Biomedical subjects

Thangavel Samikkannu

Publications and source records attributed to Thangavel Samikkannu.

6 recordsLinked to original sources

PubMed2026

Experimental approaches for investigating sirtuin-mediated mitochondrial function, cellular metabolism, and epigenetic regulation.

Cellular pathways for experimental discovery provide a comprehensive overview of sirtuin biology and its critical involvement in HIV-associated neurocognitive disorders (HAND) and related neurodegenerative diseases, highlighting the translational potential of sirtuin-targeted therapeutic strategies. As NAD+-dependent deacetylases and ADP-ribosyl transferases, sirtuins regulate diverse cellular processes, including stem cell maintenance, cellular proliferation, metabolic homeostasis, apoptosis, autophagy, oxidative stress responses, and genomic stability, all of which contribute to neuronal dysfunction and disease progression. This chapter focuses on key mammalian sirtuins, including SIRT1 and SIRT2, which are primarily localized within the nucleus and cytosol; mitochondrial sirtuins SIRT3, SIRT4, and SIRT5; and nuclear/nucleolar sirtuins SIRT6 and SIRT7. Here, a method with a detailed protocol to isolate compartment-specific sirtuin expression and activity was used: subcellular fractionation was performed using a subcellular fractionation kit to obtain cytosolic and nuclear fractions, while mitochondrial isolation was carried out using Tom20 antibody-conjugated magnetic microbeads. These approaches were applied to brain tissues from HIV-positive individuals, as well as to HIV-Tat-treated human microglial (HMC3) cells and astrocytes. This experimental framework enables accurate assessment of compartment-resolved sirtuin regulation in disease-relevant models. Collectively, the chapter highlights the protective roles of sirtuins in mitigating key pathogenic mechanisms underlying HAND and related neurodegenerative diseases. These findings support the emerging concept that sirtuins represent promising pharmacological targets for the development of novel therapeutic interventions in neurodegeneration and HIV-associated brain disorders.

Humans↗

PubMed2006

Acute effect of high glucose on long-term cell growth: a role for transient glucose increase in proximal tubule cell injury.

Although chronic exposure of renal cells to high glucose has been shown to cause cell injury, the effect of acute exposure has not been elucidated. In this study, we demonstrate that acute (10 min) exposure of human proximal tubule epithelial cells (hPTEC) to high glucose (25 mM) induces a time-dependent dual effect consisting of an early proliferation and a late apoptosis. Acute exposure of hPTEC to high glucose induced a twofold increase in DNA synthesis and cell number at 12 h. However, after 36 h, a significant decrease in cell growth is observed, followed by apoptosis. On glucose treatment, both p42/p44 mitogen-activated protein (MAP) kinases and the downstream signaling intermediate NF-kappaB were phosphorylated and translocated to the nucleus. Pretreatment of cells with MAP kinase and NF-kappaB-specific inhibitors abolished glucose-induced proliferation. However, these inhibitors were ineffective in preventing glucose-induced apoptosis. Interestingly, conditioned medium from cells exposed to high-glucose concentrations inhibited proliferation and concomitantly induced apoptosis in normal cells, suggesting that the inhibitory effect of glucose occurs through secretion of a secondary factor(s). In parallel to apoptosis, we observed an increased production of reactive oxygen species (ROS). Pretreatment of cells with the antioxidant N-acetyl cysteine reversed glucose-mediated ROS production and apoptosis, suggesting that ROS is involved in apoptosis. Our study demonstrates for the first time that a single high-glucose exposure for 10 min alone is sufficient to elicit proliferation and apoptosis in hPTEC and suggests that episodes of transient increase in glucose may contribute to cell damage leading to epithelial cell dysfunction.

Apoptosis↗

PubMed2004

alpha-thrombin rapidly induces tyrosine phosphorylation of a novel, 74-78-kDa stress response protein(s) in lung fibroblast cells.

We demonstrated previously that exposure of CCL39 lung fibroblasts to alpha-thrombin rapidly inhibits interleukin 6-induced tyrosine phosphorylation of signal transducers and activators of transcription 3 (Stat3). While studying the cross-talk between alpha-thrombin and interleukin 6, we observed that the phospho-specific (tyrosine) anti-Stat3 antibody specifically cross-reacted with a 74-78-kDa protein(s) in alpha-thrombin-treated cells. In this study, we demonstrate that in alpha-thrombin-treated CCL39 cells, the 74-78-kDa protein(s) rapidly undergoes tyrosine phosphorylation. The phosphorylation by alpha-thrombin was detected as early as 5 min and reached a maximum at 15 min; however, low levels were present at 2 h. alpha-Thrombin receptor agonist peptide (SFLLRN) induced its tyrosine phosphorylation, suggesting that alpha-thrombin mediates the effects via protease-activated receptor type 1. Anti-Stat3 antibodies specific to different regions of Stat3 failed to recognize the 74-78-kDa protein(s), suggesting that it is unrelated to Stat3. Cell fractionation experiments showed that it is localized to the cytoplasm. Mass spectrometric analysis of the immunoprecipitated protein showed that the 74-78-kDa protein(s) is related to glucose-regulated protein 75 (GRP-75), a member of the heat shock/stress-response protein family. Consistent with these data, we observed tyrosine phosphorylation of GRP-75 in alpha-thrombin-treated cells. Exposure of cells to pervanadate, a stress-inducing agent, stimulated its tyrosine phosphorylation; however, cytokines and growth factors were ineffective. This is the first report of tyrosine phosphorylation of GRP-75-related stress protein(s) by alpha-thrombin and suggests that this pathway may contribute to the ability of alpha-thrombin to prevent apoptosis in cells exposed to stress or in the injured tissue.

Animals↗

PubMed2003

Nitric oxide production by arsenite.

Arsenic can either enhance or reduce nitric oxide (NO) production, depending on the type of cell, the species and dose of arsenical tested. The mechanisms of how arsenic increases or decreases NO production remain unclear. Because NO is associated with many pathological conditions, it is conceivable that in those arsenic-target tissues, the NO production may be upregulated by continuous arsenic exposure, and a prolonged over-production of NO may cause inflammation hence a pathological condition. A prolonged interference with the normal physiological level of NO may also play a role in the initiation, promotion, and progression of arsenic-related human cancers. Suppression of NO production has been shown to reduce arsenite-induced oxidative DNA damage, inhibition of pyrimidine dimer excision, and micronuclei. However, a completely reliable story on how NO is involved in arsenic-related human disease is still lacking.

Animals↗

PubMed2003

Reactive oxygen species are involved in arsenic trioxide inhibition of pyruvate dehydrogenase activity.

Arsenite was shown to inhibit pyruvate dehydrogenase (PDH) activity through binding to vicinal dithiols in pure enzyme and tissue extract. However, no data are available on how arsenite inhibits PDH activity in human cells. The IC(50) values for arsenic trioxide (As(2)O(3)) to inhibit the PDH activity in porcine heart pure enzyme preparation and in human leukemia cell line HL60 cells were estimated to be 182 and 2 microM, respectively. Thus, As(2)O(3) inactivation of PDH activity was about 90 times more potent in HL60 cells than in purified enzyme preparation. The IC(50) values for As(2)O(3) and phenylarsine oxide to reduce the vicinal thiol content in HL60 cells were estimated to be 81.7 and 1.9 microM, respectively. Thus, As(2)O(3) is a potent PDH inhibitor but a weak vicinal thiol reacting agent in HL60 cells. Antioxidants but not dithiol compounds suppressed As(2)O(3) inhibition of PDH activity in HL60 cells. Conversely, dithiol compounds but not antioxidants suppressed the inhibition of PDH activity by phenylarsine oxide. As(2)O(3) increased H(2)O(2) level in HL60 cells, but this was not observed for phenylarsine oxide. Mitochondrial respiration inhibitors suppressed the As(2)O(3)-induced H(2)O(2) production and As(2)O(3) inhibition of PDH activity. Moreover, metal chelators ameliorated whereas Fenton metals aggravated As(2)O(3) inhibition of PDH activity. Treatment with H(2)O(2) plus Fenton metals also decreased the PDH activity in HL60 cells. Therefore, it seems that As(2)O(3) elevates H(2)O(2) production in mitochondria and this may produce hydroxyl through the Fenton reaction and result in oxidative damage to the protein of PDH. The present results suggest that arsenite may cause protein oxidation to inactivate an enzyme and this can occur at a much lower concentration than arsenite binding directly to the critical thiols.

Animals↗

PubMed2002

Endonuclease III, formamidopyrimidine-DNA glycosylase, and proteinase K additively enhance arsenic-induced DNA strand breaks in human cells.

We report here that sequential digestion with endonuclease III, formamidopyrimidine-DNA glycosylase, and proteinase K in Tris buffer markedly increased the sensitivity for detecting DNA damage in arsenic-treated cells. These three enzymes increased DNA strand breaks in an additive manner. By using this sequential-enzyme-digestion comet assay, we demonstrated that trivalent inorganic arsenic induced more DNA damage than monomethylarsonous acid, monomethylarsonic acid, and dimethylarsinic acid in human blood cell lines. However, trivalent inorganic arsenic was far less potent than monomethylarsonous acid in inhibiting pyruvate dehydrogenase activity. Therefore, different mechanisms are involved in inhibiting pyruvate dehydrogenase activity and inducing DNA damage. Our results also indicate while trivalent inorganic arsenic induced more endonuclease III-digestible adducts, monomethylarsonous acid and monomethylarsonic acid induced more proteinase K-digestible adducts. These results suggest there is a difference in the mechanism for inducing DNA damage between inorganic and organic methylated arsenic compounds.

Arsenic↗