Effects of Huang Bai (Phellodendri Cortex) on bone growth and pubertal development in adolescent female rats

Abstract

Background: To evaluate the effects of Huang Bai (Phellodendron amurense) on growth and maturation in adolescent female rats.

Methods: Female Sprague-Dawley rats (28 days old; n = 72) were divided into six daily treatment groups: control (distilled water), Huang Bai (100 and 300 mg/kg), recombinant human GH (rhGH; 20 μg/kg), estradiol (1 μg/kg), and triptorelin (100 μg). Body weight, food intake, and vaginal opening were measured daily from postnatal day (PND) 28 to PND 43. Tetracycline (20 mg/kg) was injected on PND 41. After sacrifice on PND 43, the ovaries and uterus were weighed, and the tibias were fixed in 4% paraformaldehyde. Decalcified and dehydrated tibias were sectioned at a thickness of 40 μm, and sectioned tissues were examined with a fluorescence microscope. Insulin-like growth factor (IGF)-1 and bone morphogenetic protein (BMP)-2 were detected using immunohistochemistry.

Results: Relative to controls, body weight was higher in the triptorelin group. Bone growth rate increased in the Huang Bai 100 mg/kg (354.00 ± 31.1 μm/day), rhGH (367.10 ± 27.11 μm/day), and triptorelin (374.50 ± 25.37 μm/day) groups. Expression of IGF-1 and BMP-2 in the hypertrophic zone was higher in all experimental groups. Vaginal opening occurred earlier in the estradiol group (PND 33.58 ± 1.62) than in controls and later in the triptorelin group (PND > 43). Ovarian and uterine weights were lower in the oestradiol and triptorelin groups. However, Huang Bai had nonsignificant effects on vaginal opening and the weights of ovaries and the uterus.

Conclusions: Huang Bai stimulated bone growth by upregulating IGF-1 and BMP-2 in the growth plate. However, it had no effect on pubertal development.

Keywords: Bone growth; Bone morphogenetic protein-2; Huang Bai; Insulin-like growth factor-1; Ovarian weight; Uterine weight; Vaginal opening.

Fig. 1

High-performance liquid chromatography of Huang Bai

Fig. 2

The daily longitudinal bone growth rate was calculated from the 2-day longitudinal growth plate length. Each value is shown as the mean ± SD for 12 rats, except HB300 (11 rats). Statistical analysis: *P < 0.05, **P < 0.01 compared to control (DW–administrated group). HB100: Huang Bai (100 mg/kg, p.o.), HB300: Huang Bai (300 mg/kg, p.o.), rhGH: recombinant human growth hormone (20 μg/kg, s.c.), estradiol: 17β-estradiol (1 μg/kg, s.c.), triptorelin: triptorelin (100 μg, i.p.)

Fig. 3

IGF-1 a and BMP-2 b in the growth plate was detected using immunohistochemistry. Control: DW, HB100: Huang Bai (100 mg/kg, p.o.), HB300: Huang Bai (300 mg/kg, p.o.), rhGH: recombinant human growth hormone (20 μg/kg, s.c.), estradiol: 17β-estradiol (1 μg/kg, s.c.), triptorelin: triptorelin (100 μg, i.p.). RZ: resting zone, PZ: proliferative zone, HZ: hypertrophic zone. Scale bar = 100 μm

Fig. 4

Percentage of IGF-1 a and BMP-2 b labelled chondrocytes was calculated by counting stained and total chondrocytes in two parallel columns. Each value is shown as the mean ± SD of 12 rats, except HB300 (11 rats). Statistical analysis: *P < 0.05, **P < 0.01, ***P < 0.001 compared to control (DW– administrated group). HB100: Huang Bai (100 mg/kg, p.o.), HB300: Huang Bai (300 mg/kg, p.o.), rhGH: recombinant human growth hormone (20 μg/kg, s.c.), estradiol: 17β-estradiol (1 μg/kg, s.c.), triptorelin: triptorelin (100 μg, i.p.)

Fig. 5

Percentage of female rats with vaginal opening occurring between PND 28 and 43. One rat in the Huang Bai 300 mg/kg group and all rats in the triptorelin group failed to show vaginal opening. Control: DW (p.o.), HB100: Huang Bai (100 mg/kg, p.o.), HB300: Huang Bai (300 mg/kg, p.o.), rhGH: recombinant human growth hormone (20 μg/kg, s.c.), estradiol: 17β-estradiol (1 μg/kg, s.c.), triptorelin: triptorelin (100 μg, i.p.)

Fig. 6

Ovarian and uterine indices after sacrifice were calculated as ovarian and uterine weight (g) per body weight (kg). Each value is shown as the mean ± SD of 12 rats, except HB300 (11 rats). Statistical analysis: **P < 0.01, ***P < 0.001 compared to control (DW– administrated group). HB100 Huang Bai (100 mg/kg, p.o.), HB300 Huang Bai (300 mg/kg, p.o.), rhGH recombinant human growth hormone (20 μg/kg, s.c.), estradiol 17β-estradiol (1 μg/kg, s.c.), triptorelin triptorelin (100 μg, i.p.)