Ajay K. Bauri

Attenuation of oxidative stress by Allylpyrocatechol in synovial cellular infiltrate of patients with Rheumatoid Arthritis

Abstract Free radicals are involved in the pathogenesis of Rheumatoid arthritis, a systemic autoimmune disorder characterized by unchecked synovial inflammation. Allylpyrocatechol, a phytoconstituent of Piper betle leaves, has potent anti-inflammatory activity and this study evaluated its anti-oxidant effect on the synovial infiltrate of patients with Rheumatoid arthritis. The ex vivo effect of allylpyrocatechol upon generation of reactive oxygen species in neutrophils, macrophages and lymphocytes was measured by flow cytometry using dichlorodihydrofluorescein diacetate, wherein it significantly decreased basal levels as also scavenged phorbol myristate acetate generated reactive oxygen species. Furthermore, its effect on generation of superoxide and hydroxyl radicals produced within infiltrated neutrophils was measured by cytochrome c and deoxyribose assay, respectively. Allylpyrocatechol significantly scavenged superoxide and hydroxyl radicals in infiltrated neutrophils. The effect of allylpyrocatechol on nitric oxide was measured in macrophages using 4,5-diaminofluorescein diacetate by flow cytometry wherein it decreased production of nitric oxide in infiltrated macrophages, which correlated with its in vitro nitric oxide scavenging activity. Taken together, this ex vivo study has established that allylpyrocatechol has potent scavenging activity and could be considered as an add-on therapy in the treatment of inflammation-associated disorders like Rheumatoid Arthritis.

Malabaricone-A Induces A Redox Imbalance That Mediates Apoptosis in U937 Cell Line

Background The ‘two-faced’ character of reactive oxygen species (ROS) plays an important role in cancer biology by acting both as secondary messengers in intracellular signaling cascades and sustaining the oncogenic phenotype of cancer cells, while on the other hand, it triggers an oxidative assault that causes a redox imbalance translating into an apoptotic cell death. Principal Findings Using a tetrazolium [{3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl}-2H-tetrazolium] based cell viability assay, we evaluated the cytotoxicity of a plant derived diarylnonanoid, malabaricone-A on leukemic cell lines U937 and MOLT-3. This cytotoxicity hinged on its ability to cause a redox imbalance via its ability to increase ROS, measured by flow cytometry using 5-(and-6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate and by decreasing glutathione peroxidase activity. This redox imbalance mediated apoptosis was evident by an increase in cytosolic [Ca2+], externalization of phosphatidyl serine as also depolarization of the mitochondrial membrane potential as measured by flow cytometry. There was concomitant peroxidation of cardiolipin, release of free cytochrome c to cytosol along with activation of caspases 9, 8 and 3. This led to cleavage of the DNA repair enzyme, poly (ADP-ribose) polymerase that caused DNA damage as proved by labeling with 4′,6-diamidino-2-phenylindole (DAPI); furthermore, terminal deoxy ribonucleotide transferase catalysed incorporation of deoxy uridine triphosphate confirmed DNA nicking and was accompanied by arrest of cell cycle progression. Conclusions Taken together, compounds like MAL-A having pro-oxidant activity mediate their cytotoxicity in leukemic cells via induction of oxidative stress triggering a caspase dependent apoptosis.

DNA damage dependent activation of checkpoint kinase-1 and mitogen-activated protein kinase-p38 are required in malabaricone C-induced mitochondrial cell death.

BACKGROUND Given that lung cancer is the second leading cause of cancer-related deaths with low survival rates, the project was aimed to formulate an efficient drug with minimum side effects, and rationalize its action mechanistically. METHODS Mitochondria deficient cells, shRNA-mediated BCL2 and ATM depleted cells and pharmacological inhibition of DNA-damage response proteins were employed to explore the signaling mechanism governed between nucleus and mitochondria in response to mal C. RESULTS Mal C decreased cell viability in three lung carcinoma cells, associated with DNA damage, p38-MAPK activation, imbalance in BAX/BCL2 expression, mitochondrial dysfunction and cytochrome-c release. Mitochondria depletion and p38-MAPK inhibition made A549 cells extremely resistant, but BCL2 knock-down partially sensitized the cells to mal C treatment. The mal C-induced apoptosis in A549 cells was initiated by DNA single strand breaks that led to double strand breaks (DSBs). DSB generation paralleled the induction of ATM- and ATR-mediated CHK1 phosphorylation. ATM silencing and ATR inhibition partially attenuated the mal C-induced p38-MAPK activation, CHK1 phosphorylation and apoptosis, which were completely suppressed by CHK1 inhibition. CONCLUSIONS Mal C activates the ATM-CHK1-p38 MAPK cascade to cause mitochondrial cell death in lung carcinoma cells. GENERAL SIGNIFICANCE Given that mal C has appreciable natural abundance and is non-toxic to mice, further in vivo evaluation would help in establishing its anti-cancer property.

Physicochemical insights in non-covalent interaction of a newly designed triporphyrin with fullerenes C60 and C70 in solution

The present paper reports the synthesis of free-base (H(2)-1) and zinc-triporphyrins Zn-1, and their supramolecular complexes with C(60) and C(70) in toluene. While UV-Vis studies reveal ground state interaction between fullerenes and triporphyrins, steady state fluorescence measurements establish quenching of fluorescence of triporphyrins by fullerenes. Binding constants data evoke that Zn-1 may be employed as an efficient tweezers for C(70) in toluene. Time resolved emission studies establish relatively long-lived charge separated state for the C(70)/triporphyrin complexes. Molecular mechanics force field calculations in vacuo interpret the stability difference between C(60)/and C(70)/complexes of triporphyrin.

Determination of binding strength for the supramolecular complexation of a designed bisporphyrin with C60, C70 and their derivatives employing absorption spectrophotometric, fluorescence and quantum chemical calculations

The present paper reports the synthesis of a designed bisporphyrin (1), and its supramolecular complexes with C60, C70 and their derivatives, namely, tert-butyl-(1,2-methanofullerene)-61-carboxylate (2) and [6,6]-phenyl C70 butyric acid methyl ester (3) in toluene medium. C60, C70 and their derivatives undergo ground state non-covalent interaction with 1 is evidenced from absorption spectrophotometric study in which it is observed that the intensity of the Soret absorption band of 1 decreases considerably in presence of C60, C70 and their derivatives. Steady state fluorescence studies reveal efficient quenching of fluorescence of 1 in presence of fullerenes. The binding constant (K) values of the fullerene/1 complexes follows the trend: 2/1<C(60)/1<3/1<C(70)/1. However, selectivity in K values of the bisporphyrin complexes is found to be higher for fullerene derivatives in comparison to C60 and C70. Time resolved emission studies establish relatively long-lived charge separated state for the C(70)/1 complex. Molecular mechanics calculations at force field model in vacuo evoke the single projection geometric structures of various fullerene/1 complexes and interpret their stability differences in terms of heat of formation values.