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Ganoderma lucidum polysaccharides in human monocytic leukemia cells: from gene expression to network construction.

Cheng KC, Huang HC, Chen JH, Hsu JW, Cheng HC, Ou CH, Yang WB, Chen ST, Wong CH, Juan HF - BMC Genomics (2007)

Bottom Line: In previous studies, F3, the active component of the polysaccharide extract, was found to activate various cytokines such as IL-1, IL-6, IL-12, and TNF-alpha.The apoptosis induction through the DR3 and DR4/5 death receptors was found to be one of the most significant pathways and play a key role in THP-1 cells after F3 treatment.Our results showed that F3 may induce death receptor ligands to initiate signaling via receptor oligomerization, recruitment of specialized adaptor proteins and activation of caspase cascades.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Life Science, National Taiwan University, Taipei 106, Taiwan. jerekcheng@gmail.com

ABSTRACT

Background: Ganoderma lucidum has been widely used as a herbal medicine for promoting health and longevity in China and other Asian countries. Polysaccharide extracts from Ganoderma lucidum have been reported to exhibit immuno-modulating and anti-tumor activities. In previous studies, F3, the active component of the polysaccharide extract, was found to activate various cytokines such as IL-1, IL-6, IL-12, and TNF-alpha. This gave rise to our investigation on how F3 stimulates immuno-modulating or anti-tumor effects in human leukemia THP-1 cells.

Results: Here, we integrated time-course DNA microarray analysis, quantitative PCR assays, and bioinformatics methods to study the F3-induced effects in THP-1 cells. Significantly disturbed pathways induced by F3 were identified with statistical analysis on microarray data. The apoptosis induction through the DR3 and DR4/5 death receptors was found to be one of the most significant pathways and play a key role in THP-1 cells after F3 treatment. Based on time-course gene expression measurements of the identified pathway, we reconstructed a plausible regulatory network of the involved genes using reverse-engineering computational approach.

Conclusion: Our results showed that F3 may induce death receptor ligands to initiate signaling via receptor oligomerization, recruitment of specialized adaptor proteins and activation of caspase cascades.

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CASP3 and CASP 7 were cleaved into active forms after F3 treatment in THP-1 cells. After THP-1 cells were treated with F3 for 0, 6, 12, 24 hours, we detected proforms and active forms of CASP3 and CASP7 using western blotting. CASP3 and CASP7 were activated after F3 treatment.
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Figure 6: CASP3 and CASP 7 were cleaved into active forms after F3 treatment in THP-1 cells. After THP-1 cells were treated with F3 for 0, 6, 12, 24 hours, we detected proforms and active forms of CASP3 and CASP7 using western blotting. CASP3 and CASP7 were activated after F3 treatment.

Mentions: In the pathway of apoptosis induction through DR3 and DR4/5 death receptors, 9 genes were found to be up-regulated and 2 to be down-regulated among the 27 genes in F3-treated THP-1 cells after F3 treatment for 6 hours. The 9 up-regulated genes include tumor necrosis factor (ligand) superfamily, member 10 (TNFSF10 or TRAIL), tumor necrosis factor receptor superfamily, member 10b (TNFRSF10B or DR5), caspase 10, apoptosis-related cysteine peptidase (CASP10), BH3 interacting domain death agonist (BID), CASP8 and FADD-like apoptosis regulator (CFLAR), TNFRSF1A-associated via death domain (TRADD), nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha (NFKBIA), nuclear factor of kappa light polypeptide gene enhancer in B-cells 1 (NFKB1), and caspase 7, apoptosis-related cysteine peptidase (CASP7); all of these genes are involved in cell death. Two down-regulated genes were DNA fragmentation factor, 40kDa, beta polypeptide (caspase-activated DNase, DFFB) and caspase 6, apoptosis-related cysteine peptidase (CASP6). CASP6 cleavage by caspase-3 (CASP3), caspase-8 (CASP8) or -10 (CASP10) generates the two active subunits. In the microarray gene expression results, CASP3 and CASP8 showed no significant difference after F3 treatment. However, in our Q-PCR gene expression results, CASP8 showed a significant up-regulated expression. CASP3 and CASP7 were cleaved into their active forms after F3 treatment as shown in Figure 6. In summary, F3 may bind to death receptor 4/5, and activate downstream apoptosis-related cysteine peptidase such as CASP8, CASP3 and CASP7, leading to the apoptosis of THP-1 cells.


Ganoderma lucidum polysaccharides in human monocytic leukemia cells: from gene expression to network construction.

Cheng KC, Huang HC, Chen JH, Hsu JW, Cheng HC, Ou CH, Yang WB, Chen ST, Wong CH, Juan HF - BMC Genomics (2007)

CASP3 and CASP 7 were cleaved into active forms after F3 treatment in THP-1 cells. After THP-1 cells were treated with F3 for 0, 6, 12, 24 hours, we detected proforms and active forms of CASP3 and CASP7 using western blotting. CASP3 and CASP7 were activated after F3 treatment.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2211495&req=5

Figure 6: CASP3 and CASP 7 were cleaved into active forms after F3 treatment in THP-1 cells. After THP-1 cells were treated with F3 for 0, 6, 12, 24 hours, we detected proforms and active forms of CASP3 and CASP7 using western blotting. CASP3 and CASP7 were activated after F3 treatment.
Mentions: In the pathway of apoptosis induction through DR3 and DR4/5 death receptors, 9 genes were found to be up-regulated and 2 to be down-regulated among the 27 genes in F3-treated THP-1 cells after F3 treatment for 6 hours. The 9 up-regulated genes include tumor necrosis factor (ligand) superfamily, member 10 (TNFSF10 or TRAIL), tumor necrosis factor receptor superfamily, member 10b (TNFRSF10B or DR5), caspase 10, apoptosis-related cysteine peptidase (CASP10), BH3 interacting domain death agonist (BID), CASP8 and FADD-like apoptosis regulator (CFLAR), TNFRSF1A-associated via death domain (TRADD), nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha (NFKBIA), nuclear factor of kappa light polypeptide gene enhancer in B-cells 1 (NFKB1), and caspase 7, apoptosis-related cysteine peptidase (CASP7); all of these genes are involved in cell death. Two down-regulated genes were DNA fragmentation factor, 40kDa, beta polypeptide (caspase-activated DNase, DFFB) and caspase 6, apoptosis-related cysteine peptidase (CASP6). CASP6 cleavage by caspase-3 (CASP3), caspase-8 (CASP8) or -10 (CASP10) generates the two active subunits. In the microarray gene expression results, CASP3 and CASP8 showed no significant difference after F3 treatment. However, in our Q-PCR gene expression results, CASP8 showed a significant up-regulated expression. CASP3 and CASP7 were cleaved into their active forms after F3 treatment as shown in Figure 6. In summary, F3 may bind to death receptor 4/5, and activate downstream apoptosis-related cysteine peptidase such as CASP8, CASP3 and CASP7, leading to the apoptosis of THP-1 cells.

Bottom Line: In previous studies, F3, the active component of the polysaccharide extract, was found to activate various cytokines such as IL-1, IL-6, IL-12, and TNF-alpha.The apoptosis induction through the DR3 and DR4/5 death receptors was found to be one of the most significant pathways and play a key role in THP-1 cells after F3 treatment.Our results showed that F3 may induce death receptor ligands to initiate signaling via receptor oligomerization, recruitment of specialized adaptor proteins and activation of caspase cascades.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Life Science, National Taiwan University, Taipei 106, Taiwan. jerekcheng@gmail.com

ABSTRACT

Background: Ganoderma lucidum has been widely used as a herbal medicine for promoting health and longevity in China and other Asian countries. Polysaccharide extracts from Ganoderma lucidum have been reported to exhibit immuno-modulating and anti-tumor activities. In previous studies, F3, the active component of the polysaccharide extract, was found to activate various cytokines such as IL-1, IL-6, IL-12, and TNF-alpha. This gave rise to our investigation on how F3 stimulates immuno-modulating or anti-tumor effects in human leukemia THP-1 cells.

Results: Here, we integrated time-course DNA microarray analysis, quantitative PCR assays, and bioinformatics methods to study the F3-induced effects in THP-1 cells. Significantly disturbed pathways induced by F3 were identified with statistical analysis on microarray data. The apoptosis induction through the DR3 and DR4/5 death receptors was found to be one of the most significant pathways and play a key role in THP-1 cells after F3 treatment. Based on time-course gene expression measurements of the identified pathway, we reconstructed a plausible regulatory network of the involved genes using reverse-engineering computational approach.

Conclusion: Our results showed that F3 may induce death receptor ligands to initiate signaling via receptor oligomerization, recruitment of specialized adaptor proteins and activation of caspase cascades.

Show MeSH
Related in: MedlinePlus