Effects of tumor-infiltrating lymphocytes on nonresponse rate of neoadjuvant chemotherapy in patients with invasive breast cancer.

High level of tumor-infiltrating lymphocytes (TILs) can predict the rate of total pathological complete remission (tpCR) of breast cancer patients who receive neoadjuvant chemotherapy (NACT). This study focused on evaluating the data of patients whose primary tumor and/or lymph node metastasis show nonresponse (NR) to NACT, trying to provide a basis for the clinical decision which patients will develop NACT resistance. The study included breast cancers from 991 patients who received NACT. ROC curve analysis confirmed that TILs showed significant predictive value for NR of hormone receptor (HR)+HER2- and triple-negative breast cancer (TNBC). Among HR+HER2- breast cancer, TILs ≥ 10% was an independent predictor for low NR rate. Furthermore, positive correlation of TILs with Ki67 index and Miller-Payne grade, and negative correlation with ER and PR H-scores were only identified in this subgroup. In TNBC, TILs ≥ 17.5% was an independent predictor for low NR rate. The predictive value of low TILs on NR may facilitate to screen patients with HR+HER2- or TNBC who may not benefit from NACT. HR+HER2- breast cancer with low levels of TILs should be carefully treated with neoadjuvant chemotherapy, and other alternatives such as neoadjuvant endocrine therapy can be considered.

Characteristics of recurrence, predictors for relapse and prognosis of rapid relapse triple-negative breast cancer.

Background: Triple-negative breast cancer (TNBC) patients who recur at different times are associated with distinct biological characteristics and prognoses. Research on rapid-relapse TNBC (RR-TNBC) is sparse. In this study, we aimed to describe the characteristics of recurrence, predictors for relapse, and prognosis in rrTNBC patients. Methods: Clinicopathological data of 1584 TNBC patients from 2014 to 2016 were retrospectively reviewed. The characteristics of recurrence were compared between patients with RR-TNBC and slow relapse TNBC(SR-TNBC). All TNBC patients were randomly divided into a training set and a validation set to find predictors for rapid relapse. The multivariate logistic regression model was used to analyze the data of the training set. C-index and brier score analysis for predicting rapid relapse in the validation set was used to evaluate the discrimination and accuracy of the multivariate logistic model. Prognostic measurements were analyzed in all TNBC patients. Results: Compared with SR-TNBC patients, RR-TNBC patients tended to have a higher T staging, N staging, TNM staging, and low expression of stromal tumor-infiltrating lymphocytes (sTILs). The recurring characteristics were prone to appear as distant metastasis at the first relapse. The first metastatic site was apt to visceral metastasis and less likely to have chest wall or regional lymph node metastasis. Six predictors (postmenopausal status, metaplastic breast cancer,≥pT3 staging,≥pN1 staging, sTIL intermediate/high expression, and Her2 [1+]) were used to construct the predictive model of rapid relapse in TNBC patients. The C-index and brier score in the validation set was 0.861 and 0.095, respectively. This suggested that the predictive model had high discrimination and accuracy. The prognostic data for all TNBC patients showed that RR-TNBC patients had the worst prognosis, followed by SR-TNBC patients. Conclusion: RR-TNBC patients were associated with unique biological characteristics and worse outcomes compared to non-RR-TNBC patients.

Hypoxia-inducible factor 1-alpha is a driving mechanism linking chronic obstructive pulmonary disease to lung cancer.

Patients with chronic obstructive pulmonary disease (COPD), irrespective of their smoking history, are more likely to develop lung cancer than the general population. This is mainly because COPD is characterized by chronic persistent inflammation and hypoxia, which are the risk factors for lung cancer. However, the mechanisms underlying this observation are still unknown. Hypoxia-inducible factor 1-alpha (HIF-1α) plays an important role in the crosstalk that exists between inflammation and hypoxia. Furthermore, HIF-1α is the main regulator of somatic adaptation to hypoxia and is highly expressed in hypoxic environments. In this review, we discuss the molecular aspects of the crosstalk between hypoxia and inflammation, showing that HIF-1α is an important signaling pathway that drives COPD progression to lung cancer. Here, we also provide an overview of HIF-1α and its principal regulatory mechanisms, briefly describe HIF-1α-targeted therapy in lung cancer, and summarize substances that may be used to target HIF-1α at the level of COPD-induced inflammation.

STAT3 and PD-L1 are negatively correlated with ATM and have impact on the prognosis of triple-negative breast cancer patients with low ATM expression.

INTRODUCTION: Triple-negative breast cancer (TNBC) is known for its aggressive behaviors and lacking of effective treatment. Programmed cell death ligand-1 (PD-L1) inhibitor has just been approved for using in the management of advanced TNBC. To accurately screen TNBC sensitive to anti-PD-L1 treatment and to explore the feasibility of the ataxia-telangiectasia mutation protein (ATM) inhibitor combined with PD-L1 inhibitor, radiotherapy and chemotherapy, we focus on whether ATM participates in the regulation of PD-L1 and affects the prognosis of patients through c-Src, signal transducer and activator of transcription 1&3 (STAT1 and STAT3). MATERIALS AND METHODS: We used immunohistochemical staining to explore the relationship of ATM with c-Src, STAT1, STAT3, PD-1/PD-L1, Tumor-infiltrating lymphocytes (TILs), as well as other clinicopathologic features in 86 pathological stage III TNBCs. Their impact on prognosis was also explored. RESULTS: We found ATM expression was negatively correlated with STAT1, STAT3, PD-L1, TILs and CD8 + cells in TNBC. STAT1 positively correlated the expression of PD-L1. In TNBC with ATM low expression, STAT3 was an independent factor for improved prognosis, while PD-L1 was an independent negative prognostic factor. Furthermore, in low ATM group, the phosphorylation of tyrosine at position 419 of c-Src (p-c-src Y419) was correlated with the overexpression of STAT3. CONCLUSION: Locally advanced TNBC with low ATM expression may be more likely to benefit from anti-PD-L1 inhibitors. The feasibility of ATM functional inhibitor combined with immune checkpoint blockade therapies in the treatment of TNBC is also worthy of further exploration. Our study suggests that STAT3 has different impacts on tumor progression in different tumors.

Detection of Synthetic Cannabinoid CUMYL-PEGACLONE in E-Cigarette Oil and Hair.

OBJECTIVES: To establish a combined high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) and gas chromatography-mass spectrometry (GC-MS) method to detect the synthetic cannabinoid CUMYL-PEGACLONE in e-cigarette oil and hair. METHODS: HPLC-MS/MS and GC-MS were used to establish the detection method of CUMYL-PEGACLONE, and the hair of drug-involved persons and the seized e-cigarette oil were detected. RESULTS: The main mass spectrometry characteristic ions m/z of CUMYL-PEGACLONE measured by GC-MS were 91, 179, 197, 254 and 372. CUMYL-PEGACLONE had a good linear relationship in the mass concentration range of 2-50 ng/mL, and the linear correlation coefficient (r) was greater than 0.99. The limit of detection (LOD) of CUMYL-PEGACLONE in hair was 0.01 ng/mg, and the limit of quantitation (LOQ) was 0.02 ng/mg. The LOD of CUMYL-PEGACLONE in e-cigarette oil was 1 ng/mg, and the LOQ was 2 ng/mg. The average recoveries of CUMYL-PEGACLONE under the attempt at high, intermediate and low levels in blank human hair and e-cigarette oil matrix were 98.2%-132.4% and 93.5%-110.6%, and the intraday and intraday precision were 1.2%-12.9% and 0.7%-2.9%. CUMYL-PEGACLONE was detected in the hair of 15 drug-involved persons. Except for 1 person who was lower than LOQ, the concentration of CUMYL-PEGACLONE in the hair of other 14 persons was 0.035-0.563 ng/mg. The mass fraction of CUMYL-PEGACLONE in 2 e-cigarette oil were 0.17% and 0.21%, respectively. CONCLUSIONS: The established HPLC-MS/MS and GC-MS methods are applied to the detection of HPLC-MS/MS in drug-related cases, which provides strong evidence support for the handling authority to quickly investigate these cases, and also provides a reference for the identification of such substances in future.

Coexistence of tuberculosis and squamous cell carcinoma in the right main bronchus: A case report.

BACKGROUND: Lung cancer with pulmonary tuberculosis (TB) refers to the occurrence of lesions simultaneously or sequentially in the lung(s) of the same patient, and the pathological examination and sputum TB examination diagnose them as lung cancer and TB, respectively. The occurrence of endobronchial TB (EBTB) with endobronchial tumor sequentially in the same bronchus lesion of the same patient is relatively rare. CASE SUMMARY: A 62-year-old female patient was admitted to a local hospital on June 18, 2019 after a 3-mo history of dyspnea. She was a farmer and had no history of smoking and alcohol misuse. The patient had neither family nor work contact indicating exposure to TB. Emergency chest computed tomography (CT) examination showed that the right main bronchus was occupied and malignant tumor was possible. Histopathologic examination of a bronchial biopsy showed granulomatous inflammation with caseification and the presence of acid fast bacilli (AFB). However, after 6 mo of antitubercular treatment, repeat bronchoscopy and biopsy histological examination showed squamous cell carcinoma. The patient has started on systemic chemotherapy with carboplatin. After another two cycles of therapy, chest CT showed complete resolution of the lesions. Bronchoalveolar lavage and bronchial aspirate were negative for AFB and cancer cells. CONCLUSION: It is not only more likely that a patient presenting with what appears to be TB will concurrently have a pulmonary malignancy than someone who does not have a TB infection, but also that it is of greater urgency to make an expedited diagnosis of the malignancy.

AI detection of mild COVID-19 pneumonia from chest CT scans.

OBJECTIVES: An artificial intelligence model was adopted to identify mild COVID-19 pneumonia from computed tomography (CT) volumes, and its diagnostic performance was then evaluated. METHODS: In this retrospective multicenter study, an atrous convolution-based deep learning model was established for the computer-assisted diagnosis of mild COVID-19 pneumonia. The dataset included 2087 chest CT exams collected from four hospitals between 1 January 2019 and 31 May 2020. The true positive rate, true negative rate, receiver operating characteristic curve, area under the curve (AUC) and convolutional feature map were used to evaluate the model. RESULTS: The proposed deep learning model was trained on 1538 patients and tested on an independent testing cohort of 549 patients. The overall sensitivity was 91.5% (195/213; p < 0.001, 95% CI: 89.2-93.9%), the overall specificity was 90.5% (304/336; p < 0.001, 95% CI: 88.0-92.9%) and the general AUC value was 0.955 (p < 0.001). CONCLUSIONS: A deep learning model can accurately detect COVID-19 and serve as an important supplement to the COVID-19 reverse transcription-polymerase chain reaction (RT-PCR) test. KEY POINTS: • The implementation of a deep learning model to identify mild COVID-19 pneumonia was confirmed to be effective and feasible. • The strategy of using a binary code instead of the region of interest label to identify mild COVID-19 pneumonia was verified. • This AI model can assist in the early screening of COVID-19 without interfering with normal clinical examinations.

Reprogramming of m6A epitranscriptome is crucial for shaping of transcriptome and proteome in response to hypoxia.

Hypoxia causes a series of responses supporting cells to survive in harsh environments. Substantial post-transcriptional and translational regulation during hypoxia has been observed. However, detailed regulatory mechanism in response to hypoxia is still far from complete. RNA m6A modification has been proven to govern the life cycle of RNAs. Here, we reported that total m6A level of mRNAs was decreased during hypoxia, which might be mediated by the induction of m6A eraser, ALKBH5. Meanwhile, expression levels of most YTH family members of m6A readers were systematically down-regulated. Transcriptome-wide analysis of m6A revealed a drastic reprogramming of m6A epitranscriptome during cellular hypoxia. Integration of m6A epitranscriptome with either RNA-seq based transcriptome analysis or mass spectrometry (LC-MS/MS) based proteome analysis of cells upon hypoxic stress revealed that reprogramming of m6A epitranscriptome reshaped the transcriptome and proteome, thereby supporting efficient generation of energy for adaption to hypoxia. Moreover, ATP production was blocked when silencing an m6A eraser, ALKBH5, under hypoxic condition, demonstrating that m6A pathway is an important regulator during hypoxic response. Collectively, our studies indicate that crosstalk between m6A and HIF1 pathway is essential for cellular response to hypoxia, providing insights into the underlying molecular mechanisms during hypoxia.

Heat-response patterns of the heat shock transcription factor family in advanced development stages of wheat (Triticum aestivum L.) and thermotolerance-regulation by TaHsfA2-10.

BACKGROUND: Heat shock transcription factors (Hsfs) are present in majority of plants and play central roles in thermotolerance, transgenerational thermomemory, and many other stress responses. Our previous paper identified at least 82 Hsf members in a genome-wide study on wheat (Triticum aestivum L.). In this study, we analyzed the Hsf expression profiles in the advanced development stages of wheat, isolated the markedly heat-responsive gene TaHsfA2-10 (GenBank accession number MK922287), and characterized this gene and its role in thermotolerance regulation in seedlings of Arabidopsis thaliana (L. Heynh.). RESULTS: In the advanced development stages, wheat Hsf family transcription profiles exhibit different expression patterns and varying heat-responses in leaves and roots, and Hsfs are constitutively expressed to different degrees under the normal growth conditions. Overall, the majority of group A and B Hsfs are expressed in leaves while group C Hsfs are expressed at higher levels in roots. The expression of a few Hsf genes could not be detected. Heat shock (HS) caused upregulation about a quarter of genes in leaves and roots, while a number of genes were downregulated in response to HS. The highly heat-responsive gene TaHsfA2-10 was isolated through homeologous cloning. qRT-PCR revealed that TaHsfA2-10 is expressed in a wide range of tissues and organs of different development stages of wheat under the normal growth conditions. Compared to non-stress treatment, TaHsfA2-10 was highly upregulated in response to HS, H2O2, and salicylic acid (SA), and was downregulated by abscisic acid (ABA) treatment in two-leaf-old seedlings. Transient transfection of tobacco epidermal cells revealed subcellular localization of TaHsfA2-10 in the nucleus under the normal growth conditions. Phenotypic observation indicated that TaHsfA2-10 could improve both basal thermotolerance and acquired thermotolerance of transgenic Arabidopsis thaliana seedlings and rescue the thermotolerance defect of the T-DNA insertion mutant athsfa2 during HS. Compared to wild type (WT) seedlings, the TaHsfA2-10-overexpressing lines displayed both higher chlorophyll contents and higher survival rates. Yeast one-hybrid assay results revealed that TaHsfA2-10 had transactivation activity. The expression levels of thermotolerance-related AtHsps in the TaHsfA2-10 transgeinc Arabidopsis thaliana were higher than those in WT after HS. CONCLUSIONS: Wheat Hsf family members exhibit diversification and specificity of transcription expression patterns in advanced development stages under the normal conditions and after HS. As a markedly responsive transcriptional factor to HS, SA and H2O2, TaHsfA2-10 involves in thermotolerance regulation of plants through binding to the HS responsive element in promoter domain of relative Hsps and upregulating the expression of Hsp genes.

Increased number of intratumoral IL-17+ cells, a harbinger of the adverse prognosis of triple-negative breast cancer.

INTRODUCTION: Triple negative breast cancer (TNBC) is an aggressive cancer subtype and lack of effective targeted therapies. It has been recently reported that Interleukin 17 (IL-17), a family of cytokines secreted in tumor microenvironment, affects tumor progression through a variety of molecular pathways. Its role in TNBC is so far still poorly explored. MATERIALS AND METHODS: We employed immunohistochemistry to evaluate the distribution of IL-17+ cells in TNBC with no special type features (TNBC-NST), their association with tumor microangiogenesis, as well as their impact on prognosis of the patients. RESULTS: In comparison to medullary carcinoma with triple-negative molecular features (TNBC-MC), we found a significant increase in IL-17+ cell infiltrates in intratumoral stroma and extratumoral stroma of TNBC-NST. Similarly, stromal cells with co-expression of CD4 and IL-17 were noted in intratumoral and extratumoral stroma in both TNBC-NST and TNBC-MC. In addition, intratumoral IL-17+ cells were positively associated with tumor cell expression of vascular endothelial growth factor A (VEGFA) and with intratumoral tumor microvascular density (MVD). Multivariate analysis identified that intratumoral IL-17+ cells (P = 0.018), MVD (P = 0.039), and TNM stage (P = 0.002) were independent prognostic factors for predicting poor PFS. CONCLUSION: The study indicates that IL-17 is overexpressed in intratumoral stromal cells of TNBC-NST. The overexpression of IL-17 might engage in active tumor microangiogenesis through its signal transduction pathways resulting in increased tumor secretion of VEGFA, and then promote tumor progression. IL-17 might serve as a potential new target for individualized therapy to TNBC-NST patients by development of specific antibodies. Additional study is deemed to further explore the role of IL-17+ stromal cells in breast cancer.

Resveratrol downregulates TNF-α-induced monocyte chemoattractant protein-1 in primary rat pulmonary artery endothelial cells by P38 mitogen-activated protein kinase signaling.

Background: To evaluate the effects of resveratrol to monocyte chemoattractant protein-1 (MCP-1) and the role of p38 mitogen-activated protein kinase (MAPK) in this process in vitro. Materials and methods: Animal acute pulmonary thromboembolism (PTE) model: rat model was established by infusion of an autologous blood clot into the pulmonary artery through a polyethylene catheter. One hundred and thirty-two rats were randomly and equally divided into ten groups: rats-control (untreated), rats-1% DMSO, rats-TNF-α, rats-TNF-α + resveratrol, rats-TNF-α +C1142, rats-TNF-α+SB203580, rats-TNF-α+resveratrol + SB203580, rats-resveratrol only, rats-C1142 only, and rats-SB203580 only. Rat pulmonary artery endothelial cells (RPAs) tests: RPAs were isolated from above animal and designated as: RPAs-control, RPAs-1% DMSO control, RPAs-TNF-α, RPAs-TNF-α + resveratrol, RPAs-TNF-α + C1142, RPAs-TNF-α + SB203580, RPAs-TNF-α + resveratrol + SB203580, RPAs-resveratrol only, RPAs-C1142 only, and RPAs-SB203580 only. Each group was further divided into 1, 4, and 8 hrs time point for evaluation (n=6 rats per time point) except RPAs-TNF-α + SB203580, RPAs-TNF-α + resveratrol + SB203580, RPAs-C1142 and RPAs-SB203580 only, which were evaluated at 8 hrs time point. At each time point, mRNA and protein expressions of RPAs of MCP-1 were measured. The phosphorylation of p38 MAPK (p-pMAPK) of RPAs was also detected. Results: We found that the RPAs-TNF-α elicited significant increases in MCP-1 expression and phosphorylation of p38 mitogen-activated protein kinase (p-p38 MAPK). Furthermore, the MCP-1 expressions of RPAs-Resveratrol, RPAs-C1142, and RPAs-SB203580 were significantly down-regulated, which was associated with robustly suppressed TNF-α-induced p-p38MAPK expression. Conclusion: Our findings suggested that MCP-1 was involved in the formation of TNF-α-induced inflammatory response, and resveratrol could down-regulate the expression of MCP-1 via TNF-α- inhibition, which might contribute to the decline of acute PTE-induced PH in vivo.

A Genome-Wide Association Study of α-Synuclein Levels in Cerebrospinal Fluid.

α-Synuclein is a 140-amino acid protein produced predominantly by neurons in the brain which plays a role in the regulation of neurotransmitter release, synaptic function, and plasticity, thus making it the focus in understanding the etiology of a group of neurodegenerative diseases. We conducted genome-wide association studies (GWAS) of α-synuclein levels in cerebrospinal fluid (CSF) with 209 non-Hispanic white participants from the Alzheimer's Disease Neuroimaging Initiative (ADNI-1) cohort using a linear regression model to identify novel variants associated with α-synuclein concentration. The minor allele (T) of rs7072338 in the long intergenic non-protein coding RNA 1515 (LINC01515) and the minor allele (T) of rs17794023 in clusterin-associated protein 1 (CLUAP1) were associated with higher CSF α-synuclein levels at genome-wide significance (P = 4.167 × 10-9 and 9.56 × 10-9, respectively). In addition, single nucleotide polymorphisms (SNPs) near amyloid beta precursor protein (APP) (rs1394839) (P = 2.31 × 10-7), Rap guanine nucleotide exchange factor 1 (RAPGEF1) (rs10901091) (P = 8.07 × 10-7), and two intergenic loci on chromosome 2 and 14 (rs11687064 P = 2.50 × 10-7and rs7147386 P = 4.05 × 10-7) were identified as suggestive loci associated with CSF α-synuclein levels. We have identified significantly associated SNPs for CSF α-synuclein. These associations have important implications for a better understanding of α-synuclein regulation and allow researchers to further explore the relationships between these SNPs and α-synuclein-related neurodegenerative disorders.

Tumor-driven like macrophages induced by conditioned media from pancreatic ductal adenocarcinoma promote tumor metastasis via secreting IL-8.

Tumor-associated macrophages (TAMs) are abundant population of inflammatory cells which play an essential role in remodeling tumor microenvironment and tumor progression. Previously, we found the high density of TAMs was correlated with lymph node metastasis and poor prognosis in pancreatic ductal adenocarcinoma (PDAC). Therefore, this study was designed to investigate the mechanisms of interaction between TAMs and PDAC. THP-1 monocytes were the exposure to conditioned media (CM) produced by PDAC cells; then, monocyte recruitment and macrophage differentiation were assessed. CM from PDAC attracted and polarized THP-1 monocytes to tumor-driven like macrophages. mRNA expression cytokine profiling and ELISA identified the IL-8 secretion was increasing in tumor-driven like macrophages, and STAT3 pathway was involved. Addition of exogenous recombinant human IL-8 promoted PDAC cells motility in vitro and metastasis in vivo via upregulating Twist expression, which mediated epithelial-mesenchymal transition in cancer cells. What is more, IL-8 expression level in tumor stroma by immunohistochemical analysis was related to lymph node metastasis, the number of tumor CD68 but not CD163 positive macrophages and patient outcome. Taken together, these findings shed light on the important interplay between cancer cells and TAMs in tumor microenvironment and suggested that IL-8 signaling might be a potential therapeutic target for PDAC.

[Effects of Long-term Fertilization Regimes on As Accumulation in Upland Red Earth and the Crops Growing on It].

To investigate the effects of different long-term fertilization treatments on arsenic (As) accumulation in upland red earth and the crops growing on it, different fertilization treatments, including no fertilization (CK), fertilization with chemical fertilizers containing nitrogen, phosphorus and potassium (NPK), fertilization with swine manure (M), and fertilization with chemical fertilizers plus manure (NPKM), in a long-term fertilization site located in Qiyang County, Hunan Province, were selected.. The results showed that after 26 years of fertilization, both the total and available As concentrations in the soils were significantly improved in the fertilized soils (i.e., NPK, M, and NPKM) when compared to the CK soil. Meanwhile, NPK treatment had the highest As concentration (44.68 mg·kg-1), which was higher than the national standard value (40 mg·kg-1) in acidic soils. Fertilization practices had the biggest impacts on the non-specifically-absorbed As, followed by the residual As, and the well-crystallized hydrous oxides of Fe and Al absorbed As, but minor impacts on the amorphous and poorly-crystalline hydrous oxides of Fe and Al-absorbed As, and the specifically absorbed As. Furthermore, NPKM treatment significantly decreased the concentrations of As(Ⅲ) and As(Ⅴ) in the soils, when compared to the other treatments. Also, M treatment could cause the accumulation of As in the grains and straws in corn. In contrast, M treatment could not cause the accumulation of As in the grains and straws in wheat. Furthermore, correlative synchrotron-radiation-based FTIR and micro X-ray fluorescence spectromicroscopic analysis showed that As had a similar distribution pattern with clay -OH and C-OH.

In situ measurement of perfluoroalkyl substances in aquatic systems using diffusive gradients in thin-films technique.

To better understand the environmental impact of ubiquitous perfluoroalkyl substances (PFASs) in waters, reliable and robust measurement techniques are needed. As one of the most widely used passive sampling approaches, diffusive gradients in thin-films (DGT) is not only easy to handle but also provides time-weighted analyte concentrations. Based on DGT with XAD18 as a binding agent, we developed a new methodology to measure two frequently detected PFASs in surface waters and wastewaters, i.e. perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS). Their diffusion coefficients in the diffusive gel, measured using an independent diffusion cell, were 4.37 × 10-6 and 5.08 × 10-6 cm2 s-1 at 25 °C, respectively. DGT had a high capacity for PFOA and PFOS at 196 and 246 µg per gel disk, suggesting the DGT sampler was suitable for deployment of several weeks. Time-integrated concentrations of PFOA and PFOS in a natural lake and river, and a municipal wastewater treatment plant effluent using DGT samplers deployed in situ for 12-33 d were comparable to those measured by a solid-phase extraction method coupled with high-frequency grab sampling. This study demonstrates that DGT is an effective tool for in situ monitoring of PFASs in natural waters and wastewaters.

Sulfur Oxidation in the Acidophilic Autotrophic Acidithiobacillus spp.

Sulfur oxidation is an essential component of the earth's sulfur cycle. Acidithiobacillus spp. can oxidize various reduced inorganic sulfur compounds (RISCs) with high efficiency to obtain electrons for their autotrophic growth. Strains in this genus have been widely applied in bioleaching and biological desulfurization. Diverse sulfur-metabolic pathways and corresponding regulatory systems have been discovered in these acidophilic sulfur-oxidizing bacteria. The sulfur-metabolic enzymes in Acidithiobacillus spp. can be categorized as elemental sulfur oxidation enzymes (sulfur dioxygenase, sulfur oxygenase reductase, and Hdr-like complex), enzymes in thiosulfate oxidation pathways (tetrathionate intermediate thiosulfate oxidation (S4I) pathway, the sulfur oxidizing enzyme (Sox) system and thiosulfate dehydrogenase), sulfide oxidation enzymes (sulfide:quinone oxidoreductase) and sulfite oxidation pathways/enzymes. The two-component systems (TCSs) are the typical regulation elements for periplasmic thiosulfate metabolism in these autotrophic sulfur-oxidizing bacteria. Examples are RsrS/RsrR responsible for S4I pathway regulation and TspS/TspR for Sox system regulation. The proposal of sulfur metabolic and regulatory models provide new insights and overall understanding of the sulfur-metabolic processes in Acidithiobacillus spp. The future research directions and existing barriers in the bacterial sulfur metabolism are also emphasized here and the breakthroughs in these areas will accelerate the research on the sulfur oxidation in Acidithiobacillus spp. and other sulfur oxidizers.

Lentivirus-mediated disintegrin and metalloproteinase 17 RNA interference reversed the acquired resistance to gefitinib in lung adenocarcinoma cells in vitro.

OBJECTIVE: The aim of the study is to evaluate the effects of silencing a disintegrin and metalloproteinase 17 (ADAM17) gene expression by lentivirus-mediated RNA interference (RNAi) in the gefitinib-resistant lung adenocarcinoma cells, and then to explore whether the recombinant lentivirus mediated ADAM17 RNAi reversed the acquired resistance of lung adenocarcinoma to gefitinib in vitro. METHODS: The gefitinib-resistant RPC-9 cells were established and the mutations of EGFR were detected by gene sequencing. The ADAM17 shRNA expression vectors were constructed and packaged to recombinant lentivirus. The cell proliferation viability was detected by MTT, and cellular apotosis was analyzed by flow cytometry assay. The expression levels of ADAM17, EGFR and the phosphorylated EGFR were respectively detected by reverse transcription polymerase chain reaction and western blot. TGF-α production in the supernatant was detected by enzyme-linked immunosorbent assay. RESULTS: The gefitinib-resistant RPC-9 cells in which mutated EGFR (exon 20) carried 790T > T/M mutation were established. When the concentrations of gefitinib were less than 10µmol/L, there were no significant changes in the apoptosis and cellular proliferation of RPC-9 with the dose-escalation of gefitinib. The cell proliferation viability of RPC-9 was significantly decreased by lentivirus mediated ADAM17 RNAi (P < 0.05). Gefitinib did not inhibit ADAM17 expression in both the gefitinib-sensitive PC-9 and gefitinib-resistant RPC-9 cells (P > 0.05). Gefitinib had no significant effects on TGF alpha production in the supernatants (P > 0.05). Gefitinib did not inhibit EGFR expression in gefitinib-sensitive PC-9 and gefitinib-resistant RPC-9 cells (P > 0.05). The phosphorylation of EGFR in gefitinib-sensitive PC-9 cells was significantly inhibited by gefitinib (P < 0.05), but that in gefitinib-resistant RPC-9 could not be inhibited by gefitinib (P > 0.05). Lentivirus mediated ADAM17 RNAi significantly inhibited the mRNA and protein expression of ADAM17 in gefitinib-resistant RPC-9 cells (P < 0.05), as well as TGF alpha production in the supernatants (P < 0.05). Also, the phosphorylation of EGFR was significantly reduced in gefitinib-resistant RPC-9 cells by lentivirus mediated ADAM17 RNAi (P < 0.05); however, the mRNA and protein expression of EGFR could not be inhibited. CONCLUSION: Lentivirus mediated ADAM17 RNAi may reverse the acquired resistance of lung adenocarcinoma to gefitinib via inhibiting the upstream of EGFR signal pathway, which may provide a new therapeutic target to solve the acquired resistance to EGFR tyrosine kinase inhibitors in lung adenocarcinoma. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 34:196-205, 2018.

The Two-Component System RsrS-RsrR Regulates the Tetrathionate Intermediate Pathway for Thiosulfate Oxidation in Acidithiobacillus caldus.

Acidithiobacillus caldus (A. caldus) is a common bioleaching bacterium that possesses a sophisticated and highly efficient inorganic sulfur compound metabolism network. Thiosulfate, a central intermediate in the sulfur metabolism network of A. caldus and other sulfur-oxidizing microorganisms, can be metabolized via the tetrathionate intermediate (S4I) pathway catalyzed by thiosulfate:quinol oxidoreductase (Tqo or DoxDA) and tetrathionate hydrolase (TetH). In A. caldus, there is an additional two-component system called RsrS-RsrR. Since rsrS and rsrR are arranged as an operon with doxDA and tetH in the genome, we suggest that the regulation of the S4I pathway may occur via the RsrS-RsrR system. To examine the regulatory role of the two-component system RsrS-RsrR on the S4I pathway, ΔrsrR and ΔrsrS strains were constructed in A. caldus using a newly developed markerless gene knockout method. Transcriptional analysis of the tetH cluster in the wild type and mutant strains revealed positive regulation of the S4I pathway by the RsrS-RsrR system. A 19 bp inverted repeat sequence (IRS, AACACCTGTTACACCTGTT) located upstream of the tetH promoter was identified as the binding site for RsrR by using electrophoretic mobility shift assays (EMSAs) in vitro and promoter-probe vectors in vivo. In addition, ΔrsrR, and ΔrsrS strains cultivated in K2S4O6-medium exhibited significant growth differences when compared with the wild type. Transcriptional analysis indicated that the absence of rsrS or rsrR had different effects on the expression of genes involved in sulfur metabolism and signaling systems. Finally, a model of tetrathionate sensing by RsrS, signal transduction via RsrR, and transcriptional activation of tetH-doxDA was proposed to provide insights toward the understanding of sulfur metabolism in A. caldus. This study also provided a powerful genetic tool for studies in A. caldus.

A R2R3-MYB transcription factor gene in common wheat (namely TaMYBsm1) involved in enhancement of drought tolerance in transgenic Arabidopsis.

We isolated the TaMYBsm1 genes, encoding R2R3-type MYB proteins in common wheat, aimed to uncover the possible molecular mechanisms related to drought response. The TaMYBsm1 genes, TaMYBsm1-A, TaMYBsm1-B and TaMYBsm1-D, were isolated and analyzed from the common wheat cultivar Shimai 15. Their expression patterns under PEG 6000 and mannitol were monitored by semi-quantitative RT-PCR and ß-glucuronidase (Gus) assay. The function of TaMYBsm1-D under drought stress in transgenic Arabidopsis plants was investigated, and the germination rate, water loss rate, as well as the proline and malondialdehyde (MDA) content were compared with that in wild type (WT) plants. The expression of three downstream genes (DREB2A, P5CS1 and RD29A) in TaMYBsm1-D transgenic plants was analyzed. The R2R3-MYB domains of the MYBsm1 proteins were highly conserved in plants. In addition, the TaMYBsm1 proteins were targeted to the nucleus and contained transcriptional activation domains (TADs). Gus assay and semi-quantitative RT-PCR analysis demonstrated that the TaMYBsm1 genes were up-regulated when the wheat was treated by PEG and mannitol. Compared with WT plants, the germination rates were much higher, but the water loss rates were much lower in TaMYBsm1-D overexpression plants. TaMYBsm1-D transgenic plants showed distinct higher proline contents but a lower MDA content than the WT plants. The three downstream genes were highly expressed in TaMYBsm1-D transgenic plants. We concluded from these results that TaMYBsm1 genes play an important role in plant drought stress tolerance through up-regulation of DREB2A, P5CS1 and RD29A. The increase of proline content and decrease of MDA content may also be involved in the drought response.