Lavudi Kousalya, Harika Venkata Satya, Kokkanti Rekha Rani, Patchigolla Swaroopa, Sinha Anupriya, Patnaik Srinivas, Penchalaneni Josthna
KIIT School of Biotechnology, KIIT University, Bhubaneswar, Odisha, India.
Department of Biotechnology, Sri Padmavati Mahila Visva Vidyalayam, Tirupati, Andhra Pradesh, India.
Front Bioeng Biotechnol. 2022 Aug 16;10:978846. doi: 10.3389/fbioe.2022.978846. eCollection 2022.
Our research focused on generating AgNPs using (MU) seed extracts and studied their efficacy in combating tumor growth using the 2-Dimensional method for ovarian cancer cell line-PA-1. Characterization studies including a UV-visible spectrophotometer confirmed the surface plasmon resonance peak of 436 nm. Particle size determination data validated the nanoparticle diameter of 91.8 nm. Synthesized AgNPs possess a negative charge of -28.0 mV, which was confirmed through the zeta potential study. Structural characterization studies including XRD determined the crystal phase of AgNPs at four distant peaks at 2θ (38.17, 44.36, 64.52, and 77.46) and were assigned to 111, 200, 220, and 311 planes of the FCC. FTIR studies have confirmed the presence of O-H, N-H, C=O, ethers, C-Br, and C-I groups in AgNPs respectively. DPPH study has confirmed the presence of free radicles and we observed that at 500 μg/ml concentration, 76.08% of free radicles were formed which shows their efficiency. MTT assay shows the efficacy of MU-AgNPs in reducing the cell viability. At lower concentrations of MU-AgNP, 66% viability was observed and 9% of viability was observed at higher dose. ROS production (21%) was observed using MU-AgNPs with respect to 0.45% in controls, which affirms the capacity to induce DNA damage via apoptosis. Standard drug camptothecin generated 26% of ROS production which confirms higher potential of AgNPs in inducing DNA damage in tumor cells without causing lethality to the healthy cells. Further, the Fluorescence-activated cell sorting (FACS) study using a standard Caspase-3 marker confirms the generation of apoptotic bodies using two different concentrations of MU-AgNPs. At 40 μg, 64% of apoptotic cell death was observed, whereas, using 20 μg, 23% of apoptosis was recorded via fluorescent intensity. Propidium iodide-based Cell cycle study has shown a significant decrease in G0/G1 phase compared to control (88.8%), which further confirmed the apoptotic induction. Matrix metalloproteinases (MMP) studies using JC-1 dye, showed a significant increase in green fluorescence owing to lowered membrane potential, thus ensuring the breakdown of mitochondrial potential compared to untreated and standard drugs. With the obtained results, we are concluding that MU-AgNPs has a tremendous capacity to suppress the ovarian cancer cell proliferation by inducing DNA damage and apoptosis.
我们的研究聚焦于利用锦葵（MU）种子提取物生成银纳米颗粒（AgNPs），并采用二维方法对卵巢癌细胞系PA - 1研究其抑制肿瘤生长的功效。包括紫外可见分光光度计在内的表征研究证实了表面等离子体共振峰在436 nm。粒度测定数据验证了纳米颗粒直径为91.8 nm。通过zeta电位研究证实，合成的AgNPs带 - 28.0 mV的负电荷。包括X射线衍射（XRD）在内的结构表征研究确定了AgNPs在2θ处四个不同峰（38.17、44.36、64.52和77.46）的晶相，并归属于面心立方（FCC）的111、200、220和311晶面。傅里叶变换红外光谱（FTIR）研究分别证实了AgNPs中存在O - H、N - H、C = O、醚、C - Br和C - I基团。二苯基苦味酰基自由基（DPPH）研究证实了自由基的存在，我们观察到在500 μg/ml浓度下，形成了76.08%的自由基，这显示了它们的效率。MTT法显示了锦葵 - AgNPs在降低细胞活力方面的功效。在较低浓度的锦葵 - AgNP下观察到66%的细胞活力，在高剂量下观察到9%的细胞活力。相对于对照组的0.45%，使用锦葵 - AgNPs观察到活性氧（ROS）产生率为21%，这证实了其通过凋亡诱导DNA损伤的能力。标准药物喜树碱产生了26%的ROS产生率，这证实了AgNPs在诱导肿瘤细胞DNA损伤而不致使健康细胞死亡方面具有更高的潜力。此外，使用标准半胱天冬酶 - 3标记物的荧光激活细胞分选（FACS）研究证实了使用两种不同浓度的锦葵 - AgNPs产生凋亡小体。在40 μg时，观察到64%的凋亡细胞死亡，而使用20 μg时，通过荧光强度记录到23%的凋亡率。基于碘化丙啶的细胞周期研究表明，与对照组相比，G0/G1期显著减少（88.8%），这进一步证实了凋亡诱导。使用JC - 1染料的基质金属蛋白酶（MMP）研究表明，由于膜电位降低，绿色荧光显著增加，从而确保与未处理组和标准药物相比线粒体电位的破坏。根据获得的结果，我们得出结论，锦葵 - AgNPs具有通过诱导DNA损伤和凋亡来抑制卵巢癌细胞增殖的巨大能力。
