Asthma prediction via affinity graph enhanced classifier: a machine learning approach based on routine blood biomarkers.

BACKGROUND: Asthma is a chronic respiratory disease affecting millions of people worldwide, but early detection can be challenging due to the time-consuming nature of the traditional technique. Machine learning has shown great potential in the prompt prediction of asthma. However, because of the inherent complexity of asthma-related patterns, current models often fail to capture the correlation between data samples, limiting their accuracy. Our objective was to use our novel model to address the above problem via an Affinity Graph Enhanced Classifier (AGEC) to improve predictive accuracy. METHODS: The clinical dataset used in this study consisted of 152 samples, where 24 routine blood markers were extracted as features to participate in the classification due to their ease of sourcing and relevance to asthma. Specifically, our model begins by constructing a projection matrix to reduce the dimensionality of the feature space while preserving the most discriminative features. Simultaneously, an affinity graph is learned through the resulting subspace to capture the internal relationship between samples better. Leveraging domain knowledge from the affinity graph, a new classifier (AGEC) is introduced for asthma prediction. AGEC's performance was compared with five state-of-the-art predictive models. RESULTS: Experimental findings reveal the superior predictive capabilities of AGEC in asthma prediction. AGEC achieved an accuracy of 72.50%, surpassing FWAdaBoost (61.02%), MLFE (60.98%), SVR (64.01%), SVM (69.80%) and ERM (68.40%). These results provide evidence that capturing the correlation between samples can enhance the accuracy of asthma prediction. Moreover, the obtained [Formula: see text] values also suggest that the differences between our model and other models are statistically significant, and the effect of our model does not exist by chance. CONCLUSION: As observed from the experimental results, advanced statistical machine learning approaches such as AGEC can enable accurate diagnosis of asthma. This finding holds promising implications for improving asthma management.

One panel with four single nucleotide polymorphisms for Chinese children with asthma: Integrating public data and whole exome sequencing.

BACKGROUND: Polymorphisms in susceptibility genes are a major risk factor for the development of asthma. Understanding these genetic variants helps elucidate asthma's pathogenesis, predict its onset, expedite antiasthma medication development, and achieve precise targeted individualized treatment. This study developed a test kit based on susceptibility genes for predicting asthma in Chinese children. METHODS: The present study constructed a VariantPro Targeted Library Preparation System with 72 single nucleotide polymorphism (SNP) loci associated with asthma from the ClinVar, OMIM, and SNPedia databases. These SNP loci were detected in the peripheral blood of 499 children with asthma and 500 healthy children. Significant differences were discovered for seven SNP loci. Simultaneously, whole exome sequencing of 46 children with asthma and 50 healthy children identified eight SNP loci with significant differences. The 15 SNP loci identified from Chinese children with asthma were validated in an independent population of 97 children with asthma and 93 healthy children by conducting multiplex polymerase chain reaction (PCR)-next-generation sequencing genotyping. RESULTS: Four loci (rs12422149, rs7216389, rs4065275, and rs41453444) were identified, and a single-tube multifluorescent qPCR (real-time quantitative PCR) test kit was developed using these four SNP loci. The kit was tested on 269 children with asthma and 724 children with bronchopneumonia. CONCLUSIONS: We identified four loci as susceptibility genes and developed a quantitative PCR test kit for predicting asthma development in Chinese children.

Identification and validation of biomarkers related to lipid metabolism in osteoarthritis based on machine learning algorithms.

BACKGROUND: Osteoarthritis and lipid metabolism are strongly associated, although the precise targets and regulatory mechanisms are unknown. METHODS: Osteoarthritis gene expression profiles were acquired from the GEO database, while lipid metabolism-related genes (LMRGs) were sourced from the MigSB database. An intersection was conducted between these datasets to extract gene expression for subsequent differential analysis. Following this, functional analyses were performed on the differentially expressed genes (DEGs). Subsequently, machine learning was applied to identify hub genes associated with lipid metabolism in osteoarthritis. Immune-infiltration analysis was performed using CIBERSORT, and external datasets were employed to validate the expression of these hub genes. RESULTS: Nine DEGs associated with lipid metabolism in osteoarthritis were identified. UGCG and ESYT1, which are hub genes involved in lipid metabolism in osteoarthritis, were identified through the utilization of three machine learning algorithms. Analysis of the validation dataset revealed downregulation of UGCG in the experimental group compared to the normal group and upregulation of ESYT1 in the experimental group compared to the normal group. CONCLUSIONS: UGCG and ESYT1 were considered as hub LMRGs in the development of osteoarthritis, which were regarded as candidate diagnostic markers. The effects are worth expected in the early diagnosis and treatment of osteoarthritis.

Comparative Genomics Reveals Insights into the Divergent Evolution of Astigmatic Mites and Household Pest Adaptations.

Highly diversified astigmatic mites comprise many medically important human household pests such as house dust mites causing ∼1-2% of all allergic diseases globally; however, their evolutionary origin and diverse lifestyles including reversible parasitism have not been illustrated at the genomic level, which hampers allergy prevention and our exploration of these household pests. Using six high-quality assembled and annotated genomes, this study not only refuted the monophyly of mites and ticks, but also thoroughly explored the divergence of Acariformes and the diversification of astigmatic mites. In monophyletic Acariformes, Prostigmata known as notorious plant pests first evolved, and then rapidly evolving Astigmata diverged from soil oribatid mites. Within astigmatic mites, a wide range of gene families rapidly expanded via tandem gene duplications, including ionotropic glutamate receptors, triacylglycerol lipases, serine proteases and UDP glucuronosyltransferases. Gene diversification after tandem duplications provides many genetic resources for adaptation to sensing environmental signals, digestion, and detoxification in rapidly changing household environments. Many gene decay events only occurred in the skin-burrowing parasitic mite Sarcoptes scabiei. Throughout the evolution of Acariformes, massive horizontal gene transfer events occurred in gene families such as UDP glucuronosyltransferases and several important fungal cell wall lytic enzymes, which enable detoxification and digestive functions and provide perfect drug targets for pest control. This comparative study sheds light on the divergent evolution and quick adaptation to human household environments of astigmatic mites and provides insights into the genetic adaptations and even control of human household pests.

Dynamic changes of serum metabolites associated with infection and severity of patients with acute hepatitis E infection.

Dynamic changes in metabolites may affect liver disease progression, and provide new methods for predicting liver damage. We used ultra-performance liquid chromatography-mass spectroscopy to assess serum metabolites in healthy controls (HC), and patients with acute hepatitis E (AHE) or hepatitis E virus acute liver failure (HEV-ALF). The principal component analysis, partial least squares discriminant analysis, and discriminant analysis of orthogonal projections to latent structures models illustrated significant differences in the metabolite components between AHE patients and HCs, or between HEV-ALF and AHE patients. In pathway enrichment analysis, we further identified two altered pathways, including linoleic acid metabolism and phenylalanine, tyrosine, and tryptophan biosynthesis, when comparing AHE patients with HCs. Linoleic acid metabolism and porphyrin and chlorophyll metabolism pathways were significantly different in HEV-ALF when compared with AHE patients. The discriminative performances of differential metabolites showed that taurocholic acid, glycocholic acid, glycochenodeoxycholate-3-sulfate, and docosahexaenoic acid could be used to distinguish HEV-ALF from AHE patients. The serum levels of glycocholic acid, taurocholic acid, deoxycholic acid glycine conjugate, and docosahexaenoic acid were associated with the prognosis of HEV-ALF patients. Dynamic changes in serum metabolites were associated with AHE infection and severity. The identified metabolites can be used to diagnose and predict the prognosis of HEV-ALF.

Regulatory roles of three miRNAs on allergen mRNA expression in Tyrophagus putrescentiae.

BACKGROUND: Tyrophagus putresecentiae is an important mite species in rural and urban environments, causing sensitization and allergic disease. While evidence suggests that microRNAs (miRNAs) may regulate the expression of allergen-encoding genes, no study has directly investigated this possibility. Here, this gap was addressed by profiling miRNAs and elucidating their target allergen messenger RNAs (mRNAs) in this mite species. METHODS: Small RNA and transcriptome libraries were constructed for eggs, larvae, nymphs, and adults. After deep miRNA and whole-transcriptome sequencing were performed, the miRNA and allergen-encoding mRNA regulatory networks were explored. RESULTS: A total of 540 miRNAs were identified, including 155 with expression levels differing significantly across the four mite developmental stages (p < .01), 59 of which were novel. The mRNA expression for allergens was higher for Tyr p 1 in adults than in other developmental stages; Tyr p 2-5, 7, 10, 13, 33, and 34 in immature stages; and Tyr p 28, 35, and 36 in eggs and adults. A combined miRNA and transcriptome bioinformatics analysis showed that allergen Tyr p 3 was regulated by miRNA PC-5p-5698441_1, Tyr p 4 was regulated by PC-5p-7050653_1, and Tyr p 34 was regulated by PC-5p-5534223_1 and PC-5p-5698441_1. These three allergen mRNA and three miRNAs were identified using qRT-PCR, and their regulatory roles were confirmed by double-fluorescent reporter gene system and site-directed mutagenesis technology. CONCLUSIONS: For the first time, allergen mRNA expression and miRNAs were profiled throughout the life cycle for an allergen-producing mite, and the results showed that miRNAs bind to target allergen mRNAs to regulate their expression.

Dermatophagoides pteronyssinus allergen Der p 22: Cloning, expression, IgE-binding in asthmatic children, and immunogenicity.

BACKGROUND: Dust mite extract contains multiple components that, while useful in clinical allergy diagnosis and treatment, can cause serious side effects. Defining components of dust mite extract is important their contributions to allergic disease. This study aimed to characterize a novel dust mite allergen, Der p 22. METHODS: We amplified the cDNA encoding Der p 22 from total RNA of the mite Dermatophagoides pteronyssinus, and inserted it into an expression construct for transformation to competent cells. Purified recombinant (r) Der p 22 was tested for IgE-binding reactivity in sera obtained from children with allergic asthma by the Affiliated Wuxi Children's Hospital of Nanjing Medical University (Jiangsu, China). rDer p 22 also was used to challenge BALB/c mice to assess effects on T helper cells and cytokine levels and applied to cultured lung epithelial cells to evaluate apoptosis and cytokine secretion. RESULTS: rDer p 22 bound to IgE in 93.75% of sera from pediatric allergic asthma patients. Mice challenged with rDer p 22 had altered Th1/Th2 ratios in spleen and lymph, and lower levels of cytokines IFN-γ but higher levels of IL-4 and IL-10 in alveolar lavage fluid compared with controls (p < .05). Cultured lung epithelial cells had greater apoptosis rates and exhibited higher levels of IL-6, IL-8, and GM-CSF when treated with rDer p 22 compared with control treatment (p < .05). CONCLUSIONS: Recombinant Der p 22 exhibited high IgE-binding rates in allergic children, indicating the activity of the recombinant protein and suggesting this novel allergen may be appropriate for inclusion in an allergy diagnostic workup. This finding is supported by in vitro and mouse in vivo studies showing rDer p 22 induced strong allergenic reactivity and apoptosis.

Microfluidic Point-of-Care (POC) Devices in Early Diagnosis: A Review of Opportunities and Challenges.

The early diagnosis of infectious diseases is critical because it can greatly increase recovery rates and prevent the spread of diseases such as COVID-19; however, in many areas with insufficient medical facilities, the timely detection of diseases is challenging. Conventional medical testing methods require specialized laboratory equipment and well-trained operators, limiting the applicability of these tests. Microfluidic point-of-care (POC) equipment can rapidly detect diseases at low cost. This technology could be used to detect diseases in underdeveloped areas to reduce the effects of disease and improve quality of life in these areas. This review details microfluidic POC equipment and its applications. First, the concept of microfluidic POC devices is discussed. We then describe applications of microfluidic POC devices for infectious diseases, cardiovascular diseases, tumors (cancer), and chronic diseases, and discuss the future incorporation of microfluidic POC devices into applications such as wearable devices and telemedicine. Finally, the review concludes by analyzing the present state of the microfluidic field, and suggestions are made. This review is intended to call attention to the status of disease treatment in underdeveloped areas and to encourage the researchers of microfluidics to develop standards for these devices.

Diagnostic value of YKL-40 for patients with asthma: A meta-analysis.

Objective: YKL-40 is a highly conserved and chitin-bound human glycoprotein in mammals that is associated with airway inflammation and has no enzyme activity. We aimed to conduct a meta-analysis to assess the use of YKL-40 levels as a diagnosis of asthma. Methods: A meta-analysis was conducted based on the data from medical literature data base searches with time restrictions of January 2007 to January 2021. We searched and extracted relevant information from a total of 15 studies that reported YKL-40 levels in patients with asthma and in healthy controls, and obtained a sample of 1647 patients with asthma and 1259 healthy controls. Standardized mean differences (SMD) with 95% confidence intervals (CI) were calculated for this study by using statistical software packages. Results: Serum YKL-40 levels were higher in the patients with asthma than in the healthy controls (SMD 1.36 ng/ml [95% CI, 0.82-1.89 ng/ml]). YKL-40 levels are elevated in pediatric patients with asthma (SMD 2.26 ng/ml [95% CI, 1.33-3.18 ng/ml]) and in adult patients with asthma (SMD 0.96 ng/ml [95% CI, 0.26-1.66 ng/ml]). In addition, a subgroup analysis of asthma disease status showed that YKL-40 levels were significantly increased in the patients with stable asthma (SMD 1.69 ng/ml [95% CI, 0.81-2.56 ng/ml]) and in those with acute exacerbation asthma (SMD 3.31 ng/ml [95% CI, 2.04-4.58 ng/ml]), and serum YKL-40 levels were significantly higher in patients with acute exacerbation asthma than in patients with stable asthma (SMD 1.49 ng/ml [95% CI, 0.50-2.48 ng/ml]). Conclusion: Results of this meta-analysis suggested that increased serum levels of YKL-40 in patients with asthma could be used as an emerging indicator for distinguishing individuals with asthma from healthy individuals.

Evodiamine protects against airway remodelling and inflammation in asthmatic rats by modulating the HMGB1/NF-κB/TLR-4 signalling pathway.

CONTEXT: Evodiamine, which is isolated from Evodia rutaecarpa (Rutaceae), possess strong anti-inflammatory, immunomodulatory, and antibacterial properties. OBJECTIVE: The protective effects of evodiamine in asthma were evaluated. MATERIALS AND METHODS: Thirty-two Sprague-Dawley (SD) rats were used, asthma was induced by injecting intraperitoneally with a mixture of Al(OH)3 (100 mg) and ovalbumin (OA; 1 mg/kg), further exposing them to a 2% OA aerosol for 1 week. All animals were divided into four groups: control, asthma, and evodiamine 40 and 80 mg/kg p.o. treated group. Serum levels of inflammatory cytokines, interferon gamma (IFN-γ), and immunoglobulin E (IgE) and infiltrations of inflammatory cells in the bronchoalveolar lavage fluid (BALF) of the animals were determined. The thickness of the smooth muscle layer and airway wall in the intact small bronchioles of asthmatic rats was examined as well. RESULTS: Cytokine levels in the serum and BALF were lower in the evodiamine-treated group than in the asthma group. Evodiamine treatment reduced IgE and IFN-γ levels as well as the inflammatory cell infiltrate in the lung tissue of asthmatic rats. The thickness of the smooth muscle layer and airway wall of intact small bronchioles was less in the evodiamine-treated group than in the asthma group. Lower levels of TLR-4, MyD88, NF-κB, and HMGB1 mRNA in lung tissue were measured in the evodiamine-treated group than in the asthma group. DISCUSSION AND CONCLUSION: The effect of evodiamine treatment protects the asthma, as evodiamine reduces airway inflammation and remodelling in the lung tissue by downregulating the HMGB1/NF-κB/TLR-4 pathway in asthma.

Identification of continuous immunoglobulin G epitopes of Dermatophagoides farinae allergens by peptide microarray immunoassay.

Dermatophagoides farinae, as a common house dust mite species, is one of the main sources of allergens in the world. At present, Dermatophagoides farinae is found to contain more than 30 groups of allergens. These allergens are used for allergen-specific immunotherapy (AIT) of allergic diseases. During the AIT process, immunoglobulin G (IgG) antibodies can block immunoglobulin E (IgE) antibody-induced allergic reactions in the human body. One of the mechanisms may be that IgG and IgE competitively bind to the same allergic protein, so it is necessary to explore the binding sites (epitopes) of IgG antibodies to allergens. In this study, peptide arrays were constructed to react with the serums from patients with allergic asthma to find the IgG epitopes of several allergens including major allergens (Der f 1, 2) and mid-tier allergens (Der f 4, 5, and 7), and then verified by enzyme-linked immunosorbent assay (ELISA) test. Relevant epitopic sequences were located on the tertiary structure of individual allergens, as reconstructed by homology modeling. One IgG epitope of Der f 1 (90-106aa, NVPSELDLRSLRTVTPI), five IgG epitopes of Der f 4 (61-77aa, ERYQPVSYDIHTRSGDE; 193-209aa, FRSDASTHQWPDDLRSI; 226-242aa, HPFIYHETIYYGGNGIN; 271-287aa, LRWLRNFGTEWGLVPSG; 352-368aa, NDWVGPPTDQHGNILSV), and one IgG epitope of Der f 5 (84-101aa, RYNVEIALKSNEILERDL) were identified. IgG epitopes of Der f 2, 7 were not found. There are overlaps between the IgG and IgE epitopes of Der f 1, 4, and 5. These findings not only reflect the practicality of peptide array and ELISA test in the allergen IgG epitope identification, but also provide more information for further understanding of the human immunological changes during AIT and the molecular mechanisms of IgG blocking IgE activity.

Identification of LincRNA from Dermatophagoides farinae (Acari: Pyroglyphidae) for Potential Allergen-Related Targets.

Long noncoding RNAs (lncRNAs), especially their important subclass of long intergenic noncoding RNAs (lincRNAs), have been identified in some insects. They play important roles in the regulation of biological processes, such as immune response or cell differentiation and as possible evolutionary precursors for protein coding genes. House dust mites (HDMs) are recognized as allergenic mites because allergens are found in their feces and bodies. Dermatophagoides farinae is one of the most important pyroglyphid mites because of its abundance in the household. To determine if lincRNAs can regulate allergen presentation in HDMs, we analyzed RNA-seq data for HDMs. We identified 11 lincRNAs that are related to mRNAs coding for allergens in HDMs. Using qRT-PCR, we amplified 10 lincRNAs and their putative target allergen-encoding mRNAs, confirming expression of these lincRNAs and allergen genes. The results suggest that lincRNAs might be involved in the regulation of allergen production in HDMs and might represent potential acaricidal candidates to inhibit mite allergen production.

Changes in the Th9 cell population and related cytokines in the peripheral blood of infants with recurrent wheezing.

INTRODUCTION: T helper type 9 (Th9) cells have been shown to play a key role in initiating allergic reactions and promoting airway inflammation. However, to the best of our knowledge, their role has not been analyzed in infants with recurrent wheezing. MATERIAL AND METHODS: We performed a case-control study including 34 infants with recurrent wheezing and the same number of healthy infants as controls; all subjects were aged 1- to 3-years-old. The Th9 cell populations in the peripheral blood of these subjects were analyzed using flow cytometry, along with the assessment of Th9- and Th2-related plasma cytokine levels, including interleukin (IL)-1ß, IL-4, IL-5, IL-9, IL-10, IL-13, IL-17A, and IL-33, and transforming growth factor ß1 (TGF-ß1) using a Luminex 200 immunoassay. RESULTS: Our results indicatedthat infants with recurrent wheezing had higher percentages of Th9 cells (median, 0.69%; range, 0.46-1.08%) as compared to healthy infants (median, 0.25%, range, 0.13-0.36%; p < 0.05). In addition, infants with recurrent wheezing also exhibited higher plasma levels of cytokines IL-4, IL-9, IL-10, IL-33, and TGF-ß1. Furthermore, the percentage of Th9 cells was positively correlated with the levels of IL-4 (r = 0.408, p < 0.05) and IL-9 (r = 0.644, p < 0.05) in the peripheral blood of wheezing infants. CONCLUSIONS: Our findings suggest that the percentage of Th9 cells is increased in infants with recurrent wheezing; thus, Th9 cells may play an important role in the pathogenesis of recurrent wheezing.

Transcriptomics-Based Identification of Aquaporin Diversity in the House Dust Mite Dermatophagoides farinae (Acariformes: Pyroglyphidae).

Aquaporin water channel proteins are highly conserved across many diverse species. Some evidence indicates that aquaporins in insects may contribute to insect-related mammalian diseases and inflammation, and thus these proteins may represent viable therapeutic targets. Here, we used RNA sequencing and bioinformatics to identify putative aquaporins from the house dust mite, Dermatophagoides farinae. Six putative aquaporins were identified based on sequence similarity with aquaporins from other species. These putative aquaporins, deposited in GenBank and named DerfAQP1-4 (KY231248, KY231249, KY231250, and KY231251, respectively), DerfAQP5.01, and DerfAQP5.02 (KY231252 and KY231253), were successfully cloned into a bacterial plasmid. The identification of full-length aquaporin sequences from D. farinae provides a foundation for future molecular and biochemical studies of these proteins in D. farinae and related species.

Der p 1 and Der p 2 specific immunoglobulin E measurement for diagnosis of Dermatophagoides pteronyssinus allergy: A systematic review and meta-analysis.

BACKGROUND: A clinical history of allergic symptoms and a skin-prick test with house-dust mite crude extracts are standard diagnostic procedures for Dermatophagoides pteronyssinus allergy. Specific immunoglobulin E (IgE) responses to Der p 1 and Der p 2 allergens have been used for the diagnosis of D. pteronyssinus allergy; however, evaluation of the diagnostic performance of Der p 1 and Der p 2 specific IgE (sIgE) produced inconsistent findings. We sought to evaluate the diagnostic accuracy of Der p 1 sIgE and Der p 2 sIgE measurement in the diagnosis of D. pteronyssinus allergy by performing a systematic review and meta-analysis of previously published studies. METHODS: Several medical literature electronic data bases were searched for related literature published through August 1, 2016. A bivariate model was used to pool estimates of sensitivity, specificity, diagnostic odds ratio, and area under the summary receiver operating curves as the main diagnostic measures. RESULTS: Eight studies, which involved 1095 patients, were included in our analysis. The pooled estimates of sensitivity, specificity, and diagnostic odds ratio for Der p 1 were 0.84, 0.97, and 166.57, respectively. The combined results for Der p 2 were a sensitivity of 0.87, specificity of 1.00, and a diagnostic odds ratio of 17342.35. The areas under the summary receiver operating curves for Der p 1 sIgE and Der p 2 sIgE were 0.94 and 0.98, respectively. CONCLUSION: Our results supported the use of Der p 1 and Der p 2 sIgE in the diagnosis of D. pteronyssinus allergy. Both displayed good diagnostic performance and would be useful in a clinical setting in the accurate diagnosis of dust mite allergy.

Sequential epitopes of Dermatophagoides farinae allergens identified using peptide microarray-based immunoassay.

House dust mites produce over 30 proteins proposed to induce immunoglobulin E (IgE) antibody production in patients. Continued identification of IgE-binding epitopes of these allergens is critical to advancing diagnosis and treatment of allergic disease. To identify possible sequential IgE-binding epitopes of the major- and mid-potency allergens from the house dust mite Dermatophagoides farinae by peptide microarray-based immunoassay, nucleotide sequences of D. farinae allergens (Der f) 1, 2, 4, 5, and 7 were used to generate overlapping peptides covering the full protein sequences minus signal peptides. Short peptides were printed onto microarray chips. Because asthma occurs as a symptom of mite allergy more commonly among children than adults, the peptide chips were exposed to sera pooled from six serum-positive pediatric patients with D. farinae hypersensitivity and six serum-negative control children for screening sequential IgE-binding epitopes by IgE immunolabeling. Higher-than-average immunolabel signal intensity was observed for 21 short peptides in the serum-positive group (P < 0.01). Due to sequence overlap, these 21 signals represented four fragments of Der f 1 (amino acid positions 46-53, 71-78, 99-110, 179-186), three fragments of Der f 2 (15-22, 80-89, 106-113), six fragments of Der f 4 (69-82, 107-116, 225-232, 261-268, 355-365, 483-496), one fragment of Der f 5 (102-109), and three fragments of Der f 7 (32-39, 52-64, 100-107). These findings not only demonstrate the utility of a peptide microarray immunoassay in identifying epitopes for these allergens, but also provide a foundation for future exploration of specific immunotherapies. © 2016 IUBMB Life, 68(10):792-798, 2016.

Dust mite allergen Der f 4: Expression, characterization, and IgE binding in pediatric asthma.

BACKGROUND: House dust mite hypersensitivity affects millions of people worldwide, and although many allergens produced by house dust mite species have been identified, some of the less potent allergens remain to be studied. METHODS: The full-length cDNA encoding the group 4 allergen of the house dust mite species Dermatophagoides farinae (Der f 4) was generated through degenerate primer-based PCR, 5' RACE, and 3' RACE, and the cDNA fragment was cloned into an expression vector for nucleotide sequencing. Following codon optimization and removal of the signal peptide sequence, the mature gene fragment was subcloned into pET-28b (+) and transfected into E. coli BL21 cells for expression. The recombinant protein was purified by nickel affinity chromatography, identified by SDS-PAGE, Western blotting, and MALDI-TOF, and tested by ELISA for IgE reactivity with sera from individuals with asthma. Bioinformatics analyses were used to identify features of Der f 4. RESULTS: SDS-PAGE and Western blotting of the codon-optimized expression product showed a specific band. The mature recombinant Der f 4 was characterized as a stable and hydrophilic 57.9-kDa protein, and its secondary structure comprised alpha helix (25.3%), extended strand (22.51%), and random coils (52.19%). The structure of the recombinant protein was consistent with that of α-amylase. Among 27 pediatric asthma patients, 40.74% exhibited reactivity to rDer f 4 by ELISA. CONCLUSIONS: This initial cloning and characterization of the Der f 4 allergen serves as a foundation for future studies into the clinical importance and application of this protein for house dust mite allergy.

[Effect of artenisiae scopariae and poriae powder on calpain-2 expression in liver tissue from rats with obstructive jaundice].

OBJECTIVE: To explore the eff ect of artenisiae scopariae and poriae powder (ASPD) on calpain-2 expression in liver tissue from rats with obstructive jaundice. METHODS: The rat model of obstructive jaundice was established. SD rats was divided into the control group, the obstructive jaundice group, the obstructive jaundice model plus ASPD group, the obstructive jaundice model plus saline group. Th e serum levels of TBIL, ALT, AST and other biochemical indexes were detected. The pathological changes of liver tissue were evaluated by HE staining. The calpain-2 mRNA and protein expression in liver was measured by Real-time PCR and immunohistochemistry or Western blot, respectively. RESULTS: The calpain-2 mRNA and protein expression levels were significantly up-regulated in live tissues from the rats with obstructive jaundice in a time-dependent manner. The ASPD could inhibit the calpain-2 expression in rats with obstructive jaundice concomitant with the decreased liver damage and the improved liver function, suggesting that calpain-2 was involved in endoplasmic reticulum stress-mediated cellular apoptosis and the occurrence of obstructive jaundice. CONCLUSION: ASPD could be used for patients with obstructive jaundice to promote the recovery of liver function after operation and to reduce the incidence of complications, which provide a theoretical basis for the reasonable application of traditional Chinese medicine in the peroperative period.