Identification of allergen epitopes is a key component in proper understanding of the pathogenesis of type I allergies, for understanding cross‐reactivity and for the development of mimotope ...immunotherapeutics. Phage particles have garnered recognition in the field of molecular allergology due to their value not only in competitive immunoscreening of peptide libraries but also as immunogenic carriers of allergen mimotopes. They integrate epitope discovery technology and immunization functions into a single platform. This article provides an overview of allergen mimotopes identified through the phage display technique. We discuss the contribution of phage display peptide libraries in determining dominant B‐cell epitopes of allergens, in developing mimotope immunotherapy, in understanding cross‐reactivity, and in determining IgE epitope profiles of individual patients to improve diagnostics and individualize immunotherapy. We also discuss the advantages and pitfalls of the methodology used to identify and validate the mimotopes.
In the recent decades, great progress has been made in the development of ghrelin receptor ligands. The discovery of the first in vitro only active peptide growth hormone secretagogue derived from ...Met-enkephalin was the foundation for later discoveries of the receptor and the endogenous ligand ghrelin. Since then, the scope of peptides, peptidomimetics, and small-molecules targeting the ghrelin receptor, GHS-R1a, has expanded dramatically. Numerous agonists have been tested in animals and several in humans, and a handful have progressed to clinical trials for indications such as growth hormone release, gastric emptying, and cachexia. However, with the exception of the approval of GHRP-2 for diagnostic purposes in Japan, none of the candidates have been successfully introduced into the market. More recently, the attention of researchers has been concentrated on developing antagonists and inverse agonists for pharmacological treatment of the ever-expanding obese and overweight population. In this review, we describe the development of GHS-R1a targeting agonists, antagonists, and inverse agonists. We focus on current and completed clinical trials and the therapeutic potential of currently available ligands.
Lactic acid bacteria (LAB) have become increasingly studied over the last two decades as potential delivery systems for various biological molecules to the gastrointestinal tract. This article ...presents an overview of characteristics of LAB as delivery systems and of the applications
which have already been developed. The majority of LAB strains are able to survive the intestinal passage and some are also able to persist and colonize the intestine. Several strains were in fact described as members of the human commensal flora. They can interact with their host and are
able to deliver large molecular weight biomolecules across the epithelium via M-cells or dendritic cells. The most widely applied LAB species has been Lactococcus lactis; however species from genus Lactobacillus are gaining popularity and the first examples from genus Bifidobacterium
are starting to emerge. Bacteria are mostly applied live and enable continuous delivery of the biomolecules. However, killed bacteria (e.g. gram-positive enhancer matrix), with bound biomolecules or as adjuvants, are also being developed. The techniques for genetic modification of LAB are
well known. This review focuses on the delivery of recombinant proteins and DNA, which can cause either local or systemic effects. We divide recombinant proteins into antigens and therapeutic proteins. Delivery of antigens for the purpose of vaccination represents the most abundant application
with numerous successful demonstrations of the efficacy on the animal model. Therapeutic proteins have mostly been developed for the treatment of the inflammatory bowel disease, by the delivery of anti-inflammatory cytokines, or downregulation of pro-inflammatory cytokines. Delivery of allergens
for the modulation of allergic disorders represents the second most popular application of therapeutic proteins. The delivery of DNA by LAB was demonstrated and offers exciting opportunities, especially as a vaccine. New discoveries may eventually lead to the transition of LAB as delivery
systems in clinical practice.
To evaluate brazzein production in Lactococcus lactis using the nisin-controlled expression (NICE) system. The approach is through analysis of different plasmid/strain combinations. Two ...plasmid/strain combinations of the NICE system were used in brazzein expression: L. lactis NZ9000 harbouring plasmid pNZ8148, and L. lactis IL1403 harbouring plasmid pMSP3545. The former combination proved superior, with a >800-fold increase in His-tagged brazzein expression (to 1·65 mg l⁻¹ of fermentation broth), comparable to expression levels in Escherichia coli. Improved expression resulted in a minor increase in secretion to the medium with the use of the Usp45 signal peptide. The yield of wild-type brazzein corresponded to that of His-tagged brazzein. Wild-type brazzein was partially soluble and low-intensity sweetness was detected. The plasmid/strain combination of the NICE system has a significant impact on the expression of brazzein where a >800-fold increase was achieved. The greatly increased expression of brazzein resulted in minor improvement in secretion and low-intensity sweetness. The choice of the plasmid/strain combination of the NICE system was shown to be of extreme importance in brazzein expression.
Silver fir (Abies alba) bark extract contains a mixture of bioactive polyphenols. We tested their effectiveness in the treatment of psoriasis in order to further investigate the potential topical ...anti-inflammatory activity of polyphenols by means of a randomized, double-blind,
placebo-controlled add-on clinical trial, after having examined their ability to downregulate the expression of IL-1β cytokine in monocyte/macrophage primary cell culture. 61 patients with mild psoriasis met the inclusion criteria and were willing to comply with protocol requirements,
were enrolled in the study. The severity of the disease was measured by psoriasis area severity index (PASI). Treatment efficacy was evaluated by assessing erythema (E, 0 to 4-point scale), desquamation (D, 0 to 4-point scale) and induration (I, 0 to 4-point scale) of lesions before and after
the treatment. All patients enrolled in the study had symmetrical psoriasis plaques on the skin. All patients received O/V ointment with 2% of silver fir bark extract and/or placebo, respectively. We compared medications by right/left intra-patient comparison, so that the control group was
always contralateral of the tested one. Location of the tested or control site was randomised, using a computer-generated randomisation schedule. Silver fir extract was well-tolerated. A superiority of active treatment above placebo, based on the clinical investigational PASI score system
was observed by 15 % in all volunteers and in 40% regarding the improvement of psoriasis on elbows. However, statistical analysis showed no significant differences between placebo and active treatment with the extract from silver fir bark (p < 0.05).
The neutralisation of circulating ghrelin, the only known peripheral orexigenic peptide hormone, is a promising approach for the pharmacological treatment of obesity. To select peptides with an ...affinity towards ghrelin, 4 selection procedures were carried out with random peptide phage display libraries Ph.D.-7 and Ph.D.-12. Due to the absence of a common consensus motif, a stepwise elimination approach was used. The pool of selected peptides displaying phage clones was thoroughly examined to remove any potential target-unrelated peptides. The affinity of the remaining phage clones for ghrelin was tested with ELISA. An analysis of the binding properties revealed four-phage displayed peptides that bind to ghrelin with moderate affinity, with ADTVPRH and MEMKKTHPVLGA being the most specific. Additional advantage of peptide MEMKKTHPVLGA is an indication of binding to octanoyl group on N-terminal part of ghrelin, involved in receptor interaction. Hence peptide MEMKKTHPVLGA represents the most suitable lead for further investigation.
The aim of the present study was to evaluate the effect of long-term swimming exercise in conjunction with a high fat mixed lipid (HFML) diet on colon cancer (CC) development and lipid peroxidation ...in the large bowel. We used forty male Wistar rats, which were randomly divided into one control group and four cancer groups: sedentary and swimming groups fed a standard diet (LFCO) and sedentary and swimming groups fed an HFML diet. Corticosterone was determined during the experiment. After 6 months of swimming, the rats were sacrificed and blood, heart, liver, muscle and large bowel were taken for determining the activity of serum enzymes, antioxidant capacity and CC development. The results demonstrate that exercise has a protective role in CC development. Attenuated development of CC and increased levels of malondialdehyde (MDA) in the large bowel of exercised rats show that one of the protective effects of exercise on developing CC is induction of oxidative stress. However, in terms of the combined effects of dietary fat and exercise, our results indicate that the protective role of exercise on CC development is significantly depressed by an HFML diet. An HFML diet significantly reduced the protective influence of exercise on colon carcinogenesis in rats and affected the degree of peroxidation in the large bowel during exercise, as well as concentrations of serum enzymes (LDH, α-HBDH, CK, ALT and AST). Our results indicate that an HFML diet, which reflects the composition of a Western style diet, is a significant modifier of the protective effects of exercise on CC development in rats.
The aim of this study was to test the extract from silver fir wood (Belinal) on the reduction of the blood glucose concentrations after consumption of a standard meal. 31 healthy participants ...consumed 100 g of white bread 4 times (with 1 week washout period, consequently) concomitantly
with a capsule of Belinal, capsule of chestnut wood extract, placebo or acarbose (active control). Glucose and insulin in the blood were measured before and after the meal. The area under the curve of glucose concentration in blood after the meal was 35 % lower when Belinal was added
compared with the placebo group (p = 0.019). Acarbose lowered the area for 43 % (p = 0.002). By this, we proved that the effect of Belinal might be beneficial for prevention of diabetes. This is the first study that provides a scientific rationale for use of silver fir wood extract
as food supplement for reduction of health risks connected to type 2 diabetes mellitus.
To improve the production of sweet-tasting protein brazzein in Lactococcus lactis using controlled fermentation conditions. The nisin-controlled expression system was used for brazzein expression. ...The concentration of nisin for induction and the optical density (OD) at induction were therefore optimized, together with growth conditions (medium composition, pH, aerobic growth in the presence of hemin). Brazzein was assayed with ELISA on Ni-NTA plates and Western blot. Use of the M-17 medium, containing 2·5% glucose, anaerobic growth at pH 5·9 and induction with 40 ng ml⁻¹ nisin at OD 3·0 led to an approx. 17-fold increase in brazzein per cell production compared to non-optimized starting conditions. Aerobic growth in the presence of hemin did not increase the production. Considerable increase in brazzein per cell production was obtained at optimized fermentation conditions. Optimized growth conditions could be used in application of brazzein expression in L. lactis. The importance of pH and OD at induction contributes to the body of knowledge of optimal recombinant protein expression in L. lactis. The new assay for brazzein quantification was introduced.
Several animal models of breast cancer have been developed to study various aspects of breast cancer biology. Substantial evidence suggests that the
N
-methylnitrosourea (MNU) animal model mimics ...human breast cancer in many respects. It has therefore been used extensively to evaluate preventive and therapeutic agents for human breast cancer. Chemically induced rodent models are also suitable for studying malignant progression. Recently, Liska et al. 7 established two protocols of MNU administration depending on the animal’s age and number of applications of carcinogen, with the aim of investigating the advanced stages of mammary gland tumours. We used the same protocol as Liska but have obtained substantially different results. These results are presented and discussed in the frame of suggested key drawbacks of the MNU induced breast cancer rat model, as a contribution to the debate about the suitability of that model for evaluating preventive and therapeutic agents.
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