Research Institute of Experimental and Clinical Medicine, Kharkiv National Medical University, 4 Nauky ave, 61022, Kharkiv, Ukraine.
Department of Cryobiochemistry, Institute for Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine, Kharkiv, 61015, Ukraine.
Cell Biochem Biophys. 2024 Jun;82(2):747-766. doi: 10.1007/s12013-024-01225-9. Epub 2024 Feb 9.
Aim In the current study, hemocompatibility of three major commercially available types of carrageenans (ι, κ and λ) was investigated focusing on eryptosis.
Carrageenans of ι-, κ- and λ-types were incubated with washed erythrocytes (hematocrit 0.4%) at 0-1-5-10 g/L for either 24 h or 48 h. Incubation was followed by flow cytometry-based quantitative analysis of eryptosis parameters, including cell volume, cell membrane scrambling and reactive oxygen species (ROS) production, lipid peroxidation markers and confocal microscopy-based evaluation of intracellular Ca levels, assessment of lipid order in cell membranes and the glutathione antioxidant system. Confocal microscopy was used to assess carrageenan cellular internalization using rhodamine B isothiocyanate-conjugated carrageenans.
All three types of carrageenans were found to trigger eryptosis. Pro-eryptotic properties were type-dependent and λ-carrageenan had the strongest impact inducing phosphatidylserine membrane asymmetry, changes in cell volume, Ca signaling and oxidative stress characterized by ROS overproduction, activation of lipid peroxidation and severe glutathione system depletion. Eryptosis induction by carrageenans does not require their uptake by erythrocytes. Changes in physicochemical properties of cell membrane were also type-dependent. No carrageenan-induced generation of superoxide and hydroxyl radicals was observed in cell-free milieu.
Our findings suggest that ι-, κ- and λ-types trigger eryptosis in a type-dependent manner and indicate that carrageenans can be further investigated as potential eryptosis-regulating therapeutic agents.
目的 在本研究中，主要研究了三种市售角叉菜胶（ι、κ 和 λ）的血液相容性，重点关注红细胞的溶血。
材料和方法 将 ι、κ 和 λ 型角叉菜胶在 0-1-5-10 g/L 下与洗涤后的红细胞（血球比容 0.4%）孵育 24 小时或 48 小时。孵育后，通过流式细胞术定量分析溶血参数，包括细胞体积、细胞膜混乱和活性氧（ROS）的产生、脂质过氧化标志物以及基于共聚焦显微镜的细胞内 Ca 水平评估、细胞膜中脂质有序性和谷胱甘肽抗氧化系统的评估。使用 rhodamine B 异硫氰酸酯结合角叉菜胶通过共聚焦显微镜评估角叉菜胶的细胞内内化。
结果 三种角叉菜胶均被发现可触发溶血。促溶血作用具有类型依赖性，λ-角叉菜胶的影响最强，诱导磷脂酰丝氨酸膜不对称、细胞体积变化、Ca 信号和 ROS 过度产生、脂质过氧化激活以及谷胱甘肽系统严重耗竭引起的氧化应激。角叉菜胶诱导的溶血不需要红细胞摄取。细胞膜理化性质的变化也具有类型依赖性。在无细胞环境中未观察到角叉菜胶诱导的超氧自由基和羟基自由基的产生。
结论 我们的研究结果表明，ι、κ 和 λ 型角叉菜胶以依赖于类型的方式触发溶血，并表明角叉菜胶可以进一步作为潜在的调节溶血的治疗剂进行研究。
