Insight into OroxylinA-7-<i>O</i>-β-<span style="font-variant: small-caps">d</span>-Glucuronide-Enriched <i>Oroxylum indicum</i> Bark Extract in Oral Cancer HSC-3 Cell Apoptotic Mechanism: Role of Mitochondrial Microenvironment
Sharmila Kameyanda Poonacha,
Madhyastha Harishkumar,
Madhyastha Radha,
Remya Varadarajan,
Suchetha Kumari Nalilu,
Shilpa Sharathraj Shetty,
Praveen Kumar Shetty,
Revanasiddappa Bistuvalli Chandrashekharappa,
Mahendra Gowdru Sreenivas,
Satheesh Kumar Bhandary Bavabeedu
Affiliations
Sharmila Kameyanda Poonacha
Central Research Laboratory, K.S. Hegde Medical Academy, Nitte (Deemed to Be) University, Mangaluru 575018, India
Madhyastha Harishkumar
Central Research Laboratory, K.S. Hegde Medical Academy, Nitte (Deemed to Be) University, Mangaluru 575018, India
Madhyastha Radha
Department of Cardio-Vascular Physiology, Faculty of Medicine, University of Miyazaki, Miyazaki 8891692, Japan
Remya Varadarajan
Central Research Laboratory, K.S. Hegde Medical Academy, Nitte (Deemed to Be) University, Mangaluru 575018, India
Suchetha Kumari Nalilu
Central Research Laboratory, K.S. Hegde Medical Academy, Nitte (Deemed to Be) University, Mangaluru 575018, India
Shilpa Sharathraj Shetty
Central Research Laboratory, K.S. Hegde Medical Academy, Nitte (Deemed to Be) University, Mangaluru 575018, India
Praveen Kumar Shetty
Central Research Laboratory, K.S. Hegde Medical Academy, Nitte (Deemed to Be) University, Mangaluru 575018, India
Revanasiddappa Bistuvalli Chandrashekharappa
Department of Pharmaceutical Chemistry, NGSM Institute of Pharmaceutical Sciences, Nitte (Deemed to Be) University, Mangaluru 575018, India
Mahendra Gowdru Sreenivas
Department of Pharmaceutical Chemistry, NGSM Institute of Pharmaceutical Sciences, Nitte (Deemed to Be) University, Mangaluru 575018, India
Satheesh Kumar Bhandary Bavabeedu
Central Research Laboratory, K.S. Hegde Medical Academy, Nitte (Deemed to Be) University, Mangaluru 575018, India
Oroxylum indicum, of the Bignoniaceae family, has various ethnomedical uses such as an astringent, anti-inflammatory, anti-bronchitis, anti-helminthic and anti-microbial, including anticancer properties. The druggability of OI stem bark extract was determined by its molecular docking interactions with PARP and Caspase-3, two proteins involved in cell survival and death. Note that 50 µg/mL of Oroxylum indicum extract (OIE) showed a significant (p p < 0.5%) reduction in cell number at 4 h of incubation time. Cell elongation and spindle formation was noticed when HSC-3 cells were treated with 50 µg/mL of OIE. OIE initiated DNA breakage and apoptosis in HSC-3 cells, as evident from DNA ladder assay and calcein/EB staining. Apoptosis potential of OIE is confirmed by flow cytometer and triple-staining (live cell/apoptosis/necrosis) assay. Caspase-3/7 fluorescence quenching (LANCE) assay demonstrated that 50 µg/mL of OIE significantly enhanced the RFU of caspases-3/7, indicating that the apoptosis potential of OIE is probably through the activation of caspases. Immuno-cytochemistry of HSC-3 cells treated with 50 µg/mL of OIE showed a significant reduction in mitochondrial bodies as well as a reduction in RFU in 60 min of incubation time. Immunoblotting studies clearly showed that treatment of HSC-3 cells with OI extract caused caspase-3 activation and PARP deactivation, resulting in apoptotic cell death. Overall, our data indicate that OIE is an effective apoptotic agent for human squamous carcinoma cells and it could be a future cancer chemotherapeutic target.
