNA moiety accounts for its suppressive activity. The findings also suggest that in vivo the factor may be a modulator of the immune response.

Animals↗

PubMed1994

Immune response impairment, genotoxicity and morphological transformation induced by Taenia solium metacestode.

In chronic helminthic infections such as cysticercosis, where the parasites live for years, profound modulation of the host immune response has been reported. To evaluate the genotoxicity of a drug used to treat cysticercosis, we observed the occurrence of genetic damage in cultured lymphocytes from cysticercotic swine and patients who had not been exposed to the drug. The human lymphocytes also showed a slower proliferation. These data suggested that the disease itself was promoting genetic damage in host lymphocytes which, in part, could explain the retardation of the lymphocyte proliferation observed in cysticercotic patients. Pigs infected with Taenia solium cysticerci showed an increased lymphocyte proliferation for 6-8 weeks post infection, followed by an impaired proliferation after this period. Significant induction of sister-chromatid exchanges was also observed in lymphocytes from infected pigs after the 6th week post infection. Additionally, it was found that a factor secreted by the cysticerci morphologically transformed primary fibroblasts in culture. The results strongly suggest that the parasite produces genetic instability in the host cells, which could result in immunosuppression and malignant transformation of target cells.

Animals↗

PubMed1993

Immunization against porcine cysticercosis in an endemic area in Mexico: a field and laboratory study.

An antigenic extract from Taenia solium metacestodes was evaluated for immunogenicity in pig populations from a large area of endemic porcine cysticercosis in the State of Guerrero, Mexico. A total of 3,295 pigs from 18 villages were immunized with a single dose of 250 micrograms of protein administered intramuscularly. Systematic immunization was also performed on pigs (1,076 immunizations) from two of the villages with the highest percentages of cysticercosis. A year after immunization, porcine cysticercosis decreased from 4.8% and 5.4% to 0%. Immunity against the T. solium metacestode was estimated in vitro by measurements of 3H-thymidine uptake and inhibition of leukocyte migration. Peripheral blood lymphocytes from immunized cysticercotic (pigs that had cysticercosis prior to immunization), cysticercotic immunized (pigs that acquired cysticercosis after immunization), and normal control pigs incorporated 3H-thymidine better than lymphocytes from cysticercotic pigs when stimulated with concanavalin A. A significant inhibition in the leukocyte migration inhibition test was also found in leukocytes from immunized cysticercotic pigs (P < 0.01). Histopathologic studies revealed granuloma formation surrounding the metacestodes of the immunized cysticercotic and cysticercotic immunized pigs. These metacestodes exhibited several stages of destruction. Large numbers of eosinophils were frequently observed in a close association with the degeneration and destruction of parasites. Metacestodes in control cysticercotic pigs were intact and surrounded by a minor inflammatory reaction. Finally, the rate of in vitro evagination of scolices was high in metacestodes obtained from cysticercotic pigs and low or absent in those from immunized pigs (P < 0.01).

Animals↗

PubMed1993

Effects of serum from neurocysticercosis patients on the structure and viability of Taenia solium oncospheres.

Neurocysticercosis, caused by Taenia solium, is arguably the most common parasitic disease of the central nervous system. In taeniid infections of nonhuman mammals, there is strong evidence of immunity in the intermediate host to the invasive larvae (oncospheres). This immunity, which is mediated by antibody and complement, has been exploited to develop vaccines that effectively prevent infection. To examine the immune response in humans, T. solium eggs were hatched and activated in vitro. Activated oncospheres were incubated with heat-inactivated sera from patients with neurocysticercosis with or without complement (guinea pig serum). Controls included oncospheres plus complement alone, normal human serum alone, normal serum with complement, or buffer alone. Serum from infected patients, especially with complement, markedly reduced oncosphere mobility and led to disappearance of secretory vesicles and loss of membrane integrity. Viability as assessed by staining with dimethyl-thiazolyl-diphenyl-tetrazolium was reduced from 92.5% in controls to 61.5% with immune serum and 38.8% with immune serum and complement (P < 0.01). Preliminary western blot analysis showed antigens at 22, 64, and 70 kDa recognized by all 3 sera, but not by control sera. These data suggest that sera from patients with cysticercosis can kill oncospheres in vitro and may be used to identify protective antigens.

Animals↗

PubMed1992

Host-parasite interactions in Taenia solium cysticercosis.

Human neurocysticercosis results from infestation of the central nervous system with the metacestode form (tissue cyst) of Taenia solium. Cysticercosis is being increasingly recognized as a cause of neurologic symptoms in residents and emigrants from developing countries. Taeniid parasites have developed elaborate mechanisms to persist in the tissues of their intermediate hosts. The invasive larvae, termed oncospheres, are susceptible to antibody and complement. However, by the time that the host has generated an antibody response, the parasites have begun to transform to the more resistant metacestode form. The metacestodes also have means of evading complement-mediated destruction, including paramyosin, which inhibits C1q; taeniaestatin, which inhibits both classical and alternate pathways (likely by inhibiting factor D and C3 esterase); and sulfated polysaccharides, which activate complement away from the parasite. Similarly, antibody does not seem to be able to kill the mature metacestode. The parasites may even stimulate the host to produce antibody, which could be bound via Fc receptors, and used as a source of protein. Finally, taeniaestatin and other parasite molecules may interfere with lymphocyte proliferation and macrophage function, thus paralyzing the cellular immune response. Because the symptoms of neurocysticercosis are typically associated with a brisk inflammatory response, we hypothesize that disease is primarily the result of injured or dying parasites. This hypothesis raises important questions in assessing the role of chemotherapy in the management of neurocysticercosis as well as in evaluation of clinical trials, most of which have been uncontrolled.

Animals↗

PubMed1992

Twinning in metacestodes of Taenia solium.

Two cysticerci containing 2 scolices were found among several thousand Taenia solium metacestodes dissected from swine. Microscopic study of tissue sections revealed that both worms were equally well developed in 1 bladder worm, whereas 1 member of the other pair was incompletely formed.

Animals↗

PubMed1992

Detection and preliminary characterization of Taenia solium metacestode proteases.

The metacestode of Taenia solium persists for years in the human central nervous system. As proteolytic enzymes play an important role in the survival of tissues helminths, we examined extracts of T. solium metacestodes for proteolytic activity using 9 synthetic peptide substrates and 3 proteins (hemoglobin, albumin, and immunoglobulin G). The proteolytic enzymes were classified based on their inhibitor profiles. At neutral pH, aminopeptidase(arginine-7-amino-4-trifluoromethylcoumarin) and endopeptidase(benzyloxy-carbonyl-glycine-glycine-arginine-7-amino-4- trifluoromethylcoumarin) substrates were cleaved. Hydrolysis of both substrates was inhibited by chelating agents, which inhibit metalloproteases. Peak activity with both substrates eluted in gel filtration fractions corresponding to a molecular weight of about 104 kDa. Cysteine protease activity was identified, which cleaved benzyloxy-carbonyl-phenylalanine-arginine-7-amino- 4-trifluoromethylcoumarin (Z-Phe-Arg-AFC) and hemoglobin. Cleavage of Z-Phe-Arg-AFC was maximal at acid pH, was stimulated by thiols, and was inhibited by leupeptin and Ep459. Peak cysteine protease activity eluted in gel filtration fractions corresponding to a molecular weight of 32 kDa. Aspartic protease activity was identified by specific inhibition with pepstatin of acid digestion of hemoglobin and immunoglobulin G. Immunoglobulin digestion occurred at acid pH, with preferential degradation of the heavy chain. Upon gel filtration chromatography, the aspartic protease activity eluted as a broad peak with maximal activity at about 90 kDa. No serine protease activity was detected. None of the parasite enzymes digested albumin. Proteolytic enzymes of T. solium may be important for parasite survival in the intermediate host, by providing nutrients and digesting host immune molecules.

Amino Acid Sequence↗

PubMed1990

Depressive effect of a Taenia solium cysticercus factor on cultured human lymphocytes stimulated with phytohaemagglutinin.

Cysticercosis caused by Taenia solium is associated with immunodepression of T and B lymphocytes. In order to ascertain if this parasite affects lymphocyte activity, a factor isolated from the parasite was tested on (3H) thymidine uptake by cultured human lymphocytes stimulated by phytohaemagglutinin. This dialysable factor had a molecular weight of less than 3500 Da, and was isolated from an extract of Cysticercus cellulosae. It decreased phytohaemagglutinin-stimulated uptake of (3H) thymidine. After the material was treated with RNase 'A', the suppressive activity was destroyed. It thus appears that the factor could correspond to an RNA fraction.

Animals↗

PubMed1983

Taenia solium: immunity in hogs to the Cysticercus.

Protection was induced in hogs against Taenia solium cysticercosis using an immunogenic complex obtained from its larval "bladder worm" form, Cysticercus cellulosae. Immunoelectrophoresis revealed that this complex contained at least eight antigens. In immunized hogs a total of 71 (mean 11.8) cysticerci were found, whereas in the control animals 397 (mean 74.9) were found. Histopathological studies showed that more than 40% of larvae obtained from immunized hogs were completely destroyed and the others were seen in various stages of degeneration. Eosinophils and mononuclear cells were observed infiltrating the internal structures of the larvae. Intense granulomatous reactions of eosinophils, lymphocytes, macrophages, epithelioid cells, plasma cells, and fibroblasts surrounded the larvae. Larvae from control hogs were intact and surrounded by a small inflammatory reaction. The cellular response was measured by the macrophage migration inhibition test, which was higher in immunized hogs when compared with control animals, either before the infection with T. solium eggs or before slaughter. No significant difference was found in the humoral response of immunized and control hogs.

Animals↗

PubMed1983

Taenia solium: cell reactions to the larva (Cysticercus cellulosae) in naturally parasitized, immunized hogs.

In hogs naturally infected with Taenia solium larvae (i.e., Cysticercus cellulosae), we studied the host response induced by antigens obtained from the larvae. Histopathological studies of cysticerci removed after 4 and 8 weeks of immunization showed an intense inflammatory reaction surrounding the larvae. The response was greater in the 8-week specimens. A dense layer of eosinophils was in close contact with the external membrane of the bladder wall and, in several cases, the eosinophils had infiltrated this tegument. Many eosinophils were seen in the spiral canal of larvae. This infiltration by eosinophils increased with time. Preparations from the 8-week samples showed many degenerated and disrupted eosinophils whose granules were found in close contact with the outer membrane of the larval tegument and, in some cases, had entered through the broken surface of this structure. More than 90% of the larvae were found in various stages of degeneration; the rest were completely destroyed and surrounded by a mass of eosinophils. After immunization, peripheral blood eosinophilia increased to 17%, whereas the eosinophilia of the control hog was 4% throughout the study. The larval worms removed from control hogs showed intact structures, with a low degree of infiltration by eosinophils and a discrete inflammatory reaction surrounding the bladder wall of the larvae.

Animals↗

PubMed1979

Immunogenic complexes obtained from Salmonella typhi-murium and Salmonella typhi Ty2 by the bacterial acetone powder method.

An immunogenic complex was isolated from Salmonella typhi-murium and another one from Salmonella typhi Ty2. Both were prepared by the bacterial acetone powder method which eliminated the cell wall, the DNA almost completely and the membrane phospholipids. The complexes were denominated "New Vaccines". The S. typhi-murium new vaccine induced, even at doses of 0.5 microgram dry weight per mouse, a high degree of protection against the challenge of the virulent microorganism. By immunoelectrophoresis, 21 antigen-antibody systems could be detected, two of them corresponding to O antigens. The S. typhi Ty2 new vaccine induced better protection than the standard vaccine (heat-phenol inactivated typhoid vaccine) when both vaccines were compared in the relative potency test. Moreover, the new vaccine had very low toxicity when inoculated in humans at doses of 1 microgram dry weight, able to elicite a high antibody titre (1/1,790 mean of 10 sera) in 75% of the tested population, estimated by the complement fixation test. In contrast, the standard vaccine induced a low antibody titre (1/222, mean of 5 sera) in 50% of the humans inoculated with 1 X 10(8) bacterial cells. The new vaccine did not induce undesirable effects whereas the standard vaccine induced an important inflammatory process in 100% of the cases, with intense local pain in 67% after 24 h post-first inoculation as well as other less severe symptoms.

Acetone↗