The Potentized Homeopathic Drug, Lycopodium clavatum (5C and 15C) Has Anti-cancer Effect on HeLa Cells In Vitro.

Asmita Samadder,Anisur Rahman Khuda-Bukhsh,Sreemanti Das,Jayeeta Das,Avijit Paul,Naoual Boujedaini 사단법인약침학회 2013 Journal of Acupuncture & Meridian Studies Vol.6 No.4

Cancer is a disease that needs a multi-faceted approach from different systems of medicine. The purpose of this study was to evaluate whether homeopathically-potentized ultra-high dilutions of Lycopodium Clavatum (LC-5C and LC-15C, respectively) have any anti-cancer effects on HeLa cells. Cells were exposed to either LC-5C (diluted below Avogadro's limit, i.e., 10−10) or LC-15C (diluted beyond Avogadro's limit, i.e., 10−30) (drug-treated) or to 30% succussed ethanol (“vehicle” of the drug). The drug-induced modulation in the percent cell viability, the onset of apoptosis, and changes in the expressions of Bax, Bcl2, caspase 3, and Apaf proteins in inter-nucleosomal DNA, in mitochondrial membrane potentials and in the release of cytochrome-c were analyzed by utilizing different experimental protocols. Results revealed that administration of LC-5C and LC-15C had little or no cytotoxic effect in normal peripheral blood mononuclear cells, but caused considerable cell death through apoptosis in cancer (HeLa) cells, which was evident from the induction of DNA fragmentation, the increases in the expressions of protein and mRNA of caspase 3 and Bax, and the decreases in the expressions of Bcl2 and Apaf and in the release of cytochrome-c. Thus, the highly-diluted, dynamized homeopathic remedies LC-5C and LC-15C demonstrated their capabilities to induce apoptosis in cancer cells, signifying their possible use as supportive medicines in cancer therapy.

Ameliorative Effects of Syzygium jambolanum Extract and its Poly (lactic-co-glycolic) Acid Nano-encapsulated Form on Arsenic-induced Hyperglycemic Stress: A Multi-parametric Evaluation

Asmita Samadder,Anisur Rahman Khuda-Bukhsh,Sreemanti Das,Jayeeta Das,Avijit Paul 사단법인약침학회 2012 Journal of Acupuncture & Meridian Studies Vol.5 No.6

In South East Asia, groundwater arsenic contamination has become a great menace. Chronic arsenic intoxication leads to a hyperglycemic condition in animals and man. Because of undesirable side-effects and affordability, orthodox medicine, like insulin, is not preferred by many who like natural products instead. Unfortunately, such natural products mostly lack scientific validation. Therefore, we became interested in assessing the efficacy of the ethanolic seed extract of Syzygium jambolanum (SJ), traditionally used against diabetic conditions. We also formulated poly (lactic-co-glycolic) acid (PLGA)-encapsulated nano-SJ (NSJ) and tested whether the ameliorative potentials of SJ could be enhanced by nano-encapsulation. In this study, we conducted both in vitro (in L6 cells) and in vivo (in mice) experiments to assess the relative efficacy of SJ and NSJ. We characterized the physico-chemical features of NSJ by atomic force microscopy and critically analyzed several bio-markers and signal proteins associated with arsenicinduced stress and hyperglycemia. We also determined the relative ameliorative potentials of SJ and NSJ by using standard protocols. NSJ could cross the blood brain barrier in mice. Overall results suggested that NSJ had a greater potential than that of SJ, indicating the possibility of using NSJ in the future drug design and management of arsenicinduced hyperglycemia and stress.

Enhanced Drug Carriage Efficiency of Curcumin-Loaded PLGA Nanoparticles in Combating Diabetic Nephropathy via Mitigation of Renal Apoptosis

Asmita Samadder,Banani Bhattacharjee,Sudatta Dey,Arnob Chakrovorty,Rishita Dey,Priyanka Sow,Debojyoti Tarafdar,Maharaj Biswas,Sisir Nandi,Asmita Samadder 대한약침학회 2024 Journal of pharmacopuncture Vol.27 No.1

Background: Diabetic nephropathy (DN) is one of the major complications of chronic hyperglycaemia affecting normal kidney functioning. The ayurvedic medicine curcumin (CUR) is pharmaceutically accepted for its vast biological effects. Objectives: The Curcuma-derived diferuloylmethane compound CUR, loaded on Poly (lactide-co-glycolic) acid (PLGA) nanoparticles was utilized to combat DN-induced renal apoptosis by selectively targeting and modulating Bcl2. Methods: Upon in silico molecular docking and screening study CUR was selected as the core phytocompound for nanoparticle formulation. PLGA-nano-encapsulated-curcumin (NCUR) were synthesized following standard solvent displacement method. The NCUR were characterized for shape, size and other physico-chemical properties by Atomic Force Microscopy (AFM), Dynamic Light Scattering (DLS) and Fourier-Transform Infrared (FTIR) Spectroscopy studies. For in vivo validation of nephro-protective effects, Mus musculus were pre-treated with CUR at a dose of 50 mg/kg b.w. and NCUR at a dose of 25 mg/kg b.w. (dose 1), 12.5 mg/kg b.w (dose 2) followed by alloxan administration (100 mg/kg b.w) and serum glucose levels, histopathology and immunofluorescence study were conducted. Results: The in silico study revealed a strong affinity of CUR towards Bcl2 (dock score –10.94 Kcal/mol). The synthesized NCUR were of even shape, devoid of cracks and holes with mean size of ~80 nm having –7.53 mV zeta potential. Dose 1 efficiently improved serum glucose levels, tissue-specific expression of Bcl2 and reduced glomerular space and glomerular sclerosis in comparison to hyperglycaemic group. Conclusion: This study essentially validates the potential of NCUR to inhibit DN by reducing blood glucose level and mitigating glomerular apoptosis by selectively promoting Bcl2 protein expression in kidney tissue.

Nanopharmaceutical Approach for Enhanced Anti-cancer Activity of Betulinic Acid in Lung-cancer Treatment via Activation of PARP: Interaction with DNA as a Target -Anti-cancer Potential of Nano-betulinic Acid in Lung Cancer-

Jayeeta Das,Asmita Samadder,Sreemanti Das,Avijit Paul,Anisur Rahman Khuda-Bukhsh 대한약침학회 2016 Journal of pharmacopuncture Vol.19 No.1

Objectives: This study examined the relative efficacies of a derivative of betulinic acid (dBA) and its poly (lactide- co-glycolide) (PLGA) nano-encapsulated form in A549 lung cancer cells in vivo and in co-mutagen [sodium arsenite (SA) + benzo[a]pyrene (BaP)]-induced lung cancer in mice in vivo. Methods: dBA was loaded with PLGA nanoparticles by using the standard solvent displacement method. The sizes and morphologies of nano-dBA (NdBA) were determined by using transmission electron microscopy (TEM), and their intracellular localization was verified by using confocal microscopy. The binding and interaction of NdBA with calf thymus deoxyribonucleic acid (CT-DNA) as a target were analyzed by using conventional circular dichroism (CD) and melting temperature (Tm) profile data. Apoptotic signalling cascades in vitro and in vivo were studied by using an enzyme-linked immunosorbent assay (ELISA); the ability of NdBA to cross the blood-brain barrier (BBB) was also examined. The stage of cell cycle arrest was confirmed by using a fluorescence- activated cell-sorting (FACS) data analysis. Results: The average size of the nanoparticles was ~ 110 nm. Confocal microscopy images confirmed the presence of NdBA in the cellular cytoplasm. The bio-physical properties of dBA and NdBA ascertained from the CD and the Tm profiles revealed that NdBA had greater interaction with the target DNA than dBA did. Both dBA and NdBA arrested cell proliferation at G0/G1, NdBA showing the greater effect. NdBA also induced a greater degree of cytotoxicity in A549 cells, but it had an insignificant cytotoxic effect in normal L6 cells. The results of flow cytometric, cytogenetial and histopathological studies in mice revealed that NdBA caused less nuclear condensation and DNA damage than dBA did. TEM images showed the presence of NdBA in brain samples of NdBA fed mice, indicating its ability to cross the BBB. Conclusion: Thus, compared to dBA, NdBA appears to have greater chemoprotective potential against lung cancer.

Nanopharmaceutical Approach for Enhanced Anti-cancer Activity of Betulinic Acid in Lung-cancer Treatment via Activation of PARP: Interaction with DNA as a Target -Anti-cancer Potential of Nano-betulinic Acid in Lung Cancer-

Das, Jayeeta,Samadder, Asmita,Das, Sreemanti,Paul, Avijit,Khuda-Bukhsh, Anisur Rahman KOREAN PHARMACOPUNCTURE INSTITUTE 2016 Journal of pharmacopuncture Vol.19 No.1

Objectives: This study examined the relative efficacies of a derivative of betulinic acid (dBA) and its poly (lactide-co-glycolide) (PLGA) nano-encapsulated form in A549 lung cancer cells in vivo and in co-mutagen [sodium arsenite (SA) + benzo[a]pyrene (BaP)]-induced lung cancer in mice in vivo. Methods: dBA was loaded with PLGA nanoparticles by using the standard solvent displacement method. The sizes and morphologies of nano-dBA (NdBA) were determined by using transmission electron microscopy (TEM), and their intracellular localization was verified by using confocal microscopy. The binding and interaction of NdBA with calf thymus deoxyribonucleic acid (CT-DNA) as a target were analyzed by using conventional circular dichroism (CD) and melting temperature (Tm) profile data. Apoptotic signalling cascades in vitro and in vivo were studied by using an enzyme-linked immunosorbent assay (ELISA); the ability of NdBA to cross the blood-brain barrier (BBB) was also examined. The stage of cell cycle arrest was confirmed by using a fluorescence-activated cell-sorting (FACS) data analysis. Results: The average size of the nanoparticles was ~ 110 nm. Confocal microscopy images confirmed the presence of NdBA in the cellular cytoplasm. The bio-physical properties of dBA and NdBA ascertained from the CD and the Tm profiles revealed that NdBA had greater interaction with the target DNA than dBA did. Both dBA and NdBA arrested cell proliferation at G0/G1, NdBA showing the greater effect. NdBA also induced a greater degree of cytotoxicity in A549 cells, but it had an insignificant cytotoxic effect in normal L6 cells. The results of flow cytometric, cytogenetial and histopathological studies in mice revealed that NdBA caused less nuclear condensation and DNA damage than dBA did. TEM images showed the presence of NdBA in brain samples of NdBA fed mice, indicating its ability to cross the BBB. Conclusion: Thus, compared to dBA, NdBA appears to have greater chemoprotective potential against lung cancer.

Strong Anticancer Potential of Nano-triterpenoid from Phytolacca decandra against A549 Adenocarcinoma via a Ca 2+ -dependent Mitochondrial Apoptotic Pathway

Jayeeta Das,Sreemanti Das,Avijit Paul,Asmita Samadder,Anisur Rahman Khuda-Bukhsh 사단법인약침학회 2014 Journal of Acupuncture & Meridian Studies Vol.7 No.3

We isolated a triterpenoid from an ethanolic extract of Phytolacca decandra and na- noencapsulated it with biodegradable nontoxic polymers of poly(lactide-co-glycolide) to examine if the nanoform of this hitherto unexplored betulinic-acid derivative (NdBA) could produce a stronger anticancer effect by rendering better drug bioavailability and targeted delivery than the nonencapsulated betulinic-acid derivative (dBA). The nano- particles were characterized with the help of physicochemical and morphological studies involving dynamic light scattering and atomic force microscopy. A549 cancer cells were exposed to NdBA and dBA at the IC 50 doses of 50 mg/mL and 100 mg/mL, respectively. Mitochondrial dysfunction-mediated apoptosis was determined by exam- ining the changes in the intracellular calcium content, the reactive oxygen species accumulation, the cytochrome c release, the upregulation of Bcl-2-associated-X protein (Bax) and caspase 3, the downregulation of B cell lymphoma 2, and the mitochondrial membrane potential (DJ m ) depolarization. Apoptosis was also verified by acridine or- ange staining observed under fluorescence microscopy and annexin V-fluorescein iso- thiocyanate/propidium iodide staining through flow cytometric studies. The levels of intracellular adenosine triphosphate/adenosine diphosphate ratio decreased, and the ATPase activity increased more strikingly in A549 cells exposed to NdBA than in A549 cells exposed to dBA. Overall results showed that both drugs directly target the mito- chondrial oxidative phosphorylation system, with NdBA having a stronger effect, indi- cating NdBA to be a better candidate for the development of an anticancer drug for use against lung adenocarcinomas.

Anticancer Potential of Myricanone, a Major Bioactive Component of Myrica cerifera: Novel Signaling Cascade for Accomplishing Apoptosis

Avijit Paul,Anisur Rahman Khuda-Bukhsh,Jayeeta Das,Asmita Samadder 사단법인약침학회 2013 Journal of Acupuncture & Meridian Studies Vol.6 No.4

Extract of Myrica cerifera bark has long been fruitfully used as a hepato-protective and anticancerdrug in various complementary and alternative systems of medicine. Myricanone, itsprincipal bioactive compound, had also been reported to have apoptosis-promoting ability. We evaluated its anti-cancer potential in vitro in HepG2 liver cancer cells and tried to understandthe signal cascades involved in accomplishing apoptosis. Further, we ascertainedby using a (3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay (MTT)assay if it had cytotoxic effects on normal noncancerous liver cells (WRL-68). We deployedvarious tools and protocols, like phase contrast, scanning electron and fluorescence microscopies,performed an annexinV-FITC/PI assay and cell cycle analysis, and estimated thereactive oxygen species (ROS) generation and mitochondrial membrane depolarizationthrough flow cytometry. Further, analyses of cytochrome-c translocation and of HSP70and caspase expressions were also done by using immunoblota and Enzyme linked immunosorbentassay (ELISA). Results revealed that myricanone induced apoptosis in HepG2 cellsthrough generation of ROS, depolarization of the mitochondrial membrane, early releaseof cytochrome-c, down-regulation of HSP70 and activation of a caspase cascade; it hadno, or insignificant, cytotoxic effects in WRL-68 cells in vitro and in mice in vivo. Thus, myricanonehas great potential for use in formulating an effective drug against both hepatotoxicityand hepatocellular cancer.

Apigenin, a Bioactive Flavonoid from Lycopodium clavatum, Stimulates Nucleotide Excision Repair Genes to Protect Skin Keratinocytes from Ultraviolet B-Induced Reactive Oxygen Species and DNA Damage

Sreemanti Das,Anisur Rahman Khuda-Bukhsh,Jayeeta Das,Avijit Paul,Asmita Samadder 사단법인약침학회 2013 Journal of Acupuncture & Meridian Studies Vol.6 No.5

In this study, we examined the antioxidative and the DNA protective potentials of apigenin, a flavonoid polyphenol isolated from Lycopodium clavatum, in both in-vitro (HaCaT skin keratinocytes) and in-vivo (mice) models against UV-B radiation. We used DAPI staining in UV-B-irradiated HaCaT skin keratinocytes pre-treated with and without apigenin to assess DNA damage. We also used a flow-cytometric analysis in mice exposed to UV-B radiation with or without topical application of apigenin to assess, through a comet assay, chromosomal aberrations and quanta from reactive oxygen species (ROS) generation. Data from the stability curves for the Gibb’s free energy determined from a melting-temperature profile study indicated that apigenin increased the stability of calf thymus DNA. Immunofluorescence studies revealed that apigenin caused a reduction in the number of cyclobutane pyrimidine dimers (CPDs) after 24 h, the time at which the nucleotide excision repair (NER) genes were activated. Thus, apigenin accelerated reversal of UV-Binduced CPDs through up-regulation of NER genes, removal of cyclobutane rings, inhibition of ROS generation, and down-regulation of NF-kB and MAPK, thereby revealing the precise mechanism of DNA repair.