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ORIGINAL ARTICLE
Year : 2016  |  Volume : 12  |  Issue : 46  |  Page : 228-230  

Life span and motility effects of ethanolic extracts from Sophora moorcroftiana seeds on Caenorhabditis elegans


1 Department of Immunology, School of Basic Medical Sciences, Lanzhou 730000, China
2 Department of Immunology, School of Basic Medical Sciences; Key Laboratory of Evidence Based Medicine and Knowledge Translation of Gansu Province, Lanzhou University, Lanzhou 730000, China
3 Department of Microbiology, Medical College, Northwest University for Nationalities, Lanzhou 730000, China

Date of Submission21-May-2015
Date of Decision26-Dec-2015
Date of Web Publication11-May-2016

Correspondence Address:
Kaizhong Dong
Department of Microbiology, Medical College, Northwest University for Nationalities, Lanzhou 730000
China
Xingming Ma
Department of Immunology, School of Basic Medical Sciences, Lanzhou University, Lanzhou 730000
China
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0973-1296.182152

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   Abstract 


Background: Sophora moorcroftiana is an endemic shrub species with a great value in folk medicine in Tibet, China. In this study, relatively little is known about whether S. moorcroftiana is beneficial in animals' nervous system and life span or not. Materials and Methods: To address this question, under survival normal temperature (25°C), S. moorcroftiana seeds were extracted with 95% ethanol, and Caenorhabditis elegans were exposed to three different extract concentrations (100 mg/L, 200 mg/L, and 400 mg/mL) from S. moorcroftiana seeds. Results: The 95% ethanolic extracts from S. moorcroftiana seeds could increase life span and slow aging-related increase in C. elegans and could not obviously influence the motility of C. elegans. Conclusion: Given these results by our experiment for life span and motility with 95% ethanolic extracts from S. moorcroftiana seeds in C. elegans, the question whether S. moorcroftiana acts as an anti-aging substance in vivo arises.
SUMMARY


  • The 95% ethanolic extracts from S. moorcroftiana seeds have no effect on the life span in C. elegans when extract concentrations from S. moorcroftiana seeds <400 mg/L
  • The 400 mg/L 95% ethanolic extracts from S. moorcroftiana seeds could increase life span in C. elegans
  • The 95% ethanolic extracts from S. moorcroftiana seeds could not obviously influence the motility in C. elegans.


Abbreviation used: S. moorcroftiana: Sophora moorcroftiana; C. elegan: Caenorhabditis elegan; E. coli OP50: Escherichia coli OP50; DMSO: Dimethyl sulfoxide.

Keywords: Anti-aging, Caenorhabditis elegans, motility, Sophora moorcroftiana


How to cite this article:
Li X, Han J, Zhu R, Cui R, Ma X, Dong K. Life span and motility effects of ethanolic extracts from Sophora moorcroftiana seeds on Caenorhabditis elegans. Phcog Mag 2016;12, Suppl S2:228-30

How to cite this URL:
Li X, Han J, Zhu R, Cui R, Ma X, Dong K. Life span and motility effects of ethanolic extracts from Sophora moorcroftiana seeds on Caenorhabditis elegans. Phcog Mag [serial online] 2016 [cited 2019 Sep 22];12, Suppl S2:228-30. Available from: http://www.phcog.com/text.asp?2016/12/46/228/182152




   Introduction Top


Sophora moorcroftiana is an endemic shrub species in Tibet, China, and is mainly distributed in the wide valleys in the middle reaches of several main tributaries of the Yalu Tsangpo River. Sophora moorcroftiana seeds have been used for a long time in Chinese folk medicine. The decoction of the seed has been used in Chinese folk medicine for dephlogistication (translated, in general, as anti-inflammation in modern medicine), detoxication, emetic processes, and treating infectious diseases and verminosis.[1] The alkaloids obtained from S. moorcroftiana seeds have a protoscolicidal, anti-inflammatory effect [2] and induce apoptosis of the human stomach cancer SGC-7901 cell line in vitro.[3] As a magical medicinal plant, which has not been fully studied, S. moorcroftiana may have more unknown available value in vivo. One of these organisms which can be used for life span and nervous system studies is Caenorhabditis elegans, which is readily available, easy to culture in the laboratory, has a short life span, and vast knowledge is known about this nematode.[4],[5],[6] Moreover, other studies have shown that some compounds can prolong C. elegans life span and influence motility under laboratory conditions.[7] In light of these studies, we aimed to determine the effects of different concentrations of 95% ethanolic extracts from S. moorcroftiana on life span and motility in C. elegans.

In this study, it can be reported that 95% ethanolic extracts from S. moorcroftiana seeds could increase life span and slow aging-related increase in C. elegans. In addition, 95% ethanolic extracts from S. moorcroftiana seeds have no effect on the motility in C. elegans. Together, our study suggests the existence of an anti-aging compound within ethanolic extracts from S. moorcroftiana seeds.


   Materials and Methods Top


Caenorhabditis elegans and its synchronization

The C. elegans wild-type (N2) strain and its food source Escherichia coli OP50 strain were kindly provided by Professor Li Hongyu, who obtained these from Caenorhabditis Genetic Center at the University of Minnesota (USA). A volume of 200 ìL of OP50 (overnight culture) was dropped at the center of 60 mm nematode growth medium (NGM) plates, which were allowed to dry overnight before worms were transferred. C. elegans cultivation media supplies were purchased from Solarbio (China).[8]

L1 larvae were prepared by eggs synchronization, hermaphrodites were lysed in 20% bleach, 0.5 M NaOH, until fragmented, and eggs were incubated in M9 buffer without food overnight (14–18 h) at 20°C with agitation (150 rpm), to allow larvae to hatch and arrest the development because of starvation. About 600 worms (in 1.5 ìL total volume of M9 buffer) were placed in several drops on each assay plate (60 mm across) with enriched NGM, seeded with E. coli OP50.[9]

Sophora moorcroftiana extracts preparation

The 95% ethanolic extracts were extracted from S. moorcroftiana seeds by our laboratory as described previously.[10] The 95% ethanolic extracts were tested for the presence of carbohydrates, proteins, amino acids, alkaloids, flavonoids, glycosides, saponins, quinones, and fats and oils, using standard procedures.[11],[12] The dry crude extracts of S. moorcroftiana were dissolved into the required concentrations in dimethyl sulfoxide (DMSO); the final concentration of DMSO was <2%.[13] The extracts were diluted with distillate water when it was used for experiment.

Life span assay

The life span assays were performed according to the standard protocol recently described by Swain et al.[14] The animals were kept at 25°C during growth into adult stage and then treated for 2 h with the agents tested, which was composed of distillate water and extracts (dissolved in DMSO) in proportion to the required concentration. Afterward, they were fed in NGM agar plates spotted with OP50 for life span assays. Life span assays were performed at 25°C by checking the animals at each day at a fixed time until all the animals died. The escaping animals from the Petri dishes were excluded from the study. The study was planned as four groups. One group was used as control. The others were 100 mg/L, 200 mg/L, and 400 mg/L concentrations of 95% ethanolic extracts from S. moorcroftiana, respectively. Then, worms were incubated at 25°C. Animals were counted and recorded on the day. Each experiment was done in replicates of three. Data are presented as mean ± standard deviation (SD). All statistical analyses were carried out using SPSS software (IBM SPSS Statistics, Armonk, NY, USA). Nonparametric tests were carried out, and P values were calculated using the Wilcoxon test; P < 0.05 was accepted as statistically significant.

Motility assay

The motility assays were performed according to the standard protocol recently described by Tsalik and Hobert.[15] The feeding ways and treated mode by agents tested of C. elegans are the same as the life span assay in this experiment. However, they were fed in NGM agar plates without OP50 for assays after treatment in agents tested. The body bends per 20 s and the head swings per minute were counted with zoom stereo microscope. Each experiment was done in replicates of three. Data are presented as mean ± SD. The data were pooled and analyzed using Student's t-test; P < 0.05 was accepted as statistically significant.


   Results and Discussion Top


The 95% ethanolic extracts from Sophora moorcroftiana seeds can increase life span and slow aging-related increase on Caenorhabditis elegans

Although the alkaloids obtained from S. moorcroftiana have a protoscolicidal, anti-inflammatory effect and induce apoptosis of the human stomach cancer SGC-7901 cell line in vitro,[2],[3] no previous literature has shown the effectiveness of crude extracts of S. moorcroftiana seeds on anti-aging and the nervous system. Our data demonstrated that the high concentration (400 mg/L) of 95% ethanol extract from S. moorcroftiana seeds were significantly active in the dilation of life span [Table 1] and could effectively slow down aging-related increase in C. elegans [Figure 1]. There is no concrete evidence for whether the effectiveness of 95% ethanolic extracts from S. moorcroftiana on life span depends on the concentration.
Table 1: The life span of Caenorhabditis elegans at 25°C with various interventions


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Figure 1: The survival curve of Caenorhabditis elegans at 25°C with various interventions. The supplementation with the high concentration (400 mg/L) of 95% ethanol extract from Sophora moorcroftiana seeds results in more prolonged life span extension in Caenorhabditis elegans. Each life span experiment was repeated at least three independent times with similar results

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Sophora moorcroftiana extracts have no obvious influence in motility of Caenorhabditis elegans

Motility is one of the key physiological indexes to assess the excitability of animals' nervous system. The body bends per 20 s and the head swings per minute are specific testing items of motility of C. elegans, and generally speaking their results are similar. Our data (both the body bends and the head swings) demonstrated that the effects of all experiment groups were not significantly different from those of control group [Figure 2]. In other words, 95% ethanol extracts from S. moorcroftiana seeds were not obviously effective in motility of C. elegans.
Figure 2: Motility of Caenorhabditis elegans with various interventions. The body bends per 20 s and the head swings per minute are the main outcome measures of the motility of Caenorhabditis elegans. We randomly selected 20 worms to observe in each experiment group. Each of the motility assays was repeated at least three independent times with similar results. P value was calculated using Student's t-test

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   Conclusion Top


We explored the potential application of 95% ethanolic extracts from S. moorcroftiana seeds in the present study. On the basis of our experimental evidence, we are the first to suggest that 95% ethanol extracts from S. moorcroftiana seeds could increase life span and could not obviously influence the motility in C. elegans. Therefore, it could be a new source of natural, which will play an important role in anti-aging. However, in terms of which ingredients of the crude 95% ethanol extracts of S. moorcroftiana seeds have effect on anti-aging and why this substance could influence life span, further studies are needed to explore related important issues. Noticeably, however, if the similar results would occur in humans still remain unknown.

Acknowledgments

The authors are grateful to Dr. Dejuan Zhi of Microbial and Biochemical Pharmacy laboratory in Lanzhou University for helpful discussions.

Financial support and sponsorship

The National Natural Science Foundation of China (31360604) and Lanzhou University Innovation and Entrepreneurship Fund for College Students (20141073001313).

Conflicts of interest

There are no conflicts of interest.

 
   References Top

1.
Ma XM, Li HY, Yin SF, Meng XQ, Zhou GK, LU XM. A study on bacteriostasis and inhibiting tumor cell proliferative activity of alkaloids from Sophora moorcroftiana seeds. J Lanzhou Univ 2003;39:74-7.  Back to cited text no. 1
    
2.
Ma XM, Li HY, Wang B, Yin SF. The study on killing protoscolex activity and anti-inflammatory of alkaloids extracted from Sophora moorcroftiana seeds in vitro . Chin J Parasit Dis 2004;17:217-9.  Back to cited text no. 2
    
3.
Ma XM, Li HY, Yin SF, Wang B. Apoptosis in SGC-790l cells induced by alkaloids from Sophora moorcroftiana seeds. Chin Tradit Pat Med 2004;26:654-7.  Back to cited text no. 3
    
4.
Wilson MA, Shukitt-Hale B, Kalt W, Ingram DK, Joseph JA, Wolkow CA. Blueberry polyphenols increase lifespan and thermotolerance in Caenorhabditis elegans . Aging Cell 2006;5:59-68.  Back to cited text no. 4
    
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Artal-Sanz M, de Jong L, Tavernarakis N. Caenorhabditis elegans : A versatile platform for drug discovery. Biotechnol J 2006;1:1405-18.  Back to cited text no. 5
    
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Silverman GA, Luke CJ, Bhatia SR, Long OS, Vetica AC, Perlmutter DH, et al. Modeling molecular and cellular aspects of human disease using the nematode Caenorhabditis elegans . Pediatr Res 2009;65:10-8.  Back to cited text no. 6
    
7.
Tan MW, Rahme LG, Sternberg JA, Tompkins RG, Ausubel FM. Pseudomonas aeruginosa killing of Caenorhabditis elegans used to identify P. aeruginosa virulence factors. Proc Natl Acad Sci U S A 1999;96:2408-3241.  Back to cited text no. 7
    
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Brenner S. The genetics of Caenorhabditis elegans . Genetics 1974;77:71-94.  Back to cited text no. 8
[PUBMED]    
9.
Stiernagle T. Maintenance of C. elegans . In: TCeR, editor. Community Worm Book: Worm Book. Beijing: Beijing Institute of Technology Press; 2006.  Back to cited text no. 9
    
10.
Xingming M, Hongyu L, Shaofu Y, Bo W. The study on bacteriosta- sis and anti-inflammatory activity of alkaloids from Sophora moorcroftiana seeds. Acta Chin Med Pharma 2004;32:23-5.  Back to cited text no. 10
    
11.
Xu Y. A preliminary study of Tibet Sophora moorcroftiana seed nutrients. Nat Resour 1992;4:379-82.  Back to cited text no. 11
    
12.
Li Y. Sophora moorcroftiana biological characteristics and utilization. Nat Resour 1993;5:75-9.  Back to cited text no. 12
    
13.
Cai WJ. Life extension of Caenorhabditis elegans caused by Epimendium flavoniods treatment [D]. Shanghai: Fudan University; 2008.  Back to cited text no. 13
    
14.
Swain SC, Kuesekotten K, Baumeister B. C. elegans metallothioneuns: New insights into phenotypic effects of eadmium toxicosis. J Mol Biol 2004;34:951-9.  Back to cited text no. 14
    
15.
Tsalik EL, Hobert O. Functional mapping of neurons that control locomotory behavior in Caenorhabditis elegans . J Neurobiol 2003;56:178-97.  Back to cited text no. 15
    

 
   Authors Top


Xingming Ma, is a Professor at the Department of Immunology, Lanzhou University, Lanzhou, China. His research interest is in the area of the immunopathogenesis injury molecular mechanisms of tuberculosis and immunotherapy tuberculosis, investigation on the antitumor and immunoregulation activity with extracts from Sophora moorcroftiana. Especially, his research team have reported many valuable research results on Sophora moorcroftiana.


Kaizhong Dong, is an Associate Professor at the Department of Microbiology, Northwest University for Nationalities, Lanzhou, China. His research interest is in the area of pathogen biology, immunoregulation activity with extracts from Chinese Herbal Medicine.


    Figures

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  [Table 1]



 

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