419. Rational design of bifunctional siRNAs that silence genes and recruit the innate immune system to treat ectopic pregnancies

2008 ◽  
Vol 20 (9) ◽  
pp. 99
Author(s):  
S. Tong ◽  
M. P. Gantier ◽  
M. Belhke ◽  
B. R. G. Williams
Keyword(s):  
Immune System ◽  
Gene Silencing ◽  
Rational Design ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Micro Rna ◽  
Peripheral Blood Mononuclear ◽  
Sirna Sequence ◽  
Sense Strand ◽  
Ectopic Pregnancies

RNA interference (RNAi) is a new therapeutic approach, silencing genes to disrupt diseases. However, short interfering siRNAs (molecule used in RNAi) can have off-target effects, activating the immune system through RNA sensing toll-like receptors (TLR) 3, 7 and 8. We have previously proposed that in some diseases (cancers, ectopic pregnancies) it may be useful to enhance the immune response. A novel class of immunostimulatory siRNAs could be developed, silencing genes important to disease and recruiting the immune system to further aid disease clearance. We set out to develop a rational design strategy that enhances immunostimulatory properties to any siRNA sequence but maintains effective gene silencing. We screened a set of siRNAs targeting lamin. All were of the same sequence, except for different immunostimulatory motifs on the 3′ end of the sense strand. We also investigated a different design where we added a small micro-RNA like poly-uridine bulge (potentially immunostimulatory) on the sense strand. We used human peripheral blood mononuclear cells (PBMCs) to test for immunostimulation, and HEK 293-T-cells to test for lamin gene knockdown.Of all strategies tested, the poly-uridine bulge was best. It silenced the lamin gene as effectively as control, but caused a 2–3 fold increase of IFN-α and TNF-α. We verified this approach by adding the poly-uridine bulge onto an siRNA of low immunostimulatory potential targeting GFP. The bulge markedly enhanced immunostimulation in a dose response manner, and did not compromise gene knockdown. The addition of a poly-uridine bulge to siRNAs can increase immunostimulation without affecting gene silencing efficacy. Immunostimulatory siRNAs might be particularly efficacious to treat ectopic pregnancies where there are abundant immune cells, and functional TLR 7/8 in the trophoblast (unpublished observations). We now plan to test this immunostimulatory siRNA approach in an in vivo ectopic pregnancy model using JEG-3 cells xenographted in NOD-SCID mice.

scholarly journals Sildenafil does not affect the proliferation of human lymphocytes in the in vitro transplant model

2019 ◽  
Author(s):  
Beata Kaleta
Keyword(s):  
Bone Marrow ◽  
Immune System ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Human Lymphocytes ◽  
Venous Blood ◽  
Human Immune System ◽  
Pde5 Inhibitors ◽  
Trypan Blue Exclusion ◽  
Peripheral Blood Mononuclear

Sildenafil is a selective inhibitor of type 5 phosphodiesterase (PDE5) used in the treatment of erectile dysfunction (ED) and pulmonary arterial hypertension (PAH). Numerous studies revealed beneficial effects of sildenafil use in chronic kidney disease, and also in renal, liver, heart and bone marrow transplant recipients. Some reports suggest that sildenafil modulates function of the immune system, and additionally proliferation of endothelial, bone marrow and cancer cells. Despite the fact that PDE5 inhibitors showed efficacy and safety, it is very important to know whether these drugs have immunomodulatory properties, because are used in patients after organ transplantation. The influence of sildenafil on antigen-induced proliferation of lymphocytes remains currently unknown, thus the aim of the study was to investigate the effects of the drug on human peripheral blood mononuclear cells (PBMCs) proliferation in a mixed lymphocyte reaction (MLR).PBMCs were isolated from venous blood from 30 donors. The proliferation was examined on the DNA synthesis level by measurements of 3H-thymidine incorporation. Cell viability was determined using trypan blue exclusion method.The study demonstated that sildenafil at concentrations of 0.06 µM, 0.6 µM and 6µM did not affect auto- and alloantigen-induced  proliferation of PBMCs and showed no cytotoxic effect. However, further analysis is required to fully understand the role of PDE5 inhibitors in the regulation of human immune system.

scholarly journals Rational Design of Hypoallergenic Vaccines: Blocking IgE-binding to Polcalcin Using Allergen-specific IgG Antibodies

2020 ◽  
Author(s):  
Mohsen Mohammadi ◽  
Gholamreza Khamisipour ◽  
Faezeh Soltanpour ◽  
Fatemeh Omrani ◽  
Behrooz Taheri ◽  
...  
Keyword(s):  
Rational Design ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Binding Capacity ◽  
Cell Epitope ◽  
Igg Antibody ◽  
Peripheral Blood Mononuclear ◽  
Ige Binding ◽  
Peptide Carrier ◽  
Specific Igg

Chenopodium album polcalcin (Che a 3) is characterized as a major cause of cross-reactivity inallergic patients to the Chenopodiaceae family. Therefore, the present study was conducted to develop a hypoallergenic Che a 3 derivatives as the candidate vaccine for type 1 allergy. Four derivatives were generated from Che a 3. The first was a mosaic peptide derivative computationally identified in Che a 3 which was coupled to keyhole limpet hemocyanin (KLH). The second one was a mutant Che a 3, and the other two derivatives included N- and C-terminal halves of Che a 3 that both coupled to KLH. The IgE-binding capacity of Che a 3 and its derivatives and also their ability to induce there combinant Che a 3 (rChe a 3)-specific IgG antibody, were determined using the enzyme-linked immune sorbent assay (ELISA). Moreover, the lymphopro liferative capacity of rChe a 3 or its derivatives and their pro-inflammatory cytokine response interleukin (IL)-5 and IL-13 were measured in the human peripheral blood mononuclear cells (PBMCs). Among all derivatives, the N-terminal half peptide and mosaic peptide exhibited the lowest IgE-binding capacity. In addition, in comparison to other antigens, KLH-coupled mosaic peptide induced the highest level of the recombinant Che a 3 (rChe a 3)-specific IgG antibody and ther Che a 3 specific-blocking IgG antibody in mice. Moreover, the mosaic peptide lacked lymphopro liferative capacity and down-regulated expression of pro-allergic IL-5 and IL-13 cytokines. Therefore, a peptide-carrier fusion vaccine, composed of the B-cell epitope coupled to the carrier, could be considered as one of the promising hypoallergenic vaccines to treat patients with allergy to low molecular weight allergens such as Che a 3.

Immune Modulation by cross-linked Bovine Colostrum in vitro and in vivo

Pteridines ◽  
2010 ◽  
Vol 21 (1) ◽  
pp. 94-102
Author(s):  
Ninfa R. Pedersen ◽  
Budi J. Hidayat
Keyword(s):  
Atopic Dermatitis ◽  
Immune System ◽  
Immune Modulation ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Bovine Colostrum ◽  
Human Immune System ◽  
Mature Milk ◽  
Peripheral Blood Mononuclear ◽  
Tryptophan Degradation

Abstract Bovine colostrum (BC) is the early thick yellow fluid produced by cows during the first several days after birth of the calf. BC is different from "mature" milk as it contains nutrients and immune factors, which strengthen the immune system of the newborn calf in its first week of life. BC has a long history of use in traditional medicine throughout the world and is currently also used in a topical cream for the treatment of immuno-related skin problems such as atopic dermatitis or psoriasis. However, despite a large amount of literature concerning the properties of human or bovine colostrum, there are only few reports on the effects of colostrum on the human immune system. In this study, the effects of cross-linked BC containing hyaluronic acid on the T-cell/macrophage interplay were investigated in human peripheral blood mononuclear cells (PBMC) and the results were compared with the effects of euxyl 9010, which was used to preserve the BC ingredients. The cross-linked BC preparations showed significant immunomodulatory effects on unstimulated and phytohaemagglutinin (PHA)-stimulated human PBMC by modulating tryptophan degradation and formation of neopterin in a biphasic manner. These results could be relevant for some of the beneficial effects of BC observed in the treatment of skin lesions of patients with atopic dermatitis or psoriasis. However, additional studies are needed to confirm the first positive results obtained in patients.

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