Archives: Projekti

IMI projekti

022-0222148-2142 Toxic Effects of Mycotoxins on Humans and Animals

MINISTRY OF SCIENCE, EDUCATION AND SPORTS OF THE REPUBLIC OF CROATIA (2007-2014)

Principal Investigator: Maja Peraica

SUMMARY
Mycotoxins are the metabolites of moulds with toxic, carcinogenic, genotoxic, mutagenic, and teratogenic properties. In our earlier studies, we found low concentrations of fumonisin B1 (FB1), fumonisin B2, ochratoxin A (OTA), and zearalenon in food samples collected in various regions of Croatia. OTA is considered to be involved in the development of endemic nephropathy (EN). This kidney disease of unknown etiology and high frequency of tumors of the upper urothelial tract appears in the western part of Brodska Posavina. It is speculated that, in addition to OTA, another nephrotoxic mycotoxin such as FB1 should be involved in the development of EN.
The mechanism of the toxicity of OTA and FB1 is not fully understood. It was proved on cultured cells that OTA and FB1 cause oxidative damage of lipids and DNA. However, it is not know whether low concentrations of these mycotoxins usually found in food can also cause oxidative damage of lipids, proteins and DNA in experimental animals. OTA and FB1 are frequently found together in cereals, and therefore it would be important to see whether their effect is synergistic or not. OTA is an organic anion that accumulates in the kidney cells and partially excretes in urine. OTA is partially reabsorbed due to organic anion transporters (OAT) on the epithelial cells along the nephron, which may contribute to its toxicity in specific parts of the kidney. Recent investigations have shown that the localization of specific OATs in rats is sex-related. Toxic effects of OTA are more severe in male rats, which could be due to the higher expression of OAT in males.
The proposed project will use recently developed methods, some of which will be used for the first time in the research of the mechanisms of action of mycotoxins.
The aim of this project is to clarify the role of oxidative stress caused by mycotoxins in damage of macromolecules (DNA, lipids, and proteins), as well as to improve the understanding of the mechanism of toxicity of OTA by investigating the changes in kidney OAT system. The other aim is to find out whether exposure to OTA and FB1 is involved in the development of EN.
We expect that our results will: a) show whether oxidative stress is involved in the mechanism of OTA and FB1 toxicity, and whether this effect is synergistic, b) show whether OTA affects the expression of OATs in the rat kidney and whether this effect is sex-related, and c) improve the knowledge about the mechanism of EN development.

Effects of Physical and Chemical Agents – Cytogenetic Biomarkers

The Ministry of Science, Education and Sports of the Republic of Croatia (2002-2006)

Principal Investigator: Aleksandra Fučić

SUMMARY
The aim of proposed project is evaluation of stability of cytogenetic biomarkers after long term exposures to low doses of physical and chemical agents, investigation of interindividual differences in response on carcinogens, especially in organisms under development. The significance of aneugenic substances will be investigated.

Within international projects  micronucleus assay will be standardized and research will be performed on the estimation of potential correlation between cancer and increased frequency of micronuclei and chromosome aberrations. It could be expected that results of project would contribute to understanding of significance of cytogenetic biomarkers in risk assessment of subjects exposed to carcinogens.

Cytogenetic biomarkers will be selected for children monitoring. Chromosome aberration assay, sister chromated exchange frequency, in vivo and in vitro micronucleus assay, fluorescent in situ hybridization and comet assay will be applied in project. Results of the project are of great importance in the health risk assessment, understanding of aetiology of cancer development in adult population and children caused by environmental factors, methodology standardization and for  regulative bodies in issuing directives for permissible emissions of carcinogens in environment.

Pesticides, Persistent and Volatile Organic Pollutants in the Environment

The Ministry of Science, Education and Sports of the Republic of Croatia (2002-2006)

Principal Investigator: Vlasta Drevenkar

The aim of the project is to investigate the relationship between physico-chemical properties of selected organic micropollutants and their distribution and fate in the environment and in humans. The studied compounds are widely spread organochlorine and triazine pesticides, polychlorinated biphenyls (PCB), dibenzo-p-dioxins and dibenzofurans (PCDD/F), chlorophenols, and volatile chlorinated and aromatic hydrocarbons.
The studies are focused on:

  • the analysis of persistent organochlorine compounds in air, atmospheric particles, coniferous tree needles, soil/sediments, and waters, partition of compounds between gas (air) and solid (particles) phase, absorption in needles, and possible biomagnification in air/needles/soil system;
  • occurrence and mobility of chloro-, methylthio- and methoxytriazine herbicides and their dealkylated/hydroxylated degradation products in water and soil environments;
  • sorption behaviour of triazine compounds and chlorophenols in soils/sediments and mechanisms of sorption interactions;
  • origins and levels of volatile chlorinated (dichloromethane, 1,1,1-trichloroethane, tri- and tetrachloroethylene) and aromatic (benzene, toluene, ethylbenzene, xylene isomers) hydrocarbons in drinking water and indoor air in the Zagreb city;
  • assessment of human exposure to organochlorine pesticides, PCB, PCDD/F by their analysis in milk and serum and evaluation of long-term variations in exposure of general population in Croatia;
  • assessment of sensitive and specific bioindicators of human exposure to triazine herbicides (atrazine) by analysis of specific urinary metabolites in occupationally exposed persons;
  • assessment of the exposure of the urban (Zagreb) general population to volatile chlorinated and aromatic hydrocarbons by analysis of urinary excreted species and correlation with pollutant levels in drinking water and air.

022-0222148-2146 Mammalian Renal Transporters; Gender Differences and Effects of Toxic Metals

MINISTRY OF SCIENCE, EDUCATION AND SPORTS OF THE REPUBLIC OF CROATIA (2007-2014)

Principal Investigator: Ivan Sabolić

SUMMARY
The kidney secretes various organic anions (OA) and cations (OC), including most of drugs. This secretion takes place predominantly in the proximal tubules, and is mediated by specific transporters (OAT, OCT), localized in the cell membrane. Recent studies have shown that these transporters also mediate the cellular uptake of nephrotoxic metals (Cd, Hg, cis-Pt) by molecular mimicry; the metal binds to OA or OC, and the complex is then transferred into the cell by OAT or OCT. So far several isoforms of OAT and OCT have been described in the mammalian nephron, but their detailed distribution in the cells, and their relative abundance among species are poorly known. Adult rodents (rat, mouse) express gender (sex) differences in the renal excretion of some organic substances due to different expressions of OAT and OCT. Examples of the sex-related excretion of organic ions exist also in pigs and humans, but the expression of their transporters along the nephron in these species has not been examined in detail. Since the levels of sex hormones depend on sex, age, and hormonal cycle (women), the expression of the transporters may affect the reabsorption, secretion and toxicity of drugs, metals, or other substances in the tubules. Furthermore, it is well known that the nephrotoxic metals cause damage to the renal tubules, resulting in the loss of organic and inorganic substances in urine, and that metal toxicity in female rodents is weaker than in males, but possible mechanisms of these effects via expression of OAT, OCT, and other transporters along the nephron are not known. Here we propose biochemical, immunochemical, and molecular biology studies in order to compare the presence and distribution of OAT, OCT, and other transporters of organic substances in the rat, mouse, pig, and human kidneys. Using experimental animals of different sex, age, and hormonal cycle, we will study the effects of sex hormones and nephrotoxic metals on the expression of these transporters along the nephron. We expect that the final data will: a) show the presence of gender and species differences in the expression of various transporters in the cells along the nephron, b) identify an optimal animal model for the studies relevant to humans, and c) contribute to a better understanding of sex-dependent problems in therapy, (re)absorption and secretion of various drugs, and nephrotoxicity of some drugs, metals and other substances in mammals.

Aging-related expression of membrane transporters in rat – AGEMETAR

Duration: 1 Oct 2014-31 Mar 2019

ABSTRACT
Aging is a physiological process characterized by a gradual decline in physical fitness and other functional capabilities. The mechanisms of aging and possible actions of slowing down or ameliorating aging-related processes are poorly understood. In humans, one of the characteristics of old age is polypharmacy, which elevates risks of drug-drug interactions and drug-induced adverse reactions and organ toxicity, such as hepatotoxicity and nephrotoxicity. These reactions may partially result from diminished drug metabolism and/or mainly from impaired activity/expression of various membrane transporters that are responsible for drug excretion in liver and kidneys. These both mechanisms have been poorly studied in senescent organs. Here we promote a hypothesis that the aging-induced oxidative stress, known to downregulate the activity/expression of the aging suppressor gene (Klotho) and to shorten the length of telomeres, also diminishes the activity/expression of various membrane transporters in liver and kidneys, and that these changes can be ameliorated by treating animals with antioxidants resveratrol and melatonin. The extent and the intensity of aging-related changes, and the beneficial effects of antioxidants may be sex-related. To test this hypothesis, we propose various biochemical and cell- and molecular-biological studies of aging parameters and aging-dependent expression of various membrane transporters in the mammalian liver and kidneys using a rat model of experimental aging. Possible aging-related decline in the expression of these transporters may explain a higher incidence (women > man) of drug interactions and toxicity in old people. The studies will define the underlying cellular mechanisms that contribute to generation of the aging-related pathophysiological conditions in the liver and kidneys, and will indicate possible ways of their amelioration. Positive results may point to remedies to be used for improving the health of elderly people.

PROJECT LEADERS:

  • Ivan Sabolić, MD, PhD; Head of  Molecular Toxicology Unit
  • Davorka Breljak, PhD; Head of Molecular Toxicology Unit (from 1 Oct 2015)
TEAM MEMBERS ON THE PROJECT
Molecular Toxicology Unit: Davorka Breljak, molecular biologist, PhD; Marija Ljubojević, molecular biologist, PhD; Ivana Vrhovac Madunić, molecular biologist, PhD; Dean Karaica, experimental biologist, PhD

Animal Breeding Unit: Vedran Micek, DVM, PhD student

Analytical Toxicology and Mineral Metabolism Unit: Jasna Jurasović, chemist, PhD; Tatjana Orct, chemist, PhD

Toxicology Unit: Maja Peraica, MD, PhD; Dubravka Rašić, biologist and chemist, PhD; Ivana Novak Jovanović, food technologist, PhD

Mutagenesis Unit: Goran Gajski, molecular biologist, PhD; Marko Gerić, experimental biologist, PhD student

Laboratory for Molecular and Cellular Biology (IRB): Ivica Rubelj, molecular biologist, PhD; Lucija Nanić, molecular biologist, PhD student