Prediction of the 3D conformation of a small peptide vaccine targeting Aß42 oligomers.

The original etiology of Alzheimer's disease (AD) is the deposition of amyloid-beta (Aß) proteins, which starts from the aggregation of the Aß oligomers. The optimal therapeutic strategy targeting Aß oligomer aggregation is the development of AD vaccines. Despite the fact that positive progress has been made for experimental attempts at AD vaccines, the physicochemical and even structural properties of these AD vaccines remain unclear. In this study, through immunoinformatic and molecular dynamics (MD) simulations, we first designed and simulated an alternative of vaccine TAPAS and found that the structure of the alternative can reproduce the 3D conformation of TAPAS determined experimentally. Meanwhile, immunoinformatic methods were used to analyze the physicochemical properties of TAPAS, including immunogenicity, antigenicity, thermal stability, and solubility, which confirm well the efficacy and safety of the vaccine, and validate the scheme reliability of immunoinformatic and MD simulations in designing and simulating the TAPAS vaccine. Using the same scheme, we predicted the 3D conformation of the optimized ACI-24 peptide vaccine, an Aß peptide with the first 15 residues of Aß42 (Aß1-15). The vaccine was verified once to be effective against both full-length Aß1-42 and truncated Aß4-42 aggregates, but an experimental 3D structure was absent. We have also explored the immune mechanism of the vaccine at the molecular level and found that the optimized ACI-24 and its analogues can block the growth of either full-length Aß1-42 or truncated Aß4-42 pentamer by contacting the hydrophobic residues within the N-terminus and ß1 region on the contact surface of either pentamer. Additionally, residues (D1, D7, S8, H13, and Q15) were identified as the key residues of the vaccine to contact either of the two Aß oligomers. This work provides a feasible implementation scheme of immunoinformatic and MD simulations for the development of AD small peptide vaccines, validating the power of the scheme as a parallel tool to the experimental approaches and injecting molecular-level information into the understanding and design of anti-AD vaccines.

Specific interaction from different Aß42 peptide fragments to α7nAChR-A study of molecular dynamics simulation.

CONTEXT: Existing researches confirmed that ß amyloid (Aß) has a high affinity for the α7 nicotinic acetylcholine receptor (α7nAChR), associating closely to Alzheimer's disease. The majority of related studies focused on the experimental reports on the neuroprotective role of Aß fragment (Aßx), however, with a lack of investigation into the most suitable binding region and mechanism of action between Aß fragment and α7nAChR. In the study, we employed four Aß1-42 fragments Aßx, Aß1-16, Aß10-16, Aß12-28, and Aß30-42, of which the first three were confirmed to play neuroprotective roles upon directly binding, to interact with α7nAChR. METHODS: The protein-ligand docking server of CABS-DOCK was employed to obtain the α7nAChR-Aßx complexes. Only the top α7nAChR-Aßx complexes were used to perform all-atom GROMACS dynamics simulation in combination with Charmm36 force field, by which α7nAChR-Aßx interactions' dynamic behavior and specific locations of these different Aßx fragments were identified. MM-PBSA calculations were also done to estimate the binding free energies and the different contributions from the residues in the Aßx. Two distinct results for the first three and fourth Aßx fragments in binding site, strength, key residue, and orientation, account for why the fourth fails to play a neuroprotective role at the molecular level.

Aggregation Dynamics Characteristics of Seven Different Aß Oligomeric Isoforms-Dependence on the Interfacial Interaction.

The aggregation of ß-amyloid (Aß) peptides has been confirmed to be associated with the onset of Alzheimer's disease (AD). Among the three phases of Aß aggregation, the lag phase has been considered to be the best time for early Aß pathological deposition clinical intervention and prevention for potential patients with normal cognition. Aß peptide exists in various lengths in vivo, and Aß oligomer in the early lag phase is neurotoxic but polymorphous and metastable, depending on Aß length (isoform), molecular weight, and specific phase, and therefore hardly characterized experimentally. To cope with the problem, molecular dynamics simulation was used to investigate the aggregation process of five monomers for each of the seven common Aß isoforms during the lag phase. Results showed that Aß(1-40) and Aß(1-38) monomers aggregated faster than their truncated analogues Aß(4-40) and Aß(4-38), respectively. However, the aggregation rate of Aß(1-42) was slower than that of its truncated analogues Aß(4-42) rather than that of Aßpe(3-42). More importantly, Aß(1-38) is first predicted as more likely to form stable hexamer than the remaining five Aß isoforms, as Aß(1-42) does. It is hydrophobic interaction mainly (>50%) from the interfacial ß1 and ß2 regions of two reactants, pentamer and monomer, aggregated by Aß(1-38)/Aß(1-42) rather than by other Aß isoforms, that drives the hexamer stably as a result of the formation of the effective hydrophobic collapse. This paper provides new insights into the aggregation characteristics of Aß with different lengths and the conditions necessary for Aß to form oligomers with a high molecular weight in the early lag phase, revealing the dependence of Aß hexamer formation on the specific interfacial interaction.

Identification and characterization of the conformation and size of amyloid-ß (42) oligomers targeting the receptor LilrB2.

Experimental observations revealed that the amyloid-ß 42 oligomer (AßO) can directly bind to the LilrB2 D1D2(LDD) receptor with nanomolar-affinity, leading to changes in synaptic plasticity and cognitive deficits. However, the dependence of neurotoxicity on the morphology, size, and aggregation stage (SP1, SP2) of AßO, as well as the specific molecular mechanism of AßO-LDD interaction, remain uncertain. To address these uncertainties, we investigated the interaction between the LDD neuroreceptor and AßO with different Aß42 species (nontoxic species, toxic species, and protofibril) and sizes. Our results showed that the LDD selectively binds AßO species rather than the Aß42 monomer, accommodating various Aß42 dimers and trimers as well as SP2 AßO, in a specific pose in the pocket of the LDD receptor (region I). Additionally, protofibrils with exposed ß1/ß2 regions can also bind to region I of the LDD receptor, as observed experimentally (Cao, et al., Nat. Chem., 2018, 10, 1213; and Aim et al., Nat. Commun., 2021, 12, 3451). More extensively, we identified two additional regions of the LDD receptor, regions II and III, suitable for binding to larger AßO species at the SP1 with different molecular weights and conformations, accounting for the stronger binding strength obtained experimentally. We suggest that the two regions are more competitive than region I in causing toxicity by AßO binding. The detailed and systematic characterization for the complexes generated between the LDD receptor and various AßO species, including the protofibril, offers deep insight into the dependence of neurotoxicity on the AßO size and conformation at the molecular level, and provides novel and specific targets for drug design of Alzheimer's disease.

Origin of stronger binding of ionic pair (IP) inhibitor to Aß42 than the equimolar neutral counterparts: synergy mechanism of IP in disrupting Aß42 protofibril and inhibiting Aß42 aggregation under two pH conditions.

Fibrous aggregates of beta-amyloid (Aß) is a hallmark of Alzheimer's disease (AD). Several major strategies of drugs or inhibitors, including neutral molecules, positive or negative ions, and dual-inhibitor, are used to inhibit the misfolding or aggregation of Aß42, among which a kind of dual-inhibitor composed of a pair of positive and negative ions is emerging as the most powerful candidate. This knowledge lacks the origin of the strong inhibitory effect and synergy mechanisms blocking the development and application of such inhibitors. To this end, we employed 1 : 1 ionic pairs (IP) of oppositely charged benzothiazole molecules (+)BAM1-EG6 (Pos) and (-)BAM1-EG6 (Neg) as well as equimolar neutral BAM1-EG6 (Neu) counterpart at two pH conditions (5.5 and 7.0) to bind Aß42 targets, Aß42 monomer (AßM), soluble pentamer (AßP), and pentameric protofibril (AßF) models, respectively, corresponding to the products of three toxic Aß42 development pathways, lag, exponential and fibrillation phases. Simulated results illustrated the details of the inhibitory mechanisms of IP and Neu for the AßY (Y = M, P, or F) in the three different phases, characterizing the roles of Pos and Neg of IP as well as their charged, hydrophobic groups and linker playing in the synergistic interaction, and elucidated a previously unknown molecular mechanism governing the IP-Aß42 interaction. Most importantly, we first revealed the origin of the stronger binding of IP inhibitors to Aß42 than that of the equimolar neutral counterparts, observing a perplexing phenomenon that the physiological condition (pH = 7.0) than the acidic one (pH = 5.5) is more favorable to the enhancement of IP binding, and finally disclosed that solvation is responsible to the enhancement because at pH 7.0, AßP and AßF act as anionic membranes, where solvation plays a critical role in the chemoelectromechanics. The result not only provides a new dimension in dual-inhibitor/drug design and development but also a new perspective for uncovering charged protein disaggregation under IP-like inhibitors.

Toxicity Mechanism of Aß42 Oligomer in the Binding between the GABABR1a sushi1 Domain and Amyloid Precursor Protein 9mer: A Mechanism like Substitution Reaction.

Amyloid-ß peptide (Aß), characterized by its abnormal folding into neurotoxic aggregates, impairs synaptic plasticity and causes synaptic loss associated with Alzheimer's disease (AD). The neurotoxicity of Aß oligomers via the binding to various cell-surface receptors was frequently observed experimentally; however, the toxic mechanism still remains unknown. In this paper, we study the intervention of Aß oligomers to the receptor-peptide binding in the GABABR1a sushi1-APP 9mer complex, a key node in increasing short-term synaptic facilitation in the mouse hippocampus and decreasing neuronal activity by inhibiting neurotransmitter release by molecular dynamics simulations. The residue types of Aß42 oligomers involved in the intervention and core contact areas of the receptor were first identified, by which an unprecedented toxicity mechanism of Aß42 oligomers is proposed. These involved residues of Aß42 oligomers are positively charged residues Asp and Glu, and the core area of GABABR1a sushi1 domain is the Coil one, sharing the rich negatively charged residues R19/R21/R25/R45 with the pocket, in which APP 9mer is locked. The presence of an Aß42 oligomer rather than of a monomer stretches these key residues in the core area and consequently "unlocks and releases" the APP 9mer from its initial pocket, unsteadying the sushi1 domain and taking into toxic effect. It looks like a chemical "substitution" reaction, Aß42 oligomer + GABABR1a sushi1-APP 9mer complex → Aß42 oligomer-GABABR1a sushi1 complex + APP 9mer. Further analysis reveals that the toxicity of Aß42 oligomer to GABABR1a sushi1 domain stability depends on the residue number of the contact area and the size of Aß42 oligomer, in which semi-extended trimeric Aß42 oligomer is identified as the most toxic one. This work provides a novel insight into the mechanism of Aß oligomeric toxicity to neuroreceptors and sets an important precedent for dealing with Aß oligomeric toxicity to other receptors at the molecular level.

[The expression of interferon-stimulating gene (STING/TMEM173) in liver tissue and its correlation with liver inflammation in patients with chronic hepatitis B].

Objective To investigate the relationship between the expression and distribution of interferon-stimulating gene/transmembrane protein 173(STING/TMEM173) in liver tissue and the grade of liver inflammation in patients with chronic hepatitis B, and to explore the underlying mechanisms in vitro. Methods The expression of STING/TMEM173 protein in liver tissue of 62 naive patients with chronic hepatitis B was detected by immunohistochemistry. Rank sum test and spearman correlation coefficient were used to analyze the correlation between hepatic STING/TMEM173 expression and liver inflammation grades as well as serum ALT levels. After transient or stable transfection by HBV whole genome plasmid, the expression of STING/TMEM173 in HepG2 cells was determined by Western blot analysis. The peripheral blood mononuclear cells (PBMCs) were stimulated by supernatant of HepG2.2.15 cells containing intact HBV virions, and the expression STING/TMEM173 gene was detected by real-time PCR. Results The results of immunohistochemical showed that STING/TMEM173 protein was higher in liver tissues of CHB patients and mainly expressed in inflammatory cells of liver tissue, and the expression of STING/TMEM173 protein was positively correlated with liver inflammation grade as well as serum ALT level. After transient and stable transfection by HBV whole genome plasmid, the STING/TMEM173 protein decreased significantly in HepG2 cells. In addition, HepG2.2.15 cell supernatant containing intact HBV virions promoted the expression of STING/TMEM173 in PBMC in a dose-dependent manner at RNA level. Conclusion HBV can up-regulate the expression of STING/TMEM173 protein in inflammatory cells of liver tissue, and the number of liver inflammatory cells expressing STING/TMEM173 may reflect the severity of liver inflammation.

Image-Based Artificial Intelligence in Wound Assessment: A Systematic Review.

Significance: Accurately predicting wound healing trajectories is difficult for wound care clinicians due to the complex and dynamic processes involved in wound healing. Wound care teams capture images of wounds during clinical visits generating big datasets over time. Developing novel artificial intelligence (AI) systems can help clinicians diagnose, assess the effectiveness of therapy, and predict healing outcomes. Recent Advances: Rapid developments in computer processing have enabled the development of AI-based systems that can improve the diagnosis and effectiveness of therapy in various clinical specializations. In the past decade, we have witnessed AI revolutionizing all types of medical imaging like X-ray, ultrasound, computed tomography, magnetic resonance imaging, etc., but AI-based systems remain to be developed clinically and computationally for high-quality wound care that can result in better patient outcomes. Critical Issues: In the current standard of care, collecting wound images on every clinical visit, interpreting and archiving the data are cumbersome and time consuming. Commercial platforms are developed to capture images, perform wound measurements, and provide clinicians with a workflow for diagnosis, but AI-based systems are still in their infancy. This systematic review summarizes the breadth and depth of the most recent and relevant work in intelligent image-based data analysis and system developments for wound assessment. Future Directions: With increasing availabilities of massive data (wound images, wound-specific electronic health records, etc.) as well as powerful computing resources, AI-based digital platforms will play a significant role in delivering data-driven care to people suffering from debilitating chronic wounds.

Multiclass wound image classification using an ensemble deep CNN-based classifier.

Acute and chronic wounds are a challenge to healthcare systems around the world and affect many people's lives annually. Wound classification is a key step in wound diagnosis that would help clinicians to identify an optimal treatment procedure. Hence, having a high-performance classifier assists wound specialists to classify wound types with less financial and time costs. Different wound classification methods based on machine learning and deep learning have been proposed in the literature. In this study, we have developed an ensemble Deep Convolutional Neural Network-based classifier to categorize wound images into multiple classes including surgical, diabetic, and venous ulcers. The output classification scores of two classifiers (namely, patch-wise and image-wise) are fed into a Multilayer Perceptron to provide a superior classification performance. A 5-fold cross-validation approach is used to evaluate the proposed method. We obtained maximum and average classification accuracy values of 96.4% and 94.28% for binary and 91.9% and 87.7% for 3-class classification problems. The proposed classifier was compared with some common deep classifiers and showed significantly higher accuracy metrics. We also tested the proposed method on the Medetec wound image dataset, and the accuracy values of 91.2% and 82.9% were obtained for binary and 3-class classifications. The results show that our proposed method can be used effectively as a decision support system in classification of wound images or other related clinical applications.

Fully automatic wound segmentation with deep convolutional neural networks.

Acute and chronic wounds have varying etiologies and are an economic burden to healthcare systems around the world. The advanced wound care market is expected to exceed $22 billion by 2024. Wound care professionals rely heavily on images and image documentation for proper diagnosis and treatment. Unfortunately lack of expertise can lead to improper diagnosis of wound etiology and inaccurate wound management and documentation. Fully automatic segmentation of wound areas in natural images is an important part of the diagnosis and care protocol since it is crucial to measure the area of the wound and provide quantitative parameters in the treatment. Various deep learning models have gained success in image analysis including semantic segmentation. This manuscript proposes a novel convolutional framework based on MobileNetV2 and connected component labelling to segment wound regions from natural images. The advantage of this model is its lightweight and less compute-intensive architecture. The performance is not compromised and is comparable to deeper neural networks. We build an annotated wound image dataset consisting of 1109 foot ulcer images from 889 patients to train and test the deep learning models. We demonstrate the effectiveness and mobility of our method by conducting comprehensive experiments and analyses on various segmentation neural networks. The full implementation is available at https://github.com/uwm-bigdata/wound-segmentation .

Processing Induced Nonequilibrium Behavior of Polyvinylpyrrolidone Nanofilms Revealed by Dewetting.

Polyvinylpyrrolidone (PVP) nanofilms prepared by spin-coating have vast applications in biological and microdevice fields. However, detailed knowledge of processing induced nonequilibrium behavior of PVP nanofilms and solutions for minimizing residual stresses toward high-quality films has still been lacking. In the present study, we first explored the rapid film formation process via statistics on nascent holes. Next, by employing dewetting as a major probe, we revealed that many processing conditions, particularly previously overlooked variables like the atmosphere, substrates, and immersion time, were correlated substantially with the degree of nonequilibrium of nanofilms. Proper aging temperature and time were demonstrated essential for releasing residual stresses and achieving more equilibrium nanofilms. This work offered abundant experimental evidence in the building relationship between the processing and nonequilibrium nature of polymer nanofilms, which were crucial for their preparation and application.

[Antiviral effects of IL-27 on HBV in HepG2.2.15 cells via inducing MxA by STAT1 signal transduction pathway].

Objective To investigate the effect of interleukin 27 (IL-27) on hepatitis B virus (HBV) replication and antigen secretion in HepG2.2.15 cells and related mechanisms. Methods HepG2.2.15 cells were treated with IL-27 at various doses. The levels of HBsAg and HBeAg in the supernatant of the cell cultures were measured by ELISA, and the levels of HBV DNA was detected by real-time quantitative PCR. The expression and localization of HBV core antigen (HBcAg) in HepG2.2.15 cells were observed by immunocytochemical staining. Moreover, the expression of STAT1, phosphorylated STAT1 (p-STAT1) and myxovirus-resistance protein A (MxA) were analyzed by Western blotting. Results After the treatment with different doses of IL-27, the levels of HBV-DNA, HBsAg, HBeAg and intracellular HBcAg in HepG2.2.15 cells gradually decreased. With the increase of IL-27 dose, the expression level of p-STAT1 significantly increased. There was no significant change in STAT1 expression. Further studies also showed that IL-27 induced MxA expression in a dose- and time-dependent manner in HepG2.2.15 cells. Conclusion IL-27 could induces MxA expression by activating STAT signaling pathway to exert anti-HBV activity.

Effect of Liuweibuqi capsule, a Chinese patent medicine, on the JAK1/STAT3 pathway and MMP9/TIMP1 in a chronic obstructive pulmonary disease rat model.

OBJECTIVE: To observe effect of Liuweibuqi Capsule, a Traditional Chinese Medicine (TCM), on the janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway and matrix metalloproteinases (MMPs) in a chronic obstructive pulmonary disease (COPD) rat model with lung deficiency in terms of TCM's pattern differentiation. METHODS: Rats were randomly divided into a normal group, model group, Liuweibuqi group, Jinshuibao group, and spleen aminopeptidase group (n=10). Aside from the normal group, all rats were exposed to smoke plus lipopolysaccharide tracheal instillation to establish the COPD model with lung deficiency. Models were established after 28 days and then the normal and model groups were given normal saline (0.09 g/kg), Liuweibuqi group was given Liuweibuqi capsule (0.35 g/kg), Jinshuibao group was given Jinshuibao capsules (0.495 g/kg), and the spleen group was given spleen aminopeptidase (0.33 mg/kg), once a day for 30 days. Changes in symptoms, signs, and lung histology were observed. Lung function was measured with a spirometer. Serum cytokines were detected using enzyme-linked immunosorbent assay, and changes in the JAK/STAT pathway, MMP-9, and MMPs inhibitor 1 (TIMP1) were detected by immunohistochemistry, RT-PCR, and western blotting, respectively. RESULTS: Compared with the normal group, lung tissue was damaged, and lung function was reduced in the model control group. Additionally, the levels of interleukin (IL)-1beta, gamma interferon (IFN-gamma), and IL-6 were higher, while IL-4 and IL-10 were lower in the model control group than those in the normal group. The expressions of JAK1, STAT3, p-STAT3, and MMP-9 mRNA and protein in lung tissue were higher, and TIMP1 mRNA and protein was lower in the model group compared with the normal group. After treatment, compared with the model group, the expression of inflammatory cytokines was lower in each treatment group, and expressions of JAK/STAT pathway, MMPs were lower. Compared with the positive control groups, the Jinshuibao and spleen aminopeptidase groups, lung function was better, and JAK1, STAT3, and p-STAT3 protein were lower and TIMP1 was higher in the Liuweibuqi group. CONCLUSION: Liuweibuqi capsules can improve the symptoms of COPD possibly by regulating the expression of the JAK1/STAT3 pathway and MMP9/TIMP1.

[Role of Foxp3/Treg and RORγt/Th17 cell imbalance in rat model of chronic obstructive pulmonary disease].

OBJECTIVE: To observe the changes in forkhead/winged helix transcription factor p3(Foxp3), regulatory T cells (Treg), retinoid-related orphan receptor gamma (RORγt) in rat model of chronic obstructive pulmonary disease (COPD). METHODS: Twenty Sprague-Dawley (SD) rats were randomly divided into normal control group and COPD model group, with 10 rats in each group. The COPD model was reproduced by smoke inhalation and tracheal instillation of lipopolysaccharide (LPS), and no such treatment was conducted in normal control group. Twenty-eight days after the model reproduction, the pulmonary function was determined, the pathological changes of lung tissue were observed with haematoxylin-eosin (HE) staining, interleukins (IL-6, IL-10) in serum were detected by enzyme-linked immunosorbent assay (ELISA), CD4⁺ CD25⁺ Foxp3⁺ Treg of peripheral blood was determined by flow cytometry, and the expressions of Foxp3, RORγt, IL-17 protein in lung tissue were assayed by Western Blot. RESULTS: Under light microscope, significal interstitial infiltration of inflammatory cells was found in alveoli and interstitial tissue of the lung, and destruction of alveolar tissue, alveolar wall thinning, and even rupture to fuse into bullae, and bleeding into alveoli in different degress could be observed. Compared with the normal control group, forced vital capacity (FVC), forced expiratory volume in 0.3 second (FEV0.3), FEV0.3/FVC, peak expiratory flow (PEF) in model group were significantly decreased [FVC (mL): 8.04 ± 2.03 vs. 9.97 ± 2.14, FEV0.3 (mL): 6.16 ± 2.23 vs. 8.84 ± 2.12, FEV0.3/FVC: 0.70 ± 0.09 vs. 0.85 ± 0.11, PEF (mL/s): 33.56 ± 4.76 vs. 40.14 ± 5.64, P<0.05 or P<0.01]. Serum IL-6 was obviously increased (ng/L: 93.17 ±20.96 vs. 76.28 ± 13.24, P<0.05), IL-10 was significantly decreased (ng/L: 78.62 ± 15.17 vs. 104.34 ± 19.46, P<0.01), and CD4⁺ CD25⁺ FoxP3(+)Treg was significantly diminished [(2.75 ± 0.83)% vs. (4.16 ± 1.14)%, P<0.01] in model group compared with those in the normal control group. The expression of Foxp3 protein in lung tissue in model group was significantly down-regulated compared with that in the normal control group (gray scale: 0.38 ± 0.15 vs. 0.63 ± 0.11, P<0.01), and RORγt and IL-17 protein expressions were significantly up-regulated [RORγt (gray scale): 0.96 ± 0.23 vs. 0.47 ± 0.11, IL-17 (gray scale): 1.02 ± 0.24 vs. 0.34 ± 0.08, both P<0.01]. Correlation analysis showed that FEV0.3 was positively correlated with Foxp3 (r=0.585, P<0.05), and FEV0.3/FVC was negatively correlated with IL-6 and RORγt (r=-0.655, r=-0.607, both P<0.05). PEF was positively correlated with Treg (r=0.573, P<0.05), and negatively correlated with IL-17 (r=-0.198, P<0.05). IL-6 was negatively correlated with Foxp3(r=-0.603, P<0.05),and positively correlated with RORγt (r=0.588, P<0.05). IL-10 was positively correlated with Treg (r=0.573, P<0.05). Treg was positively correlated with Foxp3 (r=0.607, P<0.05), and negatively correlated with IL-17 (r=-0.569, P<0.05). Foxp3 was negatively correlated with RORγt (r=-0.591, P<0.05). RORγt was positively correlated with IL-17 (r=0.578, P<0.05). CONCLUSIONS: There is a relationship among decreased pulmonary function, inflammation and imbalance of Foxp3/Treg and RORγt/Th17 in COPD.

[Imbalance in the expression of Foxp3, T-bet and GATA-3 is correlated with inflammation of chronic obstructive pulmonary disease].

OBJECTIVE: To observe the change of regulatory T cells(Tregs), fork head-like transcription factor 3 (Foxp3), T box expressed in T cells (T-bet) and GATA binding protein 3 (GATA3) in rat models of chronic obstructive pulmonary disease (COPD). METHODS: Thirty rats were randomly divided into control group and model group (n=15 each). The rats of model group were developed by lipopolysaccharide (LPS) and smoking. Interleukin-4 (IL-4) and γ-interferon (IFN-γ) in serum and bronchoalveolar lavage fluid (BALF) were detected 28 days after modeling by ELISA. Peripheral Tregs were detected by flow cytometry. The expressions of Foxp3, T-bet, GATA gene and protein in lung tissue were observed by reverse transcription PCR and Western blotting, respectively. RESULTS: Compared with the control group, the model group had more serious inflammation in lung tissues, expressed the higher levels of IFN-γ in serum and BALF (P<0.01), T-bet mRNA and protein in lung tissue, and the lower levels of IL-4, CD4⁺ CD25⁺ Treg, CD4⁺ CD25⁺ Foxp3⁺ Treg (P<0.05), Foxp3, GATA-3 mRNA and protein (P<0.01). Correlation analysis showed that there were correlations between T-bet, GATA-3, Foxp3 expressions and IL-4, IFN-γ levels (P<0.05). CONCLUSION: There is a relationship between inflammation and imbalance of in the expression of T-bet, GATA-3, Foxp3 in COPD.

[Regulation of Thl/Th2 cells by T cell-mediated transcription factor in rats with chronic obstructive pulmonary disease].

OBJECTIVE: To determine T box expressed in T cells (T bet), GATA binding protein-3 (GATA 3), and retinoid-related orphan nuclear receptor gammat (RORgammat) in rats with chronic obstructive pulmonary disease (COPD). METHODS: Thirty rats were randomly divided into a control and a COPD group. The COPD model was established through smoking and lipopolysaccharide (LPS) tracheal instillation. Pulmonary function of the rats was measured 28 d after the establishment of the COPD model by a spirometer. Enzyme linked immunosorbent assay was performed to detect serum γ interferon (IFN-γ), interleukin (IL)-4, and IL-17. The expressions of T-bet, GATA-3, and RORgammat protein in lung tissues were determined by Western blot. RESULTS: Compared with the controls, the COPD rats had decreased pulmonary function and expression of serum IL-4, and increased INF-γ, IL- 17, Th1/Th2, T-bet, T bet /GATA-3, and RORgammat protein (P<0. 05). Forced expiratory volume in 0. 3 seconds (FEV 0.3) was negatively correlated with INF γ and T-bet/GATA-3. Forced vital capacity (FVC) was positively correlated with IL 4. FEV0.3/FVC was negatively correlated with Thl/Th2, T-bet and T-bet/GATA-3. Peak expiratory flow (PEF) was negatively correlated with IL-17, T bet, and RORgammat (P<0. 05). Thl/Th2 was positively correlated with T bet/GATA-3. IL-17 was positively correlated with RORgammat. T bet/GATA-3 was positively correlated with RORgammat (P<0. 05). CONCLUSION: Imbalanced regulation of T bet / GATA 3 and RORgammat on Th1/Th2 and Th17 cells is associated with the occurrence of COPD.