Biological properties the novel application of N-trimethyl chitosan nanospheres as a stabilizer and preservative in tetanus vaccine

Purpose@#Chitosan is a natural polymer that has excellent properties include biocompatibility, biodegradability, no cytotoxicity, high charge density, low cost, mucoadhesive, permeation enhancing (ability to cross tight junction), and immunomodulating ability that makes the spectrum of its applicability much broader. This study was conducted to investigate the stabilizing, preservative and immunogenicity properties of N-trimethyl chitosan nanospheres (N-TMCNS). @*Materials and Methods@#The tetanus toxoid (TT) was encapsulated into N-TMCNS and then characterized by scanning electron microscope, atomic force microscope, and dynamic light scattering. For stabilizer assay of N-TMCNS after storage of TT-N-TMCNS at different temperatures for 3 weeks, they were used for immunization of mice and different temperatures groups’ anti-TT-N-TMCNS production compared with other groups. Finally, the immunized mice were challenged with tetanus toxin. The preservation activity of TT-N-TMCNS against Escherichia coli was compared with thimerosal formulated TT. @*Results@#Our results revealed that heat-treated TT-N-TMCNS could induce higher titer of neutralizing immunoglobulin G in compared to TT vaccine and was able to protect the mice better than TT vaccine in challenge test. Furthermore, N-TMCNS as a preservative inhibited the growth of E. coli more effective than thimerosal. @*Conclusion@#Overall, the obtained results indicated that the N-TMCNS is one of the best stabilizer and preservative agent that can be used in the formulation of TT vaccine.

A Therapeutic Oral Vaccine Candidate against Propionibacterium acnes: Preparation and Immunological Evaluation of Nanoparticles Encapsulated Sialidase-CAMP Fusion Protein

Abstract Acne Vulgaris is a common skin disease caused by Propionibacterium acnes, an anaerobic microbiota of human skin that plays a vital role in the pathology of acne. The aim of this study was to prepare nanoparticles containing an acne recombinant protein and determine its ability as an oral acne vaccine in mice. The recombinant Sialidase-CAMP gene was expressed and purified in a prokaryotic host. The chitosan nanoparticles containing the recombinant protein were prepared, encapsulated, and administered by both oral and subcutaneous routes to Balb/c mice. Sera IgA and IgG and stool IgA titers were measured by ELISA, and the immunized mice were challenged against P. acnes. A 65 kDa recombinant protein was confirmed by SDS-PAGE and western blot. The size and zeta potential of nanoparticles were 80 nm and +18 mV, respectively. After oral immunization, the serum IgG and IgA titers were 1:3200 and 1:16, respectively, and the stool IgA titer was 1:8. In the subcutaneous route, the serum IgG titer was 1:51200. Immunized mice showed no inflammation in the ear of challenged mice. It is the first study that examines a chitosan-nanoparticulated acne fusion protein as an applicable acne vaccine candidate with appropriate immunogenicity potential. Further studies are required to validate the clinical usefulness of this vaccine candidate.

[Prevalence of food allergens in patients with atopic dermatitis referring to the asthma and allergy clinic in Tehran in 2014]

Background and Aim: Atopic dermatitis is a common chronic inflammatory disease the skin. Food allergy is one of the main factors affecting the onset and severity of the disease. Recognition of the pattern of sensitivity to food allergens in the community can play an important role in the public awareness and prevention of recurrence of atopic dermatitis. The aim of this study was to recognize the prevalence of common food allergens in the patients with atopic dermatitis by means of skin prick test in Tehran during 2014

Material and Methods: In this cross sectional study, we examined all the patients for the signs and symptoms of allergies in the Asthma and Allergy Clinic during 2014. Skin prick test was performed for common food allergens in the patients with atopic dermatitis. SPSS version 20, was used for data analysis

Results: Out of 1012 patients with allergy, 282 cases [approximately 28%] were diagnosed as atopic dermatitis. 87[52 female and 35 male] patients with atopic dermatitis, were sensitive to at least one food allergen [positive prick test]. In patients with atopic dermatitis the prevalence rates of asthma, allergic rhinitis and urticaria were 6.8, 11.4 and 4.5 %, respectively. There was no relationship between gender and food allergy in the patients with atopic dermatitis. The most prevalent food allergens were egg yolk [38.4%], egg white [36.5%], hazelnut [33.3%], and peanut [28.7%], and the least prevalent allergens were found in rice [4.6%], barley [5.7%], and meat [6.9%]

Conclusion: The prevalence rate of atopic dermatitis in allergic patients in this study was more than expected. Sensitivity to egg and nuts as the most common allergens should be taken into consideration by families and allergy specialists

[Evaluation of antibodies produced against lipopolysaccharides of vibrio cholerae for rapid detection of cholera]

Objective: Cholera is an endemic disease in Iran. Early detection, especially in times of disease outbreaks, is of vital importance. The antibody against the lipopolysaccharide [LPS] is an important method for bacterial detection. This study intends to extract and purify the LPS of Vibrio cholerae and evaluate the cholera antibody for detection purposes

Methods: Vibrio cholerae was cultured in tryptone extract medium. LPS was extracted by the hot phenol water method, purified, and dialyzed. We measured the LPS protein and sugar content, purity, and biological activity. Antibodies were produced by injection of the killed bacteria with Freund's complete adjuvant into rabbits and then the LPS was injected three times with Freund's incomplete adjuvant. After the last booster, blood samples were taken. We used ELISA to determine the antibody titers against the Inaba and Ogawa serotypes, the LPS of these serotypes, and several other similar bacteria

Results: The amount of protein in the purified LPS was approximately zero and sugar was 0.5 mg/ml. The LPS had a titer activity of 1024, and consisted of three bands [5.2, 4, and 5.14 KD]. Antibodies produced by the rabbits identified the bacterial Inaba and Ogawa serotypes, and the purified LPS. Ogawa and Inaba serotypes cross-react with each other but not with other species of Vibrio and other bacteria. The LPS antibody titer against the Ogawa serotype was 1:32000, whereas for Inaba it was 1:16000

Conclusion: Due to the low cost of production, high sensitivity, and importance of cholera diagnosis in Iran, the antiserum produced in this study can be used as a tool for early screening of cholera and discrimination of O1 strains from non-O1 strains in immunologic tests

[Expression of optimized gene of Enterotoxigenic Escherichia coli CFA/I major subunit]

Enterotoxigenic Escherichia coli is considered as the most important agent of children diarrhea and mortality in developing countries. This bacterium causes 300-600 thousands of deaths in the children under 5 years of age per year. With difficulties in treatment as well as its wide prevalence, designing an effective vaccine against this microorganism is the objective of world Health Organization [WHO]. The CfaB protein as immunogen and major subunit of fimberia has a critical role in the bacterial attachment to small intestine epithelium and the produced antibody against this protein can prevent attachment of bacterium to epithelial surface. Hence, this molecule alone or with other virulent factors has been considered by many researchers in vaccine designing. In this study, expression of colonization factor B with the aim of studying the immunogenesity of this protein as a component of vaccine candidate was performed. cfaB gene was amplified by PCR and cloned into pET28a and its expression was evaluated. Since there was no expression, which was due to presence of rare codon, the cfaB gene was again cloned into pET28a using codon bias in E. coli and subsequently expressed. Presence of a 20KD band on SDS-PAGE gel indicated the expression of CfaB protein, which was later confirmed by immunoblotting with anti-His tag and anti CfaB antibodies and purified on Ni-NTA column. Codon optimization and expression in heterologous hosts is a useful approach for obtaining large quantities of recombinant protein