Real-Time Reverse Transcription-PCR Quantification of Cytokine mRNA Expression in Golden Syrian Hamster Infected with<i>Leishmania infantum</i>and Treated with a New Amphotericin B Formulation

Iñiguez, S. Rama; Dea-Ayuela, María Auxiliadora; Sánchez‐Brunete, J. A.; Torrado, J. J.; Alunda, José María; Bolás‐Fernández, F.

doi:10.1128/aac.50.4.1195-1201.2006

Cited by 28 publications

(15 citation statements)

References 39 publications

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“…Analysis of hamster IFN-γ expression was performed as previously described. 34 Determination of tumor-specific lymphocyte proliferation. Peripheral blood mononuclear cells from treated hamsters obtained 20 days after vector inoculation were exposed to tumor cells at 10:1 ratio and analyzed by flow cytometry 3 days after carboxyfluorescein succinimidyl ester labeling, basically as described in ref.…”

Section: Quantification Of Virus Production In Vivomentioning

confidence: 99%

Treatment of Pancreatic Cancer With an Oncolytic Adenovirus Expressing Interleukin-12 in Syrian Hamsters

et al. 2009

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Pancreatic cancer is an aggressive malignancy resistant to most conventional and experimental therapies, including conditionally replicative adenoviruses (CRAds). The incorporation of immunostimulatory genes such as interleukin-12 (IL-12) in these viruses may overcome some of their limitations, but evaluation of such vectors requires suitable preclinical models. We describe a CRAd in which replication is dependent on hypoxia-inducible factor (HIF) activity and alterations of the pRB pathway in cancer cells. Transgenes (luciferase or IL-12) were incorporated into E3 region of the virus using a selective 6.7K/gp19K deletion. A novel permissive model of pancreatic cancer developed in immunocompetent Syrian hamsters was used for in vivo analysis. We show that, in contrast with nonreplicating adenoviruses (NR-Ad), active viral production and enhanced transgene expression took place in vivo. A single intratumor inoculation of the CRAd expressing IL-12 (Ad-DHscIL12) achieved a potent antitumor effect, whereas higher doses of replication-competent adenoviruses carrying luciferase did not. Compared to a standard NR-Ad expressing IL-12, Ad-DHscIL12 was less toxic in hamsters, with more selective tumor expression and shorter systemic exposure to the cytokine. We conclude that the expression of IL-12 in the context of a hypoxia-inducible oncolytic adenovirus is effective against pancreatic cancer in a relevant animal model.

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Section: Quantification Of Virus Production In Vivomentioning

confidence: 99%

Treatment of Pancreatic Cancer With an Oncolytic Adenovirus Expressing Interleukin-12 in Syrian Hamsters

et al. 2009

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“…Differences in gene expression were calculated by the comparative C T method (43). This method compares test samples to a comparator sample and uses results obtained with a uniformly expressed control gene (HGPRT) to correct for differences in the amounts of RNA present in the two samples being compared to generate a ⌬C T value.…”

Section: Estimation Of Expression Of Mrna Cytokines By Real-time Pcr mentioning

confidence: 99%

“…Real-time quantitative PCR was conducted as per the protocol described earlier (43). Briefly, it was carried out with 12.5 l of SYBR green PCR master mix (Bio-Rad), 1 g of cDNA, and primers at a final concentration of 300 nM in a final volume of 25 l. PCR was conducted under the following conditions: initial denaturation at 95°C for 2 min followed by 40 cycles, each consisting of denaturation at 95°C for 30 s, annealing at 55°C for 40 s, and extension at 72°C for 40 s per cycle using the iQ5 multicolor real-time PCR system (Bio-Rad).…”

Section: Estimation Of Expression Of Mrna Cytokines By Real-time Pcr mentioning

confidence: 99%

Immunization with the DNA-Encoding N-Terminal Domain of Proteophosphoglycan of Leishmania donovani Generates Th1-Type Immunoprotective Response against Experimental Visceral Leishmaniasis

Samant

Gupta

Kumari

et al. 2009

The Journal of Immunology

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Leishmania produce several types of mucin-like glycoproteins called proteophosphoglycans (PPGs) which exist as secretory as well as surface-bound forms in both promastigotes and amastigotes. The structure and function of PPGs have been reported to be species and stage specific as in the case of Leishmania major and Leishmania mexicana; there has been no such information available for Leishmania donovani. We have recently demonstrated that PPG is differentially expressed in sodium stibogluconate-sensitive and -resistant clinical isolates of L. donovani. To further elucidate the structure and function of the ppg gene of L. donovani, a partial sequence of its N-terminal domain of 1.6 kb containing the majority of antigenic determinants, was successfully cloned and expressed in prokaryotic as well as mammalian cells. We further evaluated the DNA-encoding N-terminal domain of the ppg gene as a vaccine in golden hamsters (Mesocricetus auratus) against the L. donovani challenge. The prophylactic efficacy to the tune of ∼80% was observed in vaccinated hamsters and all of them could survive beyond 6 mo after challenge. The efficacy was supported by a surge in inducible NO synthase, IFN-γ, TNF-α, and IL-12 mRNA levels along with extreme down-regulation of TGF-β, IL-4, and IL-10. A rise in the level of Leishmania-specific IgG2 was also observed which was indicative of enhanced cellular immune response. The results suggest the N-terminal domain of L. donovani ppg as a potential DNA vaccine against visceral leishmaniasis.

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“…Nonliposomal AMB therapy of leishmaniasis (Ehrenfreund-Kleinman et al, 2005) increased TNF-α release from mouse peritoneal macrophages and human monocytes, but had no effect on IFN-γ and NO release. Rama-Iniguez et al (2006), who used AMB formulated in microspheres from human serum albumin (AMB-HSA) for treatment of leishmaniasis in hamsters, found that high doses of AMB-HSA (40 mg/kg) caused deactivation of the anti-inflammatory cytokine TGF-β, which in turn results in an up-regulation of the Th1 cytokines IFN-γ and TNF-α. The dose of 10 mg/kg b.w.…”

Section: Discussionmentioning

confidence: 99%

Immune response of mice to Echinococcus multilocularis infection after therapy with amphotericin B colloidal dispersion

Porubcová

Ševčíková

2007

Helminthologia

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47 SummaryThe effect of amphotericin B colloidal dispersion (ABCD) on selected immunological parameters and growth of the larval cysts in mice infected intraperitoneally with Echinococcus multilocularis protoscoleces was observed. ABCD was administered at a dose 10 mg/kg body weight twice a week from week 5 to 10 post infection (p.i.). The Echinococcus infection suppressed the proliferative response of splenic T lymphocytes to nonspecific mitogen concanavalin A throughout almost the whole course of the experiment and ABCD administration did not affect this inhibittion. The increase in the proliferative response of B lymphocytes to lipopolysaccharide was found in infected mice with ABCD treatment from week 6 to 10 p.i. ABCD induced a significant rise of the splenic CD4 T cell subpopulation in infected mice only on week 6 p.i. The CD8 T subpopulation was not influenced by the therapy. The level of serum Th1 cytokine IFN-γ in infected and ABCD treated mice was elevated only at week 8 p.i., while the level of serum Th2 cytokine IL-5 was not influenced by the therapy. The ABCD treatment inhibited the IFN-γ production by splenocytes in vitro from week 6 to 10 p.i. On the contrary, the IL-5 production in vitro was stimulated at weeks 8 and 12 p.i. None antiparasitic effect of ABCD on larval growth was determined. Results suggest that amphotericin B colloidal dispersion did not affect the inhibited Th1 immune response after parasite infection. On the contrary, ABCD advanced the Th2 immune response development, which allows the progressive growth of the parasite.

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Real-Time Reverse Transcription-PCR Quantification of Cytokine mRNA Expression in Golden Syrian Hamster Infected withLeishmania infantumand Treated with a New Amphotericin B Formulation

Cited by 28 publications

References 39 publications

Treatment of Pancreatic Cancer With an Oncolytic Adenovirus Expressing Interleukin-12 in Syrian Hamsters

Treatment of Pancreatic Cancer With an Oncolytic Adenovirus Expressing Interleukin-12 in Syrian Hamsters

Immunization with the DNA-Encoding N-Terminal Domain of Proteophosphoglycan of Leishmania donovani Generates Th1-Type Immunoprotective Response against Experimental Visceral Leishmaniasis

Immune response of mice to Echinococcus multilocularis infection after therapy with amphotericin B colloidal dispersion

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