Venkata Subbiah Harini, Ramesh Babu Polani, Subbiah Usha
Human Genetics Research Centre, Sree Balaji Dental College & Hospital, Bharath Institute of Higher Education and Research, Chennai, IND.
Centre for Materials Engineering and Regenerative Medicine, Bharath Institute of Higher Education and Research, Chennai, IND.
Cureus. 2023 Dec 21;15(12):e50880. doi: 10.7759/cureus.50880. eCollection 2023 Dec.
Introduction lipopolysaccharide (Pg-LPS) is one of the crucial virulence factors of periodontitis. Antimicrobial peptides (AMPs) are emerging as alternatives or adjuncts to antibiotics in the treatment of microbial infections. In this study, cytotoxicity, anti-inflammatory activity, anti-oxidative stress, cell cycle analysis, and apoptosis properties of AMP, β-defensin 1, were studied in Pg-LPS-stimulated THP-1 (Tohoku Hospital Pediatrics - 1) cell line. Methods The cytotoxic nature of Pg-LPS and β-defensin 1 was studied by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) method. The cytotoxic effect of β-defensin 1 on Pg-LPS-stimulated THP-1 cells was also studied by the same method. The anti-inflammatory role of β-defensin 1 against cyclooxygenase (COX), lipoxygenase (LOX), myeloperoxidase (MPO), and inducible nitric oxide synthase activities were studied. The anti-oxidative nature of β-defensin 1 was analyzed by measuring reactive oxygen species (ROS) generation by dichlorodihydrofluorescein diacetate (DCFDA) assay. Cell cycle distribution and apoptosis were studied by flow cytometry. The hemolytic nature of β-defensin 1 was predicted using the HemoPred web tool. Results The results of the study demonstrated that Pg-LPS showed dose-dependent cytotoxicity to THP-1 cells. β-Defensin 1 had dose-dependent cytotoxicity to THP-1 cells and showed a protective effect on THP-cells up to 1 µg/mL of Pg-LPS, beyond which cell viability decreased. β-Defensin 1 inhibited COX, LOX, MPO, and inducible nitric oxide synthase activities in a concentration-dependent manner. β-Defensin 1 showed anti-oxidative activity by suppressing the generation of ROS measured through fluorescence intensity. From the cell cycle analysis, it was found that β-defensin 1 was able to reduce the Pg-LPS-induced cell cycle arrest at the G0/G1 phase. From the apoptosis profile, β-defensin 1 was found to increase the live cells when compared to THP-1 cells stimulated only with Pg-LPS, indicating that β-defensin 1 provided a protective role to THP-1 cells. β-Defensin 1 was found to be hemolytic in nature by the HemoPred web tool. Conclusion β-Defensin 1 exerted multifunctional activities and can be considered a promising agent for controlling periodontitis.
引言 脂多糖（Pg-LPS）是牙周炎的关键毒力因子之一。抗菌肽（AMPs）正在成为治疗微生物感染的抗生素替代品或辅助药物。在本研究中，我们在Pg-LPS刺激的THP-1（东北医院儿科 - 1）细胞系中研究了抗菌肽β-防御素1的细胞毒性、抗炎活性、抗氧化应激、细胞周期分析和凋亡特性。方法 通过MTT（3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐）法研究Pg-LPS和β-防御素1的细胞毒性性质。还用相同方法研究了β-防御素1对Pg-LPS刺激的THP-1细胞的细胞毒性作用。研究了β-防御素1对环氧合酶（COX）、脂氧合酶（LOX）、髓过氧化物酶（MPO）和诱导型一氧化氮合酶活性的抗炎作用。通过二氯二氢荧光素二乙酸酯（DCFDA）测定法测量活性氧（ROS）生成来分析β-防御素1的抗氧化性质。通过流式细胞术研究细胞周期分布和凋亡。使用HemoPred网络工具预测β-防御素1的溶血性质。结果 研究结果表明Pg-LPS对THP-1细胞具有剂量依赖性细胞毒性。β-防御素1对THP-1细胞具有剂量依赖性细胞毒性，并且在Pg-LPS浓度高达1 µg/mL时对THP细胞显示出保护作用，超过该浓度细胞活力下降。β-防御素1以浓度依赖性方式抑制COX、LOX、MPO和诱导型一氧化氮合酶活性。β-防御素1通过抑制通过荧光强度测量的ROS生成而显示出抗氧化活性。从细胞周期分析发现，β-防御素1能够减少Pg-LPS诱导的细胞周期在G0/G1期的停滞。从凋亡图谱发现，与仅用Pg-LPS刺激的THP-1细胞相比，β-防御素1能增加活细胞数量，表明β-防御素1对THP-1细胞起到了保护作用。通过HemoPred网络工具发现β-防御素1具有溶血性质。结论 β-防御素1发挥了多种功能活性，可被认为是控制牙周炎的有前景的药物。
