Rejuvenation of aging skin fibroblast via restoration of disrupted proteostasis by Nelumbo Nucifera Germ Extract.
70
Presented by: Takushi Namba
Introduction
We have been seeking for ways to keep young since the ancient times. In particular, maintaining the beauty of the skin is a symbol of youth. One of the characteristics of skin aging is the formation of wrinkles. Wrinkles are caused by a disruption in the collagen structure in the dermis, and the one factor is a decrease the fibroblasts function such as collagen production due to the aging.
Aging is a complex multifunctional biological process that results in time-dependent attenuation of biological function and simultaneously induces the loss of various cellular functions. However, though the molecular mechanisms of its progression have been elucidated, a comprehensive understanding has yet to be reported.
Cells lose their functions when their homeostasis is disrupted due to aging process, leading to various time-dependent effects, such as protein aggregation, the production of aging-dependent advanced glycation end products (AGEs), the loss of organelle function, and the induction of DNA damage signals. Finally, aging cells stop proliferating and produce senescence-associated secretory phenotype (SASP) related factors, which impair the function of surrounding cells. However, the sequence of aging-related changes and how they affect each other remain unclear.
Mitochondria are one of the most important organelles for maintaining cellular homeostasis, and mitochondrial function is decreased over time. Some reports have showed that damaged mitochondria exhibit decreased function due to their own production of abnormal proteins, which are generated by the loss of cellular proteostasis, stimulating the cellular aging process. Thus, many studies to determine how mitochondria can be reactivated during the aging process to suppress aging are still being conducted.
Methods
NB1RGB cells were maintained in MEMα supplemented with 10% FBS, 100 U/ml penicillin, and 100 µg/ml streptomycin and maintained at 37°C with 5% CO2. We defined three types of each cell line according to the number of days in culture: NB1RGB: young cells: from 8 to 20 days, middle passage: from 30 to 40 days, and aging cells: from 60 to 70 days. Mitochondrial membrane potential was assessed using JC-1staining. We determined mRNA and protein expression level by using Real time PCR and immunobloting assay. We performed transmission electron microscopy to identify any change in the cellular or organelle structure such as autolysosomes and lipofuscin-like particles. AGE levels were measured using an OxiSelect AGE Competitive ELISA kit. 3-Dimensional collagen gel culture was performed by using type I atelocollagen gel medium. Immunohistochemical analys was peformed that atelocollagen gels were fixed in paraformaldehyde. The sections were blocked and incubated with anti-collagen I primary antibody and then incubated with FITC-conjugated rabbit IgG. Nuclei were stained with DAPI.
Results
This study reports that disruption of proteostasis attenuated mitochondrial function upon proliferative and replicative senescence before the induction of DNA damage signaling, and the clearance of abnormal substances via autophagy restored mitochondrial function in aging skin fibroblasts, which is an early stage of cellular aging. We screened 75 plant extracts that restored mitochondrial function in aging skin fibroblast and discovered that Nelumbo Nucifera Germ Extract reduced the activity of senescence-associated β-galactosidase, a marker of aging, and activated mitochondria via induction of autophagy, which degraded lipofuscin aggregates and AGEs. Moreover, the treatment of aging skin fibroblast with Nelumbo Nucifera Germ Extract stimulated collagen production and contractile ability in three-dimensional cell culture.
Discussion
The induction of autophagic signaling is an antiaging target to restore mitochondrial function by the restoration of disturbed proteostasis. Additionally, Nelumbo Nucifera Germ Extract was found to rejuvenate aging skin fibroblasts and is thus a promising new antiaging material.
We have been seeking for ways to keep young since the ancient times. In particular, maintaining the beauty of the skin is a symbol of youth. One of the characteristics of skin aging is the formation of wrinkles. Wrinkles are caused by a disruption in the collagen structure in the dermis, and the one factor is a decrease the fibroblasts function such as collagen production due to the aging.
Aging is a complex multifunctional biological process that results in time-dependent attenuation of biological function and simultaneously induces the loss of various cellular functions. However, though the molecular mechanisms of its progression have been elucidated, a comprehensive understanding has yet to be reported.
Cells lose their functions when their homeostasis is disrupted due to aging process, leading to various time-dependent effects, such as protein aggregation, the production of aging-dependent advanced glycation end products (AGEs), the loss of organelle function, and the induction of DNA damage signals. Finally, aging cells stop proliferating and produce senescence-associated secretory phenotype (SASP) related factors, which impair the function of surrounding cells. However, the sequence of aging-related changes and how they affect each other remain unclear.
Mitochondria are one of the most important organelles for maintaining cellular homeostasis, and mitochondrial function is decreased over time. Some reports have showed that damaged mitochondria exhibit decreased function due to their own production of abnormal proteins, which are generated by the loss of cellular proteostasis, stimulating the cellular aging process. Thus, many studies to determine how mitochondria can be reactivated during the aging process to suppress aging are still being conducted.
Methods
NB1RGB cells were maintained in MEMα supplemented with 10% FBS, 100 U/ml penicillin, and 100 µg/ml streptomycin and maintained at 37°C with 5% CO2. We defined three types of each cell line according to the number of days in culture: NB1RGB: young cells: from 8 to 20 days, middle passage: from 30 to 40 days, and aging cells: from 60 to 70 days. Mitochondrial membrane potential was assessed using JC-1staining. We determined mRNA and protein expression level by using Real time PCR and immunobloting assay. We performed transmission electron microscopy to identify any change in the cellular or organelle structure such as autolysosomes and lipofuscin-like particles. AGE levels were measured using an OxiSelect AGE Competitive ELISA kit. 3-Dimensional collagen gel culture was performed by using type I atelocollagen gel medium. Immunohistochemical analys was peformed that atelocollagen gels were fixed in paraformaldehyde. The sections were blocked and incubated with anti-collagen I primary antibody and then incubated with FITC-conjugated rabbit IgG. Nuclei were stained with DAPI.
Results
This study reports that disruption of proteostasis attenuated mitochondrial function upon proliferative and replicative senescence before the induction of DNA damage signaling, and the clearance of abnormal substances via autophagy restored mitochondrial function in aging skin fibroblasts, which is an early stage of cellular aging. We screened 75 plant extracts that restored mitochondrial function in aging skin fibroblast and discovered that Nelumbo Nucifera Germ Extract reduced the activity of senescence-associated β-galactosidase, a marker of aging, and activated mitochondria via induction of autophagy, which degraded lipofuscin aggregates and AGEs. Moreover, the treatment of aging skin fibroblast with Nelumbo Nucifera Germ Extract stimulated collagen production and contractile ability in three-dimensional cell culture.
Discussion
The induction of autophagic signaling is an antiaging target to restore mitochondrial function by the restoration of disturbed proteostasis. Additionally, Nelumbo Nucifera Germ Extract was found to rejuvenate aging skin fibroblasts and is thus a promising new antiaging material.