Transcriptome Profiling of A549 Xenografts of Nonsmall-cell Lung Cancer Treated with Qing-Re-Huo-Xue Formula.

Lung cancer is one of the most common malignant tumors, and non-small cell lung cancer (NSCLC) accounts for 85% of all lung cancer cases. Chinese herbal formula Qing-Re-Huo-Xue (QRHXF) has shown antitumor effects in the NSCLC xenograft mouse model of Lewis cells. However, the molecular mechanisms underlying the antitumor effects of QRHXF remain unknown. In this study, an A549 xenograft mouse model was established, and the mice were then treated with QRHXF or vehicle through oral gavage. Tumor growth was monitored. Tumors were subsequently harvested, and RNA sequencing was performed. Compared with the control group, mice treated with QRHXF showed smaller tumor size and slower tumor growth. RNA sequencing results indicated 36 differentially expressed genes between QRHXF treated and control groups. 16 upregulated and 20 downregulated genes were identified. Enrichment analysis showed four differential expression genes (DEGs) related to tumor growth pathways RASAL2, SerpinB5, UTG1A4, and PDE3A. In conclusion, this study revealed that QRHXF could inhibit tumor growth in an A549 xenograft mouse model, and the target genes of QRHXF may include PDE3A, RASAL2, SERPIB5, and UTG1A4.

Cycloastragenol alleviates airway inflammation in asthmatic mice by inhibiting autophagy.

Cycloastragenol (CAG), a secondary metabolite from the roots of Astragalus zahlbruckneri, has been reported to exert anti­inflammatory effects in heart, skin and liver diseases. However, its role in asthma remains unclear. The present study aimed to investigate the effect of CAG on airway inflammation in an ovalbumin (OVA)­induced mouse asthma model. The current study evaluated the lung function and levels of inflammation and autophagy via measurement of airway hyperresponsiveness (AHR), lung histology examination, inflammatory cytokine measurement and western blotting, amongst other techniques. The results demonstrated that CAG attenuated OVA­induced AHR in vivo. In addition, the total number of leukocytes and eosinophils, as well as the secretion of inflammatory cytokines, including interleukin (IL)­5, IL­13 and immunoglobulin E were diminished in bronchoalveolar lavage fluid of the OVA­induced murine asthma model. Histological analysis revealed that CAG suppressed inflammatory cell infiltration and goblet cell secretion. Notably, based on molecular docking simulation, CAG was demonstrated to bind to the active site of autophagy­related gene 4­microtubule­associated proteins light chain 3 complex, which explains the reduced autophagic flux in asthma caused by CAG. The expression levels of proteins associated with autophagy pathways were inhibited following treatment with CAG. Taken together, the results of the present study suggest that CAG exerts an anti­inflammatory effect in asthma, and its role may be associated with the inhibition of autophagy in lung cells.

RNA-Seq Expression Analysis of Chronic Asthmatic Mice with Bu-Shen-Yi-Qi Formula Treatment and Prediction of Regulated Gene Targets of Anti-Airway Remodeling.

Airway remodeling is one of the typical pathological characteristics of asthma, while the structural changes of the airways in asthma are complex, which impedes the development of novel asthma targeted therapy. Our previous study had shown that Bu-Shen-Yi-Qi formula (BSYQF) could ameliorate airway remodeling in chronic asthmatic mice by modulating airway inflammation and oxidative stress in the lung. In this study, we analysed the lung transcriptome of control mice and asthmatic mouse model with/without BSYQF treatment. Using RNA-sequencing (RNA-seq) analysis, we found that 264/1746 (15.1%) of transcripts showing abnormal expression in asthmatic mice were reverted back to completely or partially normal levels by BSYQF treatment. Additionally, based on previous results, we identified 21 differential expression genes (DEGs) with fold changes (FC) > (±) 2.0 related to inflammatory, oxidative stress, mitochondria, PI3K/AKT, and MAPK signal pathways which may play important roles in the mechanism of the anti-remodeling effect of BSYQF treatment. Through inputting 21 DEGs into the IPA database to construct a gene network, we inferred Adipoq, SPP1, and TNC which were located at critical nodes in the network may be key regulators of BSYQF's anti-remodeling effect. In addition, the quantitative real-time polymerase chain reaction (qRT-PCR) result for the selected four DEGs matched those of the RNA-seq analysis. Our results provide a preliminary clue to the molecular mechanism of the anti-remodeling effect of BSYQF in asthma.

The Role of T Cells and Macrophages in Asthma Pathogenesis: A New Perspective on Mutual Crosstalk.

Asthma is associated with innate and adaptive immunity mediated by immune cells. T cell or macrophage dysfunction plays a particularly significant role in asthma pathogenesis. Furthermore, crosstalk between them continuously transmits proinflammatory or anti-inflammatory signals, causing the immune cell activation or repression in the immune response. Consequently, the imbalanced immune microenvironment is the major cause of the exacerbation of asthma. Here, we discuss the role of T cells, macrophages, and their interactions in asthma pathogenesis.

Upregulation of stem cell markers ALDH1A1 and OCT4 as potential biomarkers for the early detection of cervical carcinoma.

Previous studies have reported the upregulation of stem cell biomarkers that are associated with tumorigenesis, in particular with cancer infiltration, recurrence and metastasis. Infection by human papilloma virus (HPV) is the main etiopathological factor of cervical carcinogenesis, but the expression of stem cell markers in cervical carcinoma and HPV infection have yet to be investigated so far. A total of 94 cases of fresh cervical tissues, 116 cases of paraffin-embedded cervical specimens and 72 cases of peripheral blood samples were collected from Uighur women who were either diagnosed with cervical squamous cell carcinoma (SCC) or cervical intraepithelial neoplasia (CIN) II-III, or from healthy subjects (negative controls, NC). HPV infection was detected in tissue DNA by polymerase chain reaction (PCR) with a HPV genotyping kit. The mRNA expression levels of aldehyde dehydrogenase 1 family member A1 (ALDH1A1), nanog homeobox (NANOG), POU class 5 homeobox 1 (OCT4), SRY-box 2 (SOX2) and twist family BHLH transcription factor 1 (Twist1) were determined using reverse transcription-quantitative PCR (RT-qPCR). Histological analysis was performed in order to examine the protein expression of ALDH1A1 and OCT4 in paraffin-embedded tissue specimens by immunohistochemical staining and the plasma levels of those two proteins was measured by ELISA. RT-qPCR analysis indicated a significant increase in the mRNA expression of ALDH1A1 and OCT4 in CIN II-III and SCC tissue specimens compared with NC (P<0.05). Although the expression levels of NANOG, SOX2 and Twist1 were significantly higher in SCC compared with NC (P<0.05), no significant difference was revealed in CIN II-III tissues compared with SCC or NC (P>0.05). Subsequent analysis by immunohistochemistry staining confirmed that the upregulation of ALDH1A1 and OCT4 was also significantly increased in SCC and CIN II-III compared with controls at the protein level. Notably, ELISA analysis detected significantly higher levels of ALDH1A1 and OCT4 in the peripheral blood (plasma) of patients with SCC compared with healthy subjects. The upregulation of stem cell markers ALDH1A1 and OCT4 in cervical carcinoma and its precursor lesions, in particular in the peripheral blood, indicates that ALDH1A1 and OCT4 may serve as biomarkers for the early detection of cervical carcinoma or for the monitoring of treatment of patients.

Proteomic identification of potential biomarkers for cervical squamous cell carcinoma and human papillomavirus infection.

It is known that high-risk human papillomavirus infection is the main etiological factor in cervical carcinogenesis. However, human papillomavirus screening is not sufficient for early diagnosis. In this study, we aimed to identify potential biomarkers common to cervical carcinoma and human papillomavirus infection by proteomics for human papillomavirus-based early diagnosis and prognosis. To this end, we collected 76 cases of fresh cervical tissues and 116 cases of paraffin-embedded tissue slices, diagnosed as cervical squamous cell carcinoma, cervical intraepithelial neoplasia II-III, or normal cervix from ethnic Uighur and Han women. Human papillomavirus infection by eight oncogenic human papillomavirus types was detected in tissue DNA samples using a quantitative polymerase chain reaction. The protein profile of cervical specimens from human papillomavirus 16-positive squamous cell carcinoma and human papillomavirus-negative normal controls was analyzed by proteomics and bioinformatics. The expression of candidate proteins was further determined by quantitative reverse transcriptase-polymerase chain reaction and immunohistochemistry. We identified 67 proteins that were differentially expressed in human papillomavirus 16-positive squamous cell carcinoma compared to normal cervix. The quantitative reverse transcriptase-polymerase chain reaction analysis verified the upregulation of ASAH1, PCBP2, DDX5, MCM5, TAGLN2, hnRNPA1, ENO1, TYPH, CYC, and MCM4 in squamous cell carcinoma compared to normal cervix ( p < 0.05). In addition, the transcription of PCBP2, MCM5, hnRNPA1, TYPH, and CYC was also significantly increased in cervical intraepithelial neoplasia II-III compared to normal cervix. Immunohistochemistry staining further confirmed the overexpression of PCBP2, hnRNPA1, ASAH1, and DDX5 in squamous cell carcinoma and cervical intraepithelial neoplasia II-III compared to normal controls ( p < 0.05). Our data suggest that the expression of ASAH1, PCBP2, DDX5, and hnRNPA1, and possibly MCM4, MCM5, CYC, ENO1, and TYPH, is upregulated during cervical carcinogenesis and potentially associated with human papillomavirus infection. Further validation studies of the profile will contribute to establishing auxiliary diagnostic markers for human papillomavirus-based cancer prognosis.

Identification of plasma protein markers common to patients with malignant tumour and Abnormal Savda in Uighur medicine: a prospective clinical study.

BACKGROUND: Traditional Uighur medicine shares an origin with Greco-Arab medicine. It describes the health of a human body as the dynamic homeostasis of four normal Hilits (humours), known as Kan, Phlegm, Safra, and Savda. An abnormal change in one Hilit may cause imbalance among the Hilits, leading to the development of a syndrome. Abnormal Savda is a major syndrome of complex diseases that are associated with common biological changes during disease development. Here, we studied the protein expression profile common to tumour patients with Abnormal Savda to elucidate the biological basis of this syndrome and identify potential biomarkers associated with Abnormal Savda. METHODS: Patients with malignant tumours were classified by the diagnosis of Uighur medicine into two groups: Abnormal Savda type tumour (ASt) and non-Abnormal Savda type tumour (nASt), which includes other syndromes. The profile of proteins that were differentially expressed in ASt compared with nASt and normal controls (NC) was analysed by iTRAQ proteomics and evaluated by bioinformatics using MetaCore™ software and an online database. The expression of candidate proteins was verified in all plasma samples by enzyme-linked immunosorbent assay (ELISA). RESULTS: We identified 31 plasma proteins that were differentially expressed in ASt compared with nASt, of which only 10 showed quantitatively different expression between ASt and NC. Bioinformatics analysis indicated that most of these proteins are known biomarkers for neoplasms of the stomach, breast, and lung. ELISA detection showed significant upregulation of plasma SAA1 and SPP24 and downregulation of PIGR and FASN in ASt compared with nASt and NC (p < 0.05). CONCLUSIONS: Abnormal Savda may be causally associated with changes in the whole regulation network of protein expression during carcinogenesis. The expression of potential biomarkers might be used to distinguish Abnormal Savda from other syndromes.