Cuihong Jin

Aluminium chloride impairs long-term memory and downregulates cAMP-PKA-CREB signalling in rats.

Epidemiological investigations have indicated that aluminium (Al) is an important environmental neurotoxicant that may be involved in the aetiology of the cognitive dysfunction associated with neurodegenerative diseases. Additionally, exposure to Al is known to cause neurobehavioural abnormalities in animals. Previous studies demonstrated that Al impaired early-phase long-term potentiation (E-LTP) in vivo and in vitro. Our previous research revealed that Al could impair long-term memory via the impairment of late-phase long-term potentiation (L-LTP) in vivo. However, the exact mechanism by which Al impairs long-term memory has been poorly studied thus far. This study was designed not only to observe the effects of subchronic Al treatment on long-term memory and hippocampal ultrastructure but also to explore a possible underlying mechanism (involving the cAMP-PKA-CREB signalling pathway) in the hippocampus of rats.. Pregnant Wistar rats were assigned to four groups. Neonatal rats were exposed to Al by parental lactation for 3 weeks and then fed with distilled water containing 0, 0.2%, 0.4% or 0.6% Al chloride (AlCl3) for 3 postnatal months. The levels of Al in the blood and hippocampus were quantified by atomic absorption spectrophotometry. The shuttle-box test was performed to detect long-term memory. The hippocampus was collected for ultrastructure observation, and the level of cAMP-PKA-CREB signalling was examined. The results showed that the Al concentrations in the blood and hippocampus of Al-treated rats were higher than those of the control rats. Al may impair the long-term memory of rats. Hippocampal cAMP, cPKA, pCREB, BDNF and c-jun expression decreased significantly, and the neuronal and synaptic ultrastructure exhibited pathological changes after Al treatment. These results indicated that Al may induce long-term memory damage in rats by inhibiting cAMP-PKA-CREB signalling and altering the synaptic and neuronal ultrastructure in the hippocampus.

Induction of the bystander effect in Chinese hamster V79 cells by actinomycin D.

Bystander effect (BE) can be induced by ionizing radiation and chemicals, including alkylating agents. Ionizing radiation mostly induces the bystander effect by causing double-strand DNA breakage in the exposed cells. However, the chemical-induced bystander effect is poorly studied. Here we chose actinomycin D (ACTD), a genotoxic chemotherapeutic drug, to investigate whether it could cause bystander effect in Chinese hamster V79 cells. Results are that (1) ACTD induced apoptosis in V79 cells and an optimal apoptosis model in V79 cells was established with ACTD (4 mg/L, 1h); (2) using apoptosis rate, chromosome aberration, and ultrastructure changes as endpoints of bystander effect, ACTD could induce bystander effect in V79 cells; (3) as in the exposed cells, ACTD mainly induced apoptosis in bystander V79 cells cultured in different period conditioned medium; (4) the strongest bystander effect was induced by 4 h conditioned medium collected from cells treated with ACTD. It suggests that ACTD could cause BE through the medium soluble factors excreted from exposed cells during apoptosis and ACTD-induced BE was a novel quantitative and kinetic response.

ERCC1 and ERCC2 Haplotype Modulates Induced BPDE-DNA Adducts in Primary Cultured Lymphocytes

Background Benzo[a]pyrene(B[a]P), and its ultimate metabolite Benzo[a]pyrene 7,8-diol 9,10-epoxide (BPDE), are classic DNA damaging carcinogens. DNA damage caused by BPDE is normally repaired by Nucleotide Excision Repair (NER), of which ERCC1 and ERCC2/XPD exert an indispensable role. Genetic variations in ERCC1 and ERCC2 have been related to DNA repair efficiency. In this study we used lymphocytes from healthy individuals to show that polymorphisms in ERCC1 and ERCC2 are directly associated with decreased DNA repair efficiency. Methods ERCC1 (rs3212986 and rs11615) and ERCC2 (rs13181, rs1799793 and rs238406) were genotyped in 818 healthy Han individuals from the northeast of China. BPDE induced DNA adducts in lymphocytes were assessed by high performance liquid chromatography (HPLC) in 282 randomly selected participants. The effect of ERCC1 rs3212986 and ERCC2 rs238406 on DNA damage caused by B[a]P was assessed with a modified comet assay. Results We found that the variant genotypes of ERCC1 rs3212986 and ERCC2 rs238406 were associated with the high levels of BPDE-DNA adducts. Especially ERCC1 rs3212986 A-allele variant was significantly associated with the high BPDE-DNA adducts. Haplotype analysis showed that the ERCC1 haplotype AC (OR = 2.36, 95% CI = 1.84–2.97), ERCC2 haplotype AGA (OR = 1.51, 95% CI = 1.06–2.15) and haplotype block AGAAC (OR = 5.28, 95% CI = 2.95–9.43), AGCAC (OR = 1.35 95% CI = 1.13–1.60) were linked with high BPDE-DNA adducts. In addition, we found that the combined minor alleles of ERCC1 rs3212986 and ERCC2 rs238406 were associated with a reduced DNA repair capacity. Conclusions Our results suggest that the variant genotypes of ERCC1 rs3212986 and ERCC2 rs238406 are associated with decreased repair efficiency of BPDE induced DNA damage, and may be predictive for an individual’s DNA repair capacity in response to environmental carcinogens.

Conditioned medium from actinomycin D-treated apoptotic cells induces mitochondria-dependent apoptosis in bystander cells

Chemical-induced bystander effects have been known for several years, but the underlying mechanism is still seldom investigated. Previous researchers have found that mitomycin C and phleomycin induced micronuclei in bystander cells the same as in exposed cells. We previously demonstrated the ability of actinomycin D (ACTD) to induce bystander effects in normal Chinese hamster fibroblast V79 cells and found that conditioned medium (CM) obtained from ACTD-exposed apoptotic cells induced apoptosis in bystander cells. The present study further explores the probable mechanism of apoptosis in bystander cells. The main findings of this study are: (1) ACTD-treated CM induced apoptosis in bystander cells in a time-dependent manner, which was confirmed with morphological changes. (2) ACTD-treated CM increased the mRNA and protein levels of pro-apoptotic p53 and Bax, whereas it decreased those of anti-apoptotic Bcl-2 in bystander cells; these were all time-dependent effects. Reactive oxygen species (ROS) were also involved in apoptosis of bystander cells. (3) ACTD-treated CM reduced mitochondria membrane potential and induced cytochrome c release. (4) ACTD-treated CM induced G1 cell phase arrest, which may be another response in bystander cells when cultured with CM. These results suggest that chemical-treated CM induces p53-Bcl-2/Bax-cytochrome c signaling (i.e., mitochondria pathway)-dependent apoptosis in bystander cells, which is a kinetic response.

Protective Role of tert-Butylhydroquinone Against Sodium Fluoride-Induced Oxidative Stress and Apoptosis in PC12 Cells

The neurotoxicity of fluoride is associated with oxidative stress due to imbalance between production and removal of reactive oxygen species (ROS). In contrast, induction of detoxifying and antioxidant genes through activation of NF-E2-related factor 2 (Nrf2) has been implicated in preventing oxidative stress and apoptosis in neurodegenerative diseases. The present study aimed to investigate the possible neuroprotective role of tert-butylhydroquinone (tBHQ), a general Nrf2 activator, on sodium fluoride (NaF)-induced oxidation damage and apoptosis in neuron-like rat pheochromocytoma (PC12) cells. Pretreatment with tBHQ protected PC12 cells against NaF-induced cytotoxicity as measured by MTT assay and apoptosis detection, simultaneously, inhibited NaF-induced overproduction of intracellular ROS and reduction of total glutathione content. Furthermore, NaF or tBHQ induced the stabilization of Nrf2, and enhanced expression of heme oxygenase-1 (HO-1) and γ-glutamylcysteine synthetase (γ-GCS) as a consequence of Nrf2 inducing. These findings indicated that tBHQ pretreatment conferred protective effect on PC12 cells against NaF-induced apoptotic cell death and oxidation-redox imbalance through stabilization of Nrf2 and elevation of downstream HO-1 and γ-GCS expressions.

The effect of nuclear factor erythroid 2-related factor/antioxidant response element signalling pathway in the lanthanum chloride-induced impairment of learning and memory in rats.

Lanthanum exerts adverse effects on the central nervous system. However, the mechanism underlying these adverse effects has not been clarified. It is known that oxidative stress plays an important role in neurological injuries induced by harmful factors. Nuclear factor erythroid 2‐related factor (Nrf2) is very important in the response to oxidative stress in tissues and cells. The purpose of this study was to explore the effect of lanthanum chloride (LaCl3) on the spatial learning and memory of rats and to determine whether the Nrf2/antioxidant response element pathway acts in the hippocampus. Four groups of Wistar rats were exposed to 0 mM, 9 mM, 18 mM or 36 mM LaCl3 through their drinking water from the day of birth to 2 months after weaning. The results showed that LaCl3 impaired the spatial learning and memory of the rats, damaged the neuronal ultrastructure, increased reactive oxygen species levels and significantly down‐regulated Nrf2 as well as the mRNA and protein expression of Nrf2‐regulated genes, including NADP(H): dehydrogenase quinone 1, haeme oxygenase‐1, superoxide dismutase 2, glutathione peroxidase 1, glutathione‐S‐transferase, γ‐glutamine cysteine synthase and glutathione reductase, in the hippocampus. This study suggests that LaCl3 can impair the spatial learning and memory of rats, possibly by perturbing the Nrf2/antioxidant response element signalling pathway.

Bystander Effect Induced by UVC Radiation in Chinese hamster V79 cells

In past decades, researches on radiation‐induced bystander effect mainly focused on ionizing radiation such as α‐particle, β‐particle, X‐ray and γ‐ray. But few researches have been conducted on the ability of ultraviolet (UV) radiation‐induced bystander effect, and knowledge of UVC‐induced bystander effect is far limited. Here, we adopted medium transfer experiment to detect whether UVC could cause bystander effect in Chinese hamster V79 cells. We determined the cell viability, apoptosis rate, chromosome aberration and ultrastructure changes, respectively. Our results showed that: (1) the viability of UVC‐irradiated V79 cells declined significantly with the dosage of UVC; (2) similar to the irradiated cells, the main death type of bystander cells cultured in irradiation conditioned medium (ICMs) was also apoptosis; (3) soluble factors secreted by UVC‐irradiated cells could induce bystander effect in V79 cells; (4) cells treated with 4 h ICM collected from 90 mJ cm−2 UVC‐irradiated cells displayed the strongest response. Our data revealed that UVC could cause bystander effect through the medium soluble factors excreted from irradiated cells and this bystander effect was a novel quantitative and kinetic response. These findings might provide a foundation to further explore the exact soluble bystander factors and detailed mechanism underlying UVC‐induced bystander effect.

The ERCC2/XPD Lys751Gln polymorphism affects DNA repair of benzo[a]pyrene induced damage, tested in an in vitro model.

Nucleotide excision repair (NER) is an important defense mechanism of the body to exogenous carcinogens and mutagens, such as benzo[a]pyrene (B[a]P). Genetic polymorphisms in ERCC2/XPD, a critical element in NER, are thought to be associated with individuals cancer susceptibility. Although ERCC2/XPD Lys751Gln (rs13181) is the most studied polymorphism, the impact of this polymorphism on DNA repair capacity to carcinogen remains unclear. In the present study, cDNA clones carrying different genotypes of ERCC2/XPD (Lys751Gln) were introduced into an ERCC2/XPD deficient cell line (UV5) in a well-controlled biological system. After B[a]P treatment, cell growth inhibition rates and DNA damage levels in all cells were detected respectively. As expected, we found that the DNA repair capacity in UV5 cells was restored to levels similar to wildtype parent AA8 cells upon introduction of the cDNA clone of ERCC2/XPD (Lys751). Interestingly, after B[a]P treatment, transfected cells expressing variant ERCC2/XPD (751Gln) showed an enhanced cellular sensitivity and a diminished DNA repair capacity. The wildtype genotype AA (Lys) was found to be associated with a higher DNA repair capacity as compared to its polymorphic genotype CC (Gln). These data indicate that ERCC2/XPD Lys751Gln polymorphism affects DNA repair capacity after exposure to environmental carcinogens such as B[a]P in this well-controlled in vitro system and could act as a biomarker to increase the predictive value to develop cancer.

Effects of Aluminium on Long-Term Memory in Rats and on SIRT1 Mediating the Transcription of CREB-Dependent Gene in Hippocampus

Epidemiological investigations have shown that aluminium (Al) is an important neurotoxicant which can be absorbed by organisms via various routes. Previous studies have confirmed that exposure to Al could cause neurodegenerative diseases, decline CREB phosphorylation and then down‐regulate the transcription and protein expression of its target genes including BDNF. However, recent studies revealed that CREB activation alone was far from enough to activate the expression of long‐term memory (LTM)‐related genes; there might be other regulatory factors involved in this process. Several studies showed that TORC1 might be involved in regulating the transcription of downstream target genes as well. Also, TORC1 could be mediated by SIRT1 during the formation of LTM. However, the role of CREB regulating system in Al‐induced LTM impairment was still not utterly elucidated till now. This study was designed to establish the rat model of subchronic Al exposure to observe the neuroethology, regulatory factor levels and molecular biological alterations in hippocampal cells. The results showed that, with the increasing AlCl3 dose, blood Al content increased gradually; morphology of the hippocampus and neuronal ultrastructure were aberrant; in the Morris water maze test, the escape latency and distance travelled became longer, swimming traces turned more complicated in the place navigation test; intracellular Ca2+, cAMP levels declined significantly in AlCl3‐treated rats, followed by abated nuclear translocation of TORC1 and decreased SIRT1, TORC1 and pCREB levels. These results indicate that SIRT1 and TORC1 might play an important mediating role in Al‐induced LTM impairment.

Genetic polymorphisms in 19q13.3 genes associated with alteration of repair capacity to BPDE-DNA adducts in primary cultured lymphocytes.

Benzo[a]pyrene(B[a]P), and its ultimate metabolite Benzo[a]pyrene 7,8-diol 9,10-epoxide (BPDE), are classic DNA damaging carcinogens. DNA damage in cells caused by BPDE is normally repaired by Nucleotide Excision Repair (NER) and Base Excision Repair (BER). Genetic variations in NER and BER can change individual DNA repair capacity to DNA damage induced by BPDE. In the present study we determined the number of in vitro induced BPDE-DNA adducts in lymphocytes, to reflect individual susceptibility to Polycyclic aromatic hydrocarbons (PAHs)-induced carcinogenesis. The BPDE-DNA adduct level in lymphocytes were assessed by high performance liquid chromatography (HPLC) in 281 randomly selected participants. We genotyped for 9 single nucleotide polymorphisms (SNPs) in genes involved in NER (XPB rs4150441, XPC rs2228001, rs2279017 and XPF rs4781560), BER (XRCC1 rs25487, rs25489 and rs1799782) and genes located on chromosome 19q13.2-3 (PPP1R13L rs1005165 and CAST rs967591). We found that 3 polymorphisms in chromosome 19q13.2-3 were associated with lower levels of BPDE-DNA adducts (MinorT allele in XRCC1 rs1799782, minor T allele in PPP1R13L rs1005165 and minor A allele in CAST rs967571). In addition, a modified comet assay was performed to further confirm the above conclusions. We found both minor T allele in PPP1R13L rs1005165 and minor A allele in CAST rs967571 were associated with the lower levels of BPDE-adducts. Our data suggested that the variant genotypes of genes in chromosome 19q13.2-3 are associated with the alteration of repair efficiency to DNA damage caused by Benzo[a]pyrene, and may contribute to enhance predictive value for individuals DNA repair capacity in response to environmental carcinogens.