Effect of ligustrum fruit extract on reproduction in experimental diabetic rats

Shi-Liang FENG, Shu-Hua LI¹, Yan WANG², Chang-Chun CHEN¹, Bin GAO¹

Liaoning Provincial Diabetes Treatment Center, Shenyang 110015 , China 
¹Shengyang Diabetes Institute, Shengyang, China
²Department of Histology, China Medical University, Shengyang, China

Asian J Androl  2001 Mar; 3: 71-73

Keywords: Fructus Ligustri Lucidi; diabetes; seminiferous tubules; LH; FSH; testosterone

Abstract

Aim: To study the effect of ligustrum fruit on spermatogenesis and blood gonadal hormones in diabetic rats. Methods: Experimental diabetes was induced in male Wistar rats with streptozotocin. Ligustrum fruit extract was given by gastric gavage at a dose of crude drug 30 gkg^-1d^-1 for 110 days. The serum gonadadotropic hormones and testosterone were determined on d 60 and testicular histology examined on d 110. Results: In the control diabetic rats, the seminiferous tubules were dilated and the spermatogenic cells irregularly arranged. Spermatogenesis was arrested with the number of spermatids highly reduced and spermatozoa not observed. In the treated rats, all types of spermatogenic cells were practically normal. The serum luteinizing hormone (LH), follicle-stimulating hormone (FSH) and testosterone levels were higher in the treated than in the control rats, but the difference was insignificant. Conclusion: In experimental diabetic rats, ligustrum fruit extract protects the damaging effect of experimental diabetes on spermatogenesis.

1 Introduction

2 Materials and methods

2.1 Ligustrum fruit preparation

The fruit we used were cultivated in Henan and bought from the Haozhou Drug Company. First soak 1 kg of the fruit in 2000 mL of tap water for one hour, then boil for 30 minutes. Add more water to supplement the water loss and boil for another 30 minutes. Finally, condense the decoction to a concentration of 4 g crude drug per mL. The principal active constituent in the preparation is oleanolic acid and the content is 2.93%.

2.2 Animals and treatments

Male Wistar rats (body weight 24020 g) were supplied by the Laboratory Animal Centre of the China Medical University. The animals were separated at random into two groups, 10 normal controls and 20 experimental diabetic rats. To the latter, streptozotocin (Sigma, USA) 60 mg/kg was injected intraperitoneally. Seven days later, blood was sampled from the orbital vein and the glucose concentration was determined by the glucose-oxidase method following the procedures described in the kit (Exquisite Chemical Industry Co., Beijing). Animals with blood glucose higher than 16.7 mmol/L were used for the experiment and divided at random into the Diabetic Treated and Diabetic Control groups of 5 animals each. Rats of the Diabetic Treated group were gavaged ligustrum fruit preparation (4.5 mLkg^-1d^-1, equivalent to crude drug 30 gkg^-1d^-1) for 110 days and the two Control groups, the same quantity of water. On day 60 of treatment, the serum levels of FSH, LH and testosterone were determined by radioimmuno-method according to the procedures provided in the kit (North Biochemistry Kit Center, Beijing). On day 110, rats were sacrificed and the testes routinely fixed, sectioned into discontinuous slices (8 m) and stained with hematoxylin/eosin for morphological examination under light microscope.

2.3 Data processing

3 Results

3.1 Hormonal changes

After 60 days of treatment the changes in the serum hormone levels were shown in Table 1. It can be seen that: the hormonal levels in the Diabetic Control rats were lower than those of the Normal controls; the LH, FSH and T levels were higher in the Diabetic Treated rats than in the Diabetic Control rats; and the LH and FSH levels were even higher in the Diabetic Treated rats than in the Normal Control rats. The difference between any of the two groups was statistically insignificant.

Table 1. Ligustrum effect on serum hormones in normal and experimental diabetic rats (s).

3.2 Testicular morphology

3.2.1 Diabetic Treated group and Normal Control group

After 110 days of treatment, all kinds of spermatogenic cells were practically normal. Cells were arranged regularly in 8-9 layers. There were numerous spermatids and grown-up spermatozoa could be seen scattered here and there (Figures 1, 2). The morphology of the Normal Control rats was more or less the same.

Figure 1. Seminiferous tubules of rats in experimental group. Thick arrow shows spermatid and thin arrow shows sperm. HE Stain (400).
Figure 2.  Seminiferous tubules of normal rat. HE Stain (400).

3.2.2 Diabetic Control group

In this group the spermatogenesis was noticeably disturbed. The seminiferous tubules were dilated and the spermatogenic cells were irregularly arranged in 3-5 layers. The number of spermatogonium, primary spermatocyte, secondary spermatocyte and spermatids were markedly decreased and grown-up spermatozoa were not found (Figures 3, 4).

Figure 3.  Seminiferous tubules of rat in the control group: the spermatogenic cell are sparsely separated, spermatids unnoticed, no sperm present (200) HE stain.
Figure 4.  Seminiferous tubules of rats in control group, under high power microscope: notably thinly scattered spermatogenic cells (400). HE stain.

4 Discussion

Ligustrum fruit is a kind of herb possessing nourishing effects on the Liver and Kidney. According to the TCM theory, Kidney is the origin of the innate constitution storing vital essence and in charge of growth, development and reproduction. The Kidney also strengthens the bone and helps producing the marrow. Li^[4] indicated that herbal drugs nourishing the Kidney could regulate female reproductive function. In the present study we observed that in rats ligustrum fruit counteracted the damaging effect of experimental diabetes on spermatogenesis. Data also suggested that the fruit might also revert the inhibitory effect of experimental diabetes on serum FSH, LH and testosterone levels, however, it is not conclusive. It is a pity that we did not assay the hormonal profiles on d 110 of treatment, which have been scheduled in a future study. The toxicity of the glossy privet fruit was very low. Rabbit taking orally fresh fruit 75 g did not show any toxic manifestation^[7]. The LD50 of liguustrum in mice is 600 mg/kg (Liu HS, restricted publication, 1993).

References

[1] Jiang GY. Practical Diabetes. 1st Edition. Beijing: People's Health Publishing House; 1996. p 295.
[2] Li SH, Bai SY, Gao B. Experimental study on effect of ligustrum fruit on blood glucose in experimental diabetic rats. Asia Chin Med Drugs 1997; 8 (Suppl): 512-3.
[3] Hai ZQ, Hang BQ, Wang Y. Studies on the Yin nourishing effect of Erzhi pill (containing ligustrum fruit). New Chin Med 1990; 11: 51-3.
[4] Li CJ, Yu J, Guo HR, Sun YL. A preliminary study on the mechanism of kidney in regulating reproduction and ovulation. J Traditional Chin Med 1982; 6: 69-71.
[5] Wu GL, Wang HY, Chang YM, Xu RJ. Studies on the relationship between changes in sex hormones and sexual function. Chin J Endocrin Metab 1987; 3: 148-50.
[6] Zhong XL. Chinical Diabetology, 1st Edition. Shanghai: Shanghai Technology Publishing House; 1989. p 248.
[7] Wang YS. Pharmacology and Application of Chinese Drugs. Beijing: People's Health Publishing House; 1983. p 130.

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Correspondence to: Dr Shi-Liang FENG, Liaoning Provincial Diabetes Treatment Center, 48 First Quanyuan Road, Shenyang, China.
Tel: +86-24-2482 1334    Fax: +86-24-2423 9287
Received 2000-07-10     Accepted 2000-12-28