血清IL-13、IL-17、IL-37与慢性阻塞性肺疾病急性加重期无创机械通气患者预后不良的关系分析

摘要 目的：分析慢性阻塞性肺疾病急性加重期（AECOPD）无创机械通气患者血清白细胞介素（IL）-13、IL-17、IL-37的变化并探讨其与患者预后不良的关系。方法：选取2019年1月～2022年2月我院收治的102例接受无创机械通气治疗的AECOPD患者（AECOPD组）,根据预后情况分为预后不良亚组41例和预后良好亚组61例,另选取68例慢性阻塞性肺疾病稳定期（SCOPD）患者作为SCOPD组和57例体检健康者作为对照组。采用酶联免疫吸附法检测血清IL-13、IL-17、IL-37水平。采用单因素和多因素Logistic回归分析AECOPD无创机械通气患者预后不良的影响因素,受试者工作特征（ROC）曲线分析血清IL-13、IL-17、IL-37水平对AECOPD无创机械通气患者预后不良的预测价值。结果： 对照组、SCOPD组、AECOPD组血清IL-13、IL-17、IL-37水平依次升高（P＜0.05）。预后不良亚组血清IL-13、IL-17、IL-37水平高于预后良好亚组（P＜0.05）。单因素分析显示,预后不良亚组年龄大于预后良好亚组,心功能不全比例和COPD评估测试（CAT）评分、C反应蛋白高于预后良好亚组（P＜0.05）,而两组性别、体质指数、AECOPD病程、吸烟、糖尿病、高血压、冠心病、慢性肾功能不全、第1秒用力呼气容积/用力肺活量（FEV₁/FVC）、白细胞计数、降钙素原组间对比无差异（P＞0.05）。多因素Logistic回归分析显示,FEV₁％增加为AECOPD无创机械通气患者预后不良的独立保护因素,年龄增加、CAT评分增加和IL-13、IL-17、IL-37升高独立危险因素（P＜0.05）。ROC曲线分析显示,血清IL-13、IL-17、IL-37水平联合预测AECOPD无创机械通气患者预后不良的曲线下面积大于IL-13、IL-17、IL-37水平单独预测。结论：AECOPD无创机械通气患者血清IL-13、IL-17、IL-37水平升高与预后不良密切相关,血清IL-13、IL-17、IL-37可作为AECOPD无创机械通气患者预后评估的潜在生物学指标。     

Relationship between cytotoxicity and induction of sister-chromatid exchanges in mouse foetal cells exposed to several doses of carcinogenic and non-carcinogenic chemicals.

The increase in sister-chromatid exchanges induced by 5 chemicals, with different DNA damaging and carcinogenic activities, was studied in short-term foetal-mouse cultures. A significant increase in SCE was induced by N-methyl-N'-nitro-N-nitrosoguanidine, N-diazoacetylglycine-amide, azaserine and methotrexate. k-Strophantin, on the contrary, was totally inactive. On a molar basis, MNNG was the most active chemical followed by MTX, AZS and DGA, in that order. At equitoxic concentrations (D37), the order of SCE-inducing abilities was MNNG, DGA, AZS and MTX. Compared with previous data, at equitoxic concentrations, the most DNA-damaging agents were also the most effective in inducing SCE. The SCE increase seems to correlate not with unspecific cytotoxicity but more with DNA damage or other damage at the genome level. MTX, a non-mutagen, which induced SCE only at toxic levels, could be considered a false positive because this positivity may reflect an enhancement of incorporation of 5-BrdUrd into DNA. The positive results obtained with AZS suggest a sufficient sensitivity of the method for detecting relatively weak carcinogens.     

Methionine transport in S37 cells. Substrate-dependent function of amino acid transport system A in exchange processes

Methionine had been observed to interact with two principal transport systems for amino acids in mammalian cells, the A and L systems. The present study of methionine transport and of exchange processes through system A arose in the course of a study to define the specificity of a transinhibition effect caused by cysteine.Methionine uptake through two transport systems in the S37 cell was confirmed by the occurrence of a biphasic double-reciprocal plot for labeled methionine uptake. Preloading cells with methionine stimulated labeled histidine uptake through both systems A and L. Efflux of labeled methionine from cells was stimulated by histidine in a biphasic manner, so that both systems A and L can be used for exchange when methionine is the intracellular amino acid. Aminocycloheptanecarboxylic acid elicited exchange efflux of labeled methionine only through system L. α-Aminoisobutyric acid and $\text{N-}\text{methyl}\text{-α-}\text{aminoisobutyric acid}$ both stimulated efflux of labeled $\text{N-}\text{methyl}\text{-α-}\text{aminoisobutyric acid}$ from S37 cells. These findings are interpreted a showing that transport system A is capable of functioning as an exchange system depending upon the identity of intracellular and extracellular substrates available.     

Preparation of Developing and Adult Drosophila Brains and Retinae for Live Imaging

The Drosophila brain and visual system are widely utilized model systems to study neuronal development, function and degeneration. Here we show three preparations of the brain and visual system that cover the range from the developing eye disc-brain complex in the developing pupae to individual eye and brain dissection from adult flies. All protocols are optimized for the live culture of the preparations. However, we also present the conditions for fixed tissue immunohistochemistry where applicable. Finally, we show live imaging conditions for these preparations using conventional and resonant 4D confocal live imaging in a perfusion chamber. Together, these protocols provide a basis for live imaging on different time scales ranging from functional intracellular assays on the scale of minutes to developmental or degenerative processes on the scale of many hours.     

Platyphylloside,a potential inhibitor from epicarp of B. aegyptiaca against CYP450 protein in T. rubrum – In vitro and in silico approaches

Trichophyton rubrum is one of the major disease causing pathogens in human; mainly it causes tinea pedis, tinea cruris and tinea corporis. Cytochrome P450 which considered to be an important protein that can impact ergosterol biosynthesis pathway. B. aegyptiaca is rich source of secondary metabolites with tremendous medicinal values and it has sweet pulp, leaves with spine, strong seed and oily kernel. The epicarp of the fruit was taken for this study to inhibit T. rubrum using in vitro and in silico techniques. The epicarp portion was extracted using various solvents and water. The anti-dermatophytic activity on T. rubrum of these extracts was assessed utilizing poison plate technique with 5 individual concentrations. The fractioned chloroform extract of epicarp had fully inhibited the growth of T. rubrum at 3 mg/ml. Further, the chloroform extract was subjected to LC-MS analysis, in total, 40 compounds were elucidated. Then, the derived compounds were included for predicting ADMETox properties using Qikprop module. From the analysis 40 compounds were identified to be eligible for docking process. Then the desirable compounds, drug Ketoconazole were subjected to docking analysis using Glide module of Schrödinger. It shows that Platyphylloside has better docking result than other compounds and drug Ketoconazole. Further, MD simulation was carried out for Ketoconazole-Cyp450 and Platyphylloside-CYP450 complexes using Desmond, Schrödinger. MD simulation study also confirmed that the Platyphylloside-CYP450 complex more stable. This study suggests that Platyphylloside may act as potential inhibitor and it could be further subjected to experimental analysis to inhibit the T. rubrum growth.     

Exposure of protein kinase motifs that trigger binding of Hsp90 and Cdc37

Hsp90 and its co-chaperone Cdc37 are required for the activity of numerous eukaryotic protein kinases. c-Jun N-terminal kinases (JNKs) appear to be Hsp90-independent kinases, as their activity is unaffected by Hsp90 inhibition. It is currently unknown why some protein kinases are Hsp90- and Cdc37-dependent for their function, while others are not. Therefore, we investigated what structural motifs within JNKs confer or defer Hsp90 and Cdc37 interaction. Both Hsp90 and Cdc37 recognized structural features that were exposed or destabilized upon deletion of JNK1alpha1's N-terminal non-catalytic structural motif, while only Hsp90 bound JNK when its C-terminal non-catalytic structural motif was deleted. Mutations in JNK's activation loop that are known to constitutively activate or inactivate its kinase activity had no effect on JNK's lack of interaction with Hsp90 and Cdc37. Our findings suggest a model in which Hsp90 and Cdc37 each recognize distinct features within the catalytic domains of kinases.     

Differential effects of Hsp90 inhibition on protein kinases regulating signal transduction pathways required for myoblast differentiation

As derivatives of the Hsp90-inhibitor and tumoricidal agent geldanamycin move into phase II clinical trials, its potential for triggering adverse effects in non-tumor cell populations requires closer examination. In this report, the effect of geldanamycin on the differentiation and survival of C2C12 myoblasts was investigated. Treatment of differentiating C2C12 myoblasts with geldanamycin blocked myogenin expression, inhibited myotubule formation, and led to the depletion of three Hsp90-dependent protein kinases, ErbB2, Fyn, and Akt, and induction of apoptosis. ErbB2 levels declined rapidly, while Fyn and Akt levels decreased at a slower rate. Geldanamycin blocked the interaction of Hsp90 and its "kinase-specific" co-chaperone Cdc37 with Fyn, indicating that Fyn is an Hsp90-dependent kinase. Pulse-chase experiments indicated that geldanamycin caused newly synthesized Akt and Fyn to be degraded rapidly, but geldanamycin had little effect on the turnover rate of mature Fyn and Akt. Curiously, total cellular Src (c-Src) protein levels and the turnover rate of newly synthesized c-Src were unaffected by geldanamycin. While, geldanamycin had no effect on the levels of the putative Hsp90 client protein MyoD expressed in C2C12 cells, geldanamycin disrupted the interaction of Cdc37 with MyoD. Thus, inhibition of Hsp90 caused C2C12 cells to become depleted of multiple signal transduction proteins whose functions are essential for myoblast differentiation, and muscle cell survival, suggesting that geldanamycin derivatives may have the prospective of adversely affecting the physiology of certain sensitive muscle cell populations in vivo.     

侧脑室注射肾上腺髓质素对大鼠脑内心血管相关核团儿茶酚胺神经元及c-fos表达的影响

目的和方法:利用Fos蛋白和酪氨酸羟化酶(TH)的双重免疫组化方法,观察侧脑室注射肾上腺髓质素对大鼠心血管相关核团中儿茶酚胺神经元及c fos表达的影响,以探讨肾上腺髓质素的中枢效应是否通过激活脑内儿茶酚胺能神经元而诱发。结果:①侧脑室注射肾上腺髓质素(3nmol/kg)诱发脑干、下丘脑及前脑等多个部位的心血管中枢出现大量Fos样免疫反应神经元。②侧脑室注射肾上腺髓质素引起最后区(AP)、孤束核(NTS)、巨细胞旁外侧核(PGL)和蓝斑核(LC)内Fos TH双标神经元明显增加。③降钙素基因相关肽受体拮抗剂CGRP8-37(30nmol/kg)可明显减弱肾上腺髓质素的效应。结论:肾上腺髓质素可兴奋脑干、下丘脑及前脑等多个部位心血管相关核团的神经元,其中枢效应通过激活儿茶酚胺能神经元而诱发,降钙素基因相关肽受体介导这一效应。     

Iron and iron chelating agents modulate Mycobacterium tuberculosis growth and monocyte-macrophage viability and effector functions

Excess of iron promotes Mycobacterium tuberculosis infection, its replication and progression to clinical disease and death from tuberculosis. Chelation of iron may reduce M. tuberculosis replication, restore host defence mechanisms and it could constitute an application in the prevention and treatment strategies where both iron overload and tuberculosis are prevalent. We investigated the effect of iron and iron chelating agents, like desferrioxamine and silybin, individually and in combination with iron on mycobacterial number, viability in culture and after recovery from monocyte-macrophages, together with monocyte-macrophages viability and oxidative defence. Mycobacterial number and viability in culture were assessed using real-time quantitative PCR of H37Rv IS6110 DNA, 16S rRNA and 85B mRNA, whereas the microplate AlamarBlue(TM) assay was used to detect viability in culture post-infection. Mitochondrial membrane potential and phosphatidyl serine exposure of monocyte-macrophages, detected using Mitotracker Red fluorescence and Annexin V binding, respectively, served as indicators of host cell viability. Superoxide generation served as marker of monocyte-macrophage effector functions. Extracellular H37Rv showed a significant increase in number and viability in presence of excess iron and, by large, a significant decrease in number and viability in presence of the iron chelating agents, silybin and desferrioxamine, compared to cultivation without supplementation. Intracellularly, excess iron increased H37Rv viability significantly but reduced monocyte-macrophages mitochondrial membrane potential and compromised superoxide production. Desferrioxamine had little influence on intracellular parameters, but consistently prevented effects of excess iron, while silybin significantly altered most intracellular parameters and mostly failed to prevent effects of excess iron. These findings suggest that chelation therapy should be considered in conditions of iron overload and that effective chelating agents like desferrioxamine, with limited intracellular access might need to be used in combination with lypophilic chelating agents.