Browsing by Author "M.K. Thakur"

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ADP-ribosylation of HMG proteins and its modulation by different effectors in the liver of aging rats
(1990) M.K. Thakur; S. Prasad
The in vitro ADP-ribosylation of high mobility group (HMG) non-histone proteins and its modulation by spermine, butyrate, dexamethasone and 3-aminobenzamide were studied in the liver of young (14 weeks) and old (113 weeks) male rats. ADP-ribosylation of HMG 1 was similar in both ages, whereas that of HMG 2 and 14 decreased but HMG 17 increased in the old. HMG 1 was ADP-ribosylated to a greater extent in young but to a lower level in the old by different effectors except spermine which showed no influence in old age. ADP-ribosylation of HMG 2 was stimulated by spermine, butyrate and dexamethasone in old but only by spermine in young rats. Other effectors decreased the ADP-ribosylation of HMG 2 in young. The ADP-ribosylation of HMG 14 was stimulated by spermine in the old but that of HMG 17 was reduced by butyrate in young and by spermine in the old. Dexamethasone decreased the ADP-ribosylation of both HMG 14 and 17 in young, whereas this showed no change in old age. Aminobenzamide inhibited ADP-ribosylation of only HMG 2 in young but all HMGs except HMG 2 in the old. Such alteration in the ADP-ribosylation of HMG proteins may affect various cellular and nuclear functions of rat liver during aging. © 1990.
Age- and sex-related expression of norbin in the brain cortex of mice
(2001) S.T. Mani; R.C. Kumar; M.K. Thakur
Norbin is a novel neuron specific protein that extends the neurites of neuronal cells. It is expressed in neural tissues like brain cortex, hippocampus, spinal cord and cerebellum. In this paper, we have studied the expression of norbin mRNA and protein in the brain cortex of male and female mice of different ages. Northern blot analysis showed that the level of norbin mRNA increased in both sexes during aging. However, Western blotting revealed that the protein increased in male but decreased in female with advancing age. These findings suggest that norbin is involved in brain function which is dependent on age and sex. © 2001 Elsevier Science Ireland Ltd.
Age-Dependent Decline of Nogo-A Protein in the Mouse Cerebrum
(Springer Science and Business Media, LLC, 2014) Anita Kumari; M.K. Thakur
Nogo-A, a myelin-associated neurite growth inhibitory protein, is implicated in synaptic plasticity. It binds to its receptor namely the Nogo-66 receptor1 (NgR1) and regulates filamentous (F) actin dynamics via small GTPases of the Rho family, RhoA kinase (ROCK), LimK and cofilin. These proteins are associated with the structural plasticity, one of the components of synaptic plasticity, which is known to decline with normal aging. So, the level of Nogo-A and its receptor NgR1 are likely to vary during normal brain aging. However, it is not clearly understood how the levels of Nogo-A and its receptor NgR1 change in the cerebrum during aging. Several studies show an age- and gender-dependent decline in synaptic plasticity. Therefore, the present study was planned to analyze the relative changes in the mRNA and protein levels of Nogo-A and NgR1 in both male and female mice cerebrum during normal aging. Western blot analysis has shown decrease in Nogo-A protein level during aging in both male and female mice cerebrum. This was further confirmed by immunofluorescence analysis. RT-PCR analysis of Nogo-A mRNA showed no significant difference in the above-mentioned groups. This was also supported by in situ hybridization. NgR1 protein and its mRNA expression levels showed no significant alteration with aging in the cerebrum of both male and female mice. Taken together, we speculate that the downregulation of Nogo-A protein might have a role in the altered synaptic plasticity during aging. © 2014, Springer Science+Business Media New York.
Age-dependent decrease in the interaction of β-tubulin with estrogen receptor alpha transactivation domain in mouse brain
(2009) Swati Ghosh; M.K. Thakur
In the previous paper, we reported that four proteins of 100 kDa, 80 kDa, 68 kDa and 50 kDa from the mouse brain interacted with estrogen receptor (ER) α-transactivation domain (TAD) and 68 kDa protein showed age and sex dependent changes. Here, we describe the identification of 50 kDa protein as β-tubulin and changes in its interaction with age and sex in mouse brain. It is a microtubule-associated protein which binds to activation function (AF)-1 region of ERα-TAD and is involved in estrogen signaling. The extent of interaction of mouse ERα-TAD with β-tubulin was higher in adult female as compared to old female and adult male. However, the expression of β-tubulin showed no significant change with age and sex. Such age and sex dependent alteration in the interaction of β-tubulin might account for the estrogen-mediated brain functions during aging. © 2009 Elsevier Ireland Ltd. All rights reserved.
Age-dependent effects of sodium butyrate and hydrocortisone on acetylation of high mobility group proteins of rat liver.
(1988) S. Prasad; M.K. Thakur
The in vitro acetylation of high mobility group (HMG) proteins and its modulation by sodium butyrate and hydrocortisone have been studied using liver slices of young (13-) and old (114-week-old) rats. Acetylation of total HMG proteins was significantly higher in young than old rats. HMG 1, in particular, showed greater acetylation than others. Whereas acetylation of HMG 1 and 2 decreased drastically, that of HMG 14 and 17 increased in old age. In young rats, sodium butyrate and hydrocortisone stimulated acetylation of HMG 14 and 17, and decreased that of HMG 2. Butyrate had no effect on HMG 1, but hydrocortisone decreased it. In old rats, butyrate and hydrocortisone decreased acetylation of all HMGs, except HMG 17, which was stimulated to a slight extent by butyrate.
Age-related analysis of EcoRI generated satellite DNA-containing chromatin of rat liver
(Taylor and Francis Inc., 1996) Pratima Chaurasia; Shibani Mukherjee; M.K. Thakur
EcoRI digestion of nuclei and their subsequent lysis with EDTA solubilizes 45% and 36% of chromatin DNA from the liver of young (18 ± 2 weeks) and old (100 ± 5 weeks) rats, respectively. After hybridization with 185 bp rat satellite I DNA, these soluble fractions are found to be enriched in specific DNA sequences such as satellite DNA. Besides regular repeat pattern, a major portion of the satellite chromatin forms higher order organization. Digestion kinetics confirms condensation of satellite DNA-containing chromatin similar to that of bulk chromatin in old age. Furthermore, densitometric scanning of the slot-blot of soluble chromatin fractions reveals loss of satellite DNA in the old. However, an increase in the linker histone H1 and its subfraction H10 in the satellite DNA-enriched fraction of chromatin from old rats suggests greater compaction. These results provide the first evidence that the satellite DNA-containing chromatin differs in the liver of young and old rats.
Age-related binding of proximal region of apoe promoter to nuclear proteins of mouse cerebral cortex
(2011) Sarika Singh; M.K. Thakur
Mouse ApolipoproteinE (ApoE) gene has maximum promoter activity in the proximal region (-212 to ?54) which includes different regulatory elements. These elements bind to specific protein factors and influence the expression of genes which are involved in key brain functions that decline with age. As there is no information on the binding of apoE promoter to nuclear proteins as a function of age, we have analyzed the binding of USF, AP1 and one negative element sequence present in ApoE proximal promoter to nuclear proteins of the cerebral cortex of mice of different ages. The findings show the formation of one complex with USF and two complexes with AP1 and negative element. The intensity of these complexes varies with age, indicating differential binding of protein factors to specific elements of apoE promoter, which reflect age-related regulation of apoE -mediated brain functions. © Springer Science+Business Media, LLC 2011.
Age-related changes in the structure and function of chromatin: A review
(1984) M.K. Thakur
Due to rapid advancement in biochemical and biophysical techniques during the last decade, extensive studies have been undertaken to understand the structure and function of chromatin. Several interesting results have benn reported regarding the changes in basic organization and function of chromatin during the life time of a eukaryotic cell. The data accumulated so far have been obtained with different organs and organisms and widely differing methods, and the conclusions drawn from them are sometimes contradictory. In this paper, therefore, the available data on the age-associated alterations in the composition, structure and function of chromatin have been discussed, and an attempt has been made to correlate the structural changes in chromatin with alteration in gene expression during aging. © 1984.
Age-related expression of Neurexin1 and Neuroligin3 is correlated with presynaptic density in the cerebral cortex and hippocampus of male mice
(Kluwer Academic Publishers, 2015) Dhiraj Kumar; M.K. Thakur
Neurexin1 (Nrxn1) and Neuroligin3 (Nlgn3) are cell adhesion proteins, which play an important role in synaptic plasticity that declines with advancing age. However, the expression of these proteins during aging has not been analyzed. In the present study, we have examined the age-related changes in the expression of these proteins in cerebral cortex and hippocampus of 10-, 30-, 50-, and 80-week-old male mice. Reverse transcriptase polymerase chain reaction (RT-PCR) analysis indicated that messenger RNA (mRNA) level of Nrxn1 and Nlgn3 significantly increased from 10 to 30 weeks and then decreased at 50 weeks in both the regions. However, in 80-week-old mice, Nrxn1 and Nlgn3 were further downregulated in cerebral cortex while Nrxn1 was downregulated and Nlgn3 was upregulated in hippocampus. These findings were corroborated by immunoblotting and immunofluorescence results. When the expression of Nrxn1 and Nlgn3 was correlated with presynaptic density marker synaptophysin, it was found that synaptophysin protein expression in cerebral cortex was high at 10 weeks and decreased gradually up to 80 weeks, whereas in hippocampus, it decreased until 50 weeks and then increased remarkably at 80 weeks. Furthermore, Pearson’s correlation analysis showed that synaptophysin had a strong relation with Nrxn1 and Nlgn3 in cerebral cortex and with Nlgn3 in hippocampus. Thus, these findings showed that Nrxn1 and Nlgn3 are differentially expressed in cerebral cortex and hippocampus which might be responsible for alterations in synaptic plasticity during aging. © 2015, American Aging Association.
Age-specific methylation of high-mobility-group proteins of the rat liver and its modulation by spermine and sodium butyrate
(1989) M.K. Thakur; S. Prasad
Liver slices from young (20 weeks) and old (117 weeks) rat were incubated with [methyl-14C]methionine in the absence or presence of spermine or sodium butyrate. The high-mobility-group (HMG) non-histone proteins were extracted from the liver with perchloric acid and separated by acid-urea polyacrylamide slab gel electrophoressi. Methylation of HMG proteins decreased drastically in old rats. Whereas spermine inhibited the methylation of total HMG proteins in young rats, it had no effect in old age. On the contrary, sodium butyrate did not change the incorporation of methyl groups into total HMG proteins of young rats, but inhibited that of old rats. Particularly, the incorporation of [14C]methyl groups into HMG 2 was enhanced but into other HMGs it was reduced by both effectors in young and old age. Such discrepancies in the methylation of HMG proteins and their differential modulation by spermine and butyrate might affect the higher-order organization of chromatin and consequential destabilize the expression of genes during aging. © 1989.
Aging of brain: Role of estrogen
(2006) M.K. Thakur; P.K. Sharma
The brain undergoes many structural and functional changes during aging. Some of these changes are regulated by estrogens which act mainly through their intracellular receptors, estrogen receptor ERα and ERβ. The expression of these receptors is regulated by several factors including their own ligand estrogen, and others such as growth hormone and thyroid hormone. The levels of these factors decrease during aging which in turn influence estrogen signaling leading to alterations in brain functions. In the present paper, we review the effects of aging on brain structure and function, and estrogen action and signaling during brain aging. The findings suggest key role of estrogen in the maintenance of brain functions during aging. © 2006 Springer Science+Business Media, LLC.
Alzheimer's disease - A challenge in the new millennium
(2000) M.K. Thakur
Alzheimer's disease (AD) is a neurodegenerative disorder of the central nervous system that begins in middle to late life, and results in severe dementia and ultimately death. It is the fourth major cause of death after heart diseases, cancer and stroke. AD is a complex genetic disorder with mutations in different genes resulting in the same clinical and pathological phenotype. It is incurable and progressively debilitating leading to death due to secondary infections and diseases. The management cost of AD is huge and it becomes an enormous social and economic burden on support services. Therefore, AD is an intellectually challenging and socially important problem for health care of the elderly. Its importance has recently attracted the attention of scientists and social workers due to ever-increasing proportion of the elderly population throughout the world. There is an urgent need for the development of drugs that can ameliorate or delay the onset of symptoms of the disease. This article focuses on the current concept of genetic basis, possible risk factors and therapeutic approaches for AD. The understanding of sequential steps in the cascade of events that mediate the effect of risk factors and gene mutations has provided potential therapeutic targets that may prevent the occurrence, slow the progress or improve the condition of AD patients. Such scientific advancements in the new millennium would be of significant value for the health of the elderly.
Amplification of exons 4 and 5 of Androgen Receptor gene by testosterone in aged female mouse brain cortex
(2000) M.K. Thakur; A. Asaithambi; Shibani Mukherjee
We have investigated the effect of testosterone on the amplification of androgen receptor (AR) gene in the brain cortex of aging female mice. For this purpose, high molecular weight (HMW) DNA purified from the brain cortex of intact, gonadectomized, testosterone- and estradiol-treated adult and old female mice was digested with different restriction enzymes and used for Southern hybridization with 32P-labeled AR cDNA fragments representing different domains of AR. The results reveal that only exons 4 and 5 corresponding to amino-terminal part of the hormone binding domain of AR are amplified in testosterone-treated old female but not in adult mice. Densitometric analysis further shows that testosterone increases the copy number of exons 4 and 5 of mouse AR gene by four-fold. Reprobing of slot blots with estrogen receptor and cathepsin D cDNA as probes supports the observation that amplification occurs only in AR gene. The tissue specificity is also confirmed when the slot blot hybridization of mouse liver HMW DNA with AR cDNA fails to show similar amplification. As the restriction map analysis of Southern blots does not show restriction fragment length polymorphism, the possibility of structural rearrangement leading to amplification of AR gene is ruled out. Thus our results suggest that the in vivo induction of mouse AR gene amplification by testosterone is tissue- and age-specific, and might contribute to the progress of genetic instability in the brain of aged female mice. © 2000 Kluwer Academic Publishers.
Amyloid Precursor Protein (APP) mRNA level is higher in the old mouse cerebral cortex and is regulated by sex steroids
(2011) Thamil Mani Sivanandam; M.K. Thakur
Alzheimer's disease (AD) is a neurodegenerative disorder characterized by the accumulation of amyloid beta peptides, which are produced by the proteolytic cleavage of amyloid precursor protein (APP). As the incidence of AD is higher in females and sex steroids are implicated in this disease, we have examined the effect of sex steroids (testosterone and 17β-estradiol) on the expression of APP mRNA and protein in the cerebral cortex of adult and old mice of both sexes. Northern blot analysis detected APP mRNA as a single 3.5-kb band and its level is increased in old as compared to adult. Following gonadectomy, its level was upregulated in female mice but downregulated in male mice. Supplementation with testosterone or estradiol decreased its levels in female mice of both ages. Testosterone supplementation increased the mRNA levels in both adult and old male mice. Estrogen supplementation decreased its level in adult but increased in old male mice. Western blot analysis detected APP specific bands ranging from 95 to 125 kD. The level of 95 kDa band representing APP695 protein showed difference in levels with age or hormone treatment. These results provide evidence for increase in APP mRNA level in the cerebral cortex of old mice and its regulation by sex steroids during aging. © Springer Science+Business Media, LLC 2010.
An insight into plant polyphenols in prevention of brain aging
(Elsevier, 2022) Padmanabh Singh; Arpita Konar; M.K. Thakur
With a global increase in the elderly population, brain aging-associated cognitive deficits and neurodegenerative pathologies have emerged as a major concern for healthcare and maintaining the quality of life. Age-associated neurodegenerative diseases are characterized by oxidative damage, inflammation, DNA damage, protein aggregation, and neuronal death leading to alteration of physiological and cognitive functions. Such complex etiology renders difficulty in treating neurological ailments and faces numerous pharmacological limitations, including adverse side effects, disturbance of body homeostasis, and high cost. In this regard, nutraceutical interventions including plant-derived dietary polyphenols have gained prominence in preventing cognitive aging and counteracting disease conditions. In this chapter, we provide comprehensive information on epidemiological and experimental studies supporting the pro-cognitive and neuroprotective potential of polyphenolic compounds during brain aging. The chapter also highlights the molecular mechanisms underlying neuro-nutraceutical potential of polyphenols, including gene expression, epigenetics, protein synthesis, and metabolic pathways. © 2023 Elsevier Inc. All rights reserved.
Analysis in vitro of uterine estrogen receptor conformation of young and old rats
(Kluwer Academic Publishers, 1991) M.K. Thakur; J. Kaur
The conformation of estrogen receptor (ER) and its in vitro transformation by RNase, Urea and ATP were analysed using the uteri of young (16 weeks) and old (92 weeks) rats. Following the digestion of ER with proteolytic enzymes like trypsin and chymotrypsin and the analysis of cleaved fragments by SDS-PAGE, similar pattern is observed in both ages. In vitro transformation of ER by RNase, Urea and ATP shows that the degree of transformation is lower in old than young. Furthermore, the transformed ER from old is less capable of binding to DNA than that from young. Thus our results show that the conformation of ER probably does not change with age, but the degree of transformation and the ability of transformed receptor to bind to DNA decrease with age. © 1991 Kluwer Academic Publishers.
Analysis of age-associated alteration in the synthesis of HMG nonhistone proteins of the rat liver
(Kluwer Academic Publishers, 1991) M.K. Thakur; S. Prasad
HMG proteins were extracted with 5% PCA or 0.35 M NaCl from whole tissue, nuclei or chromatin of the liver of young (19 weeks) and old (118 weeks) male rats. They were resolved on acetic acid-urea polyacrylamide gel. The electrophoretic patterns of the major HMG proteins 1, 2, 14 and 17 of both ages are similar. The in vitro synthesis of HMG 1 and 2 decreases, but that of HMG 14 and 17 increases considerably in the liver of old rats. The synthesis of different HMG proteins is modulated differentially by spermine, butyrate, dexamethasone and 3-aminobenzamide in the liver of young and old rats. These findings suggest that HMG proteins contribute to alterations in the organization of chromatin and expression of genes during aging. © 1991 Kluwer Academic Publishers.
Analysis of nucleosome arrangement on satellite DNA of rat liver chromatin
(Springer India, 1987) M.K. Thakur
The arrangement of nucleosomes on the nucleotide sequence of satellite DNA of Oceanian rat (Rattus rattus) has been studied. Nucleosome cores were prepared from rat liver nuclei with micrococcal nuclease, exonuclease III and nuclease Sl. From the total population of core DNA fragments, the satellite-containing fragments were selected by molecular cloning and the complete nucleotide sequence of these clones was determined. The data show that nucleosomes occupy a number of preferred positions on satellite DNA. These positions are strictly defined. Thus location of nucleosomes along the satellite sequence is non-random. Such finding may have important biological significance. © 1987 Printed in India.
Analysis of Presenilin 1 and 2 interacting proteins in mouse cerebral cortex during development
(Elsevier Ltd, 2014) Ashish Kumar; M.K. Thakur
In our previous report, we showed that Presenilin (PS)1 and 2 have differential expression profile from early embryonic stages till adulthood in mouse cerebral cortex, suggesting that both of these proteins are crucial for brain development. Genetic manipulation studies have also shown the involvement of PS1 in brain development, but PS2 remains largely unexplored. In order to understand how PS1 and 2 mediate developmental functions, we have investigated the interaction of PS1 and 2 with proteins of mouse cerebral cortex during development. Co-immunoprecipitation (Co-IP) combined with MALDI-MS/MS analysis revealed 12 interacting partners of PS1 and 11 partners of PS2. The interacting proteins were different for PS1 and 2, and involved in cell division, glycolysis, cell adhesion and protein trafficking. Densitometric analysis of protein bands visualized after SDS-PAGE separation of Co-IP proteins revealed variation in their amount and degree of interaction during different developmental stages of mice. Further, immunoblot based validation of PS1 interacting protein Notch-1 showed maximum interaction at embryonic day (E) 12.5, decline at E18.5, upregulation from postnatal day 0 (P0) to P20 and thereafter reduction at P45 and 20 weeks. In-silico analysis of PS and its interacting proteins indicated conformation based interaction through common type of secondary structures having alpha helical, extended beta strand and random coil, and CK2, PKC phosphorylation and myristoylation motifs. Taken together, our study showed that PS1 and PS2 interact to varying extent with different proteins of mouse cerebral cortex and suggests their interaction based on specific conformation and involvement in diverse functions essential for the brain development. © 2014 ISDN.
Androgen receptor mRNA is inversely regulated by testosterone and estradiol in adult mouse brain
(2004) R.C. Kumar; M.K. Thakur
Androgen receptor (AR) is expressed in different tissues including the brain and is under regulation by sex steroid hormones. It mediates the action of androgen which plays a key role in learning, memory, and other brain functions that deteriorate with increasing age. We have correlated the expression of AR mRNA with its promoter methylation and their regulation by testosterone and estradiol in the brain cortex of adult and old male and female mice. Results revealed that (i) AR mRNA expression was significantly higher in male than in female mice. (ii) In both sexes, AR mRNA level was down-regulated by testosterone in adult and old, but up-regulated by estradiol only in adult mice. (iii) Methylation of AR core promoter was increased by testosterone, but decreased by estradiol. These findings show that AR mRNA expression and its core promoter methylation are inversely regulated by testosterone and estradiol in the adult mice brain cortex. Such regulation of AR expression might influence androgen action during aging of the mice brain. © 2003 Elsevier Inc. All rights reserved.