Protein and Antibody Engineering: Suppressing Degranulation of the Mast Cells and Type I Hypersensitivity Reaction.

With an increase in atopic cases and owing to a significant role of mast cells in type I hypersensitivity, a therapeutic need to inhibit degranulation of mast cells has risen. Mast cells are notorious for IgE-mediated allergic response. Advancements have allowed researchers to improve clinical outcomes of already available therapies. Engineered peptides and antibodies can be easily manipulated to attain desired characteristics as per the biological environment. A number of these molecules are designed to target mast cells in order to regulate the release of histamine and other mediators, thereby controlling type I hypersensitivity response. The aim of this review paper is to highlight some of the significant molecules designed for the purpose.

Interfering Effects of In Vitro Fertilization and Vitrification on Expression of Gtl2 and Dlk1 in Mouse Blastocysts.

BACKGROUND: Embryo vitrification is a key instrument in assisted reproductive technologies (ARTs). However, there is increasing concern that vitrification adversely affects embryo development. This study intends to assess the effect of vitrification on developmental competence, in addition to expressions of long non-coding RNA (lncRNA) gene trap locus 2 (Gtl2) and its reciprocal imprinted gene delta-like homolog 1 (Dlk1), in mouse blastocysts. MATERIALS AND METHODS: In this experimental study, we have designed three experimental groups: control (fresh blastocysts collected from superovulated mice), in vitro fertilization (IVF; blastocysts derived from IVF) and vitrification (IVF derived blastocysts subjected to vitrification/warming at the 2-cell stage). Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was performed to assess the expression levels of Gtl2 and Dlk1 in the blastocysts. RESULTS: The results showed that vitrification group had significantly lower blastocyst and hatching rates compared to the IVF group (P<0.037) and (P<0.041), respectively. Gtl2 was down-regulated and Dlk1 was up-regulated following the IVF and vitrification (P<0.05). CONCLUSION: These results suggested that IVF and vitrification disturbed genomic imprinting and lncRNA gene expressions, which might affect the health of IVF children.

The Effect of Increased miR-16-1 Levels in Mouse Embryos on Epigenetic Modification, Target Gene Expression, and Developmental Processes.

Changes in microRNA (miRNA) levels are present in numerous diseases. Although these changes are particularly noted in male infertility, little is known about the effects of increased miR-16-1 in sperm from infertile men. In this study, we assessed the effects of increased mir-16-1 expression on the developmental process, epigenetic changes, and target gene expressions. IVF embryos, 6 h after insemination, were divided into three groups: control, control negative (CN), and miR-16-1 harboring plasmid microinjection. The developmental rates of these embryos were recorded after 24, 48, 72, and 96 h of culture. The levels of histone H3 lysine 4 tri-methylation (H3K4me3) and histone H3 lysine 27 tri-methylation (H3K27me3) were assessed in the 2-cell and blastocyst stages by immunofluorescence staining. Expression profiles of the miR16-1, Bax, Bcl-2, Suz12, and Kmt2a genes were measured by quantitative real-time polymerase chain reaction (qRT-PCR). There was a significant decrease from the 8-cell stage to the blastocyst stage of embryo development in the miR-16-1 harboring plasmid microinjection group. We observed substantial reductions in the amounts of H3K4me3 and H3K27me3 in the 2-cell and the blastocyst stages in the miR-16-1 harboring plasmid microinjection group (P ≤ 0.05). The miR-16-1 level in the miRNA group was higher than the control group in the 2-cell and the blastocyst stages. There was a significant increase (P ≤ 0.05) in Bax and decreases in Bcl2, Suz12, and Kmt2a following the injection of the miR-16-1 harboring plasmid. These results suggest that a change in miR-16-1 expression can significantly affect embryo development, epigenetic changes, and target gene expressions.

Isolation of Polyclonal Single-Chain Fragment Variable (scFv) Antibodies Against Venomous Snakes of Iran and Evaluation of Their Capability in Neutralizing the Venom.

Several species of dangerous snakes are found in Iran and, according to the Emergency Response Center of Iran from 2002 to 2011, 53,787 Iranians have suffered from snakebite. Although the mortalities caused by snakebite are very low, snakebite-related amputations are still a major concern. Currently, anti-venom polyclonal antibodies derived from animals, such as horses are used to treat snakebites; however, in some cases they can cause anaphylactic shock and serum sickness. In line with this premise, generation of recombinant anti-venom antibodies can be considered as an alternative strategy. Single-chain fragment variable (scFv) antibodies offer several advantages compared to the whole antibodies, including ease of production, high affinity and specificity. In the present study, scFv antibodies were selected against the venom of the most poisonous snakes in Iran using phage display technology. Phage particles harboring anti-venom specific scFv were separated and scFv antibodies were produced in bacteria. In-vitro assay showed that polyclonal scFvs specifically bind to the venom. Furthermore, in-vivo experiment in mice BALB/c indicated effective toxin neutralization using 20 µg of polyclonal scFv. Our study indicates the neutralizing capacity of anti-venom polyclonal scFvs, although further neutralization assays are needed to confirm their effectiveness.

Production of human single-chain fragment antibody (ScFv) against human epidermal growth factor receptor-2 (HER-2) by phage display technology.

Breast cancer with more than 1.7 million diagnoses per year has been known as one of the most prevalent cancers among women worldwide. Despite the availability of advanced treatment options, cancer progression and metastasis is observed in 20% of patients. Human epidermal growth factor receptor-2 (HER-2) is considered as an important prognostic and diagnostic tumor marker for breast cancer. While HER-2 is expressed on the surface of normal cells, its overexpression occurs in 20-25% on breast cancer tumor cells. This type of tumor which is referred to as HER-2+ is the most aggressive type of breast cancer and shows more resistance to radiotherapy. Single-chain fragment antibodies (ScFvs) offer several advantages in comparison to conventional whole antibodies due to their small size. Particularly, ScFv fragments show improved diffusion and solid tumor penetration. In this study, a human ScFv antibody library was used to isolate anti-HER-2 ScFv antibodies through cell panning and mix antigen-cell panning strategies. Analysis of the binding activity and specificity of isolated ScFv antibodies against HER-2-expressing cell lines and recombinant HER-2 antigen indicated the higher efficiency of the cell panning strategy in isolation of ScFv antibody fragments.