| Compared with ordinary materials,nanomaterials have unique physical and chemical properties and were widely used in many aspects of social production and life.Compared with other nanomaterials,silver nanoparticles have superior antibacterial and antiviral properties and become one of the most widely used metal nanomaterials.Silver nanoparticles enter the human body mainly through the digestive tract,respiratory tract and skin,which can cause damage to multiple organ systems of human body and threaten human health.The physical and chemical properties of silver nanoparticles such as particle size,material shape and surface modification resolved their biological activities.Studies have shown that silver nanoparticles cause neurotoxicity by destroy the blood-brain barrier,damage neurons and glial cells.The model organism caenorhabditis elegans has a complete nervous system and has a unique advantage in the study of neurotoxicity,although its body structure is uncomplicated.The present experimental evidence showed that heavy metals such as nickel,nano-polystyrene and various carbon-based nanomaterials can cause neurotoxicity to nematodes by damaging related neurons.In this study,two kinds of nanosilver with different surface properties(20 nm PVP-Ag NPs and 20 nm Ag NPs)were exposed to the model biological C.elegans to investigate the neurotoxicity of two nano-silver on C.elegans at low concentrations(0.01 mg/L,0.1 mg/L,1 mg/L and 10 mg/L)and longer-term exposure(prolonged exposure 48 h or chronic exposure 6 d).The effect of different coated on the neurotoxicity of silver nanoparticles were compared.The relationship between the neurotoxicity caused by silver nanoparticles and neuronal and neurotransmitter receptor damage were investigated and the mechanism of neurotoxicity of silver nanoparticles was further explored.1、Characterization of silver nanoparticles and detection of silver content in nematodes: The results of characterization showed that the particle size of 20 nm PVP-Ag NPs and 20 nm Ag NPs were 22.08 nm and 20.14 nm,respectively;the hydrated particle sizes of 20 nm PVP-Ag NPs and 20 nm Ag NPs in ultrapure water were 52.29 nm and 450.8 nm;and the Zeta potentials were-16.78 m V and-1.89 m V,respectively.20 nm PVP-Ag NPs dispersed well and gathered lessly,20 nm Ag NPs disperse less and gatherd severely.Both two kinds of silver nanoparticles were spherical.After the wild-type caenorhabditis elegans(N2)exposure to 20 nm Ag NPs or 20 nm PVP-Ag NPs at the concentration of 0.01 mg/L,0.1 mg/L,1 mg/L and 10mg/L for 48 h or 6 d respectively,the silver content in nematodes was detected by graphite furnace atomic absorption spectrometry.When the exposure time was prolonged to 6 d,the silver content in nematode was significantly higher than that in exposed 48 h,suggesting that silver nanoparticles may accumulate in nematode.At the same exposure time and exposure dose,the silver content of 20 nm PVP-Ag NPs in each group of nematodes was significantly higher than that in 20 nm Ag NPs group,which indicated that PVP coating enhanced the dispersion of nano-silver and made it easier to enter nematodes.2.Study on the basic toxic effect of silver nanoparticles on C.elegans: Detected the life span,body length and body breadth of C.elegans after it were exposured to two kinds of silver nanoparticles from L1-larvae,respectively.After exposure to silver nanoparticles for 48 h,compared with the control group,the relative mean life span of1 mg/L and 10 mg/L groups of 20 nm PVP-Ag NPs and 20 nm Ag NPs decreased to87.52%,79.91% and 94.18%,86.72% respectively.Both two nano-silver exposure reduced the maximum and relative mean lifespan of nematodes,and the longer the exposure time,the greater the damage.The results of body length and body breadth measurement showed that after exposure for 48 h,the body length decreased and the body breadth increased in the groups with 1 mg/L and 10 mg/L of 20 nm PVP-Ag NPs and group with 10 mg/L of 20 nm Ag NPs.After exposure for 6 d,the body length decreased and the body breadth increased in the groups with 1 mg/L and 10 mg/L of20 nm PVP-Ag NPs,furthermore,the body breadth increased in the groups with 0.1mg/L of 20 nm PVP-Ag NPs.For 20 nm Ag NPs,the body length decreased,the body width increased in the groups with 10 mg/L and the body breadth alos increased in the groups with 1 mg/L.The above results showed that the effect of two kinds of nano-silver exposure on body length and the body breadth of nematodes were not consistent.Effects of two kinds of silver nanoparticles on life span and growth and development of nematodes were 20 nm PVP-Ag NPs>20 nm Ag NPs.3.Neurotoxic effects of silver nanoparticles in nematodes: The movement behavior,feeding and defecation behavior,learning and memory behavior of nematodes were observed and detected after exposure to silver nanoparticles.After exposure for 48 h,compared with control group,the headtrash frequency,body bends frequency,pharyngeal pumping frequency,chemotaxis index(CI),IT curve of C.elegans were decreased and the defecation interval increased,in the groups with 1mg/L and 10 mg/L of 20 nm PVP-Ag NPs and group with 10 mg/L of 20 nm Ag NPs.After exposure for 6 d,compared with control group,the headtrash frequency,body bends frequency,pharyngeal pumping frequency,chemotaxis index(CI),IT curve of C.elegans were decreased and the defecation interval increased,in the groups with 1mg/L and 10 mg/L of both 20 nm PVP-Ag NPs and 20 nm Ag NPs,furthermore,the defecation interval of C.elegans were also increased in the groups with 0.1 mg/L of20 nm PVP-Ag NPs.The above results showed that silver nanoparticles exposure to the nematodes cause neurotoxicity,which affects local movement,feeding and defecation,and learning and memory of nematodes.Neurotoxicity of two silver nanoparticles is 20 nm PVP-Ag NPs > 20 nm Ag NPs.4.Effect of silver nanoparticles on neurons and neurotransmitter receptors in C.elegans: Fluorescence intensity in transgenic strains BZ555,EG1285,DA1240 and LX929 were detected after exposure to two kinds of silver nanoparticles respectively.The results of relative fluorescence intensity showed that after exposure for 48 h,compared with control group,the relative fluorescence intensity of BZ555 nematodes were significantly decreased in the groups with 10 mg/L of 20 nm PVP-Ag NPs and20 nm Ag NPs,the relative fluorescence intensity of EG1285 nematodes were significantly decreased in the groups with 1 mg/L and 10 mg/L of 20 nm PVP-Ag NPs and groups with 10 mg/L of 20 nm Ag NPs.Moreover,the relative fluorescence intensity of LX929 nematodes were significantly decreased only in the group with 10mg/L of 20 nm PVP-Ag NPs while there were not any significant changesthe in the relative fluorescence intensity of DA1240 nematodes in all exposure groups.After exposure for 6 d,compared with control group,the relative fluorescence intensity of BZ555 nematodes were significantly decreased in the groups with 1 mg/L and 10mg/L of 20 nm PVP-Ag NPs and groups with 10 mg/L 20 nm Ag NPs,the relative fluorescence intensity of EG1285 nematodes were significantly decreased in the groups with 1 mg/L and 10 mg/L of both 20 nm PVP-Ag NPs and 20 nm Ag NPs.For the LX929 nematodes the relative fluorescence intensity were significantly decreased in the group with 1 mg/L and 10 mg/L of 20 nm PVP-Ag NPs and group with 10 mg/L of 20 nm Ag NPs,besides there were not any significant changesthe in the relative fluorescence intensity of DA1240 nematodes in all exposure groups.The expression of γ-aminobutyric acid and dopamine receptor genes in nematodes showed that after exposure for 48 h,compared with the control group,the expression of gab-1、ggr-1、dop-1 were up-regulated and the expression of ggr-2 was down-regulated in the group with 0.01 mg/L and 0.1 mg/L of 20 nm PVP-Ag NPs and the expression of ggr-2、ggr-3and dop-3 were down-regulated in the group with 10 mg/L of 20 nm PVP-Ag NPs.After C.elegans exposure to 20 nm Ag NPs for 48 h,only the expression of dop-1was up-regulated in the group with 0.01 mg/L and 0.1 mg/L while the expression level of other genes had no significant change.After exposure for 6 d,the expression of gab-1、ggr-1、dop-1 and dop-2 were up-regulated in the group with 0.01mg/L of 20 nm PVP-Ag NPs and the expression of gab-1and dop-1 were up-regulated in the group with 0.1 mg/L of 20 nm PVP-Ag NPs.The expression of gab-1,dop-3and dop-6 were down-regulated in the group with 1 mg/L of 20 nm PVP-Ag NPs and the expression of gab-1,ggr-1,ggr-3,dop-3,dop-4,dop-5 and dop-6 were down-regulated were up-regulated of 20 nm PVP-Ag NPs.For 20 nm Ag NPs,the expression of dop-1and dop-2 were up-regulated in the group of 0.01 mg/L and the expression of ggr-2、ggr-3 and dop-5 were down-regulated in the group of 10 mg/L.The above studies show that nano-silver exposure can cause damage to Caenorhabditis elegans neurons,cause abnormal development of neurons,cause nerve cable breakage,neuronal deletion,and show sensitivity differences among different neurons.At the same time,nano-silver exposure affects the expression level of neurotransmitter receptor genes in caenor elegans.Among the two kinds of nano-silver,20 nm PVP-Ag NPs has a more obvious effect on the expression of transmitter receptor genes.In conclusion,silver nanoparticles had obvious toxic effect on Caenorhabditis elegans,which could lead to shortened life span,abnormal body development,decreased body length and larger body width.At the same time,it has many neurotoxic effects on Caenorhabditis elegans,which can lead to dyskinesia movement,abnormal feeding and defecation and decreased learning and memory ability.Both two kinds of silver nanoparticles had obvious neurotoxicity to nematode at 1 mg/L and above exposure dose,and silver nanoparticles had no obvious neurotoxicity to nematode at 0.1 mg/L and below exposure dose.The results of detection of related neurons and receptor genes showed that nano-silver damaged many neurons of C.elegans and also affected the neurotransmitter receptors of C.elegans.This study provides a dose reference for the study of chronic neurotoxicity of silver nanoparticles and provides a clue to the mechanism of neurotoxicity of silver nanoparticles. |
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