| The pulmonary neuroendocrine cells (PNEC) system consists of solitary cells and distinctive cell clusteres termed neuroepithelial bodies (NEB). The pulmonary neuroendocrine system is still regarded as an oxygen sensitive chemoreceptor with local and reflex-mediated regulatory functions, and as a regulator of airway reactivity, and as a regulator of lung growth and development. A novel role of PNEC or NEB is emerging as guardians of lung stem cell niches. This has potential implications for carcinogenesis and injury repair. These functions are always played a part in afferent sensory fibers of vagal originating with cell bodies residing in the nodose ganglion.NEB is under dual sensory innervation. A vagal sensory component with origin in the nodose ganglion revealed calbinding D28K and P2X3 purinoreceptor immunoreactive nerve fibers that were myelinated. Another sensory nerve fiber system is consisted of thin unmyelinated, nonvagal substance P/CGRP immunoreactive nerves that have their origin in dorsal root ganglia. CGRP was synthetized in neural ganglion and transported to nerve ending, the secretion of CGRP was regulated afferent neuron of vagus. There is a decreasing nerve density from proximal to more distal conducting airways, while there remains a frequent interchange of single nerve fibers across epithelial and subepithelial compartments without termination. NEB and solitary PNEC may be actually interconnected via fine submucosal nerve fibers. Unilateral vagotomy led to degeneration and disappearance of ipsilateral NEB,and NEB and solitary PNEC may be changed.The nerve fiber of broncho-mucosa and muscles would be cut in the process of tracheoplasty, bronchoplasty and lung transplantation in clinic. The response of neurostimulation would be changed, likewise neuro-immunity and the function of neuroendocine. These changes would be bought about extremely significant effect in the process of trachea-mucosal and broncho-mucosal repair. Hence, it will be important to investigate the effects and significance of amount, distribution and their secreting function of pulmonary neuroendocrine cells (PNEC) following denervation.AIM:To study the effects and changes of amount, distribution, secretion of pulmonary neuroendocrine cells after vagotomy and reconstruction of bronchus to explore the function of PNEC and the role of PNEC in damage and repair of airway after denervation.METHODS:1. 30 male sprague dawley rats were randomly divided into three groups according to sample taking time after left cervical vagotomy:1 week, 1 month, 3 months with 10 animals in each, and another 10 rats without vagotomy were used as normal control. The ultramicrostructure of PNEC were observed with electron microscope. The quantity and distribution of PNEC were studied by Grimelius silver stain. Both serotonin (5-HT) positive PNEC and calcitonin gene-related peptide (CGRP) positive PNEC were studied by immunohistochemistry. The serous concentration of CGRP was detected by radio-immunity analysis method. The level of expression of CGRP mRNA was measured by RT-PCR method.2. 30 male sprague dawley rats were randomly divided into three groups according to sample taking time after left reconstruction of bronchus: 1 week, 1 month, 3 months with 10 animals in each, and another 10 rats without surgery were taken as normal control. The broncho-healing states were observed with HE staining slice after after left reconstruction of bronchus. The quantity and distribution of PNEC were studied by Grimelius silver stain. Both serotonin (5-HT) positive PNEC and calcitonin gene-related peptide (CGRP) positive PNEC were studied by immunohistochemistry. The serous concentration of CGRP was detected by radio-immunity analysis method. The level of expression of CGRP mRNA was measured by RT-PCR method.RESULTS:1. The electron microscopic studies have shown that PNEC were mainly found in the epithelium of trachea, bronchus and bronchioles. The cells are usually flask, oval or pyramidalshaped with most of them concentrated at the lumina of respiratory tract from basal lamina or extended along the lamina of the contiguous epitheliums. Occasionally, the lateral dendriteike cytoplasmic processes extend between adjacent epithelial cells. The apical region of the cell reached the lumen where the microvilli were revealed on surface of some cells. The identification of small, cytoplasmic, generally spherical granules, otherwise known as dense-core vesicles (DCV), has been served as the major ultrastructural characteristic of the PNEC. The appearance of DCV was shown as globular. The diameters of these DCVs can range from 70 to 200 nm. The DCV contain a high electronic density core, there is a thin, electron-lucent zone or"halo"betweem core and membrane. Other ultrastructural characteristics associated with PNEC are variable amounts of free ribosomes,smooth and rough endoplasmic reticulum, bundles of cytoplasmic filaments and mitochondria. The latter are usually smaller than those of adjacent cells. The Golgi apparatus is usually visible and present in a supranuclear position.2. PNECs were mainly found in the epithelium of intra pulmonary bronchus and terminal bronchioles and some of them were assembled to form into neuroepithelial bodies (NEBs). 1 week and 1month after vagotomy, the number of PNEC was significantly increased in larger bronchi, and nearly approached the lever of normal controls after 3 months later. Except their location in the larger bronchi, the changes of 5-HT and CGRP positive PNECs presented the same as PNEC in number. The NEBs were formed by 5-HT positive PNEC or CGRP positive PNEC also.3. At 1 week and 1 month after vagotomy, the serous concentration of CGRP was significantly decreased than that in the normal controls (P﹤0.01) and gradually increased nearly to the normal level in 3 months after vagotomy. The expression of CGRP mRNA was significantly higher than that in the normal control at 1 week and 1 month after vagotomy (P﹤0.01) and gradually declined to the normal control after 3 months (P﹥0.05).4. Light microscopic studies have shown granulation tissue hyperblastosis on broncho-shama after reconstruction of bronchus, discontinued Mucosa of bronchus and stuffed bundles of hyperplasia endothelial cells and expanded blood capillary. There were massive fibroblast and inflammatory cells around blood capillaries.5. The changes of PNEC and 5-HT and CGRP positive PNECs in number, location, formation of NEB after left reconstruction of bronchus were similar as after vagotomy.6. At 1 week and 1 month after left reconstruction of bronchus, the serous concentration of CGRP was significantly decreased than that in the normal controls (P﹤0.01) and gradually increased nearly to the normal level in 3 months after left reconstruction of bronchus. The expression of CGRP mRNA was significantly higher than that in the normal control at 1 week and 1 month after left reconstruction of bronchus (P﹤0.01) and gradually declined to the normal control after 3 months (P﹥0.05).CONCLUSION:1. DCV has served as the major ultrastructural characteristic of the PNEC.2. The quantity and distribution and their secreting function of PNEC and 5-HT positive PNEC and CGRP positive PNEC were significantly changed after vagotomy. It indicated that PNEC might be dominated and their secreting function might be regulated by vagus fibres.3. The serous concentration of CGRP and expression of CGRP mRNA were significantly changed after vagotomy. It indicated that the expression of CGRP mRNA and the secretion of CGRP were dominated and regulated by vagus.4. The quantity of PNEC and 5-HT positive PNEC and CGRP positive PNEC were significantly increased after reconstruction of bronchus. It indicated that PNEC might be dominated by vagus fibres. Meanwhile, it deduced that PNEC might be taken part in damage and repair of airway.5. The serous concentration of CGRP and expression of CGRP mRNA were significantly changed after reconstruction of bronchus. It indicated that the expression of CGRP mRNA and the secretion of CGRP were dominated and regulated by CGRP-immunoreactive (IR) fibers of vagus. Meanwhile, it indicated that CGRP was considered as local regulatory factor to stimulate hyperplasia endothelialis and participated in damage and repair of airway. |
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