Traditional Chinese medicine (TCM) is characterized by its unique system of therapies and theories. The theory of exterior-interior correlation between the lung and large intestine is one of the most important components of the TCM Zang-Fu theory, which has been widely applied in TCM clinical practice, particularly in the treatments of the lung-intestine diseases, such as COPD and asthma [
Recent studies have proved the relationship between lung and large intestine from various perspectives, such as embryonic development, lymph circulation, mucosal immunity, micro ecology, and inflammatory harass [
Relaxing the bowels with laxative is one of the common treating methods for lung diseases based on the theory of exterior-interior correlation between lung and large intestine. Our previous studies found that Chinese Rhubarb (Dahuang) could not only effectively improve the intestinal obstruction in COPD patients and rats with COPD but also effectively improve the dyspnea and gas exchange function [
Mangxiao and Dahuang were purchased from a traditional Chinese medicinal store (Tongrentang) in Beijing, China, and were authenticated by Professor Yue-qi Wang, Basic Theory and Key Technology Research Center, Beijing University of Chinese Medicine. Mangxiao is an osmotic laxative, acting on large intestine, and its effective ingredient is sodium sulfate [
Male Wistar rats (
Rat COPD-like model was prepared by cigarette smoking and lipopolysaccharides (LPS, Sigma, St. Louis, MO, USA) stimuli, as previously described [
Sensitization and challenge of mice were performed as previously described [
Airway high reactivity (AHR) as an indicator of decline in lung function was detected in 24 hrs after the final exposure to aerosol by using an AniRes 2005 Lung Function system (Bestlab 2.0, Beijing, China) according to manufacturer’s instructions. Anesthesia was induced by intraperitoneal injection of 95 mg/kg pentobarbital sodium. A connection was made by a computer-controlled ventilator
For staining with H&E, lungs were inflated and fixed with 10% buffered formalin. Samples were embedded in paraffin, then sectioned (4
After the serum sample collection, lung, large intestine, stomach, kidney, spleen, heart, brain, and liver tissues were rinsed in ice cold phosphate-buffered saline (PBS: pH 7.5), then dried, and weighed. Afterwards, the tissues were homogenized by using a glass homogenizer on ice, with 10 mL/g of ice cold PBS. Next, homogenates were centrifuged at 5000 ×g, for 5 min at 4°C, and supernatants were collected for detection. The tissue levels of SP and VIP were measured by using enzyme-linked immuno sorbent assay (ELISA) kits according to manufacturer’s instructions (Cusabio).
After the serum sample collection, lung, large intestine, stomach, heart, and brain tissues were rinsed in ice cold phosphate-buffered saline. Then, the rinsed tissues were homogenized in 1 mL PBS and stored overnight at −20°C. After two freeze-thaw cycles, the tissues were performed to break the cell membranes, the homogenates were centrifuged at 5000 ×g, for 5 minutes at 4°C, and the supernatants were collected for detection.
Levels of NK1R in the tissues of lung, large intestine, stomach, heart, and brain, VIPR1 and VIPR2 levels in the tissues of lung, large intestine, stomach, and brain of COPD rats, and levels of NK1R, VIPR1, and VIPR2 in the tissues of lung, large intestine, stomach, and heart of allergic asthma mice were measured by using a commercial ELISA kit, following the manufacturer’s instructions (Cusabio).
Levels of NKA and NKB in lung, large intestine, stomach, kidney, spleen, heart, brain, and liver tissues were measured by using a commercial ELISA kit, following the manufacturer’s instructions (Cusabio).
Data were expressed as mean values ± standard deviation. Statistical comparisons were performed by using one-way analysis of variance. Significant levels were set at
Compared with the control group, the levels of SP in lung, stomach, and brain tissues were significantly increased, while the levels of SP in large intestine and heart tissues were decreased in the model group (
SP levels in lung, large intestine, stomach, kidney, spleen, heart, brain, and liver tissues of COPD-like rats (pg/ug, mean ± SD).
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Lung | Large intestine | Stomach | Kidney | Spleen | Heart | Brain | Liver |
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Control group | 6 |
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Model group (LPS + cigarette) | 7 |
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Mangxiao group (LPS + cigarette + Mangxiao) | 6 |
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Dahuang group (LPS + cigarette + Dahuang) | 8 |
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Means labeled with superscripts were significantly different. *
The levels of VIP in lung, large intestine, and brain tissues in the model group were notably lower than that in the control group (
VIP levels in lung, large intestine, stomach, kidney, spleen, heart, brain, and liver tissues of COPD-like rats (pg/ug, mean ± SD).
Groups |
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Lung | Large intestine | Stomach | Kidney | Spleen | Heart | Brain | Liver |
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Control group | 6 |
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Model group (LPS + cigarette) | 7 |
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Mangxiao group (LPS + cigarette + Mangxiao) | 6 |
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Dahuang group (LPS + cigarette + Dahuang) | 8 |
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Means labeled with superscripts were significantly different. *
Compared with the control group, the level of NK1R in lung tissues was distinctively increased, while the levels of NK1R in large intestine and brain tissues were decreased in the model group. Intervention with Mangxiao or Dahuang significantly reduced the level of NK1R in lung tissue (
NK1R levels in lung, large intestine, stomach, heart, and brain tissues of COPD-like rats (pg/ug, mean ± SD).
Groups |
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Lung | Large intestine | Stomach | Heart | Brain |
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Control group | 6 |
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Model group (LPS + cigarette) | 7 |
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Mangxiao group (LPS + cigarette + Mangxiao) | 6 |
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Dahuang group (LPS + cigarette + Dahuang) | 8 |
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Means labeled with superscripts were significantly different. *
The levels of VIPR1 and VIPR2 in lung and large intestine tissues in the model group were significantly lower than that in the control group. Intervention with Mangxiao or Dahuang increased the levels of VIPR1 and VIPR2 in lung tissues, compared with the model group, while the levels of VIPR1 and VIPR2 in stomach and brain tissues had no obvious changes (Tables
VIPR1 levels in lung, large intestine, stomach, and brain tissues of COPD-like rats (pg/ug, mean ± SD).
Groups |
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Lung | Large intestine | Stomach | Brain |
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Control group | 6 |
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Model group (LPS + cigarette) | 7 |
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Mangxiao group |
6 |
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Dahuang group |
8 |
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Means labeled with superscripts were significantly different. *
VIPR2 levels in lung, large intestine, stomach, and brain tissues of COPD-like rats (pg/ug, mean ± SD).
Groups |
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Lung | Large intestine | Stomach | Brain |
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Control group | 6 |
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Model group (LPS + cigarette) | 7 |
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Mangxiao group |
6 |
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Dahuang group |
8 |
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Means labeled with superscripts were significantly different. *
Compared with the control group, a considerable increase in the level of SP in lung tissues and a decrease in the levels in large intestine, heart, and liver tissues were observed in the model group. Intervention with Mangxiao markedly lowered the level of SP in lung tissues and increased the levels of SP in large intestine and stomach tissues (Table
SP levels in lung, large intestine, stomach, kidney, spleen, heart, brain, and liver tissues of asthma mice (pg/mg,
Groups | Lung | Large intestine | Stomach | Kidney | Spleen | Heart | Brain | Liver |
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Control group |
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Model group (OVA) |
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Mangxiao group |
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Means labeled with superscripts were significantly different. *
Compared with the control group, the levels of VIP in lung, large intestine, heart, and brain tissues were reduced significantly (
VIP levels in lung, large intestine, stomach, kidney, spleen, heart, brain, and liver tissues of asthma mice (pg/mg,
Groups | Lung | Large intestine | Stomach | Kidney | Spleen | Heart | Brain | Liver |
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Control group |
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Model group (OVA) |
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Mangxiao group |
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Means labeled with superscripts were significantly different. *
Compared with the control group, the level of NK1R in lung and stomach tissues was up (
NK1R levels in lung, large intestine, stomach, and heart tissues of OVA allergic asthma mice (pg/ug,
Groups | Lung | Large intestine | Stomach | Heart |
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Control group |
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Model group (OVA) |
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Mangxiao group (OVA + Mangxiao) |
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All values are expressed as mean ± SD. Means labeled with superscripts were significantly different. *
Compared with the control group, the levels of VIPR1 and VIPR2 in lung tissue were significantly lower (
VIPR1 levels in lung, large intestine, stomach, and heart tissues of asthma mice (pg/ug,
Groups | Lung | Large intestine | Stomach | Heart |
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Control group |
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Model group (OVA) |
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Mangxiao group |
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Means labeled with superscripts were significantly different. *
VIPR2 levels in lung, large intestine, stomach, and heart of asthma mice (pg/ug,
Groups | Lung | Large intestine | Stomach | Heart |
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Control group |
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Model group (OVA) |
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Mangxiao group (OVA + Mangxiao) |
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Means labeled with superscripts were significantly different. *
The levels of NKA and NKB in lung, large intestine, stomach, kidney, and spleen tissues were similar between the control group and the model group (
NKA levels in lung, large intestine, stomach, kidney, spleen, heart, brain, and liver tissues of asthma mice (pg/mg,
Groups | Lung | Large intestine | Stomach | Kidney | Spleen | Heart | Brain | Liver |
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Control group |
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Model group (OVA) |
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Mangxiao group |
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Means labeled with superscripts were significantly different. *
NKB levels in lung, large intestine, stomach, kidney, spleen, heart, brain, and liver tissues of asthma mice (pg/mg,
Groups | Lung | Large intestine | Stomach | Kidney | Spleen | Heart | Brain | Liver |
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Control group |
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Model group (OVA) |
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Mangxiao group |
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Means labeled with superscripts were significantly different. *
As the results of hematoxylin-eosin staining shown in Figure
The effects of Mangxiao or Dahuang on COPD-like rats’ lung histopathology (H&E stain ×100). Histological examination of lung tissues was performed 48 h after the final administration. Lung tissues were fixed, sectioned at 4
After excitation by different concentrated MCH, airway resistance changes are displayed in Figure
The development of airway hyperresponsiveness analysis (mean ± SD,
Hematoxylin-eosin staining displayed that alveolar epithelial cells arranged orderly, alveolar walls had structure integrity, no hyperemia, hemorrhage, or inflammatory cell infiltration were present in the control group. However, OVA-challenged lung tissues displayed bronchial epithelial cells which arranged disorderly. Compared to the control group, part of epithelium damage and infiltration of inflammatory cells into the airway were observed around the bronchi, bronchioles, and alveoli. Moreover, the majority of leukocytes were eosinophils and lymphocytes. Infiltration of inflammatory leukocytes in OVA-challenged mice treated with Mangxiao was significantly attenuated, compared with that in OVA-challenged mice (Figure
The effects of Mangxiao on mice’s lung histopathology (H&E stain ×400). Histological examination of lung tissues was performed 48 h after the final OVA challenge. Lung tissues were fixed, sectioned at 4
This study explored the specific link between lung and large intestine. Rat COPD-like model was prepared by cigarette smoking and LPS stimuli. Intragastric administration of Mangxiao (an osmotic laxative) or Dahuang (a stimulant laxative) was to stimulate the large intestine. Mouse model of allergic asthma was prepared by ovalbumin (OVA) sensitization and challenge. Intragastric administration of Mirabilite was to stimulate the large intestine. The tissue (including lung, and large intestine) levels of neuropeptides and the receptors, such as SP, VIP, NKA, NKB, NK1R, VIPR1, and VIPR2, were measured by using ELISA kits. The findings indicate that stimulating intestine with Mangxiao or Dahuang can specifically regulate the secretion of SP, VIP, and the receptors in lung tissues. These findings provide a new perspective to interpret the TCM theory of exterior-interior correlation between lung and large intestine.
The theory of exterior-interior correlation between the lung and large intestine was first recorded in
The theory of exterior-interior correlation between the lung and large intestine suggests that they connect with each other directly by meridians, and the connection between the lung meridian and the large intestine meridian is further strengthened by collaterals and branches of the two meridians [
Mangxiao and Dahuang, as an osmotic laxative and a stimulant laxative, respectively, have been used in TCM clinical practice for nearly 2000 years. Both of them have an intense effect on contracting the intestinal smooth muscle and increasing peristalsis [
Our findings indicate that by stimulating the large intestine by Mangxiao or Dahuang in rats with COPD, the levels of SP in lung, stomach, and brain tissues and the level of NK1R in lung were reduced. The levels of SP and NK1R in large intestine tissue were increased after stimulating by Dahuang. The levels of VIP in lung and large intestine tissues and VIPR1 and VIPR2 in lung tissue were increased. The level of VIP in brain tissue was reduced. Stimulating the large intestine by Mangxiao in mice with asthma significantly reduced the levels of SP and NK1R in lung tissues but increased the levels of SP and NK1R in large intestine tissues. The levels of VIP in lung, large intestine, and heart tissues and VIPR1 in lung tissues were increased. The levels of NKA and NKB in lung and large intestine tissues of mice with asthma were similar between the control and the model groups.
The results confirmed that after stimulating the large intestine by Mangxiao or Dahuang, SP, NK1R, VIP, VIPR1, and VIPR2 were all significantly increased in large intestine tissue of rats with COPD and mice with asthma. At the same time, the SP and NK1R were markedly decreased, while VIP, VIPR1, and VIPR2 became much higher in lung tissue. Abnormal secretion of SP and VIP could be observed in other organs such as stomach tissues of rats or heart and stomach tissues of mice; however, the receptors did not change obviously, while the NKA and NKB levels were similar in lung tissues of mice with asthma among groups. It indicates that stimulating large intestine with Mangxiao or Dahuang can specifically regulate the secretion of SP, VIP, and the receptors in lung tissues. Our findings provide some new lines of evidence to interpret the theory of exterior-interior correlation between the lung and large intestine.
In conclusion, we have found some new lines of evidence which suggest that a signal conditioning way of
The authors declare that they have no conflict of interests.
Xiang-Gen Zhong and Feng-Jie Zheng have equally contributed to this paper. Yu-Hang Li and Si-Hua Gao contributed to the conception and design of the study. Xiang-Gen Zhong and Feng-Jie Zheng analysed the data and drafted the paper. All by-line authors read and approved the final paper.
This study was funded by China National key basic research plan (no. 2009CB522704). The authors are grateful to The Ministry of Science and Technology of the People’s Republic of China for funding this study. Thanks to Professor Yong Qing Yang, Shanghai Research Institute of Acupuncture and Meridian, Yueyang Hospital, Shanghai University of Traditional Chinese Medicine, for the presentation of the guide.