A Systematic Review of In Vivo Studies of the Efficacy of Herbal Medicines for Anti-Aging in the Last Five Years

Background: The world’s population is rapidly aging, and attention to and research on the increase in life expectancy and age-related diseases are needed. This study aimed to review the in vivo studies on the anti-aging effects of herbal medicines. Methods: In vivo studies of single or complex herbal medicines for anti-aging that were published in the last five years were included in this review. The following databases were used: PubMed, Scopus, ScienceDirect, Web of Science and EMBASE. Results: A total of 41 studies were considered eligible for the review. The articles were classified into body organs and functions, experimental country, herbal medicine, extraction method, administration route, dosage, duration, animal model, aging-induced method, sex, number of animals per group, and outcomes and mechanisms A single herbal extract was used in a total of 21 studies including Alpinia oxyphylla Miq., Acanthopanax senticosus and Lyceum barbarum, and a multi-compound herbal prescription was used in a total of 20 studies, including Modified Qiongyu paste, Wuzi Yanzong recipe, etc. Each herbal medicine had anti-aging effects on learning and memory, cognition, emotion, internal organs, gastrointestinal tracts, sexual functions, musculoskeletal function and so on. The common mechanisms of action were antioxidant and anti-inflammatory, and various effects and mechanisms for each organ and function were identified. Conclusions: Herbal medicine exhibited beneficial effects on anti-aging in various parts of the body and its function. Further investigation of the appropriate herbal medicine prescriptions and their components is recommended.


Introduction
According to the World Health Organization (WHO), by 2050, 22% of the world's population (approximately 2 billion people) will be over the age of 60, and the problem of deteriorating health in old age is expected to increase significantly, owing to the increase in life expectancy and age-related diseases. Increased life expectancy and the desire for "healthy aging" to maintain good health in old age for as long as possible have led to the recognition of aging as a treatable disease, and interest in the field of research called "Anti-aging Medicine" is growing [1].
The WHO recently recognized aging as a "major disease risk factor" rather than a "natural phenomenon" and changed the "Senility (R54)" code of the 10th edition of the International Classification of Diseases (ICD-10) to the "Old age (MG2A)" code. As the ICD code is a prerequisite for the registration of all new drugs and treatments, it can be considered as laying the groundwork for the development of therapeutic interventions and prevention strategies targeting aging and age-related diseases.
Herbal medicine is attracting attention as it can play a role as an anti-aging treatment, and various experimental studies have been conducted on herbal medicines, such as A total of 960 studies were screened in the initial electronic search, and 185 studies were excluded owing to duplicate publications. After screening the titles and abstracts, 628 studies were excluded for the following reasons: not anti-aging (n = 426), not herbal medicine (n = 140), in vitro (n = 61) and clinical trials (n = 1). After reviewing 147 full-text articles, 106 were excluded for the following reasons: studies performed with only some ingredients or compounds isolated from herbs (n = 48), specific disease-induced model studies (n = 22), review articles (n = 17), conference abstracts (n = 2), other analysis methods (e.g., network pharmacology, n = 10) and articles not written in English (n = 7). After this process, 41 studies were considered eligible for the review. A flow diagram of the article selection process is shown in Figure 1.

Characteristics of Included Studies
In total, 41 rodent studies using herbal medicines showed the efficacy or mechanisms of anti-aging. The classification, experimental country, herbal medicine, extraction method, administration route, dosage, duration, animal model, aging-induced method, sex, number of animals per group, and outcomes and mechanisms are summarized in Table 1.

Characteristics of Included Studies
In total, 41 rodent studies using herbal medicines showed the efficacy or mechanisms of anti-aging. The classification, experimental country, herbal medicine, extraction method, administration route, dosage, duration, animal model, aging-induced method, sex, number of animals per group, and outcomes and mechanisms are summarized in Table 1.  The 41 studies were classified into the following categories: brain, learning and memory, cognition, emotion, cardiac function, liver and brain, liver, liver and kidney function, gastrointestinal tract, sexual function, musculoskeletal function, skin, skin and sexual organs, lifespan extension, physiology, gut microbiota and metabolome, and metabolomic analysis.
Several of the studies identified antioxidant enzymes, such as superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX), as well as cytokines, such as interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α). In addition, the evaluation indicators and mechanisms were specific to each organ or function. The anti-aging effect on the brain (learning and memory, and cognition) was confirmed, mainly through memory function tests and observation of hippocampal neurogenesis. In studies on internal organs, the functions of each organ, blood tests and cytokines were observed. Studies related to the gastrointestinal tract were characterized by observing changes in the gastric histomorphology or intestinal flora. Most studies have confirmed changes in various hormones related to sexual function, such as GnRH, LH, testosterone and FSH. Studies examining the anti-aging effect on musculoskeletal function confirmed that these preparations had a positive effect, including improvement of gastrocnemius muscle and motor function.

Discussion
We reviewed in vivo (rodent) studies on the use of herbal medicines for anti-aging that were published in the last five years. A total of 41 articles were analyzed according to the described research selection criteria. As a result, it was confirmed that many single herbal medicines and complex herbal medicines have anti-aging effects on various organs and functions.
Four studies were conducted on the brain (cognition and emotion were separately classified), with two of those studies focusing on single herbal medicines and two focusing on complex herbal medicines. In addition to antioxidant effects, memory and motor functions were improved, and the effect on hippocampal neurogenesis was also confirmed. Gastrodiae Rhizoma is a single herbal medicine widely used for central nervous system (CNS) diseases, such as headache, dizziness, epilepsy and stroke [51]. Lycium barbarum and Modified Qiongyu paste have been used as promising anti-aging agents for a long time; in particular, a number of studies have been conducted on Lycium barbarum [52]. Therefore, these herbal medicines may be used for brain-related anti-aging.
Nine research papers have been published on learning and memory, cognition and emotion [12][13][14][15][16][17][18][19][20]. Seven of these focused on single herbal medicines [12][13][14][16][17][18][19], and two focused on complex herbal medicines [15,20]. Scutellaria baicalensis Georgi has traditionally been used to reduce fever and has been found to have anti-inflammatory, antiviral, antioxidant and antibacterial effects [53]. As this herb has been proven to be effective in memory impairment models in several studies [54][55][56], it can be widely used for anti-aging purposes through cognitive improvement and memory impairment prevention in the future. Dushen Tang is a prescription for the decoction of Panax ginseng. Panax ginseng is widely used as an effective herbal medicine, and many studies related to brain senescence have already been conducted [57]. Therefore, Panax ginseng alone can be used as an anti-aging remedy; it can also be added to a prescription in consideration of certain health conditions. The Morris water maze test is the most frequently used method to evaluate learning and memory, and some studies have confirmed the mechanism of reducing hippocampal neuronal damage and increasing synaptic density. Even changes at the mRNA level have been observed. These results provide the basis for the use of herbal medicines for cognitive improvement and dementia prevention.
The elderly population shows depressive symptoms that are different from those of general depression and may be accompanied by decreased concentration and memory, insomnia and personality changes. Kososan (Xiang-Su-San in Chinese; Hyangsosan in Korean), which was used in the study by Ito et al. [20], is a prescription that can be used for patients who experience no motivation, feeling down or poor digestive function. Depressive behavior was improved in the SAM8 mouse model, and this improvement was confirmed by observing the hippocampus and tau, which can be expected to help improve depression and cognitive function in the elderly.
There were a total of eight papers related to internal organs, such as the heart, liver and kidney, as well as the gastrointestinal tract [21][22][23][24][25][26][27][28]. There were seven studies on the use of a single herbal medicine [21][22][23][24][25][26][27] and one on complex herbal medicine [28]. Both studies were conducted in Taiwan [21,22], and it was confirmed that the administration of Alpinia oxyphylla Miq. reduced aging-related heart damage and improved heart function. Alpinia oxyphylla Miq. has been used for the purpose of enhancing cardiac function, and it has been shown to improve dementia through several experiments [58]. Recently, however, results of extending lifespan in a Caenorhabditis elegans (C. elegans) model have been reported [59]. Based on these results, it is thought that Alpinia oxyphylla Miq. may be useful for anti-aging in cardiac functions.
Chrysanthemum indicum Linne has been used for various diseases, such as high blood pressure and headache, and has been found to have hepatoprotective effects [60]. In a study by Zhang et al. [23], Chrysanthemum indicum Linne restored body weight and lowered ALT and AST levels in aging-induced mice. In addition, it alleviated the abnormal alterations in the structure and function of the brain and liver. Therefore, Chrysanthemum indicum Linne may be used to prevent aging in cases related to the liver. Angelica sinensis has been used for tonifying, invigorating blood, replenishing and treating female menstrual disorders [61]. Angelica sinensis is a hepatoprotective herbal medicine, and its mechanisms have been confirmed [62]. It is often used in combination with Astragalus membranaceus, and the combined effect of these two herbal medicines on nephrotic syndrome has been studied [63]. In a study by Mo et al. [26], Angelica sinensis improved various outcomes related to the liver and kidneys of aging-induced rats. Therefore, Angelica sinensis is effective for anti-aging, and it can be considered for the deterioration of liver and kidney function in the elderly.
There were two papers related to the gastrointestinal tract, one on single herbal medicine [27] and one on complex herbal medicine [28]. Codonopsis pilosula is a herbal medicine that strengthens the spleen and lungs [64]. Meng et al. [27] confirmed that Codonopsis pilosula improves gastrointestinal function and gastric histomorphological changes in aging mice. This is an experimentally proven clinical effect based on the medical literature, and it is possible to consider Codonopsis pilosula in elderly patients with reduced gastrointestinal function.
Regarding sexual function, there were three studies conducted on the use of single herbal medicines [31,33,34], and the remaining seven studies were conducted on the use of complex herbal medicines [30,32,[35][36][37][38]. Testicular function, hypogonadism and ovarian function were shown to be restored by the administration of herbal medicine through sex hormones, morphological observation and gene level analysis using male and female aging models, respectively. The effect and mechanism of action of the Wuzi Yanzong recipe (Ojayeonjonghwan in Korean) on testicular dysfunction and hypogonadism were confirmed [35]. This prescription has been used to treat male infertility and has been reported to be effective against oligoasthenozoospermia [65]. Aging decreases sperm count and viability; moreover, changes in hormone secretion, such as testosterone and FSH, reduce sexual function. Samul-tang (Siwu-tang in Chinese; Shimotsu-to in Japanese) is a basic prescription that is mainly used for women with blood deficiency conditions and menstrual and uterine diseases. According to a study by Kim et al. [37], various mechanisms by which Samul-tang improved age-related decline in ovarian function have been identified. Yu Linzhu is a complex herbal medicine that is widely prescribed for infertility and habitual abortion, and its effects and mechanisms of restoring ovarian function have been confirmed [38]. It has also been confirmed that various herbal medicines inhibit sexual dysfunction. This is an important mechanism and effect for anti-aging.
There was a total of three articles that were classified by musculoskeletal function, one single herbal medicine study [39] and two complex prescription studies [40,41]. Juzentaihoto (Shi-quan-da-bu-tang in Chinese; Sipjeondaebo-tang in Korean) administration improved gastrocnemius muscle and motor function in SAMP8 mice [40]. Juzentaihoto is a complex herbal prescription that is mainly used for fatigue and loss of appetite. Recently, many studies on cancer have been conducted [66,67]. Therefore, if body function is weakened, it is possible to recover this function through the use of Juzentaihoto and to pursue its anti-aging effects.
In some articles, there were omissions, such as herbal medicine extraction methods [8,11,25,32,[36][37][38]41,44,46], and in most studies, there was no description of the herbal medicine dosage calculation or aging-inducing drug dosage setting. In a model in which aging was induced by the administration of d-galactose, the administration concentration and duration were not certain; therefore, a guide may be necessary in future studies.

Conclusions
In this review, a variety of single and complex herbal medicines exhibited beneficial effects on anti-aging in various parts of the body and function. The common main mechanisms of action of herbal medicines are antioxidant and anti-inflammatory, and various mechanisms have been identified according to each organ and function. Learning and memory were shown to be improved through memory function tests and hippocampal neurogenesis, identifying the anti-aging mechanisms of the brain. Aging-associated cardiac damage, elevated liver enzyme levels and intestinal inflammation were decreased. The levels of GnRH, LH, testosterone, FSH, etc. improved. In addition, the mechanisms of action on each organ and its function were confirmed. Thus, the possibility that herbal medicine can be used in various ways for the purposes of anti-aging has been confirmed. In clinical practice, these herbal medicines can be used for treating the aging process.
However, since the experiments have been conducted using inconsistent models and methods, it is difficult to compare the results, which is a limitation of our analysis. Future studies to confirm and compare the anti-aging effects and mechanisms of various herbal medicines in the same model are recommended. Further investigation of the appropriate herbal medicine prescriptions and their components is also recommended. Accordingly, it is hoped that humans will live a long and healthy life by developing new natural products and conducting clinical research.

Data Availability Statement:
No new data were created or analyzed in this study. Data sharing is not applicable to this article.

Conflicts of Interest:
The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.