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Background: Clinical and experimental analyses indicate a pathognomonic role for allergen IgE crosslinking through epitope–paratope interactions as a major initial step in the cascade leading to effector cell activation and clinical manifestations of lgE-mediated food allergies. We aimed to undertake the initial development and assessment of Ara h 2-specific IgE epitope-like peptides that can bind to allergenspecific IgE paratopes and suppress effector cell activation. Methods: We performed biopanning, screening, IgE binding, selection and mapping of peptides. We generated synthetic peptides for use in all functional experiments. ImmunoCAP inhibition, basophil and mast cell activation tests, with LAD2 cells, a human mast cell line were performed. Twenty-six children or young adults who had peanut allergy were studied. Results: We identified and selected three linear peptides (DHPRFNRDNDVA, DHPRYGP and DHPRFST), and immunoblot analyses revealed binding to lgE from peanut-allergic individuals. The peptide sequences were aligned to the disordered region corresponding to the loop between helices 2 and 3 of Ara h 2, and conformational mapping showed that the peptides match the surface of Ara h 2 and h 6 but not other peanut allergens. In ImmunoCAP inhibition experiments, the peptides significantly inhibit the binding of IgE to Ara h 2 (p < .001). In basophil and mast cell activation tests, the peptides significantly suppressed Ara h 2-induced effector cell activation (p < .05) and increased the half-maximal Ara h 2 effective concentration (p < .05). Binding of the peptides to specific IgEs did not induce activation of basophils or mast cells. Conclusions: These studies show that the indicated peptides reduce the allergenic activity of Ara h 2 and suppress lgE-dependent basophil and mast cell activation. These observations may suggest a novel therapeutic strategy for food allergy based on epitope–paratop blocking.

The year 2020 will undoubtedly be marked by the coronavirus disease 2019 (covid-19) pandemic, caused by the novel coronavirus SARS-CoV-2. It has been shown that in diabetes patients, covid-19 occurs in a more severe form, with these patients being more prone to the need for mechanical ventilation and having higher mortality rates than non-diabetic patients. In the present article, we describe possible pathophysiological mechanisms that could explain a more severe course of covid-19 in diabetes patients. We also describe the recommendations for use and discontinuation of anti-diabetic drugs during infection, and continue by explaining how to adjust the management of this chronic disease during an epidemic. In conclusion, our own experience in organizing outpatient diabetes clinic during the covid-19 epidemic at the University Medical Centre Ljubljana is described. Covid-19 still carries many unknowns, so it will not surprise us, if we soon realize that the findings described so far, are outdated. Nonetheless, the present recommendations for the treatment of diabetes patients during the epidemic will remain in force and will hopefully help to improve and optimize the treatment of our patients, even after the pandemic.

Type 2 diabetes (T2D) management has reached a point where not only optimal glycaemic control is necessary, but also additional interventions with proven cardiovascular risk reduction benefit. Subcutaneous semaglutide has been shown to provide cardiovascular protection, but its use may be limited by its injection formulation. To overcome this limitation, an oral semaglutide tablet has been developed, which could potentially be of the same value as its injection counterpart, but in a much wider group of patients with T2D, thereby allowing for broader cardiovascular risk reduction in this vulnerable patient population.A total of 100 consecutive patients with T2D and a disease duration of up to 10 years, without manifest cardiovascular disease, who are treated with metformin (± sulphonylurea) and optimal cardioprotective therapy, will be recruited in a single-blinded, randomized trial named "Semaglutide Anti-atherosclerotic Mechanisms of Action Study (SAMAS)." After 1:1 randomization, patients will receive either oral semaglutide 14 mg daily or placebo for 1 year. The primary outcome comprises changes in atherosclerosis-related structural and functional characteristics of the arterial wall, namely: reduction of the carotid intima-media thickness, improvement of endothelial function and decrease in arterial stiffness. Secondary outcomes are changes in atherogenic small dense low-density lipoproteins, glucose control (HbA1c) and inflammatory markers (hsCRP). Possible correlations between primary endpoints and changes in lipids, HbA1c and high-sensitivity C-reactive protein will be sought.This is the first study to investigate the direct and indirect anti-atherosclerotic mechanisms of oral semaglutide. The results are expected to confirm the position of oral semaglutide in the multifactorial management of T2D with an emphasis on cardiovascular disease prevention.ClinicalTrials.gov Identifier: NCT05147896.

Bee venom immunotherapy is the main treatment option for bee sting allergy. Its major limitations are the high percentage of allergic side effects and long duration, which are driving the development of novel therapeutic modalities. Three general approaches have been evaluated including the use of hypoallergenic allergen derivatives, adjunctive therapy, and alternative delivery routes. This article reviews preclinical and clinical evidence on the therapeutic potential of these new therapies. Among hypoallergenic derivatives, hybrid allergens showed a markedly reduced IgE reactivity in mouse models. Whether they will offer therapeutic benefit over extract, it is still not known since clinical trials have not been carried out yet. T cell epitope peptides have proven effective in small clinical trials. Major histocompatibility complex class II restriction was circumvented by using long overlapping or promiscuous T cell epitope peptides. However, the T cell-mediated late-phase adverse events have been reported with both short and longer peptides. Application of mimotopes could potentially overcome both T cell- and IgE-mediated adverse events. During this evolution of vaccine, there has been a gain in safety. The efficacy was further improved with the use of Toll-like receptor-activating adjuvants and delivery systems. In murine models, the association of allergen Api m 1 with cytosine-guanosine rich oligonucleotides stimulated strong T-helper type-1 response, whereas its encapsulation into microbubbles protected mice against allergen challenge. An intralymphatic administration of low-dose vaccine has shown the potential to decrease treatment from 5 years to only 12 weeks. Bigger clinical trials are needed to follow up on these results.

Ragweed is a prominent cause of seasonal allergies. Thus far, information on IgE-binding sites of major allergen in ragweed pollen, Amb a 1, is very limited. A powerful experimental method to gain insights on the allergen epitopes is the selection of peptides from biological libraries that bind to anti-allergen antibodies. In this work, we aimed to map IgE epitopes of Amb a 1 using epitope-mimicking short peptides % mimotopes that were affinity-selected from phage-displayed random peptide libraries. The peptides weakly aligned with the Amb a 1 primary sequence, thus suggesting that the epitopes are conformational. When the peptides were mapped onto the surface of Amb a 1 homology model, the EpiSearch analysis predicted the location of four potential epitopic sites on surface patches centred at residues K104, S110, H214, and W312. The peptides matching to the predicted epitopes bound selectively to the IgE from pool of ragweed-allergic patients% sera and therefore represent mimetics of Amb a 1 IgE epitopes. The knowledge of IgE epitopes is a prerequisite for the rational design of molecular-based approaches to diagnosis and immunotherapy of allergic diseases. Ambrozija je pomemben vzrok sezonskih alergij. Do sedaj imamo zelo malo informacij o IgE vezavnih mestih na glavnem alergenu iz cvetnega prahu ambrozije, Amb a 1. Afinitetna selekcija peptidov iz bioloških knjižnic z uporabo specifičnih protiteles proti alergenu je uporabna laboratorijska metoda za določevanje epitopov. V tej raziskavi smo s pomočjo mimotopov, kratkih peptidov, ki posnemajo epitope, izoliranih iz bakteriofagnih knjižnic naključnih peptidov, določali IgE epitope Amb a 1. Izbrani peptidi so se le šibko ujemali s primarnim zaporedjem Amb a 1, kar je nakazovalo, da so epitopi konformacijski. Da bi jih določili, smo izdelali homologni model tridimenzionalne strukture Amb a 1, na katerega smo prilegali izbrane peptide. S pomočjo programa EpiSearch smo identificirali štiri potencialne epitope na površini alergena okoli aminokislinskih ostankov K104, S110, H214 in W312. Peptidni mimetiki predvidenih epitopov so se specifično vezali na IgE iz zmesi serumov za ambrozijo alergičnih pacientov in tako predstavljajo mimetike IgE epitopov Amb a 1. Poznavanje IgE epitopov je predpogoj za racionalno načrtovanje molekularnih pristopov v diagnostiki in imunoterapiji alergijskih bolezni.

Ragweed is a prominent cause of seasonal allergies. Thus far, information on IgE-binding sites of major allergen in ragweed pollen, Amb a 1, is very limited. A powerful experimental method to gain insights on the allergen epitopes is the selection of peptides from biological libraries that bind to anti-allergen antibodies. In this work, we aimed to map IgE epitopes of Amb a 1 using epitope-mimicking short peptides - mimotopes that were affinity-selected from phage-displayed random peptide libraries. The peptides weakly aligned with the Amb a 1 primary sequence, thus suggesting that the epitopes are conformational. When the peptides were mapped onto the surface of Amb a 1 homology model, the EpiSearch analysis predicted the location of four potential epitopic sites on surface patches centred at residues K104, S110, H214, and W312. The peptides matching to the predicted epitopes bound selectively to the IgE from pool of ragweed-allergic patients' sera and therefore represent mimetics of Amb a 1 IgE epitopes. The knowledge of IgE epitopes is a prerequisite for the rational design of molecular-based approaches to diagnosis and immunotherapy of allergic diseases.

Introduction. Cystic fibrosis (CF) is a genetic disorder of cystic fibrosis transmembrane conductance regulator (CFTR) resulting in production of thick mucus. The latest novelty in treatment of CF patients is triple combination of CFTR modulators (elexacaftor-tezacaftor-ivacaftor) that greatly improves CFTR function in epithelial cells leading to better pulmonary function. Objectives. CFTR is also located on endothelial cells and might be associated with endothelial dysfunction in CF. The aim of our single case study to explore the influence of CFTR modulators on endothelial function, which has yet to be investigated. Methods. Flow mediated dilation (FMD) is a well-established method for assessing endothelial function. FMD values below 4% are considered pathological. We evaluated FMD before and 1 month after introduction of elexacaftor-tezacaftor-ivacaftor in a 26-year-old female with advanced CF, who was among first patients in Slovenia to receive this treatment. Results. Patient’s condition quickly improved. She coughed up less mucus, her physical performance and pulmonary function improved (FEV1 increased from 26% to 38%), she no longer needed oxygen supplement during exercise and her need for pancreatic enzymes replacement therapy decreased. She reported no side effects. Before treatment her FMD value was dimished to 1.99%, one month after the initiation, the FMD value rose to 5.21%, reaching normal values for her age. Relative FMD improvement was as much as 261.8%. Conclusion. Endothelial function of our patient was severely impaired, as expected in CF. Its normalization was noted one month after initiation of the new treatment. The latter speaks in favour of the systemic effects of this drug, outside epithelial cells, specifically on endothelial cells.

Diabetes mellitus is a major healthcare problem. It is not only characterized by hyperglycemia and chronic complications, but in longer lasting diabetes and a longer living population, it is also associated with accelerated arterial ageing, which importantly contributes to cardiovascular complications. The accelerated arterial ageing in patients with diabetes should be considered separately from arterial ageing in patients without diabetes. Basic and clinical research have allowed better insight into the mechanisms of arterial ageing. In a simplified mechanistic way, it could be considered that the three tightly connected cornerstone characteristics of arterial ageing in patients with diabetes are: phenotypic presentation as endothelial dysfunction and arterial stiffness, and the underlying basic ageing-facilitating mechanism represented as the impaired expression of genetic longevity pathways. Currently, specific drugs for preventing/treating arterial ageing are not available. Therefore, we aimed to review the capacity of available drugs, particularly antidiabetic drugs, to interfere with the arterial ageing process. In the near future, these characteristics could help to guide therapy in patients with diabetes. Overall, it appears that arterial ageing could become a new target in diabetes. The expanding knowledge regarding the capability of antidiabetic drugs and other available drugs to inhibit/delay arterial aging is therefore essential.

Background: Favourable cardiovascular effects of SGLT2 inhibitors have been clearly revealed, whereas the underlying mechanism(s) still remain unknown. We explored the anti-oxidative action of empagliflozin (alone and in combination with metformin) in type 1 diabetes mellitus (T1DM) patients. Methods: Forty T1DM male patients (age 44.7 ± 2.5 years, BMI 28.3 ± 0.6 kg/m2) were equally randomised into four groups: 1) control (placebo), 2) empagliflozin (25 mg daily), 3) metformin (2000 mg daily) and 4) combination (empagliflozin 25 mg daily and metformin 2000 mg daily). Oxidative stress parameters were assessed at inclusion and after 12 weeks of treatment, comprising of total antioxidative status (TAS), superoxide dismutase (SOD), glutathione peroxidase (GPx), isoprostane, interleukin 6 (IL6) and advanced oxidation protein products (AOPP). One-way analysis of variance (ANOVA) with Bonferroni post-test was used for statistical analysis. Results: Empagliflozin in combination with metformin significantly improved antioxidative status parameters by increasing TAS for up to 1.07-fold (P Conclusion: We found that empagliflozin in combination with metformin provided powerful antioxidative defense in T1DM patients. The results could at least partially explain the beneficial effects of the two drugs beyond glucose control. Antioxidant activity should be focused on in further large clinical studies. Disclosure M. Janic: None. M. Lunder: None. M. Sabovic: None. A. Janez: None.

Background: Optimal glycemic control and proper basal insulin initiation timing when non-insulin treatment becomes insufficient is critical in type 2 diabetes mellitus (T2DM). The DUNE study investigated the effectiveness and safety of newly initiated basal insulin in T2DM patients compared to those who already received insulin. The impact of background therapy with sulfonylureas (SU) on insulin dose and hypoglycemia occurrence was also explored. Methods: The DUNE study was a 12-week, prospective, single-arm, observational study of adults with T2DM. The present sub-analysis included the patients from the Balkan region (Slovenia and Serbia). The first group (n=83) included patients with newly initiated basal insulin and the second group (n=94) included patients who were already treated with basal insulin for up to 12 months. Patients also received other non-insulin antihyperglycemic drugs. Results: The proportion of patients receiving SUs was higher in the group with newly initiated basal insulin (72.3% vs. 55.3%; p Conclusions: Continuation of SUs in T2DM patients, both newly initiated and already initiated with basal insulin therapy (NPH or first-generation insulin analog), should be considered with caution due to the potentially enhanced effect on the risk of hypoglycemia. Disclosure A. Janez: None. M. Lunder: None. M. Janic: None. N. Grulovic: None. D. Zdravkovic: None. A.Z. Jotic: None. K. Lalic: Speaker’s Bureau; Self; AstraZeneca, Boehringer Ingelheim, Merck, Novo Nordisk, Sanofi. N. Lalic: None.

Background Statins and sartans can, in therapeutic doses, induce pleiotropic cardiovascular effects. Similar has recently been shown also for sub-therapeutic doses. We thus explored and compared the cardiovascular pleiotropic efficacy of sub-therapeutic vs. therapeutic doses. Methods Wistar rats were randomly divided into 7 groups receiving fluvastatin, valsartan and their combination in sub-therapeutic and therapeutic doses, or saline. After 6 weeks, the animals were euthanised, their hearts and thoracic aortas isolated, and blood samples taken. Endothelium-dependent relaxation of the thoracic aortae and ischaemic-reperfusion injury of the isolated hearts were assessed along with the related serum parameters and genes expression. Results Fluvastatin and valsartan alone or in combination were significantly more effective in sub-therapeutic than therapeutic doses. The sub-therapeutic combination greatly increased thoracic aorta endothelium-dependent relaxation and maximally protected the isolated hearts against ischaemia-reperfusion injury and was thus most effective. Beneficial effects were accompanied by increased levels of nitric oxide (NO) and decreased levels of asymmetric dimethylarginine (ADMA) in the serum (again prominently induced by the sub-therapeutic combination). Furthermore, nitric oxide synthase 3 (NOS3) and endothelin receptor type A (EDNRA) genes expression increased, but only in both combination groups and without significant differences between them. In the therapeutic dose groups, fluvastatin and valsartan decreased cholesterol values and systolic blood pressure. Conclusion Sub-therapeutic doses of fluvastatin and valsartan are more effective in expressing cardiovascular pleiotropic effects than therapeutic doses of fluvastatin and/or valsartan. These results could be of significant clinical relevance.

Previously we revealed the effectiveness of a new therapeutic approach with a short-term, very-low dose fluvastatin-valsartan combination on the improvement of arterial function in type 1 diabetes mellitus patients (T1DM). In this study we explored whether this approach influences inflammation and oxidative stress and explored any association of these effects with arterial function improvement.This was a supplementary analysis of the two previous double blind randomized studies (included 44 T1DM patients). Treatment group received very-low dose fluvastatin-valsartan, the control group received placebo. Blood samples were collected and inflammation parameters: high-sensitivity CRP (hsCRP), interleukin 6 (IL-6), vascular cell adhesion molecule-1 (VCAM-1) and oxidative stress parameter total antioxidant status (TAS) were measured.Treatment decreased hsCRP values (by 56.5%, P0.05) and IL-6 values (by 33.6%, P0.05) and increased TAS values (by 21.1%; P0.05) after 30days of treatment. High sensitivity CRP and TAS remained decreased 3months after treatment discontinuation. Importantly, the anti-inflammatory and anti-oxidative action significantly correlated with arterial function improvement.The approach consisting of short-term (30days) treatment with a very low-dose fluvastatin-valsartan combination acts anti-inflammatory and anti-oxidative in T1DM patients. These observations along with the improvement of arterial function support the assumption that this approach could have an important clinical benefit in T1DM patients.

Objective: Integrative diabetes care encompasses Patient Reported Outcome Measures (PROMs); instruments to measure patients’ perceptions of treatment options. In T1D and T2D patients with well controlled diabetes on MDI, introduction of new means of glucose monitoring (e.g., CGM) should not impose additional psychological burden regardless supposed added benefit in glucose control. The aim of present study was to investigate CGM as a tool for optimizing and personalizing treatment goals in elderly. Methods: Elderly patients (n=25, age 67±6 years, HbA1c=7.1±0.7%) treated with MDI were instructed to use a CGM device. Blinded CGM was followed by real-time CGM use. Glycemic control metrics (time in range (TIR), glycemic variability expressed by Coefficient of Variation (CV)) were compared by paired samples T-test. PROMs were measured by questionnaires. Results: Significant improvements in TIR (3.9 - 10.0 mmol/L) (66,3±12,4% vs. 76,9±14,1%; p Conclusion: Introduction of CGM use in well controlled diabetes patients on MDI results in more stable glycemia and more time spent in target range; without additional diabetes treatment related emotional burden detected. Despite confronted with novel diabetes technology, the elderly reported of positive subjective outcomes and were keen to adopt CGM. Disclosure S. Volcansek: None. M. Lunder: None. A. Janez: None.

The incidence of aging-related disorders may be decreased through strategies influencing the expression of longevity genes. Although numerous approaches have been suggested, no effective, safe, and easily applicable approach is yet available. Efficacy of low-dose fluvastatin and valsartan, separately or in combination, on the expression of the longevity genes in middle-aged males, was assessed. Stored blood samples from 130 apparently healthy middle-aged males treated with fluvastatin (10 mg daily), valsartan (20 mg daily), fluvastatin-valsartan combination (10 and 20 mg, respectively), and placebo (control) were analyzed. They were taken before and after 30 days of treatment and, additionally, five months after treatment discontinuation. The expression of the following longevity genes was assessed: SIRT1, PRKAA, KLOTHO, NFE2L2, mTOR, and NF-κB. Treatment with fluvastatin and valsartan in combination significantly increased the expression of SIRT1 (1.8-fold; p < 0.0001), PRKAA (1.5-fold; p = 0.262) and KLOTHO (1.7-fold; p < 0.0001), but not NFE2L2, mTOR and NF-κB. Both fluvastatin and valsartan alone significantly, but to a lesser extent, increased the expression of SIRT1, and did not influence the expression of other genes. Five months after treatment discontinuation, genes expression decreased to the basal levels. In addition, analysis with previously obtained results revealed significant correlation between SIRT1 and both increased telomerase activity and improved arterial wall characteristics. We showed that low-dose fluvastatin and valsartan, separately and in combination, substantially increase expression of SIRT1, PRKAA, and KLOTHO genes, which may be attributed to their so far unreported pleiotropic beneficial effects. This approach could be used for prevention of ageing (and longevity genes)–related disorders.

BACKGROUND Octocrylene (OCT) is one of the most widespread chemical UV filters used in sunscreens and cosmetic products. Despite the use of sunscreens and personal care products over decades, melanoma as the most serious and aggressive form of skin cancer is still a cause of concern. Hence the aim of this study was to investigate any potential influence of OCT on metabolic activity, cytotoxicity and ABCB5 mRNA expression in melanoma cells. The ABCB5 transmembrane protein was tested due to its well-known role in the initiation, invasion and metastatic spread of various cancers, including melanoma. METHODS Metastatic melanoma cell line WM-266-4 (ATCC) was incubated with selected concentrations of OCT and for different time intervals. The MTT and LDH assays to measure the cells' metabolic activity and cytotoxicity were used respectively. Target gene (ABCB5) expression was detected by quantitative real-time PCR (qRT-PCR), using TaqMan® chemistry. RESULTS Our results suggest decreased metastatic melanoma cells' metabolic activity, increased cytotoxicity and increased ABCB5 mRNA expression (P

Pojavnost kardiometabolnih bolezni je dosegla pandemične razsežnosti, zato se čedalje večja pozornost posveča tudi z življenjskim slogom pogojenemu zdravju prebivalstva, še posebej načinu prehranjevanja in vnosu določenih skupin živil. Številne epidemiološke raziskave so dokazale, da je uživanje pijač z dodanim sladkorjem povezano z nastankom debelosti, sladkorne bolezni in srčno-žilnih bolezni, dodatno je tudi dober pokazatelj posameznikovega nezdravega življenjskega sloga. Pijače z dodanim sladkorjem so vir tako imenovanih »praznih kalorij«, ki nimajo hranilne vrednosti in predstavljajo največji dodatni vir energije in vnosa dodanih sladkorjev, še posebej fruktoze. Zmanjšanje vnosa pijač z dodanim sladkorjem dokazano vodi v znižanje telesne mase in zmanjšanje tveganja za nastanek kardiometabolnih bolezni. Uživanje pijač z dodanim sladkorjem je uvrščeno kar med 15 najpogostejših dejavnikov tveganja med kazalci nezdravega življenjskega sloga. Posledično je smiselno intenzivno ozaveščanje o pomembnosti opustitve uživanja pijač z dodanim sladkorjem, še posebej pri osebah s povečanim tveganjem za presnovne bolezni in mladih z nezdravim življenjskim slogom. V prispevku so opredeljene pijače z dodanim sladkorjem, opisana je njihova povezava z debelostjo, sladkorno boleznijo in srčno-žilnimi boleznimi ter možni zdravi nadomestki.

Background : Silver fir trunk extract (SFTE) is a complex mixture of antioxidative polyphenols (lignans and phenolic acids) from the trunks of silver fir trees ( Abies alba , lignum). In our previous study, we have shown that SFTE exerts strong antioxidative and protective effects against atherogenic, diet-induced arterial wall damage. Objective : The aim of the present study was to test the potential protective effects of SFTE and its compounds, two phenolic acids (p-coumaric and protocatechuic acids) in ischemia–reperfusion injury of isolated rat hearts. Design : Isolated hearts of Wistar rats aged 4–8 weeks were exposed to perfusion, ischemia, and reperfusion periods. The experiments were performed using the following five groups: control, SFTE (10 µg/L), SFTE (100 µg/L), protocatechuic acid, and p-coumaric. Aortas were isolated to measure vascular responses in the presence of N ω -Nitro- L -arginine. Results : SFTE dose-dependently reduced ischemic-reperfusion heart damage, which was indicated as the decrease in the lactate dehydrogenase (LDH) release rate and arrhythmias duration by 80% and an increase in coronary flow rate during the reperfusion period. Two tested compounds (p-coumaric and protocatechuic acids) acted less cardioprotective, since they decreased the duration of arrhythmias only by 40 and 45%, respectively, and did not decrease LDH release rates during the reperfusion period. Only p-coumaric acid increased coronary flow rates, whereas protocatechuic acid did not. Conclusions : We conclude that the SFTE exerted the strongest cardioprotective effect, whereas its constituents (the p-coumaric and protocatechuic acids) were less effective in inducing cardioprotection. Keywords: Abies alba; silver fir trunk extract; cardioprotective; rat heart; p-coumaric acid; protocatechuic acid (Published: 18 October 2016) Citation: Food & Nutrition Research 2016, 60: 29623 - http://dx.doi.org/10.3402/fnr.v60.29623

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.

Background Deteriorated arterial function and high incidence of cardiovascular events characterise diabetes mellitus. Metformin and recent antidiabetic drugs, SGLT2 inhibitors, reduce cardiovascular events. We explored the possible effects of empagliflozin’s effect on top of metformin treatment on endothelial function and arterial stiffness parameters in type 1 diabetes mellitus (T1DM) patients. Methods Forty T1DM patients were randomised into three treatment groups: (1) empagliflozin (25 mg daily), (2) metformin (2000 mg daily) and (3) empagliflozin/metformin (25 mg daily and 2000 mg daily, respectively). The fourth group received placebo. Arterial function was assessed at inclusion and after 12 weeks treatment by: endothelial function [brachial artery flow-mediated dilation (FMD), reactive hyperaemia index (RHI)], arterial stiffness [pulse wave velocity (PWV) and common carotid artery stiffness (β-stiffness)]. For statistical analysis one-way analysis of variance with Bonferroni post-test was used. Results Empagliflozin on top of metformin treatment significantly improved endothelial function as did metformin after 12 weeks of treatment: FMD [2.6-fold (P Conclusion Empagliflozin on top of metformin treatment significantly improved arterial stiffness compared to metformin in T1DM patients. Endothelial function was similarly improved in all treatment groups. Empagliflozin seems to possess a specific capacity to decrease arterial stiffness, which could support its cardioprotective effects observed in large clinical studies. Trial registration Clinical trial registration: NCT03639545

The diet rich in fruits and vegetables reduces the risk of metabolic syndrome, including diabetes development by various mechanisms of action, mainly due to the presence of polyphenolic compounds. Extracts from different conifer species are known to be a rich source of various polyphenols. In the present study we elucidated the in vitro mechanism of anti-diabetic activity of silver fir (Abies alba) wood and bark extracts and compared their activity to non-coniferous sweet chestnut wood extract and standardized maritime pine bark extract. Extracts and lignans were tested for their inhibitory activity of enzymes involved in the regulation of blood glucose in vitro. The ability of extracts to protect against oxidative stress in high glucose environment was tested on mouse myoblast cell line. Silver fir wood and bark extracts were shown to be effective inhibitors of α-glucosidase, α-amylase and dipeptidyl peptidase 4, three enzymes involved in the regulation of blood glucose levels. Coniferous extracts also showed protection against oxidative stress generated in high glucose environment. Lignans, particularly pinoresinol diglucoside, isolariciresinol and secolariciresinol were shown to be important contributors of antihyperglycemic activity through inhibition of dipeptidyl peptidase 4. This corroborates previously published in vivo results on blood glucose level obtained with silver fir wood extract and supports the use of silver fir wood and bark extracts as food supplements or functional foods in borderline diabetes.

Sponges (Porifera) currently represent one of the richest sources of natural products and account for almost half of the pharmacologically active compounds of marine origin. However, to date very little is known about the pharmacological potential of the sponges from polar regions. In this work we report on screening of ethanolic extracts from 24 Antarctic marine sponges for different biological activities. The extracts were tested for cytotoxic effects against normal and transformed cell lines, red blood cells, and algae, for modulation of the activities of selected physiologically important enzymes (acetylcholinesterase, butyrylcholinesterase, and α-amylase), and for inhibition of growth of pathogenic and ecologically relevant bacteria and fungi. An extract from Tedania (Tedaniopsis) oxeata was selectively cytotoxic against the cancer cell lines and showed growth inhibition of all of the tested ecologically relevant and potentially pathogenic fungal isolates. The sponge extracts from Isodictya erinacea and Kirkpatrickia variolosa inhibited the activities of the cholinesterase enzymes, while the sponge extracts from Isodictya lankesteri and Inflatella belli reduced the activity of α-amylase. Several sponge extracts inhibited the growth of multiresistant pathogenic bacterial isolates of different origins, including extended-spectrum beta-lactamase and carbapenem-resistant strains, while sponge extracts from K. variolosa and Myxilla (Myxilla) mollis were active against a human methicillin-resistant Staphylococcus aureus strain. We conclude that Antarctic marine sponges represent a valuable source of biologically active compounds with pharmacological potential.

Diet, rich in plant polyphenols prevents atherogenesis that manifests as reduced vascular relaxation and formation of plaques.Atherosclerosis could be reduced by the intake of silver fir (Abies alba) extract (SFTE), rich in polyphenols.Chronic, in vivo treatment animal study.Guinea pigs (Cavia porcellus) were fed for 8 weeks with one of the following three diets: atherogenic, basic or atherogenic + SFTE. After isolation, we measured the relaxation and contractile responses of the thoracic aorta. Additionally, we measured the area of fatty plaques on the aortic walls.Compared to the basic diet, the atherogenic diet decreased the ability of the aorta to relax by 63% (p0.001). The addition of SFTE to the atherogenic diet improved the aorta relaxation response compared to that of the atherogenic diet without SFTE (the decrease relative to the basic diet was 26%, p0.001). The aorta contractility did not differ between the groups. The SFTE group generated significantly fewer atherosclerotic plaques than did the atherogenic group. The areas of atherosclerotic plaques were 7.4, 0.3 and 1.6% in the aortas of guinea pigs receiving atherogenic, basic or atherogenic + SFTE diets, respectively.In a guinea pig model, prolonged treatment with antioxidative polyphenol-rich SFTE prevents aortic functional and morphological changes caused by an atherogenic diet.

Improvement of arterial wall (AW) characteristics decreases cardiovascular risk. In a previous study, it was observed that AW characteristics in patients with diabetes mellitus type 1 are significantly improved by short-term treatment with a low-dose combination of fluvastatin and valsartan. Additionally, a unique phenomenon of prolonged effect after treatment discontinuation was suggested. The present study tested whether repeated treatm ent after a certain period results in the same beneficial effect, th ereby advancing the hypothesis that cyclic treatment can provide a long-term improvement of AW characteristics. A total of 44 patients with diabetes mellitus type 1 that participated in the previous study were recruited. Six months after the discontinuation of the initial treatment, the same treatment with a low-dose fluvastatin (10 mg daily) and valsartan (20 mg daily) combination (n=22) or placebo (n=22) was repeated. Brachial artery flow-mediated dilation (FMD), pulse wave velocity (PWV) and carotid artery β-stiffness were measured. It was found that the beneficial effect achieved with an initial 1-month treatment was completely regained following treatment repetition: FMD improved by 50.9% (P

Aims/Introduction Acute glucose fluctuations during the postprandial period pose great risk for cardiovascular complications and thus represent an important therapeutic approach in type 2 diabetes. In the present study, screening of peptide libraries was used to select peptides with an affinity towards mammalian intestinal alpha-glucosidase as potential leads in antidiabetic agent development. Materials and Methods Three phage-displayed peptide libraries were used in independent selections with different elution strategies to isolate target-binding peptides. Selected peptides displayed on phage were tested to compete for an enzyme-binding site with known competitive inhibitors, acarbose and voglibose. The four best performing peptides were synthesized. Their binding to the mammalian alpha-glucosidase and their effect on enzyme activity were evaluated. Results Two linear and two cyclic heptapeptides with high affinity towards intestinal alpha-glucosidase were selected. Phage-displayed as well as synthetic peptides bind into or to the vicinity of the active site on the enzyme. Both cyclic peptides inhibited enzyme activity, whereas both linear peptides increased enzyme activity. Conclusions Although natural substrates of glycosidase are polysaccharides, in the present study we successfully isolated novel peptide modulators of alpha-glucosidase. Modulatory activity of selected peptides could be further optimized through peptidomimetic design. They represent promising leads for development of efficient alpha-glucosidase inhibitors.

In Diabetes Mellitus (DM), hyperglycaemia and insulin resistance progressively lead to both microvascular and macrovascular complications. Whereas the incidence of microvascular complications is closely related to tight glycaemic control, this does not apply to macrovascular complications. Hyperglycaemia influences many interweaving molecular pathways that initially lead to increased oxidative stress, increased inflammation and endothelial dysfunction. The latter represents the initial in both types of vascular complications; it represents the “obligatory damage” in microvascular complications development and only “introductory damage” in macrovascular complications development. Other risk factors, such as arterial hypertension and dyslipidaemia, also play an important role in the progression of macrovascular complications. All these effects accumulate and lead to functional and structural arterial wall damage. In the end, all factors combined lead to the promotion of atherosclerosis and consequently major adverse cardiovascular events. If we accept the pivotal role of vascular wall impairment in the pathogenesis and progression of microvascular and macrovascular complications, treatment focused directly on the arterial wall should be one of the priorities in prevention of vascular complications in patients with DM. In this review, an innovative approach aimed at improving arterial wall dysfunction is described, which may show efficacy in clinical studies. In addition, the potential protective effects of current treatment approaches targeting the arterial wall are summarised.

Metformin represents the first line of treatment and is the most widely prescribed antihypergycemic drug in type 2 diabetic patients. It can be used as monotherapy or in combination with other oral antihyperglycemic drugs or insulin. Additionally, it is also prescribed in type 1 diabetic patients, it proved to be effective in prediabetes and also provided beneficial effects in other insulin resistant states, for example in polycystic ovary syndrome. Nevertheless, the exact molecular mechanism of its action remains unknown. It was shown that it inhibits liver gluconeogenesis, facilitates glucose uptake into peripheral tissues, such as striated muscle; it also acts in the gut. Besides antihyperglycemic effects, metformin was also shown to possess several beneficial, protective effects, so-called pleiotropic effects: particularly on the cardiovascular system and in cancer patients. Metformin has only few side effects, the most serious being metformin-associated lactic acidosis. The latter appears in rare clinical cases with pre-existing chronic kidney disease or advanced heart failure with tissue hypoperfusion, which consequently represent relative contraindications for metformin use. In the past, metformin treatment was usually discontinued when performing iodine contrast imaging, however recently there is evidence of its safety even in patients with higher stages of chronic kidney disease. All in all, metformin is a drug with a long tradition and a promising future.

New preventive strategies for atherosclerosis are needed. In this study, we tested whether a new therapeutic approach consisting of low-dose treatment with a statin and sartan combination could prevent atherogenic diet-induced impairment of the arterial wall in guinea pigs. Twenty-five Dunkin-Hartley guinea pigs were randomly assigned to five experimental groups: 1) normal diet; 2) atherogenic diet (AD); 3) AD + a low-dose atorvastatin and valsartan combination (5mg/kg/day and 2.4mg/kg/day, respectively); 4) AD + low-dose atorvastatin (5mg/kg/day); 5) AD + low-dose valsartan (2.4mg/kg/day). After 8 weeks of treatment, the animals were killed, blood samples collected and thoracic and abdominal aortas isolated. The atherogenic diet significantly impaired maximal thoracic aorta endothelium-dependent relaxation by 40.1% relative to the normal diet. The low-dose combination, compared to the separate drugs, completely preserved thoracic aorta endothelium-dependent relaxation at the level of the group receiving normal diet. This substantial effect was associated with a significant change in the expression of NOS3 (R=0.93; P=0.0002) and IL1b (R=-0.79; P=0.003) genes. In addition, treatment with the low-dose combination or the separate drugs also prevented atherosclerotic plaque formation. We found that treatment with the low-dose atorvastatin and valsartan combination has the capability to completely protect the arterial wall from atherogenic diet-induced damage in the guinea pig model. Further studies evaluating this new therapeutic approach are desirable.

Background Statin use is frequently associated with muscle-related symptoms. Coenzyme Q10 supplementation has yielded conflicting results in decreasing statin myopathy. Herein, we tested whether coenzyme Q10 supplementation could decrease statin-associated muscular pain in a specific group of patients with mild-to-moderate muscle symptoms. Material/Methods Fifty patients treated with statins and reporting muscle pain were recruited. The Q10 group (n=25) received coenzyme Q10 supplementation over a period of 30 days (50 mg twice daily), and the control group (n=25) received placebo. The Brief Pain Inventory (BPI) questionnaire was used and blood testing was performed at inclusion in the study and after 30 days of supplementation. Results The intensity of muscle pain, measured as the Pain Severity Score (PSS), in the Q10 group was reduced from 3.9±0.4 to 2.9±0.4 (P