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Journal of Chinese Integrative Medicine ›› 2013, Vol. 11 ›› Issue (3): 195-205.doi: 10.3736/jintegrmed2013029

• Research Article • Previous Articles     Next Articles

Constituents of the anti-asthma herbal formula ASHMITM synergistically inhibit IL-4 and IL-5 secretion by murine Th2 memory cells, and eotaxin by human lung fibroblasts in vitro

Bolleddula Jayaprakasama,Nan Yanga, Ming-Chun Wenb, Rong Wangc, Joseph Goldfarbd, Hugh Sampsona, Xiu-Min Lia()   

  1. a. Center for Chinese Herbal Therapy for Asthma and Allergy, Department of Pediatrics, Icahn School ofMedicine at Mount Sinai, New York, NY 10029, USA
    b. Department of Pneumology, Weifang Asthma Hospital, Weifang 261041, Shandong Province, China
    c. Department of Human Genetics, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA
    d. Department of Pharmacology and Systems Therapeutics, Icahn School of Medicine at Mount Sinai, NewYork, NY 10029, USA
  • Received:2013-01-22 Accepted:2013-04-07 Online:2013-05-10 Published:2013-05-15

Objective

Anti-asthma herbal medicine intervention (ASHMITM), a combination of three traditional Chinese medicinal herbs developed in our laboratory, has demonstrated efficacy in both mouse models of allergic asthma, and a double-blind placebo-controlled clinical trial in patients with asthma. This study was designed to determine if the anti-inflammatory effects of individual herbal constituents of ASHMITM exhibited synergy.


Methods

Effects of ASHMI and its components aqueous extracts of Lingzhi (Ganoderma lucidum), Kushen (Sophora flavescens) and Gancao (Glycyrrhiza uralensis), on Th2 cytokine secretion by murine memory Th2 cells (D10.G4.1) and eotaxin-1 secretion by human lung fibroblast (HLF-1) cells were determined by measuring levels in culture supernatants by enzyme-linked immunosorbent assay. Potential synergistic effects were determined by computing interaction indices from concentration-effect curve parameters.


Results

Individual Lingzhi, Kushen and Gancao extracts and ASHMI (the combination of individual extracts) inhibited production of interleukin (IL)-4 and IL-5 by murine memory Th2 cells and eotaxin-1 production by HLF-1 cells. The mean 25%-inhibitory-concentration (IC25) values (mg/mL) for ASHMI, Lingzhi, Kushen and Gancao for IL-4 production were 30.9, 79.4, 123, and 64.6, respectively; for IL-5 production were 30.2, 263, 123.2 and 100, respectively; for eotaxin-1 were 13.2, 16.2, 30.2, and 25.1, respectively. The IC50 values (mg/mL) for ASHMI, Lingzhi, Kushen and Gancao for IL-4 production were 158.5, 239.9, 446.7, and 281.8, respectively; for eotaxin-1 were 38.1, 33.1, 100, and 158.5, respectively. The interaction indices of ASHMI constituents at IC25 were 0.35 for IL-4, 0.21 for IL-5 and 0.59 for eotaxin-1. The interaction indices at IC50 values were 0.50 for IL-4 and 0.62 for eotaxin-1 inhibition. Inhibition of IL-5 did not reach IC50 values. All interaction indices were below 1 which indicated synergy.


Conclusion

By comparing the interaction index values, we find that constituents in ASHMITM synergistically inhibited eotaxin-1 production as well as Th2 cytokine production.

Key words: Traditional Chinese medicine, Herbal medicine, Plant extracts, Anti-asthma herbal medicine intervention (ASHMI), Anti-asthmatic agents, Chemokine CCL11, Interleukin-4, Interleukin-5, in vitro

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Figure 1

Consistency of HPLC fingerprints and major chemical constituents in different preparations of ASHMIA: HPLC fingerprints generated from standard ASHMITM, experimental ASHMI, and individual herb extracts. B: Characterization of chemical constituents of major corresponding peaks between standard ASHMITM, experimental ASHMI and individual herb extracts. Peaks 16 and 32 correspond to liquiritin and glycyrrhizin from Gancao; peaks 7 and 19 correspond to matrine and kushenol O from Kushen; peaks 26 and 27 correspond to ganoderic acid D and ganoderic acid A from Lingzhi. HPLC: High pressure liquid chromatography."

Figure 2

Effects of A) ASHMI; B) Lingzhi; C) Kushen; and D) Gancao on conalbumin-induced interleukin-4D10 cells were cultured for 72 h with or without herbal extracts in the presence of antigen conalbumin and antigen-presenting cells. Interleukin-4 level was measured from the supernatant by enzyme-linked immunosorbent assay and the percentage of inhibition was calculated with respect to untreated cells. Three replications were run at each concentration. Concentration-response curves were constructed using Prism 4.0 (GraphPad Software, San Diego, CA, USA). The dotted lines indicate the 95% confidence intervals."

"

Figure 3

Effects of A) ASHMI; B) Lingzhi; C) Kushen; and D) Gancao on conalbumin-induced interleukin-5D10 cells were cultured for 72 h with or without herbal extracts in the presence of antigen conalbumin-presenting cells. Interleukin-5 level was measured from the supernatant by enzyme-linked immunosorbent assay and percent inhibition was calculated with respect to negative control. Three replicates were run at each concentration. Concentration-response curves were constructed using Prism 4.0 (GraphPad Software, San Diego, CA, USA). The dotted lines indicate the 95% confidence intervals."

"

Figure 4

Effects of A) ASHMI; B) Lingzhi; C) Kushen; and D) Gancao on eotaxin-1 production by human lung fibroblastsHuman lung fibroblasts were cultured with or without herbal extracts for 48 h. Eotaxin-1 level in the culture supernatants was measured by enzyme-linked immunosorbent assay. The percentage of inhibition was calculated with respect to untreated cells. Three replicates were run at each concentration. Concentration-response curves were constructed using Prism 4.0 (GraphPad Software, San Diego, CA, USA). The dotted lines indicate the 95% confidence intervals."

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