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Journal of Chinese Integrative Medicine ›› 2006, Vol. 4 ›› Issue (4): 368-373.doi: 10.3736/jcim20060410

• Original Experimental Research • Previous Articles     Next Articles

Pre-administ ration of Ganoderma lucidum spore reduces incidence ofneural tube defects induced by retinoic acid in pregnant mice

Wei Zhang, Yuan-Shan Zeng, Yi Xiong, Sui-Jun Chen, Zhi-qiang Zhong   

  1. Division of Neurosciences, Depar tment of Histology and Embryology , Zhongshan Medical College , Sun Yat-sen University ,Guangzhou , Guangdong Province 510080 , China
  • Online:2006-07-31 Published:2006-07-15

Objective

To explore whether pre-administrat ion of Ganoderma lucidum spore (GASP) can reduce incidence of neural tube defects (NTDs) induced by all-trans retinoic acid ( ATRA) in pregnant mice.

Methods

Twenty pregnant mice were randomly divided into four groups: normal control group , solventtreated group, ATRA-induced group , and GASP-treated plus ATRA-induced group . GASP solut ion, which wasprepared with solvent ( sodium carboxymethyl cellulose) , was fed to the pregnant mice in the GASP-treated plus ATRA-induced group twice a day from embryo (E) 0 d to E10 .5 d . The same dose of solvent was given to the pregnant mice in the solvent- treated group . At E7 .75 d, ATRA ( 50 mg/ kg) was given to the pregnant mice in both ATRA-induced group and GASP-treated plus ATRA-induced group for single t ime . Embryos were sampled from pregnant mice at E10 .5 d . Then the incidence rate of NTDs in mouse embryo was calculated and the crown-rump length of mouse embryo was measured . The positive rate of nestin expression and the distribution of cell cycle of embryonic neural tube neuroepithelial cells were detected by histochemical stainingtechnique and flow cytometry respectively .Reverse transcription-polymerase chain reaction method was used to detect the gene expressions of cyclin-dependent protein kinase 2 (Cdk2) and Cdk4 mRNAs .

Results

The incidence rates of NTDs in mouse embryos in the ATRA-induced group and the GASP-treated plus ATRA-induced group were 79 .41% and 21 .67% respectively , while the crown-rump length of mouse embryos in these two groups were (3 .62±1 .27) mm and (5 .84±0 .92) mm respectively .The positive rate of nestin expression in embryonic neural tube neuroepi thelial cells of mouse embryo at E10 .5 d in the ATRA-induced group was 32 .44% , while that in the GASP-treated plus ATRA-induced group was 77 .65% . The cell cycle of embryonic neural tube neuroepithelial cells was obviously arrested at G0/ G1 phase in the ATRA-induced group as compared wi th that in the GASP-treated plus ATRA-induced group . The Cdk4 mRNA was transcripted at a high level in embryonic neural tube in the GASP-treated plus ATRA-induced group, but the Cdk2 mRNA wasnot detected in this group.

Conclusion

Pre-administrat ion of GASP can reduce the incidence of NTDs induced by ATRA in pregnant mice .

Key words: Neural tube defects, Ganoderma lucidum spore, Ret inoic acid, Nestin, Cyclin-dependent kinase

CLC Number: 

  • R714.431

Figure 1

Mouse embryo at E10.5 d in solvent-treated groupA: Entirety morphology of mouse embryo at E10.5 d; B: Horizontal section along the line in A, immunofluorescence histochemical staining (red) of nestins in embryonic neural tube neuroepithelial cells (the scale bar represents 1 mm); C: Partial enlarged details from Figure 1-B (the scale bar represents 0.05 mm)."

Figure 2

Mouse embryo at E10.5 d in ATRA-induced groupA: Entirety morphology of mouse embryo at E10.5 d; B: Horizontal section along the line in A, immunofluorescence histochemical staining (red) of nestins in embryonic neural tube neuroepithelial cells (the scale bar represents 1 mm); C: Partial enlarged details from Figure 2-B (the scale bar represents 0.05 mm)."

"

Table 1

Incidence rates of NTDs and crown-rump length of mouse embryos at E10.5 d in four groups"

Group n Total number
of embryos
Number of
dead embryos
Number of
abnormal embryos
Incidence rate
of NTDs (%)
Crown-rump
length(?x±s, mm)
Normal control 5 53 2 0 3.77* 5.62±0.44
Solvent-treated 5 62 5 0 8.06* 5.96±0.55
ATRA-induced 5 68 20 34 79.41* 3.62±1.27*
GASP-treated plus ATRA-induced 5 60 7 6 21.67 5.84±0.92

Table 2

Positive rates of nestin expressions in embryonic neural tube neuroepithelial cells of mouse embryos at E10.5 d and their cell cycle distributions in four groups (ヌ±S,%)"

Group n Positive rate of nestin expression Cell cycle distribution
G0/G1 G2/M S
Normal control 5 74.73±10.49△ 65.62±10.99* 6.60±4.40 27.76±12.20*
Solvent-treated 5 77.83±18.88△ 65.94±1.90* 10.02±3.90 24.02±5.26*
ATRA-induced 5 32.44±4.64* 82.80±7.39* 4.00±2.13 13.20±5.35*
GASP-treated plus ATRA-induced 5 77.65±5.81 53.74±3.15 7.12±3.73 39.18±4.05

Figure 4

Expression of nestin in neural tube neuroepithelial cells of mouse embryo at E10.5 d detected by flow cytometry"

Figure 5

Electrophoregram of RT-PCR products of Cdk2 mRNAM: Marker; 1: Normal control group; 2: Solvent-treated group; 3: ATRA-induced group; 4: GASP-treated plus ATRA-induced group."

Figure 6

Electrophoregram of RT-PCR products of Cdk4 mRNAM: Marker; 1: Normal control group; 2: Solvent-treated group; 3: ATRA-induced group; 4: GASP-treated plus ATRA-induced group."

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