Toxic effects of lead, cadmium, mercury and arsenic mixture in a subacute exposure model for rats: protective effect of probiotics

[K. S. Živančević]

227 str.

Farmacija - Toksikologija / Pharmacy -Toxicology

Sažetak U okviru ove doktorske disertacije ispitivana je subakutna toksičnost smeše metal(oid)a olova (Pb), kadmijuma (Cd), žive (Hg) i arsena (As) kod pacova, kao i utvrđivanje potencijalnog protektivnog efekta probiotika na umanjenje štetnih efekata ispitivane smeše metal(oid)a. Studija je sprovedena na mužjacima Wistar pacova podeljenim u 8 grupa po 5 jedinki od kojih je jedna grupa bila kontrolna, a 7 grupa je tretirano oralnim putem tokom 28 dana rastućim dozama smeše (MIX: Pb+Cd+Hg+As), od najnižeg nivoa doze - MIX 1 do najvišeg nivoa doze - MIX 5 (mg/kg t.m./dan) (Pb: 0,003, 0,01, 0,1, 0,3, 1; Cd: 0,01, 0,03, 0,3, 0,9, 3; Hg: 0,0002, 0,0006, 0,006, 0,018, 0,06; As: 0,002, 0,006, 0,06, 0,18, 0,6), kao i monokomponentnim (5,16×10⁸ CFU/kg/dan; Saccharomyces cerevisiae var. boulardii) i multikomponentnim probiotikom (8,78×10⁸ CFU/kg/dan; Saccharomyces cerevisiae var. boulardii, Lactobacillus rhamnosus, Lactobacillus plantarum LP 6595, i Lactobacillus plantarum HEAL9) uz smešu najvišeg doznog nivoa MIX 5. Ispitani su nivoi toksičnih metal(oid)a u krvi, hematološki, biohemijski, hormonski i parametri redoks statusa, kao i nivoi bioelementa, dok su organi analizirani histopatološki, uz određivanje toksičnih metal(oid)a, bioelemenata i redoks parametara, kao i aktivnosti acetilholinesteraze u mozgu. Modelovan je odnos doza-odgovor i određene su Benchmark doze za toksične efekte pojedinačnih metal(oid)a u smeši. Značajan porast nivoa Pb, Cd Hg i As u krvi uočen je samo u grupi kojoj je davana najviša doza, dok je akumulacija toksičnih metala dobijena prvenstveno u jetri, bubrezima, mozgu i femuru. Izlaganje najnižoj dozi ispitivane smeše metala dovodi do značanjih promena parametara anemije u krvi, odnosno sniženja broja eritrocita, hemoglobina, hematokrita, i nivoa gvožđa, kao i značajnog povećanja parametra oksidativnog stresa, O2 .-, PAB i AOPP. Tkiva ispoljavaju različitu osetljivost na oksidativni stres izazvan rastućim dozama smeše toksičnih metala (Pb, Cd, Hg, As). Najizraženije promene zabeležene su mozgu i plućima, dok je u jetri dobijeno smanjenje SOD i GSH pri većini doznih nivoa, dok je MDA snižen u srednjem i dva najviša dozna nivoa u bubrezima. Dodatno, više doze smeše Pb, Cd, Hg i As značajno su smanjile aktivnost enzima AChE u mozgu. Histopatološka analiza ukazuje na progresivna oštećenja većine organa pri izloženosti smeši metal(a), sa uočljivijim efektima pri većim doznim nivoima. Benchmark analiza utvrdila je postojanje doza-odgovor pri čemu je najniža BMD vrednost za nivo srednjeg volumena eritrocita (MCV), koji predstavlja kritični toksični efekat za svaki metal(oid) u ispitivanoj smeši i iznosi 3,15E-04 mg Pb/kg t.m./dan, 1,05E-03 mg Cd/kg t.m./dan, 2,10E-05 mg Hg/kg t.m./dan i 2,10E-04 mg As/kg t.m./dan. Probiotici, monokomponentni i mulitkomponentni, uticali su značajno na smanjenje Cd, Hg i As u krvi, kao i umanjenje pojedinih metala u jetri, bubrezima, plućima i femuru. Primena oba probiotika dovodi do normalizovanja izmenjenih vrednosti hematoloških parametara i TAS parametra u krvi, dok primena multikomponentnog probiotika normalizuje vrednosti TAS u jetri, IMA u plućima, TOS i O2.- u pankreasu i MDA u testisima i aktivnost enzima acetilholinesteraze u mozgu koji su značajno promenjeni pod uticajem smeše ispitivanih metal(oid)a. Dobijeni rezultati ukazuju da je mutikomponentni probiotik efikasniji u umanjenju štetnih efekata ispitivane smeše olova, kadmijuma, žive i arsena u odnosu na monokomponentni probiotik..

In this doctoral dissertation, the subacute toxicity of the metal(oid) mixture of lead (Pb), cadmium (Cd), mercury (Hg) and arsenic (As) in rats was examined, as well as the determination of the potential protective effect of probiotics on reducing the harmful effects of the investigated metal(oid) mixture. The study was conducted on male Wistar rats divided into 8 groups of 5 individuals, one group was a control group, and 7 groups were treated orally for 28 days with increasing doses of the mixture (MIX: Pb+Cd+Hg+As), from the lowest dose level - MIX 1 to the highest dose level - MIX 5 (mg/kg bw/day) (Pb: 0.003, 0.01, 0.1, 0.3, 1; Cd: 0.01, 0.9, 3; Hg: 0.0006, 0.018, 0.06; As: 0.002, 0.006, 0.18, 0.6). (5.16×10⁸ CFU/kg/day; Saccharomyces cerevisiae var. boulardii) and a multicomponent probiotic (8.78×10⁸ CFU/kg/day; Saccharomyces cerevisiae var. boulardii, Lactobacillus rhamnosus, Lactobacillus plantarum LP 6595, and Lactobacillus plantarum HEAL9) with the mixture of the highest dose level MIX 5. The levels of toxic metal(oids) in the blood, hematological, biochemical, hormonal and redox status parameters were examined, as well as the levels of bioelements, while the organs were analyzed histopathologically, with the determination of toxic metal(oids), bioelements and redox parameters, as well as acetylcholinesterase activity in the brain. A dose-response relationship was modeled and Benchmark doses for the toxic effects of individual metal(oids) in the mixture were determined. A significant increase in the levels of Pb, Cd, Hg and As in the blood was observed only in the group that received the highest dose, while the accumulation of toxic metals was obtained primarily in the liver, kidneys, brain and femur. Exposure to the lowest dose of the examined metal mixture leads to significant changes in the parameters of anemia in the blood, i.e. a decrease in the number of erythrocytes, hemoglobin, hematocrit, and iron levels, as well as a significant increase in the parameters of oxidative stress, O2.-, PAB and AOPP. Tissues exhibit different sensitivity to oxidative stress caused by increasing doses of a mixture of toxic metals (Pb, Cd, Hg, As). The most pronounced changes were recorded in the brain and lungs, while SOD and GSH were reduced in the liver at most dose levels, while MDA was decreased in the middle and two highest dose levels in the kidneys. Additionally, higher doses of a mixture of Pb, Cd, Hg and As significantly decreased the activity of the AChE enzyme in the brain. Histopathological analysis indicates progressive damage to most organs upon exposure to the metal(s) mixture, with more noticeable effects at higher dose levels. Benchmark analysis established the existence of a dose-response, where the lowest BMD value for the mean erythrocyte volume (MCV) level, which represents the critical toxic effect for each metal(oid) in the tested mixture, is 3.15E-04 mg Pb/kg bw/day, 1.05E-03 mg Cd/kg bw/day, 2.10E-05 mg Hg/kg bw/day and 2.10E-04 mg As/kg body weight/day. Probiotics, monocomponent and multicomponent, had a significant effect on the reduction of Cd, Hg and As in the blood, as well as the reduction of certain metals in the liver, kidneys, lungs and femur. The use of both probiotics leads to the normalization of the changed values of hematological parameters and the TAS parameter in the blood, while the use of the multicomponent probiotic normalizes the values of TAS in the liver, IMA in the lungs, TOS and O2.- in the pancreas and MDA in the testicles and the activity of the acetylcholinesterase enzyme in the brain, which were significantly changed under the influence of the mixture of investigated metal(oids). The obtained results indicate that the multi-component probiotic is more effective in reducing the harmful effects of the examined mixture of lead, cadmium, mercury and arsenic compared to the mono-component probiotic.

smeša olova, kadmijuma, žive i arsena, subakutna toksičnost, mono- i multikomponentni probiotik, protektivni efekat, Benchmark doze

mixture of lead, cadmium, mercury and arsenic, subacute toxicity, mono- and multicomponent probiotic, protective effect, Benchmark doses

615.9(043.3)

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