Recombinant Klotho Protects Human Periodontal Ligament Stem Cells by Regulating Mitochondrial Function and the Antioxidant System during H₂O₂-Induced Oxidative Stress

Abstract

Human periodontal ligament stem cells (hPDLSCs) are a favourable source for tissue engineering, but oxidative stress conditions during cell culture and transplantation could affect stem cell viability and stemness, leading to failed regeneration. The aim of this study was to evaluate the antioxidant and protective effects of Klotho, an antiageing protein, against cell damage and the loss of osteogenesis in hPDLSCs in H2O2-induced oxidative environments. H2O2 was used as an exogenous reactive oxygen species (ROS) to induce oxidative stress. Recombinant human Klotho protein was administered before H2O2 treatment. Multitechniques were used to assess antioxidant activity, cell damage, and osteogenic ability of hPDLSCs in oxidative stress and the effects of Klotho on hPDLSCs. Mitochondrial function was analyzed by an electron microscopy scan of cellular structure, mitochondrial DNA copy number, and cellular oxygen consumption rate (OCR). Furthermore, we explored the pathway by which Klotho may function to regulate the antioxidant system. We found that pretreatment with recombinant human Klotho protein could enhance SOD activity and reduce intracellular oxidative stress levels. Klotho reduced H2O2-induced cellular damage and eventually maintained the osteogenic differentiation potential of hPDLSCs. Notably, Klotho promoted mitochondrial function and activated antioxidants by negatively regulating the PI3K/AKT/FoxO1 pathway. The findings suggest that Klotho protein enhanced the antioxidative ability of hPDLSCs and protected stem cell viability and stemness from H2O2-induced oxidative stress by restoring mitochondrial functions and the antioxidant system.

Improvement of ECM-based bioroot regeneration via N-acetylcysteine-induced antioxidative effects

“…With regard to osteogenic differentiation, most studies suggest that ROS inhibit osteogenic differentiation [ 27 ]. In this study, we found that NAC increased the osteogenic differentiation of hDFCs under oxidative stress, promoting mineralized nodule deposition and Col I, ALP, RunX2 expression, which is consistent with a previous study [ 11 ]. The mRNA expression pattern of osteoblast-related genes is characteristic of the process of osteogenic differentiation.…”

“…Tissue necrosis caused by both oxidative stress and the inflammatory response further prevents nutrients and oxygen from reaching viable cells, which contributes to expanding the necrotic area and leads to implant failure [ 7 ]. Therefore, the unfavourable microenvironment after isolation and transplantation reduces the stemness and inhibits the therapeutic effects of surviving seed cells in intragrafts [ 11 ]. Thus, specific control of the balance within this cellular microenvironment may represent an efficient and novel strategy to increase the success rate of transplantation.…”

Klotho inhibits H2O2‐induced oxidative stress and apoptosis in periodontal ligament stem cells by regulating UCP2 expression

“…Periodontitis aggravates inflammation and insulin secretion in diabetic mice through interleukin‐12 (IL‐12) regulation by klotho. Klotho can serve as an IL‐12 inhibitor to decrease immune stimulation and insulin secretion in diabetic mice with periodontitis 18,19 . Recombinant human klotho was confirmed to resist oxidative stress and mitochondrial dysfunction by negatively regulating the PI3K/AKT/FoxO1 pathway in PDLSCs 20 .…”

“…The ability of UCP2 to regulate glucose is strongly associated with cellular proliferation and apoptosis 17 . Additionally, the UCP2 upregulation might significantly suppress the proliferation and migration of vascular smooth muscle cells by inhibiting oxidative stress 18,19 . Studies have shown that periodontal lesions in chronic periodontitis depend on periodontal ligament stem cells (PDLSCs) apoptosis through mitochondrial pathways 20 .…”

Quercetin Prevents Oxidative Stress-Induced Injury of Periodontal Ligament Cells and Alveolar Bone Loss in Periodontitis

“… 9 Studies have found that PDLCs show decreased proliferation, increased apoptosis and autophagy, impaired osteogenesis, and even cellular senescence under OS. 10−12 Antioxidant therapies can effectively relieve the oxidative damage of PDLCs and restore their osteogenic and regenerative abilities. 13 , 14 Therefore, it is important to maintain the balance between oxidation and antioxidation in periodontal tissues, especially in PDLCs.…”