Sambucus ebulus - from traditional medicine to recent studies

Sambucus ebulus L. (Dwarf elder) is an herbaceous plant well-known in traditional medicine of Bulgaria and other countries from Central Europe to the Middle East. It is used for treatment of inflammation-related gastrointestinal disorders, influenza, kidney and lung diseases, and rheumatoid arthritis as well as of wounds, snake and insect bites. Recent research of Dwarf elder shows a high content of total polyphenols, anthocyanin, quercetin and vitamin C, which may be related to its antioxidant activity. Compounds with antiinflammatory and antinociceptive action such as chlorogenic and ursolic acids in the herb have been established, too. Some studies reveal anticancer properties of the herb, and plant ebulins are considered as the main compounds with anticancer properties. Fruit extracts modulate the gene expression of antioxidant enzymes in a preadipocyte cell culture model. Data available indicate that Dwarf elder is a good source of phytotherapeutic means to treat pathological conditions related to inflammation and oxidative stress.Scripta Scientifica Medica 2013; 45(2): 36-42

Antidiabetic Potential of Plants Used in Bulgarian Folk Medicine and Traditional Diet

The idea of this chapter is that currently available antidiabetic drugs specifically target several points of the T2D pathophysiology but they do not cover all aspects of the disease. In addition, many adverse effects of synthetic antidiabetic agents have been reported. The suggested manuscript is an overview of the available scientific literature focused on antiobesity and antidiabetic potential of selected 42 medicinal and edible plants of the Bulgarian flora. Most of the reports reveal the effect of extracts or their active components on specific biochemical mechanisms. Mechanistic data about hypoglycemic and hypolipidemic action are presented for some of the plants. An essential part of this review is dedicated to the target mechanisms behind the effects of the selected plant species. The authors hope that this review will serve as a starting point for future investigations with a contribution to the prevention and therapy of diabetes

Changes in COX-2, iNOS and GCL gene expression in 3T3-L1 preadipocytes incubated in macrophage conditioned medium

PURPOSE: Obesity is recognized as a major risk factor for a number of diseases, including metabolic syndrome and type 2 diabetes mellitus. It is characterized with increased macrophage infiltration in adipose tissue. Macrophages secrete numerous inflammatory mediators and contribute to the increased inflammatory status of adipose tissue which is associated with increased reactive oxygen species generation and oxidative stress. In cell biology conditioned nutrient mediums (CMs) are used to assess the complex impact of the secretory products of a particular cell type on the behaviour of other cells. The aim of the present study was to examine the effect of CM obtained from J744A.1 macrophage cell culture on expression in 3T3-L1 preadipocytes of some enzymes related to antioxidant defense and the inflammatory response.MATERIAL AND METHODS: mRNA levels of inducible cyclooxygenase (COX-2), inducible NO synthase (iNOS) and glutamate-cysteine-ligase (GCL) were measured. 3T3-L1 cells were treated with increasing CM concentrations (10-30%) for 24 hours. Changes in gene expression levels were analyzed by real-time qPCR method. Calculations were performed using 2-ΔΔCt method.RESULTS: The higher CM concentration contributed to increased expression levels of the genes examined. Treatment with CM in concentrations of 10, 20 and 30% resulted in 2.3-, 2.6- and 2.7-fold increase in COX-2 expression (p<0.01), respectively. Both 10% and 20% CM up-regulated iNOS by 1.7 times (p<0.001). The highest CM concentration of 30% stimulated the enzyme by two times (p<0.01). GCL mRNA levels were by 2.5 times higher than these of the untreated controls when stimulated with 30% CM (p<0.05).CONCLUSION: CM from J744A.1 cell culture may induce inflammatory and oxidative stress-related response in 3T3-L1 preadipocytes thus demonstrating a possible link between macrophage infiltration and local inflammation in adipose tissue.Scripta Scientifica Medica 2013; 45(2): 32-35

Ethanol induces IL-6 and TNF-α cytokine and iNOS and COX-2 enzyme gene expression in 3T3-L1 preadipocytes

t is known that alcohol influences on human health and its effects are widely studied. Adipose tissue (AT) may be an important target of ethanol action. Data about the effect of ethanol on cytokine and pro-inflammatory enzymes` gene expression in adipocytes are scarce. The aim of the present study was to establish its effect on transcription of inflammatory markers IL-6, TNF-α, COX-2 and iNOS in cultured 3T3-L1 cells. MTT test was performed in order to determine ethanol effect on cell viability. Expression of selected genes was measured using two step quantitative PCR. Results were analyzed using 2-ΔΔCt method. Exposure of 3T3-L1 cells to increasing ethanol concentrations (0.125%-0.625%) resulted in gradual cell viability decline. Expression of all the genes studied was up-regulated upon ethanol treatment. In cells treated with 0.5% ethanol, there was a 4.8- (p < 0.001), 6.3- (p<0.001), 5.5- (p<0.05) and 3.6- (p<0.05) fold increase of mRNA levels of IL-6, TNF-α, iNOS and COX-2, respectively, as compared with untreated controls. mRNA levels of the respective genes highly correlated with the applied ethanol concentration. In conclusion, our results show that ethanol treatment may increase the inflammatory potential of AT up-regulating expression of inflammatory cytokines IL-6 and TNF-α and inflammatory enzymes iNOS and COX-2 on transcription level.Scripta Scientifica Medica 2012; 44(2): 31-35

Increasing detection sensitivity of low copy number transcripts through preamplification of cDNA molecules

With the advances in the field of molecular biology and its applications in the clinical practice during the recent years, it has become crucial to perform molecular analyses on limited amount of tissue obtained through biopsies. Additionally, tissue is fixed in formalin and further embedded in paraffin (FFPE), a pro­cedure which causes extensive degradation of nucleic acids, mainly RNAs. Furthermore, when studying gene expression profiles of a set of genes, which present physiologically low expression, the number of transcripts is low and cannot be detected. In this study we focus on amplifying effectively cDNA molecules synthesized from small amounts of initial RNA before analyzing the expression levels through quantitative polymerase chain reaction (qPCR). By introducing the preamplification step in the procedure, we achieved highly effi­cacious detection and quantification of expression levels of low-expression genes

Improving silica-based spin-column efficiency for RNA isolation from FFPE tissue

jkhkjhRNA isolation from formalin-fixed paraffin embedded tissue has been a target for improvement for many years. The major limitation is the highly degraded RNA, which in turn increases the demand of the highest yields possible during isolation procedures. In this study, we compared the purification efficacy of RNA between the traditional ethanol precipitation and a modified protocol, which includes the use of silica-based spin-columns for RNA purification and recovery. We also modified and assessed the efficacy of a protocol for effective decontamination and regeneration of spin-columns

Cytoprotective and cytoproliferative effects of ethanolinfusions from Morus nigra and Cotinus coggygria heartwood in 3T3-L1 preadipocytes

PURPOSE: The purpose of the present study was to examine the effect of two ethanol infusions from the heartwood of mulberry tree (Morus nigra) and smoke tree (Cotinus coggygria) on viability of preadipocyte cells. MATERIAL ANDMETHODS: Preadypocyte 3T3-L1 cells were treated with 40% ethanol infusions from the heartwood of both species, as well as with 40% ethanol as a control. MTT-test was applied to assess cell viability.RESULTS: Ethanol infusions increased cell viability as compared to ethanol-treated controls. Along with the cytoprotective effect, also cytoproliferative effect was established for both infusions.CONCLUSION: Estimated cytoprotective percentage values of infusion in the nutrient medium could be used in further investigations aimed to explore biological effects of ethanol heartwood infusions from M. nigra and C. coggygria

Stress-tolerance tests and postprandial low-grade inflammation response

Both tumor necrosis factor alpha (TNF alpha) and interleukin 6 (IL-6) are considered as proinflammatory cytokines which display marked daily and postprandial variations. The low grade inflammation is known to contribute to the development of certain states such as type 2 diabetes, metabolic syndrome and related disorders. Oral glucose tolerance test (OGTT) has been applied to diagnose diabetes and is widely used to estimate the insulin sensitivity in the postprandial phase. Application of tolerance tests with high lipid content such as oral lipid tolerance test (OLTT), high-fat meal (HFM) and mixed meal tests are informative for the metabolic response to complex diets. Postprandial hyperlipidemia and hyperglycemia are important and residual risk factors especially in patients with diabetes mellitus and related metabolic disturbances. There are data suggesting that low grade inflammation could be mediated by hyperglycemia and dyslipidemia. The aim of the review is to summarize how different challenge tests influence the postprandial circulation levels of IL-6 and TNF alpha

Challenge tests and their application in nutrition research to investigate phenotypic flexibility

От гледна точка на метаболизма оптималното здраве се характеризира като способността на организма да поддържа и възстановява хомеостаза в непрекъснато променящата се околна среда и особено в отговор на широк спектър от стресови фактори. Капацитетът на организма да се адаптира във времето и пространството към всички променливи, особено към вариациите в диетата, се нарича `фенотипна пластичност`. Проследяването на метаболитните промени в биологични проби в хода на стрес тестове би предоставило информация за способността на организма да се справя с ежедневните промени в храненето. Стрес тестове, базирани на суплементиране с въглехидрати, липиди, протеини и/или комбинация от тях, се използват в клиничната практика за диагностика на метаболитни нарушения. Общото в тях е, че те водят до временно нарушаване на хомеостазата на тялото. Степента на нарушенията и скоростта, с която се възстановява хомеостазата, се използват като индикатори за здравния статус. Счита се, че нормалният глюкозен толеранс е показател за добра фенотипна пластичност. Оралният глюкозотолерантен тест (ОГТТ) е най-често прилаганият стрес тест, използван от десетилетия за диагностициране на диабет. Въпреки че триглицеридите на гладно са независим рисков фактор за някои заболявания, редица автори предлагат проследяването на постпрандиалните им нива и промяната в метаболитите в хода на приложен орален липидотолерантнен тест (ОЛТТ) за по-точна диагностика. Настоящото проучване разглежда възможността и потенциала на ОГТТ, ОЛТТ, тестове със смесено хранене и техни вариации да бъдат използвани като индикатори за оценка на фенотипната пластичност при проучвания в областта на храненето и оценка на здравния статус на индивида.From a metabolic point of view, the optimum health is the ability of the organism to maintain and regain homeostasis in ever-changing environmental conditions and to a wide spectrum of stressors. The capacity of the organism to adapt to all the variables, especially to the dietary variations is called `phenotypic flexibility`. Metabolic profiling of biological samples during a challenge response phase should offer a profound view on the flexibility of a phenotype to cope with daily stressors. Stress tests based on supplementation with carbohydrates, lipids, proteins and/or combination aim temporary disturbance of body`s homeostasis. Clinical diagnostics takes advantage of such tests using them to detect metabolic impairment. The degree of disturbance and the rate of recover are indicative for the health status. The normal glucose tolerance is considered as a sign for a good phenotypic flexibility. The oral glucose tolerance test (OGTT) is the most commonly used stress test and has been applied in diabetes diagnosis for decades. Although the fasting triacylglycerol levels are an independent risk factor for certain conditions, some authors suggest measuring the postprandial levels of metabolites with an oral lipid tolerance test (OLTT) for more precise diagnostics. Thus current study explores the possible application and potential of OGTT, OLTT, mixed-meal challenge and combination of them to assess phenotypic flexibility in nutrition research and to evaluate individual`s health status.

Mechanisms of Cyanotoxin Toxicity—Carcinogenicity, Anticancer Potential, and Clinical Toxicology

Cyanoprokaryotes are distributed worldwide and they produce various bioactive compounds, including cyanotoxins. The major route of human exposure to cyanotoxins is the oral intake by using contaminated drinking water, by incidental intake of contaminated water during recreational and professional activities, and by consuming contaminated food or dietary supplements prepared from cyanobacteria. The prolonged chronic exposure to low concentrations of cyanotoxins provokes cell damage and may increase the risk for cancer development. Due to the variety of cyanotoxin chemical structures, different mechanisms of their toxic effects are known. At the same time, some of the natural compounds produced by cyanoprokaryotes have anticancer potential and are promising sources for the development of novel drugs. This chapter is dedicated to the target mechanisms behind the effects of the widely distributed cyanotoxins with an impact on human health, microcystins, nodularins, and cylindrospermopsin