Dataset Information

Cyclin D1, Caspase 9 and P53 Expressions in T47D Cell Lines after Treatment of Plectranthus amboinicus, (Lour.) Spreng. Leaves Ethanolic Extract Nanoparticles.

ABSTRACT: BACKGROUND:The application of nanotechnology is aimed to enhance the capability of the chemical compounds of Plectranthus amboinicus (Lour.) Spreng. leaves activity. The contact area of the particle surface becomes larger in nano size, which can increase the amount of the isolated active substance. The nanoparticles of this extract exhibited the most effective impact in decreasing the cell number of T47D by induction of apoptosis and arresting the cell cycle. AIM:To evaluate the expressions of cyclin D1, caspase-9 and p53 in T47D cell lines treated by Plectranthus amboinicus, (Lour.) Spreng. Leaves Ethanolic Extract Nanoparticles (PAEEN). METHODS:The inhibition of cyclin D1 was done by flow cytometry assay. The expression of cyclin D1, caspase-9 and p53 were observed by immunocytochemistry assay. RESULTS:Plectranthus amboinicus, (Lour.) Spreng. Leaves Ethanolic Extract Nanoparticles (PAEEN) on concentration IC50 (89.2 µg/mL) inhibited the expression of cyclin D1 around 5.59%. Further, the immunocytochemistry assay indicated that there was an inhibition of cyclin D1, upregulation of caspase-9 and restoration of p53, indicated by their expression. CONCLUSIONS:The Plectranthus amboinicus, (Lour.) Spreng. Leaves Ethanolic Extract Nanoparticles could be an effective anticancer by improving its downregulation on cyclinD1, upregulation caspase-9 and p53.

SUBMITTER: Hasibuan PAZ

PROVIDER: S-EPMC7048340 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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Cyclin D1, Caspase 9 and P53 Expressions in T47D Cell Lines after Treatment of <i>Plectranthus amboinicus</i>, (Lour.) Spreng. Leaves Ethanolic Extract Nanoparticles.

Hasibuan Poppy Anjelisa Zaitun PAZ Sumaiyah Sumaiyah S

Open access Macedonian journal of medical sciences 20191114 22

<h4>Background</h4>The application of nanotechnology is aimed to enhance the capability of the chemical compounds of <i>Plectranthus amboinicus</i> (Lour.) Spreng. leaves activity. The contact area of the particle surface becomes larger in nano size, which can increase the amount of the isolated active substance. The nanoparticles of this extract exhibited the most effective impact in decreasing the cell number of T47D by induction of apoptosis and arresting the cell cycle.<h4>Aim</h4>To evaluat ...[more]

PMID: 32127976

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Project description:Plectranthus amboinicus (Lour.) Spreng is an aromatic medicinal herb known for its therapeutic and nutritional properties attributed by the presence of monoterpene and sesquiterpene compounds. Up until now, research on terpenoid biosynthesis has focused on a few mint species with economic importance such as thyme and oregano, yet the terpene synthases responsible for monoterpene production in P. amboinicus have not been described. Here we report the isolation, heterologous expression and functional characterization of a terpene synthase involved in P. amboinicus terpenoid biosynthesis. A putative monoterpene synthase gene (PamTps1) from P. amboinicus was isolated with an open reading frame of 1797 bp encoding a predicted protein of 598 amino acids with molecular weight of 69.6 kDa. PamTps1 shares 60-70% amino acid sequence similarity with other known terpene synthases of Lamiaceae. The in vitro enzymatic activity of PamTps1 demonstrated the conversion of geranyl pyrophosphate and farnesyl pyrophosphate exclusively into linalool and nerolidol, respectively, and thus PamTps1 was classified as a linalool/nerolidol synthase. In vivo activity of PamTps1 in a recombinant Escherichia coli strain revealed production of linalool and nerolidol which correlated with its in vitro activity. This outcome validated the multi-substrate usage of this enzyme in producing linalool and nerolidol both in in vivo and in vitro systems. The transcript level of PamTps1 was prominent in the leaf during daytime as compared to the stem. Gas chromatography-mass spectrometry (GC-MS) and quantitative real-time PCR analyses showed that maximal linalool level was released during the daytime and lower at night following a diurnal circadian pattern which correlated with the PamTps1 expression pattern. The PamTps1 cloned herein provides a molecular basis for the terpenoid biosynthesis in this local herb that could be exploited for valuable production using metabolic engineering in both microbial and plant systems.

Project description:Linalool and nerolidol are terpene alcohols that occur naturally in many aromatic plants and are commonly used in food and cosmetic industries as flavors and fragrances. In plants, linalool and nerolidol are biosynthesized as a result of respective linalool synthase and nerolidol synthase, or a single linalool/nerolidol synthase. In our previous work, we have isolated a linalool/nerolidol synthase (designated as PamTps1) from a local herbal plant, Plectranthus amboinicus, and successfully demonstrated the production of linalool and nerolidol in an Escherichia coli system. In this work, the biochemical properties of PamTps1 were analyzed, and its 3D homology model with the docking positions of its substrates, geranyl pyrophosphate (C10) and farnesyl pyrophosphate (C15) in the active site were constructed. PamTps1 exhibited the highest enzymatic activity at an optimal pH and temperature of 6.5 and 30 °C, respectively, and in the presence of 20 mM magnesium as a cofactor. The Michaelis-Menten constant (Km) and catalytic efficiency (kcat/Km) values of 16.72 ± 1.32 µM and 9.57 × 10-3 µM-1 s-1, respectively, showed that PamTps1 had a higher binding affinity and specificity for GPP instead of FPP as expected for a monoterpene synthase. The PamTps1 exhibits feature of a class I terpene synthase fold that made up of α-helices architecture with N-terminal domain and catalytic C-terminal domain. Nine aromatic residues (W268, Y272, Y299, F371, Y378, Y379, F447, Y517 and Y523) outlined the hydrophobic walls of the active site cavity, whilst residues from the RRx8W motif, RxR motif, H-α1 and J-K loops formed the active site lid that shielded the highly reactive carbocationic intermediates from the solvents. The dual substrates use by PamTps1 was hypothesized to be possible due to the architecture and residues lining the catalytic site that can accommodate larger substrate (FPP) as demonstrated by the protein modelling and docking analysis. This model serves as a first glimpse into the structural insights of the PamTps1 catalytic active site as a multi-substrate linalool/nerolidol synthase.

Project description:Induction chemotherapy has been previously demonstrated to downgrade locally advanced or aggressive cancers and increase the likelihood of primary lesion eradication. Based on our previous phase 3 trial on TPF (docetaxel, cisplatin and fluorouracil) induction chemotherapy in patients with oral squamous cell carcinoma (OSCC), in which short-term prognostic and predictive values of cyclin D1 expression were reported, the present study aimed to determine the long-term predictive value of cyclin D1 expression in the same patients with OSCC who were eligible to receive TPF induction chemotherapy. In addition, the present study investigated the potential association between cyclin D1 expression and chemosensitivity to TPF agents during OSCC cell intervention, and the underlying apoptotic mechanism of action. In total, 232 patients with locally advanced OSCC from our previous trial with a median follow-up of 5 years were included for survival analysis using the Kaplan-Meier method and the log-rank test in the present study, where cyclin D1 expression in their tissues was detected by immunohistochemistry. Cyclin D1 knockdown, cytotoxicity assays assessing the efficacy of the TPF chemotherapeutic agents and measurements of caspase-3 and PARP activity in HB96, CAL27 and HN30 cell lines were performed. Patients with OSCC in the low cyclin D1 expression group exhibited significantly superior long-term clinical outcomes compared with those in patients in the high cyclin D1 expression group [overall survival (OS), P=0.001; disease-free survival, P=0.003; local recurrence-free survival, P=0.004; distant metastasis-free survival (DMFS), P=0.001]. Furthermore, patients with stage clinical nodal stage 2 (cN2) OSCC in the high cyclin D1 expression group benefitted from TPF induction chemotherapy (OS, P=0.024; DMFS, P=0.024), whilst patients with cN2 OSCC in the low cyclin D1 expression group did not benefit from this chemotherapy. Overexpression of cyclin D1 expression was found to enhance chemosensitivity to TPF chemotherapeutic agents in OSCC by mediating caspase-3-dependent apoptosis. Based on these findings, TPF induction chemotherapy can benefit patients with cN2 OSCC and high cyclin D1 expression in terms of long-term survival from compared with standard treatment. In addition, OSCC cell lines overexpressing cyclin D1 are more sensitive to TPF chemotherapeutic agents in a caspase-3-dependent manner (clinical trial. no. NCT01542931; February 2012).

Dataset Information

Cyclin D1, Caspase 9 and P53 Expressions in T47D Cell Lines after Treatment of Plectranthus amboinicus, (Lour.) Spreng. Leaves Ethanolic Extract Nanoparticles.

Publications

Cyclin D1, Caspase 9 and P53 Expressions in T47D Cell Lines after Treatment of <i>Plectranthus amboinicus</i>, (Lour.) Spreng. Leaves Ethanolic Extract Nanoparticles.

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