Biomedical subjects

C Boulard

Publications and source records attributed to C Boulard.

At least 19 recordsLinked to original sources

PubMed2003

Hypodermin A, a new inhibitor of human complement for the prevention of xenogeneic hyperacute rejection.

BACKGROUND: Hyperacute rejection (HAR) of discordant xenografts in the pig-to-human combination can be prevented using tranplants expressing transgenic molecules that inhibit human complement. Hypodermin A (HA), a serine esterase that degrades C3, was tested in the guinea-pig-to-rat and in the pig-to-human combinations. METHODS: Hypodermin A was tested in vitro, ex vivo, and in vivo models of HAR in the guinea-pig-to-rat combination. Hamster ovary cells (CHO) and a line of porcine aortic endothelial cells (PAEC11) were transfected with HA complementary DNA (cDNA). RESULTS: The pattern of degradation of rat and human C3 by HA was different (multiple bands lower than 40 kDa) from the physiologic pattern observed after spontaneous degradation of rat C3 or physiologic activation of human C3. The CH50 activity in serum was significantly lower in rats treated with 3.2 mg HA/kg than in untreated rats (45 +/- 16 U/ml vs. 700 +/- 63 U/ml, P < 0.05). Sera from rats injected with 3.2 mg/kg of HA were less effective in lysing guinea-pig endothelial cells (12 +/- 7%) than normal rat sera (79 +/- 3%; P < 0.001). Ex vivo, guinea-pig hearts perfused by rat serum supplemented with HA survived longer than those perfused by non-treated serum (210 +/- 34 and 154 +/- 71 min, respectively; P < 0.05). In vivo, guinea-pig hearts transplanted into HA treated rats survived longer than in non-treated rats (27 +/- 5 min vs. 13 +/- 4 min; P < 0.001). In the presence of human serum, smaller amounts of C6 and C5b-9 were deposited onto HA-transfected CHO cells than onto control cells. The mHA-PAEC11 cells were significantly more resistant to lysis by human C than control PAEC11 cells. CONCLUSIONS: These data suggest that transgenic HA could be used to prevent hyperacute xenogeneic rejection.

Acute Disease↗

PubMed2002

Early hepatic cytokine mRNA expression in experimental rat fasciolosis.

We studied the development of the cellular response, particularly with respect to Th1 and Th2 cytokine mRNA levels, in rat liver during the first 14 days of experimental infection with Fasciola hepatica. We analysed the panel of cytokines involved in initiation of the inflammatory and immune response. The levels of various mRNAs, particularly those primarily associated with the acute inflammatory response, and those commonly associated with T-cell proliferation and differentiation, were assessed by reverse transcription-polymerase chain reaction (RT-PCR) in liver samples. We also investigated the immune and inflammatory mediators balance in the liver, draining lymph node and spleen, by RT-competitive PCR quantification of mRNA levels for IL-4, IL-10 and IFN-gamma. Our data provide the first evidence that, in the early phase of infection, the inflammatory response in the liver of infected animals is transiently depressed or delayed. A Th0 profile was initially observed in the liver and hepatic lymph node, which developed into a Th2 profile 2 weeks after infection in the liver only. In the spleen, cytokine down-regulation was initiated and maintained during this period, suggesting that the parasite acts differently locally and in the periphery.

Animals↗

PubMed2001

[A new model of isolated perfused heart. Study of hypodermin A in hyperacute xenograft rejection].

AIMS OF THE STUDY: Isolated perfused heart (IPH) system and heart transplantation in the guinea-pig/rat combination represent a good model for the study of hyperacute xenograft rejection (HAR) in which the component plays a central role. Hypodermin A (HA), a protease cleaving the component, could be used to delay the HAR. METHODS: Creation of an original IPH working with rat serum (30 mL) and ex vivo study of HAR and I'HA. RESULTS: Study of HAR is possible with this IPH system. The mean guinea-pig heart survival after perfusion by normal rat serum was 38 +/- 7 min and was lower than survival observed after perfusion by guinea-pig serum (210 +/- 34 min) (p < 0.001), by decomplemented rat serum (177 +/- 45 min) (p < 0.001), and by rat serum with 20 micrograms/mL of HA (154 +/- 71 min) (p < 0.001). CONCLUSION: We developed an original system of isolated perfused heart allowing ex vivo study of HAR. HA delayed the occurrence of the HAR and confirmed the central role of the component in the HAR.

Acute Disease↗

PubMed2000

Allopatric combination of Fasciola hepatica and Lymnaea truncatula is more efficient than sympatric ones.

Parasites are capable of rapid evolutionary changes relative to their hosts, due to short life cycle, short generation time, and high fecundity. The direction of the evolution of parasite virulence can be studied in cross-transfer experiments, combining hosts and parasites from different localities, and comparing the outcome of established (sympatric and potentially locally adapted) and novel (allopatric) combinations of hosts and parasites. We aimed to compare the compatibility with snails hosts, the infectivity of metacercariae in rabbits and rats, and the fitness among different combinations (French-FF and Spanish-SS sympatries and allopatry-FS). The first isolate of Fasciola hepatica and its corresponding intermediate host, Lymnaea truncatula originated from Lugo's northwestern Spain. The second isolate of parasite and snail was collected in the Limoges area in central France. The Spanish snails were more susceptible to their sympatric trematode than the French snails. The Spanish flukes were more infective to intermediate hosts (snails) than the French flukes, but subsequent definitive hosts (rats or rabbits) infections remained similar. The estimated fitness was low in sympatric infections and highly similar (from 4.7 to 5.3). The fitness similarity corresponds, however, to different variations in life-history traits that could represent different strategies among the host-parasite local combinations. The infection rate in snails, metacercarial productivity, metacercarial infectivity, and the estimated fitness were better for allopatric combination (FS). The susceptibility data showed a higher efficiency of flukes in the allopatric snail population than in their local snail population. However, our results were obtained after one generation and from a single isolate and it remains to be determined if all allopatric fluke-snail isolates may present a better fitness. Nevertheless our results indicate that introduction of liver fluke-infected cattle should be monitored carefully, as it could result in the introduction of more efficient parasites.

Animals↗

PubMed2000

Fasciola hepatica: influence of gender and liver biotransformations on flukicide treatment efficacy of rats infested and cured with either clorsulon/ivermectin or triclabendazole.

Two fasciolicide preparations have been compared in 130 rats experimentally infected with Fasciola hepatica. Parasitological, immunological, and biochemical parameters have been followed to monitor the efficacy of the treatments. While Fascinex (triclabendazole) efficiently cured both male and female rats when administered as soon as 4 weeks postinfection, treatment with Ivomec-D (clorsulon + ivermectin) displayed a low efficacy on either male or female rats at this time point (54 and 0%, respectively). Moreover, when administered 8 weeks postinfection, the Ivomec-D treatment proved highly efficient on male rats while it displayed little effect on the female population (100 and 53%, respectively). This unexpected result has been related to an overexpression of a P4503A isoform that is observed only in females that have been treated with Ivomec-D. The influence of this P4503A cytochrome on drug metabolism and the need for the incorporation of both genders in clinical trials are discussed.

Animals↗

PubMed2000

Comparison of hepatic and renal drug-metabolising enzyme activities in sheep given single or two-fold challenge infections with Fasciola hepatica.

The activity of drug-metabolising enzymes was compared in liver and kidneys of adult sheep given single or two-fold fluke infection. Fascioliasis was induced by oral administration of 200 metacercariae of Fasciola hepatica to female sheep either 10 or 20 weeks (mono-infections) or 10 and 20 weeks (bi-infection) before killing. The parasitic pathology was ascertained at autopsy and by clinical observation of animals. In the liver of both mono- and bi-infected animals, significant decreases (P<0.05) (17-44%) were observed in the microsomal content of cytochrome P450 and in the two measured P450-dependent monooxygenase activities, benzphetamine and ethylmorphine N-demethylations. Moreover, Western blot analysis of microsomes demonstrated a decrease in the expression of cytochrome P4503A subfamily correlative with that of its presumed corresponding activity ethylmorphine N-demethylase. By contrast, the conjugation of chloro-dinitrobenzene to glutathione remained unchanged in liver cytosolic fractions prepared from all these animals. In kidneys, a significant decrease (P<0.05) (30%) in microsomal cytochrome P450 level of 10-week mono-infected sheep was observed whereas there was no change in the other groups of animals. The inflammatory origin and the consequences in terms of pathology and animal productivity of the fascioliasis-induced decreases in tissue-oxidative drug metabolism are discussed, particularly in the case of adult sheep suffering repetitive infections.

Animals↗

PubMed2000

Local hepatic immune response in rats during primary infection with Fasciola hepatica.

The distribution of lymphocyte subpopulations (TCD4+, TCD8+, TCD43+ and Ig+ cells), macrophages and eosinophils were analysed in the inflammatory infiltrates associated with hepatic lesions and in hepatic lymph nodes (HLN) from rats experimentally infected with F. hepatica and necropsied 1, 2, 3, 4, 6 and 8 week post infection (WPI). We also investigated the fixation of immunoglobulin isotypes on migrating flukes in the liver. As early as 1WPI, portal tract areas surrounding migratory tunnels were infiltrated with immune and inflammatory cells. The dominant cells were eosinophils and to lesser extent, macrophages and lymphocytes (TCD4+, TCD8+ and B). Most of the inflammatory and immune cells reached the posterior part of flukes, whereas in front of the parasites these cells were fewer in number. Except for eosinophils, no immune cells penetrated through granuloma consisting of hepatic necrotic cells. As early as 1WPI, IgM could be detected in the liver, and to a lesser extent IgA, IgG2a and IgG2b. At 2WPI, IgE and IgG1 began being detected. IgG2c was detectable at 3WPI. In HLN, we observed numerous microscopic follicles in the cortical zone with proliferation of germinal centres and medullary cords. The protective role of infiltrating cell populations and immunoglobulin isotypes and possible mechanisms of immune evasion by the parasite are discussed.

Animals↗

PubMed1999

Serodiagnosis of goat warble fly infestation by Przhevalskiana silenus with a commercial ELISA kit.

Cross-reactivity between Hypoderma lineatum antigen and anti-Przhevalskiana silenus antibodies has been demonstrated by an ELISA technique. To evaluate the applicability of a commercial ELISA kit for the immunodiagnosis of goat warble fly infestation, different dilutions of serum and conjugate were tested, the development of antibody to P silenus in naturally infested goats was studied, and the results were compared with an ELISA technique using an antigen extracted from the first instar larvae of H lineatum. The best results were obtained with a serum dilution of 1:50; with both techniques the highest antibody concentration were recorded in October, November and December. In view of the confirmed cross-reactivity between H lineatum antigen and anti-P silenus antibodies, and the simplicity and rapidity of the assay, the commercial ELISA kit can be considered as a useful tool for the diagnosis of goat warble fly infestation.

Animals↗

PubMed1998

Comparison of three ELISA tests for seroepidemiology of bovine fascioliosis.

The aim of the present study was to compare the sensitivity, specificity and usefulness of the DIG-ELISA, DOT-ELISA and Indirect ELISA tests for determining the seroprevalence of fasciolosis in cattle under tropical conditions in Mexico. To standardize the tests, positive and negative sera to F. hepatica from 88 Holstein Freisian adult cows located in an enzootic area of fascioliosis and 88 crossbred adult cattle from a fluke-free area were used. For the epidemiological study, 85 crossbred cattle between 1 to 7 years of age were used. Animals were bled every two months, from March 1995 to September 1996 and the sera obtained were stored at -70 degrees C, until used. Indirect ELISA showed a sensitivity of 96.5% and a specificity of 98.8%, DIG-ELISA 97.5% and 80.0% and DOT-ELISA 93.1% and 95.4%, respectively. During 1995, Indirect ELISA yielded the highest levels of IgG anti-F. hepatica antibodies. However, in 1996, after animal treatment with triclabendazole, DIG-ELISA tended to show higher percentages of antibody-positive animals, but it was not significantly different (p>0.05) from the other tests. Comparisons made in parallel to the faecal sedimentation test demonstrated that all serological tests detected higher percentages of positive animals. Only one serum out of ten (10%) of Paramphistomum spp. cross-reacted with the DOT-ELISA test, but no cross-reaction was observed with sera from animals with other parasites. All ELISA tests were highly sensitive and specific; they may be recommended for use in seroepidemiological surveys for F. hepatica.

Animals↗

PubMed1998

Activity of moxidectin 1% injectable solution against first instar Hypoderma spp. in cattle and effects on antibody kinetics.

The activity of the moxidectin as an 1% w/v injectable solution on first instar Hypoderma spp. has been evaluated in sixteen naturally infested young cattle. The animals were selected on the basis of their serological status and allocated to two groups of eight animals. At the end of November, one group was treated with moxidectin at a dose rate of 0.2 mg/kg via the subcutaneous route and the non treated control calves injected with the vehicle. The serological status was assessed 1, 2, 4, 8 and 12 weeks post treatment and the presence of Hypoderma lumps determined every two weeks from February to June. A 100% efficacy of the injectable formulation was demonstrated. A progressive fall of the antibody levels was observed in the treated calves for one month following treatment, suggesting a progressive action of the test compound and a limited risk of hypersensitivity.

Animals↗

PubMed1998

IFN gamma and IL-10 production by hepatic lymph node and peripheral blood lymphocytes in Fasciola hepatica infected sheep.

The present study was designed to determine whether IFN gamma and IL-10 were secreted by peripheral blood mononuclear cells (PBMC) and hepatic lymph node mononuclear cells (HLNMC) from 17 Fasciola hepatica infected sheep after in vitro stimulation by Fasciola hepatica excretory-secretory products (FhESP) during the first six weeks of two different infections. The IFN gamma concentration in mononuclear cells (MC) culture supernatants was determined by sandwich ELISA. The presence of IL-10 in MC culture supernatants was determined by the capacity of these to inhibit, with or without monoclonal antibodies to IL-10, the NO production of recombinant bovine IFN gamma (rBoIFN gamma) activated monocytes. FhESP-stimulated PBMC and HLNMC produced IFN gamma in culture supernatants from DPI 7 (day post-infestation 7) to DPI 14. Culture supernatants of PBMC stimulated with FhESP first induced an increase in the NO production of rBoIFN gamma activated monocytes. After 2 or 3 weeks of infection, MC culture supernatant reduced the NO production of rBoIFN gamma activated monocytes. From DPI 35, anti-IL-10 antibodies addition restored or enhanced NO production. During sheep fasciolosis, IFN gamma production increased only until the first two weeks post infection. IL-10 could be secreted throughout the first six weeks of infection and could inhibit monocytes from DPI 35.

Animals↗

PubMed1997

Enzymatic effect of hypodermin A, a parasite protease, on bovine lymphocyte membrane antigens.

The protease hypodermin A (HA) is produced by the parasitic warble-fly larva and is implicated in the modulation of the bovine immune system. This study examines the effect of this enzyme on the cell surface markers of bovine lymphocytes. HA interfered with the binding of all anti-lymphocyte receptor antibodies tested. Anti-BoCD2 and CD5 staining was completely abolished. But the mean fluorescence intensity (MFI) only was diminished for antibodies against BoCD4, CD8 and CD18. On the contrary, the MFI for anti-MHC Cl I molecules staining was increased. This effect of HA began as early as one h, and was reversed by removal of HA. Heating or PMSF treatment, which both inhibit protease activity, abolished the action of HA on the surface antigens. The HA concentrations (100 micrograms/ml) needed to alter antibody binding were similar to those that inhibited phytohaemagglutinin (PHA)-induced proliferation. These results show that enzymatic activity of HA on lymphocyte surface markers may be implicated in the inhibition of lymphocyte proliferation.

Animals↗

PubMed1997

[Diagnosis of bovine fascioliasis using serology of pools of sera. Interpretation in field conditions].

The efficacy of a serological test (ELISA with excretory-secretory products of Fasciola hepatica as an antigen) for the diagnosis of bovine fascioliasis was assessed under field conditions using pools of sera from 170 herds. Blood samples were collected from six to ten heifers and from five to ten adults in each herd. In 92 of the herds (Group 1) the animals had not been treated for fascioliasis; in 78 herds (Group 2), a fasciolicide had been used 3 to 6 months earlier. In each herd, three serum pools were examined: aliquots from all the animals sampled in a herd (GM); aliquots from heifers only (PM); and aliquots from the cows only (PM). Individual sera and the pools (GM and PM) were analyzed similarly and the specific antibody titer was calculated by comparison with laboratory standards. The antibody titer for pooled sera were correlated with the proportion of positive sera in the pool (P < 0.0001); but these could not be used to estimate the prevalence of seropositivity in a herd. Diagnosis based on pooled sera was less effective for the detection of low prevalences of seropositivity in Group 2 than in Group 1 because the antibody titer decreases after fasciolicide treatment. Infection was always detected using pooled sera in herds with a prevalence of seropositivity of more than 60% (GM) in Group 1, or with a prevalence of seropositivity of more than 70% in Group 2. The best method for herd diagnosis of fascioliasis was based on the presence of a positive test in either of the two PM groups. Using this method, all infected herds with a prevalence of seropositivity of more than 30% in Group 1, or 50% in Group 2, were accurately detected.

Animals↗

PubMed1997

The prevalence of goat hypodermosis in Greece.

Goat hypodermosis caused by Przhevalskiana silenus is common in Greece causing quite serious losses on animal production and breeding, but no data regarding the prevalence of this parasitic disease were available within the last 17 years. Towards this end, at the slaughter house of the area of Thessaloniki, the number of affected hides due to skin penetration by P. silenus larvae was recorded and the prevalence was assessed to be 54.2%. The mean number of holes found per hide was 27.07 (ranged from 0 to 162 holes per hide). Moreover, a total of 4900 goat blood serum samples were collected from different parts of the country and tested for hypodermosis using the ELISA method. Based on the results of this test, the prevalence of the infection was found to be 49.2%, varying from place to place between 4.6 and 85.1%. The range of the infection per flock was from 0 up to 100%.

Animal Husbandry↗

PubMed1997

The prevalence of bovine hypodermosis in Greece.

Hypodermosis is a common parasitic disease of great importance on cattle production and breeding worldwide. In Greece, no reliable data are available regarding the extent of this infection and therefore, as expected, no national or regional control programme is being applied. In order to cover this lack of information, a total of 4200 cattle blood sera were collected from different parts of the country and tested for hypodermosis using the ELISA method. The prevalence of the infection was found to be 37.4%, varying from place to place between 0.7 and 78.8%. The range of the infection per flock was from 0 up to 100%.

Animals↗

PubMed1996

Sero-surveillance of hypodermosis in a herd under therapeutic control. Effect of a low level of infestation.

A cattle herd from the experimental farm of INRA in Nouzilly has been treated for hypodermosis since October 1990. Additionally, a regional eradication scheme has been implemented in this area since autumn 1992. Bi-monthly warble counts were performed between March and July each year on an average of 200 animals. No warble was recorded in this herd from 1991 to 1994 with the exception of two dairy cows in 1993. In autumn 1994, therapeutic control measures were stopped. Serological surveys were performed in the autumn of each year from 1991 until 1995. Anti-Hypoderma antibodies were found in 25%, 27.2%, 4.3%, 3.2% and 0% of the animals respectively. An experimental low infestation was conducted in the summer of 1994. During the spring 1994, third instars of Hypoderma bovis were collected from naturally infested animals. From a total of 13 pupae, six adults (four males and two females) emerged and were released in the herd of Nouzilly on 24 June and 4 July. In October 1994 serological investigations revealed two animals seropositive for hypodermosis. This number increased to six in January 1995. The antibody kinetics of these six animals remained parallel throughout the next 6 months: the titres increased up to April and started to fall in May to return to negative values in August. Manual examinations of the animals at weekly intervals between April and July revealed the presence of four warbled animals with one, one, two and three warbles respectively. The two other seropositive animals remained warble free. One other animal showed antibody titre fluctuations between negative and low positive values, but was warble free in the spring. In October 1995 all the animals of the herd were seronegative. The interpretation and the value of a sensitive immunodiagnosis in a large eradication programme are discussed and compared with warble counts, especially in the case of a low level of infestation.

Animals↗

PubMed1996

Local immune response to experimental Fasciola hepatica infection in sheep.

Macrophages, eosinophils, neutrophils and lymphocyte subpopulations (OvCD5+, OvCD4+, OvCD8+, OvWC1+ and lg+) were identified in sections of hepatic tissue and hepatic lymph nodes from sheep experimentally infected with F. hepatica and necropsied 14, 28, 42 or 56 days post infection. The migratory tunnels produced by juvenile flukes appeared as local areas of necrosis surrounded by infiltrating inflammatory cells, particularly numerous macrophages, eosinophils and OvCD4+ lymphocytes n addition, B lymphocyte proliferation was observed in hepatic lymph nodes and in hepatic portal tracts. Only three juvenile flukes were identified in the sections. They where partially in contact with healthy tissue and partially with neutrophils, macrophages and eosinophils; they were covered by IgM Host-parasite interactions resulting from immune response regulation by helper T lymphocytes and from immune evasion by the parasite are discussed.

Animals↗

PubMed1996

Cross-reactive, stage-specific antigens in the Oestridae family.

The larval stages of different economically important Oestridae species were studied for their antigenicity and cross reactivity, using ELISA and immunoblotting. The immune sera of cattle from Algeria, Belgium, France and Switzerland were compared for their capacity to recognize the stage-specific antigens of their specific parasites Hypoderma bovis and H lineatum, originating from different populations. This comparison was extended to other Hypoderminae species responsible for economic losses in European animal production: H diana in roe deer and H tarandi in reindeer. The specific host for each of these parasites recognized common epitopes of hypodermin C, a collagenolytic enzyme previously well characterized in the first instar of H bovis and H lineatum. No cross reactivity was found with other Oestridae, such as Oestrus ovis or Gasterophilus intestinalis. The specificity of hypodermin C was evaluated using sera from animals harbouring other arthropods such as ticks, helminths, including Fasciola hepatica and Haemonchus contortus, or protozoa such as Anaplasma sp, and no reaction was observed. The use of hypodermin C is therefore suggested as an antigen in the immuno-survey of economically-important Hypoderminae.

Abattoirs↗