目的:在之前的研究中,我们报道了在高压给药无菌盐水等渗溶液(HPPpSIS)治疗的患者中,鼻神经生长因子(NGF)的升高.在这里,我们根据先天免疫反应和细胞因子特征来表征鼻粘膜,包括抗病毒特性。还讨论了HppSIS的潜在NGF和抗病毒益处。
方法:二十(20)名患者(11名男性,9名女性;年龄范围30-75岁)接受了HPPpSIS,并在治疗前后收集了鼻腔样本。鼻刮片用于形态学(涂片和QuickMay-GrunwaldGiemsa染色,MGG),生物化学(组胺,血清素;ELISA)和分子(信使RNA,mRNA)分析。Toll样受体(TLR)3(TLR3)特异性转录本的扩增,TLR7,TLR9,白细胞介素-(IL)18(IL18),IL13,IL12,嗜酸性粒细胞衍生的神经毒素(EDN),嗜酸性粒细胞阳离子蛋白(ECP),γ干扰素(γIFN),使用两步实时逆转录聚合酶链反应(RT-PCR)进行类胰蛋白酶和5-羟色胺。对临床和实验室数据进行分析比较。
结果:临床评估显示我们的治疗具有保护作用。涂片显示存在白细胞,嗜酸性粒细胞(EOS)和肥大细胞(MC),和增加的免疫反应性ECP/RNase3和EDN后HPSIS。ELISA显示血清素和EDN水平升高,P物质(SP)和组胺水平不变。通过原位荧光分析证实了嗜酸性粒细胞衍生的神经毒素(EDN)水平的增加。HPSIS诱导TLR3、TLR7和TLR9转录物上调,虽然没有观察到细胞间粘附分子1(ICAM1)的变化,IL18、白细胞介素-15(IL15)和IL12转录物,白细胞介素-6(IL6)和IL13也是如此。γIFN和EDN/RNase2转录本也没有观察到变化,而ECP/RNase3转录本在HPSIS后显著上调。最后,HppSIS后,类胰蛋白酶转录本没有变化,而5-羟色胺转录本明显增加。
结论:由于HpSS治疗而在鼻粘膜观察到的临床和生物分子变化表明先天性监测的激活,通过TLR转录,和可能的抗病毒反应,由于EDN上调。还有待验证,如果NGF,已知在HpSIS治疗后局部释放,可能部分负责此本地激活。
In a previous study, we reported an increase of nasal nerve growth factor (NGF) in patients treated with high-pressure administration of sterile saline isotonic solution (HPpSIS). Herein we characterized the nasal mucosa in terms of innate immune response and cytokine signature, including antiviral properties. Potential NGF and antiviral benefits of HPpSIS were also discussed.
Twenty (20) patients (11 males, 9 females; age range 30-75 years old) underwent HPpSIS and nasal samples were collected before and after treatment. Nasal scraping was used for morphological (smears and Quick May-Grunwald Giemsa staining, MGG), biochemical (Histamine, Serotonin; ELISA) and molecular (messenger RNA, mRNA) analyses. Amplification of transcripts specific for Toll-like receptor (TLR) 3 (TLR3), TLR7, TLR9, Interleukin-(IL) 18 (IL18), IL13, IL12, eosinophil-derived neurotoxin (EDN), Eosinophil Cationic Protein (ECP), γ Interferon (γIFN), tryptase and serotonin was performed using the 2-step real-time Reverse Transcription Polymerase Chain Reaction (RT-PCR). Clinical and laboratory data were analyzed and compared.
The clinical evaluation showed a protective effect of our therapy. Smears showed the presence of leucocytes, eosinophils (EOs) and mast cells (MCs), and increased immunoreactivity for ECP/RNase3 and EDN after HPpSIS. ELISA showed increased levels of Serotonin and EDN associated with unchanged levels of substance P(SP) and histamine. Increased eosinophil-derived neurotoxin eosinophil-derived neurotoxin (EDN) levels were confirmed by in situ fluorescent analysis. HPpSIS induced the upregulation of TLR3, TLR7 and TLR9 transcripts, while no changes were observed for Intercellular Adhesion Molecule 1 (ICAM1), IL18, Interleukin-15 (IL15) and IL12 transcripts nor for Interleukin-6 (IL6) and IL13. No changes were also observed for γIFN and EDN/RNase2 transcripts, while ECP/RNase3 transcripts were significantly upregulated after HPpSIS. Finally, tryptase transcripts were unchanged while serotonin transcripts were significantly increased after HPpSIS.
The clinical and biomolecular changes observed at the nasal mucosa due to HpSS treatment suggest the activation of an innate surveillance, by means of TLR transcription, and a possible anti-viral response due to EDN upregulation. It remains to be verified if NGF, known to be released locally upon HpSIS treatment, might in part be responsible for this local activation.