Role of PNEC and GABA in pulmonary neuroendocrine tumors / 解放军医学杂志

Objective To investigate the role of pulmonary neuroendocrine cells(PNEC)and γ-aminobutyric acid(GABA)in patients with pulmonary neuroendocrine tumors(PNET).Methods The pathological specimens of 29 cases of PNET treated in the eighth Medical Center of Chinese PLA General Hospital from October 2018 to January 2022 were collected.The morphological characteristics were observed by HE staining,and the expression levels of synaptophysin(Syn),chromogranin A(CgA),CD56,Ki-67,CD86 and CD163 were observed by immunohistochemical staining.Calcitonin gene-related peptide(CGRP)and glutamic acid decarboxylase(GAD)65/67 in different types of PNETs were detected by double antibody immunofluorescence co-staining,and the correlation between GAD65/67 positive PNEC and macrophage polarization was analyzed.Results The results of HE staining showed that all four types of PNET tissues had neuroendocrine(NE)characteristics:rosette structure and organ nesting or palisade pattern,but they were different,and the proportion of mitotic cells from low to high was typical carcinoid(TC),atypical carcinoid(AC),large cell neuroendocrine carcinoma(LCNEC)and small cell lung cancer(SCLC).The results of immunohistochemical staining showed that the positive expression rate of Syn and CgA and the positive degree of Syn,CgA and CD56 in carcinoid(TC and AC)were significantly higher than those in LCNEC and SCLC(P＜0.05).The Ki-67 indices of the four types of PNET are:TC＜5%,AC 5%-20%,LCNEC and SCLC＞75%respectively.The number of PNEC in carcinoid was significantly higher than that in LCNEC,SCLC and paratumoral tissues(P＜0.05),but there was no significant difference in the number of PNEC between LCNEC and SCLC and para-tumor tissues(P＞0.05).The results of immunofluorescence staining showed that the number of GAD65/67 positive cells co-expressing GAD65/67 in 95%PNEC was significantly higher than that in LCNEC,SCLC and para-tumor tissues(P＜0.05),but there was no significant difference between LCNEC and SCLC GAD65/67 positive cells and para-tumor tissues(P＞0.05).The results of immunohistochemical staining also showed that the number of CD86 positive M1 macrophages was significantly higher than that of CD163 positive M2 macrophages in para-tumor tissues(P＜0.05),while M2 macrophages were significantly more than M1 macrophages in AC,LCNEC and SCLC(P＜0.01).Correlation analysis showed that the number of GAD65/67 positive PNEC cells in PNET was negatively correlated with the number of CD163 positive M2 macrophages in tumor stroma(r=-0.6336,P=0.0174).Conclusions PNEC is the main source of GABA in lung tissue and plays an immunomodulatory role in the lung,which may be involved in the progression of PNET.

Effect of EBR on Seed Germination， Seedling Growth and Key Enzyme Activities for Flavonoid Biosynthesis in Astragalus membranaceus var. mongholicus Under PEG-induced Drought Stress / 中国实验方剂学杂志

Objective:To study the effects of 24-epibrassinolide （EBR） at different concentrations on seed germination， seedling growth and key enzyme activities for flavonoid biosynthesis in <italic>Astragalus membranaceus </italic>var.<italic> mongholicus</italic> under polyethylene glycol （PEG）-induced drought stress， in order to provide theoretical reference for standardizing the drought-resistant cultivation techniques of <italic>A. membranaceus </italic>var.<italic> mongholicus</italic>. Method:The seeds of <italic>A. membranaceus </italic>var.<italic> mongholicus </italic>were soaked in EBR solution at different concentrations（0.001， 0.01， 0.1， 1， 10 μmol·L^-1） followed by foliar spraying to explore their effects on seed germination， seedling growth， photosynthesis， chlorophyll and malondialdehyde （MDA） contents， and key enzyme activities for flavonoid biosynthesis under drought stress induced by 20% polyethylene glycol （PEG）-6000. Result:Compared with the control（CK）group， PEG-induced drought stress led to an obvious decrease in germination potential， germination rate， germination index， vigor index， relative germination rate， plant height， root length， aboveground and root dry weight， net photosynthetic rate （<italic>Pn</italic>）， stomata conductance （<italic>Gs</italic>）<italic>， </italic>transpiration rate （<italic>Tr</italic>）， chlorophyll contents and chlorophyll a/b ratio， while a significant increase in intercellular carbon dioxide concentration （<italic>Gi</italic>）， MDA， flavonoids contents， and key enzyme activities for flavonoid biosynthesis such as phenylalanine ammonia lyase （PAL）. The treatment with exogenous EBR solution at the suitable concentration significantly enhanced the adaptation of <italic>A. membranaceus </italic>var.<italic> mongholicus </italic>seeds and seedlings to PEG-induced drought stress， manifested as significantly elevated germination potential， germination rate， germination index， vigor index， relative germination rate， plant height， root length， aboveground and root dry weight， <italic>Pn</italic>， <italic>Gs， Tr</italic>， chlorophyll a/b ratio， chlorophyll and flavonoids contents and key enzyme activities for flavonoid biosynthesis like PAL while lowered <italic>Ci</italic> and MDA contents. The optimal concentration of EBR solution was 0.1 μmol·L^-1. Conclusion:Exogenous EBR solution at the suitable concentration ameliorates the inhibitory effect of 20% PEG stress against seed germination and seedling growth of <italic>A. membranaceus </italic>var.<italic> mongholicus</italic>， reduces the oxidative damage in leaves， and improves the stress resistance to a certain extent by up-regulating the key enzyme activities and promoting flavonoid synthesis.