Lymph-Node-Based CD3+ CD20+ Cells Emerge from Membrane Exchange between T Follicular Helper Cells and B Cells and Increase Their Frequency following Simian Immunodeficiency Virus Infection.

CD4+ T follicular helper (TFH) cells are key targets for human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) replication and contribute to the virus reservoir under antiretroviral therapy (ART). Here, we describe a novel CD3+ CD20+ double-positive (DP) lymphocyte subset, resident in secondary lymphoid organs of humans and rhesus macaques (RMs), that appear predominantly after membrane exchange between TFH and B cells. DP lymphocytes are enriched in cells displaying a TFH phenotype (CD4+ PD1hi CXCR5hi), function (interleukin 21 positive [IL-21+]), and gene expression profile. Importantly, expression of CD40L upon brief in vitro mitogen stimulation identifies, by specific gene-expression signatures, DP cells of TFH-cell origin versus those of B-cell origin. Analysis of 56 RMs showed that DP cells (i) significantly increase following SIV infection, (ii) are reduced after 12 months of ART in comparison to pre-ART levels, and (iii) expand to a significantly higher frequency following ART interruption. Quantification of total SIV-gag DNA on sorted DP cells from chronically infected RMs showed that these cells are susceptible to SIV infection. These data reinforce earlier observations that CD20+ T cells are infected and expanded by HIV infection, while suggesting that these cells phenotypically overlap activated CD4+ TFH cells that acquire CD20 expression via trogocytosis and can be targeted as part of therapeutic strategies aimed at HIV remission. IMPORTANCE The HIV reservoir is largely composed of latently infected memory CD4+ T cells that persist during antiretroviral therapy and constitute a major barrier toward HIV eradication. In particular, CD4+ T follicular helper cells have been demonstrated as key targets for viral replication and persistence under ART. In lymph nodes from HIV-infected humans and SIV-infected rhesus macaques, we show that CD3+ CD20+ lymphocytes emerge after membrane exchange between T cells and B cells and are enriched in phenotypic, functional, and gene expression profiles found in T follicular helper cells. Furthermore, in SIV-infected rhesus macaques, these cells expand following experimental infection and after interruption of ART and harbor SIV DNA at levels similar to those found in CD4+ T cells; thus, CD3+ CD20+ lymphocytes are susceptible to SIV infection and can contribute to SIV persistence.

The Potential Role of Cytokines and Growth Factors in the Pathogenesis of Alzheimer's Disease.

Alzheimer's disease (AD) is one of the most prominent neurodegenerative diseases, which impairs cognitive function in afflicted individuals. AD results in gradual decay of neuronal function as a consequence of diverse degenerating events. Several neuroimmune players (such as cytokines and growth factors that are key players in maintaining CNS homeostasis) turn aberrant during crosstalk between the innate and adaptive immunities. This aberrance underlies neuroinflammation and drives neuronal cells toward apoptotic decline. Neuroinflammation involves microglial activation and has been shown to exacerbate AD. This review attempted to elucidate the role of cytokines, growth factors, and associated mechanisms implicated in the course of AD, especially with neuroinflammation. We also evaluated the propensities and specific mechanism(s) of cytokines and growth factors impacting neuron upon apoptotic decline and further shed light on the availability and accessibility of cytokines across the blood-brain barrier and choroid plexus in AD pathophysiology. The pathogenic and the protective roles of macrophage migration and inhibitory factors, neurotrophic factors, hematopoietic-related growth factors, TAU phosphorylation, advanced glycation end products, complement system, and glial cells in AD and neuropsychiatric pathology were also discussed. Taken together, the emerging roles of these factors in AD pathology emphasize the importance of building novel strategies for an effective therapeutic/neuropsychiatric management of AD in clinics.

Okadaic Acid and Hypoxia Induced Dementia Model of Alzheimer's Type in Rats.

Alzheimer's disease (AD) is the most common cause of progressive decline of memory function in aged humans. To study about a disease mechanism and progression, animal models for the specific disease are needed. For AD, although highly valid animal models exist, none of the existing models recapitulates all aspects of human AD. The pathogenic mechanisms involved in AD are diverse and thus it is difficult to recapitulate human AD in model organisms. Intracerebroventricular (ICV) injection of okadaic acid (OKA), a protein phosphatase 2A (PP2A) inhibitor, in rats causes neurotoxicity associated with neurofibrillary degeneration. However, this model lacks amyloid pathology as observed in AD. We aimed at combining two different treatments and hence producing a better animal model of AD which may mimic most of the neuropathological, neurobehavioral, and neurochemical changes observed in AD. For this, OKA (200 ng) was microinjected bilaterally into the hippocampus of male Wistar rats followed by exposure of same rats to hypoxic conditions (10%) for 3 days. The result of which, the combination model exhibited tau hyperphosphorylation along with Aß upregulation as evident by western blotting and immunohistochemistry. The observed changes were accompanied with dysfunction of neurotransmitter system, i.e., decreased acetylcholine activity and expression. This combinatorial model also exhibited cognitive deficiency which was assessed by Morris water maze and avoidance tests along with enhanced oxidative stress which is thought to be a major player in AD pathogenesis. Taken together, we established an easily reproducible and reliable rat model for sporadic dementia of Alzheimer's type in rats which allows effective testing of new therapeutic strategies.

Cell cycle activation in p21 dependent pathway: An alternative mechanism of organophosphate induced dopaminergic neurodegeneration.

In the previous study, we demonstrated that dichlorvos induces oxidative stress in dopaminergic neuronal cells and subsequent caspase activation mediates apoptosis. In the present study, we evaluated the effect and mechanism of dichlorvos induced oxidative stress on cell cycle activation in NGF-differentiated PC12 cells. Dichlorvos exposure resulted in oxidative DNA damage along with activation of cell cycle machinery in differentiated PC12 cells. Dichlorvos exposed cells exhibited an increased expression of p53, cyclin-D1, pRb and decreased expression of p21suggesting a re-entry of differentiated cells into the cell cycle. Cell cycle analysis of dichlorvos exposed cells revealed a reduction of cells in the G0/G1 phase of the cell cycle (25%), and a concomitant increase of cells in S phase (30%) and G2/M phase (43.3%) compared to control PC12 cells. Further, immunoblotting of cytochrome c, Bax, Bcl-2 and cleaved caspase-3 revealed that dichlorvos induces a caspase-dependent cell death in PC12 cells. These results suggest that Dichlorvos exposure has the potential to generate oxidative stress which evokes activation of cell cycle machinery leading to apoptotic cell death via cytochrome c release from mitochondria and subsequent caspase-3 activation in differentiated PC12 cells.

Understanding Aspects of Aluminum Exposure in Alzheimer's Disease Development.

Aluminum is a ubiquitously abundant nonessential element. Aluminum has been associated with neurodegenerative diseases such as Alzheimer's disease (AD), amyotrophic lateral sclerosis, and dialysis encephalopathy. Many continue to regard aluminum as controversial although increasing evidence supports the implications of aluminum in the pathogenesis of AD. Aluminum causes the accumulation of tau protein and Aß protein in the brain of experimental animals. Aluminum induces neuronal apoptosis in vivo and in vitro, either by endoplasmic stress from the unfolded protein response, by mitochondrial dysfunction, or a combination of them. Some, people who are exposed chronically to aluminum, either from through water and/or food, have not shown any AD pathology, apparently because their gastrointestinal barrier is more effective. This article is written keeping in mind mechanisms of action of aluminum neurotoxicity with respect to AD.

TLR5-mediated sensing of gut microbiota is necessary for antibody responses to seasonal influenza vaccination.

Systems biological analysis of immunity to the trivalent inactivated influenza vaccine (TIV) in humans revealed a correlation between early expression of TLR5 and the magnitude of the antibody response. Vaccination of Trl5(-/-) mice resulted in reduced antibody titers and lower frequencies of plasma cells, demonstrating a role for TLR5 in immunity to TIV. This was due to a failure to sense host microbiota. Thus, antibody responses in germ-free or antibiotic-treated mice were impaired, but restored by oral reconstitution with a flagellated, but not aflagellated, strain of E. coli. TLR5-mediated sensing of flagellin promoted plasma cell differentiation directly and by stimulating lymph node macrophages to produce plasma cell growth factors. Finally, TLR5-mediated sensing of the microbiota also impacted antibody responses to the inactivated polio vaccine, but not to adjuvanted vaccines or the live-attenuated yellow fever vaccine. These results reveal an unappreciated role for gut microbiota in promoting immunity to vaccination.

Effect of Resveratrol and Nicotine on PON1 Gene Expression: In Vitro Study.

Dietary and lifestyle factors have been shown to have a profound effect on paraoxonase-1 (PON1) activity. Cigarette smoke has been shown to inhibit its mass and activity where as resveratrol has been shown to enhance it. We exposed hepatoma derived cell line (HepG2) to resveratrol and nicotine in varying doses and measured PON1 enzymatic activity and PON1 gene expression. In addition, total protein content of HepG2 cells was also measured. Resveratrol in a dose of 15 µmol/l or above significantly increased the PON1 enzyme activity (p > 0.001) where as nicotine in a dose of 1 µmol/l or higher significantly reduced it (p < 0.05). The resveratrol in this dose also enhanced the PON1 gene expression whereas nicotine decreased it as compared to controls. However, the protein conent of cells was not changed suggesting that they were not cytotoxic in the doses used. Till date the antioxidant vitamins have shown disappointing results against LDL oxidation and cardiovascular protection. However, the effect of resveratrol on PON1 gene expression and activity was significant, suggesting increase in PON1 activity and enhanced gene expression may be its alternative mechanism for offering protection against cardiovascular disease and may be an potential pharmacological agent which can be used for this.

4-Hydroxy TEMPO attenuates dichlorvos induced microglial activation and apoptosis.

Microglial cells have been implicated in various neurodegenerative diseases. Previous studies from our lab have shown that dichlorvos (an organophosphate) could induce Parkinson's like features in rats. Recently, we have shown that dichlorvos can induce microglial activation, and if not checked in time could ultimately induce neuronal apoptosis. However, this activation does not always pose a threat to the neurons. Activated microglia also secrete various neuronal growth factors, suggesting that they have beneficial roles in CNS repair. Therefore, it is essential to control their detrimental functions selectively. Here, we tried to find out how microglial cells behave when exposed to dichlorvos in either the presence or absence of potent nitric oxide scavenger and superoxide dismutase mimetic, 4-hydroxy TEMPO (4-HT). Wistar rat pups (1 day) were used to isolate and culture primary microglial cells. We found 4-HT pretreatment successfully attenuated the dichlorvos mediated microglial activation. Moreover, 4-HT pretreatment decreased the up-regulated levels of p53 and its downstream effector, p21. The expression of various cell cycle regulators such as Chk2, CDC25a, and cyclin A remained close to their basal levels when 4-HT pretreatment was given. DNA fragmentation analysis showed significant reduction in the DNA damage of 4-HT pretreated microglia as compared to dichlorvos treated cells. In addition to this, we found 4-HT pretreatment prevented the microglial cells from undergoing apoptotic cell death even after 48 h of dichlorvos exposure. Taken together, our results showed 4-HT pretreatment could successfully ameliorate the dichlorvos induced microglial cell damage.

Attenuation of dichlorvos-induced microglial activation and neuronal apoptosis by 4-hydroxy TEMPO.

The neurotoxic consequences of acute high-level as well as chronic low-level organophosphates exposure are associated with a range of abnormalities in nerve functions. Previously, we have shown that after 24 h of dichlorvos exposure, microglia become activated and secrete pro-inflammatory molecules like nitric oxide, tumour necrosis factor-α and interleukin-1ß. Here, we extended our findings and focused on the neuronal damage caused by dichlorvos via microglial activation. For this, neurons and microglia were isolated separately from 1-day-old Wistar rat pups. Microglia were treated with dichlorvos for 24 h and supernatant was collected (dichlorvos-induced conditioned medium, DCM). However, when 4-hydroxy TEMPO (4-HT) pretreatment was given, we observed significant attenuation of dichlorvos-induced microglial activation; we also collected the supernatant of this culture (4-HT + DCM, TDCM). Next, we checked the effects of DCM on neurons and found heavy loss in viability as evident from NF-H immunostaining and MTT results, whereas dichlorvos alone-treated neurons showed comparatively less damage. However, we observed significant increase in neuronal viability when cells were treated with TDCM. Semi-quantitative PCR and western blot results revealed significant increase in p53, Bax and cytochrome c levels along with caspase 3 activation after 24 h of DCM treatment. However, TDCM-treated neurons showed significant decrease in the expression of these pro-apoptotic molecules. Taken together, these findings suggest that 4-HT can significantly attenuate dichlorvos-induced microglial activation and prevent apoptotic neuronal cell death.

Quercetin protects against chronic aluminum-induced oxidative stress and ensuing biochemical, cholinergic, and neurobehavioral impairments in rats.

In this study, we investigated the protective effect of chronic quercetin (a natural flavanoid) administration against Al-induced cognitive impairments, oxidative damage, and cholinergic dysfunction in male Wistar rats. Al lactate (10 mg/kg b.wt./day) was administered intragastrically to rats which were pre-treated with quercetin (10 mg/kg b.wt./day, intragastrically) for 12 weeks. At the end of 6 or 12 weeks of the study, several behavioral parameters were carried out to evaluate cognitive functions. Further after 12 weeks of exposure, various biochemical tests and H&E staining were performed to assess the extent of oxidative damage and neurodegeneration, respectively. Al levels were also estimated in HC and CS regions of rat brain. Chronic administration of quercetin caused significant improvement in the muscle coordination, cognition, anxiety, locomotion, and initial exploratory patterns in Al-treated rats. Quercetin supplementation to Al-treated animals also reduced oxidative stress, decreased ROS production, increased MnSOD activity and glutathione levels with decreased lipid peroxidation and protein oxidation. It increased AChE activity and ATP levels in HC and CS regions of rat brain compared to Al-treated rats. Quercetin administration ameliorates Al-induced neurodegenerative changes in Al-treated rats as seen by H&E staining. Further with the help of atomic absorption spectrophotometer, we found that quercetin supplementation to Al-treated rats also decreases the accumulation of Al in the HC and CS regions of rat brain. Taken together the results of this study show that quercetin offers neuroprotection against Al-induced cognitive impairments, cholinergic dysfunction, and associated oxidative damage in rats.

Effect of acute aluminum phosphide exposure on rats: a biochemical and histological correlation.

Aluminum phosphide (AlP), a widely used fumigant and rodenticide leads to high mortality if ingested. Its toxicity is due to phosphine liberated when it comes in contact with moisture. The exact mechanism of action of phosphine is not known. In this study male Wistar rats were used. The animals received a single dose (20mg AlP/kg body weight i.g.) orally. Basic serum biochemical parameters, activity of mitochondrial complexes, antioxidant enzymes and parameters of oxidative stress, individual mitochondrial cytochrome levels were measured along with tissue histopathology and immunostaining for cytochrome c and compared with controls. The serum levels of creatinine kinase-MB, lactate dehydrogenase, magnesium and cortisol were higher (p<0.01); the activities of mitochondrial complexes I, II, IV were observed to be significantly decreased in liver tissue in treated rats (p<0.01). The activity of catalase was lower (p<0.05) with a significant increase in lipid peroxidation (p<0.05) whereas superoxide dismutase and glutathione peroxidase were unaffected in them. There was a significant decrease in all the cytochromes in brain and liver tissues (p<0.05) with the exception of cytochrome b in brain, the levels of which remained same. Histopathology revealed congestion in most organs with centrizonal hemorrhagic necrosis in liver. Ultra structural changes indicating mitochondrial injury was observed in heart, liver and kidney tissues. There was also a marked reduction in the cytochrome-c immunostaining compared to the controls. Toxicity due to AlP appears to result as a consequence of both-energy insufficiency and oxidative stress, with a possible and preferential interaction with the tissue cytochromes.

Dichlorvos exposure results in activation induced apoptotic cell death in primary rat microglia.

Dichlorvos [2,2-dichlorovinyl dimethyl phosphate] is one of the most common in-use organophosphate (OP) in developing nations. Previous studies from our lab have shown chronic Dichlorvos exposure leads to neuronal cell death in rats. However, the extent of damage caused by Dichlorvos to other cells of the central nervous system (CNS) is still not clear. Microglial cells are the primary threat sensors of CNS which become activated in many pathological conditions. Activation of microglial cells results in reactive microgliosis, manifested by increased cellular damage in the affected regions. Using rat primary microglial cultures, here we show that Dichlorvos exposure can activate and induce apoptotic cell death in microglia. We observed significant up-regulation of pro-inflammatory molecules like nitric oxide, TNF-α, and IL-1ß when microglia were treated with Dichlorvos (10 µM). Significant up-regulation of CD11b, microglial specific activation marker, was also observed after 24 h of Dichlorvos treatment. The activated microglial cells eventually undergo cell death after 48 h of Dichlorvos treatment. The DNA fragmentation pattern of Dichlorvos treated microglia along with increased expression of Bax in mitochondria, cytochrome c release from mitochondria, and caspase-3 activation led us to assume that microglia were undergoing apoptosis. Thus, the present study showed that Dichlorvos can induce microglial activation and ultimately apoptotic cell death. These findings gave new perspective to the current knowledge of Dichlorvos (OPs) mediated CNS damage and presents microglial activation as a potential therapeutic target for preventing the OP induced neuronal damage.

siRNA against presenilin 1 (PS1) down regulates amyloid ß42 production in IMR-32 cells.

BACKGROUND: One of the pathological hallmarks of Alzheimer's disease (AD) is the deposition of the ~4 kDa amyloid ß protein (Aß) within lesions known as senile plaques. Aß is also deposited in the walls of cerebral blood vessels in many cases of AD. A substantial proportion of the Aß that accumulates in the AD brain is deposited as Amyloid, which is highly insoluble, proteinaceous material with a ß-pleated-sheet conformation and deposited extracellularly in the form of 5-10 nm wide straight fibrils. As γ-secretase catalyzes the final cleavage that releases the Aß42 or 40 from amyloid ß -protein precursor (APP), therefore, it is a potential therapeutic target for the treatment of AD. γ-Secretase cleavage is performed by a high molecular weight protein complex containing presenilins (PSs), nicastrin, Aph-1 and Pen-2. Previous studies have demonstrated that the presenilins (PS1 and PS2) are critical components of a large enzyme complex that performs γ-secretase cleavage. METHODS: In this study we used RNA interference (RNAi) technology to examine the effects of small-interfering RNA (siRNA) against PS1 on expression levels of PS1 and Aß42 in IMR-32 Cells using RTPCR, western blotting and immunofluorescence techniques. RESULTS: The results of the present study showed down regulation of PS1 and Aß42 in IMR32 cells transfected with siRNA against PS1. CONCLUSION: Our results substantiate the concept that PS1 is involved in γ-secretase activity and provides the rationale for therapeutic strategies aimed at influencing Aß42 production.

Aluminum phosphide poisoning: an unsolved riddle.

Aluminum phosphide (ALP), a widely used insecticide and rodenticide, is also infamous for the mortality and morbidity it causes in ALP-poisoned individuals. The toxicity of metal phosphides is due to phosphine liberated when ingested phosphides come into contact with gut fluids. ALP poisoning is lethal, having a mortality rate in excess of 70%. Circulatory failure and severe hypotension are common features of ALP poisoning and frequent cause of death. Severe poisoning also has the potential to induce multi-organ failure. The exact site or mechanism of its action has not been proved in humans. Rather than targeting a single organ to cause gross damage, ALP seems to work at the cellular level, resulting in widespread damage leading to multiorgan dysfunction (MOD) and death. There has been proof in vitro that phosphine inhibits cytochrome c oxidase. However, it is unlikely that this interaction is the primary cause of its toxicity. Mitochondria could be the possible site of maximum damage in ALP poisoning, resulting in low ATP production followed by metabolic shutdown and MOD; also, owing to impairment in electron flow, there could be free radical generation and damage, again producing MOD. Evidence of reactive oxygen species-induced toxicity owing to ALP has been observed in insects and rats. A similar mechanism could also play a role in humans and contribute to the missing link in the pathogenesis of ALP toxicity. There is no specific antidote for ALP poisoning and supportive measures are all that are currently available.

Mitochondrial energy metabolism impairment and liver dysfunction following chronic exposure to dichlorvos.

Although the effects of acute pesticide poisoning are well known but, hardly any data exists regarding the health effects after long-term low-level exposure. Major unresolved issues include the effect of moderate exposure in the absence of poisoning. The present study elucidates a possible mechanism by which chronic organophosphate exposure (dichlorvos 6 mg/kg b.wt., s.c. for 12 weeks) causes liver dysfunction. Mitochondria, a primary site of cellular energy generation and oxygen consumption represent a likely target for organophosphate poisoning. Therefore, the objective of the current study was planned with an aim to investigate the effect of chronic dichlorvos exposure on liver mitochondrial electron transport chain (ETC), mitochondrial calcium uptake and its implications on the induction of liver enzymes and liver dysfunction in rodent model. Our results indicated decreased mitochondrial electron transfer activities of cytochrome oxidase along with altered mitochondrial complexes I and II activity. This decrease in the activities of electron transport complexes in turn affected the ATP synthesis and ATP levels adversely in the mitochondria isolated from dichlorvos (DDVP) treated rat liver. Mitochondrial preparation from DDVP treated rat liver demonstrated significant increase in mitochondrial Ca(2+) uptake and increase ROS levels. The alterations in the mitochondrial calcium uptake, mitochondrial electron transfer enzyme activities and increase ROS levels in turn might have caused an increase in liver enzymes (ALT, AST and ALP). The electron micrographs of liver cells depicted morphological changes in mitochondria as well as nucleus following 12 weeks of exposure to DDVP. These studies provide an evidence of impaired mitochondrial bioenergetics which may lead to liver dysfunction after chronic exposure to dichlorvos.

Impaired mitochondrial energy metabolism and kinetic properties of cytochrome oxidase following acute aluminium phosphide exposure in rat liver.

The present study was designed with an aim to analyze the effect of acute aluminium phosphide (ALP) exposure (10mg/kg b.wt, intragastrically) on the kinetic characteristics of cytochrome oxidase and energy metabolism in male Wistar rat liver mitochondria. Liver mitochondrial preparations from ALP-treated rats exhibited significant decrease (66%) in the activity of cytochrome oxidase suggesting that there was a decrease in the catalytic efficiency of the active oxidase molecules on ALP treatment. The decreased activity of cytochrome oxidase with altered NADH and succinic dehydrogenase activities might have contributed towards a significant decline in state 3 and state 4 respiration as observed. These alterations in the electron transport chain complexes in turn adversely affected the ATP synthesis rate as well as ATP levels in the mitochondria isolated from treated rats. The alterations in the respiratory chain, was followed by enhanced lipid peroxidation in rat liver mitochondria which might have further contributed to change in the fluidity of membrane as depicted by decreased fatty acid content of liver mitochondria. However, no significant change was observed in cholesterol and phospholipids content in our study. The present study thus highlights the significance of altered mitochondrial respiratory chain functions and membrane integrity after acute ALP exposure.

Neurobehavioral impairments, generation of oxidative stress and release of pro-apoptotic factors after chronic exposure to sulphur mustard in mouse brain.

Recent global events have focused attention on the potential threat of international and domestic chemical terrorism, as well as the possibility of chemical warfare proliferation. Sulphur mustard (SM) is one of the potent chemical warfare agents (CWA), which initiates a cascade of events that converge on the redox mechanisms common to brain injury. The present study was designed to examine the effects of chronic SM exposure on neurobehavioral impairments, mitochondrial oxidative stress in male Swiss Albino mice and its role in inducing apoptotic neuronal cell death. The animals were divided into four groups (control, low, medium and high dose) of 5 animals each. Exposure to SM was given percutaneously daily for 12 weeks. The results demonstrated impairment in neurobehavioral indices viz. rota rod, passive avoidance and water maze tests in a dose dependent manner. There was a significant increase in lipid peroxidation and protein carbonyl content whereas, decrease in the activity of manganese superoxide dismutase (MnSOD), glutathione reductase and glutathione peroxidase suggesting impaired antioxidant defense system. Immunoblotting of cytochrome c, Bcl-2, Bax and activation of caspase-3 suggest induction of apoptosis in a dose dependent manner. Finally, increased p53 expression suggests that it may target the mitochondrial pathway for inducing apoptosis in response to DNA damage signals. In conclusion, chronic SM exposure may have the potential to generate oxidative stress which may trigger the release of cytochrome c as well as caspase-3 activation in neurons leading to cell death by apoptosis in a dose dependent manner which may in the end be responsible for the disruption of cognitive functions in mice.

Aluminium-induced oxidative DNA damage recognition and cell-cycle disruption in different regions of rat brain.

The present study was undertaken to reveal the effects of chronic aluminium exposure (10 mg/kg/b.wt, intragastrically for 12 weeks) on the oxidative DNA damage and its implication on the expression of p53 and other cell-cycle regulatory proteins in male Wister rats. Chronic aluminium exposure resulted in increased formation of 8-hydoxydeoxyguanosine in the mitochondrial DNA isolated from different regions of rat brain. The agarose gel electrophoresis revealed the DNA fragmentation pattern in aluminium-treated rat brain regions. Increased expression of p53 demonstrated that aluminium induces DNA damage. Western blot and mRNA expression analysis demonstrated increased expression of cyclin D1, suggesting disruption of cell cycle. The immunohistochemical studies showed nuclear localization of p53; however, the localization of cyclin D1 was both cytoplasmic and nuclear in aluminium-treated rat brain regions. Thus, the findings of the present study reveal that aluminium-induced oxidative damage to DNA may be involved in the neurodegeneration via increase in p53 expression and activation of cell cycle.

Impairment of mitochondrial energy metabolism in different regions of rat brain following chronic exposure to aluminium.

The present study was designed with an aim to evaluate the effects of chronic aluminium exposure (10 mg/kg b.wt, intragastrically for 12 weeks) on mitochondrial energy metabolism in different regions of rat brain in vivo. Mitochondrial preparations from aluminium treated rats revealed significant decrease in the activity of various electron transport complexes viz. cytochrome oxidase, NADH cytochrome c reductase and succinic dehydrogenase as well, in the hippocampus region. The decrease in the activity of these respiratory complexes was also seen in the other two regions viz. corpus striatum and cerebral cortex, but to a lesser extent. This decrease in the activities of electron transport complexes in turn affected the ATP synthesis and ATP levels adversely in the mitochondria isolated from aluminium treated rat brain regions. We also studied the spectral properties of the mitochondrial cytochromes viz. cyt a, cyt b, cyt c1, and cyt c in both control and treated rat brains. The various cytochrome levels were found to be decreased following 12 weeks of aluminium exposure. Further, these impairments in mitochondrial functions may also be responsible for the production of reactive oxygen species and impaired antioxidant defense system as observed in our study. The electron micrographs of neuronal cells depicted morphological changes in mitochondria as well as nucleus only from hippocampus and corpus striatum regions following 12 weeks exposure to aluminium. The present study thus highlights the significance of altered mitochondrial energy metabolism and increased ROS production as a result of chronic aluminium exposure in different regions of the rat brain.