CD69 is a TGF-β/1α,25-dihydroxyvitamin D3 target gene in monocytes

CD69 is a transmembrane lectin that can be expressed on most hematopoietic cells. In monocytes, it has been functionally linked to the 5-lipoxygenase pathway in which the leukotrienes, a class of highly potent inflammatory mediators, are produced. However, regarding CD69 gene expression and its regulatory mechanisms in monocytes, only scarce data are available. Here, we report that CD69 mRNA expression, analogous to that of 5-lipoxygenase, is induced by the physiologic stimuli transforming growth factor-β (TGF-β) and 1α,25-dihydroxyvitamin D3 (1α,25(OH)2D3) in monocytic cells. Comparison with T- and B-cell lines showed that the effect was specific for monocytes. CD69 expression levels were increased in a concentration-dependent manner, and kinetic analysis revealed a rapid onset of mRNA expression, indicating that CD69 is a primary TGF-β/1α,25(OH)2D3 target gene. PCR analysis of different regions of the CD69 mRNA revealed that de novo transcription was initiated and proximal and distal parts were induced concomitantly. In common with 5-lipoxygenase, no activation of 0.7 kb or ~2.3 kb promoter fragments by TGF-β and 1α,25(OH)2D3 could be observed in transient reporter assays for CD69. Analysis of mRNA stability using a transcription inhibitor and a 3′UTR reporter construct showed that TGF-β and 1α,25(OH)2D3 do not influence CD69 mRNA stability. Functional knockdown of Smad3 clearly demonstrated that upregulation of CD69 mRNA, in contrast to 5-LO, depends on Smad3. Comparative studies with different inhibitors for mitogen activated protein kinases (MAPKs) revealed that MAPK signalling is involved in CD69 gene regulation, whereas 5-lipoxygenase gene expression was only partly affected. Mechanistically, we found evidence that CD69 gene upregulation depends on TAK1-mediated p38 activation. In summary, our data indicate that CD69 gene expression, conforming with 5-lipoxygenase, is regulated monocyte-specifically by the physiologic stimuli TGF-β and 1α,25(OH)2D3 on mRNA level, although different mechanisms account for the upregulation of each gene

Establishment of novel long-term cultures from EpCAM positive and negative circulating tumour cells from patients with metastatic gastroesophageal cancer

Circulating tumour cell (CTC) enumeration and profiling has been established as a valuable clinical tool in many solid malignancies. A key challenge in CTC research is the limited number of cells available for study. Ex vivo CTC culture permits expansion of these rare cell populations for detailed characterisation, functional assays including drug sensitivity testing, and investigation of the pathobiology of metastases. We report for the first time the establishment and characterisation of two continuous CTC lines from patients with gastroesophageal cancer. The two cell lines (designated UWG01CTC and UWG02CTC) demonstrated rapid tumorigenic growth in immunodeficient mice and exhibit distinct genotypic and phenotypic profiles which are consistent with the tumours of origin. UWG02CTC exhibits an EpCAM+, cytokeratin+, CD44+ phenotype, while UWG01CTC, which was derived from a patient with metastatic neuroendocrine cancer, displays an EpCAM−, weak cytokeratin phenotype, with strong expression of neuroendocrine markers. Further, the two cell lines show distinct differences in drug and radiation sensitivity which match differential cancer-associated gene expression pathways. This is strong evidence implicating EpCAM negative CTCs in metastasis. These novel, well characterised, long-term CTC cell lines from gastroesophageal cancer will facilitate ongoing research into metastasis and the discovery of therapeutic targets

Factorizations of Elements in Noncommutative Rings: A Survey

We survey results on factorizations of non zero-divisors into atoms (irreducible elements) in noncommutative rings. The point of view in this survey is motivated by the commutative theory of non-unique factorizations. Topics covered include unique factorization up to order and similarity, 2-firs, and modular LCM domains, as well as UFRs and UFDs in the sense of Chatters and Jordan and generalizations thereof. We recall arithmetical invariants for the study of non-unique factorizations, and give transfer results for arithmetical invariants in matrix rings, rings of triangular matrices, and classical maximal orders as well as classical hereditary orders in central simple algebras over global fields.Comment: 50 pages, comments welcom

Retrospective evaluation of the use of pembrolizumab in malignant mesothelioma in a real-world Australian population

Introduction: We investigated the efficacy and toxicity of pembrolizumab in patients with mesothelioma from a real-world Australian population. We aimed to determine clinical factors and predictive biomarkers that could help select patients who are likely to benefit from pembrolizumab. Method: Patients with mesothelioma who were treated with pembrolizumab as part of the Insurance and Care New South Wales compensation scheme were included. Clinical information was collected retrospectively. Tumor biomarkers such as programmed death-ligand 1 (PD-L1), BAP1, and CD3-positive (CD3+) tumor-infiltrating lymphocytes (TILs) were examined using archival formalin-fixed paraffin-embedded tumor samples. Results: A total of 98 patients were included with a median age of 70 years (range, 46–91 y); 92% were men; 76% had epithelioid subtype; 21% had an Eastern Cooperative Oncology Group (ECOG) performance status of 0. Pembrolizumab was used as second-line or subsequent-line treatment in 94 patients and as first-line treatment in four patients. The overall response rate was 18%, and the disease control rate was 56%. The median progression-free survival (PFS) was 4.8 months (95% confidence interval: 3.6–6.2), and the median overall survival (OS) was 9.5 months (95% confidence interval: 6.6–13.7). Immune-related adverse events occurred in 27% of patients, of which nine (9%) were of grade 3 or higher. In the multivariable analysis, factors independently associated with longer PFS included baseline ECOG status of 0 (median PFS: 12 mo versus 4 mo, p < 0.01) and PD-L1 tumor proportion score of greater than or equal to 1% (median PFS: 6 mo versus 4 mo, p < 0.01). Baseline platelet count of less than or equal to 400 × 109/liter was independently associated with longer PFS and OS (median PFS: 6 mo versus 2 mo, p = 0.05; median OS: 10 mo versus 4 mo, p = 0.01), whereas lack of pretreatment dexamethasone was independently associated with OS but not PFS (median OS: 10 mo versus 3 mo, p = 0.01). The odds of response were higher for patients with baseline ECOG status of 0 (p = 0.02) and with greater than or equal to 5% CD3+ TILs in the tumor (p < 0.01). PD-L1 expression, BAP1 loss, and CD3+ TILs in the stroma were not significantly associated with the overall response rate. Conclusions: Immunotherapy is a reasonable treatment option for patients with mesothelioma. Our results are comparable to other clinical trials investigating pembrolizumab in mesothelioma in terms of response. Good performance status assessment remains the most robust predictor for patient outcomes. CD3+ TILs in the tumor may help select patients that are likely to respond to pembrolizumab, whereas factors such as PD-L1 expression, baseline platelet count, and lack of pretreatment dexamethasone may help predict survival outcomes from pembrolizumab treatment

The effectiveness of sewage treatment processes to remove faecal pathogens and antibiotic residues

Pathogens and antibiotics enter the aquatic environment via sewage effluents and may pose a health risk to wild life and humans. The aim of this study was to determine the levels of faecal bacteria, and selected antibiotic residues in raw wastewater and treated sewage effluents from three different sewage treatment plants in the Western Cape, South Africa. Sewage treatment plant 1 and 2 use older technologies, while sewage treatment plant 3 has been upgraded and membrane technologies were incorporated in the treatment processes. Coliforms and Escherichia coli (E. coli) were used as bioindicators for faecal bacteria. A chromogenic test was used to screen for coliforms and E. coli. Fluoroquinolones and sulfamethoxazole are commonly used antibiotics and were selected to monitor the efficiency of sewage treatment processes for antibiotic removal. Enzyme Linked Immunosorbent Assays (ELISAs) were used to quantitate antibiotic residues in raw and treated sewage. Raw intake water at all treatment plants contained total coliforms and E. coli. High removal of E. coli by treatment processes was evident for treatment plant 2 and 3 only. Fluoroquinolones and sulfamethoxazole were detected in raw wastewater from all sewage treatment plants. Treatment processes at plant 1 did not reduce the fluoroquinolone concentration in treated sewage effluents. Treatment processes at plant 2 and 3 reduced the fluoroquinolone concentration by 21% and 31%, respectively. Treatment processes at plant 1 did not reduce the sulfamethoxazole concentration in treated sewage effluents. Treatment processes at plant 2 and 3 reduced sulfamethoxazole by 34% and 56%, respectively. This study showed that bacteria and antibiotic residues are still discharged into the environment. Further research needs to be undertaken to improve sewage treatment technologies, thereby producing a better quality treated sewage effluent

Post-Transcriptional Regulation of 5-Lipoxygenase mRNA Expression via Alternative Splicing and Nonsense-Mediated mRNA Decay

5-Lipoxygenase (5-LO) catalyzes the two initial steps in the biosynthesis of leukotrienes (LT), a group of inflammatory lipid mediators derived from arachidonic acid. Here, we investigated the regulation of 5-LO mRNA expression by alternative splicing and nonsense-mediated mRNA decay (NMD). In the present study, we report the identification of 2 truncated transcripts and 4 novel 5-LO splice variants containing premature termination codons (PTC). The characterization of one of the splice variants, 5-LOΔ3, revealed that it is a target for NMD since knockdown of the NMD factors UPF1, UPF2 and UPF3b in the human monocytic cell line Mono Mac 6 (MM6) altered the expression of 5-LOΔ3 mRNA up to 2-fold in a cell differentiation-dependent manner suggesting that cell differentiation alters the composition or function of the NMD complex. In contrast, the mature 5-LO mRNA transcript was not affected by UPF knockdown. Thus, the data suggest that the coupling of alternative splicing and NMD is involved in the regulation of 5-LO gene expression