Neutrophils and monocytes belong to the first line of immune defence cells and are recruited to sites of inflammation during infection or sterile injury. Both cells contain huge amounts of the ...heterodimeric protein S100A8/A9 in their cytoplasm. S100A8/A9 belongs to the Ca
binding S100 protein family and has recently gained a lot of interest as a critical alarmin modulating the inflammatory response after its release (extracellular S100A8/A9) from neutrophils and monocytes. Extracellular S100A8/A9 interacts with the pattern recognition receptors Toll-like receptor 4 (TLR4) and Receptor for Advanced Glycation Endproducts (RAGE) promoting cell activation and recruitment. Besides its biological function, S100A8/A9 (also known as myeloid related protein 8/14, MRP8/14) was identified as interesting biomarker to monitor disease activity in chronic inflammatory disorders including inflammatory bowel disease and rheumatoid arthritis. Furthermore, S100A8/A9 has been tested successfully in pre-clinical imaging studies to localize sites of infection or sterile injury. Finally, recent evidence using small molecule inhibitors for S100A8/A9 also suggests that blocking S100A8/A9 activity exerts beneficial effects on disease activity in animal models of autoimmune diseases including multiple sclerosis, systemic lupus erythematosus, rheumatoid arthritis and inflammatory bowel disease. This review will provide a comprehensive and detailed overview into the structure and biological function of S100A8/A9 and also will give an outlook in terms of diagnostic and therapeutic applications targeting S100A8/A9.
► Biotechnologically relevant aspects of Pichia pastoris promoters are reviewed. ► The regulation of the alcohol oxidase 1 promoter (PAOX1) is discussed. ► Known alternative promoters such as PTEF1, ...PPHO89, PTHI11 or PAOD are described. ► Regulatory networks involving Hac1, Fep1, Yap1 and Rop are highlighted.
The methylotrophic yeast Pichia pastoris is a widely used host for heterologous protein production. Along with favorable properties such as growth to high cell density and high capacities for protein secretion, P. pastoris provides a strong, methanol inducible promoter of the alcohol oxidase 1 (AOX1) gene. The regulation of this promoter has been extensively studied in recent years by characterizing cis-acting sequence elements and trans-acting factors, revealing insights into underlying molecular mechanisms. However, new alternative promoters have also been identified and characterized by means of their transcriptional regulation and feasibility for protein production using P. pastoris. Besides the often applied GAP promoter, these include a variety of constitutive promoters from housekeeping genes (e.g. TEF1, PGK1, TPI1) and inducible promoters from particular biochemical pathways (e.g. PHO89, THI11, AOD). In addition to these promoter sequence/function based studies, transcriptional regulation has also been investigated by characterizing transcription factors (TFs) and their modes of controlling bioprocess relevant traits. TFs involved in such diverse cellular processes such as the unfolded protein response (UPR) (Hac1p), iron uptake (Fep1p) and oxidative stress response (Yap1p) have been studied. Understanding of these natural transcriptional regulatory networks is a helpful basis for synthetic biology and metabolic engineering approaches that enable the design of tailor-made production strains.
In recent years, local authorities in the UK have begun to adopt a variety of “smart” technological changes to enhance service delivery. These changes are having profound impacts on the structure of ...public administration. Focusing on the particular case of artificial intelligence, specifically autonomous agents and predictive analytics, a combination of desk research, a survey questionnaire, and interviews were used to better understand the extent and nature of these changes in local government. Findings suggest that local authorities are beginning to adopt smart technologies and that these technologies are having an unanticipated impact on how public administrators and computational algorithms become imbricated in the delivery of public services. This imbrication is described as algorithmic bureaucracy, and it provides a framework within which to explore how these technologies transform both the socio‐technical relationship between workers and their tools, as well as the ways that work is organized in the public sector.
The locoregional interventional oncological treatment approach is an accepted modality for liver neoplasms, especially for hepatocellular carcinoma (HCC) and oligonodular liver metastases.
The main ...aim of ablation therapies like microwave ablation (MWA) is to eradicate all malignant cells in a minimally invasive technique under imaging guidance while preserving the healthy tissue with a sufficient safety margin (at least 5 mm) surrounding the ablated lesion.
Ablation therapy can be performed via a percutaneous, laparoscopic or intraoperative approach under ultrasound, MRI or CT guidance for adequate localization and monitoring of the ablation process.
Ablation is the method of choice for oligonodular HCCs ≤ 3 cm. The technical success rate varies from 88 % to 98 % and progression-free survival (PFS) at 3 years from 27 % to 91.7 %. The same criteria apply to the therapy of liver metastases.
· Careful selection of patients proves to be essential for optimum results of MWA. · Interventionists should be familiar with all aspects of complication and rapid assessment of imaging methods in order to evaluate induced damage by thermal ablation. · MWA seems to have some advantages over radiofrequency ablation, like shorter ablation time, less pain, less heat sink effect; however, scientific proof is needed.
· Vogl TJ, Nour-Eldin A, Hammerstingl RM et al. Microwave Ablation (MWA): Basics, Technique and Results in Primary and Metastatic Liver Neoplasms - Review Article. Fortschr Röntgenstr 2017; 189: 1055 - 1066.
In this article, the authors discuss the technical background and summarize the current body of literature regarding virtual monoenergetic (VM) images derived from dual-energy CT data, which can be ...reconstructed between 40 and 200 keV. Substantially improved iodine attenuation at lower kiloelectron volt levels and reduced beam-hardening artifacts at higher kiloelectron volt levels have been demonstrated from all major manufacturers of dual-energy CT units. Improved contrast attenuation with VM imaging at lower kiloelectron volt levels enables better delineation and diagnostic accuracy in the detection of various vascular or oncologic abnormalities. Low-kiloelectron-volt VM imaging may be useful for salvaging CT studies with suboptimal contrast material delivery or providing additional information on the arterial vasculature obtained from venous phase acquisitions. For patients with renal impairment, substantial reductions in the use of iodinated contrast material can be achieved by using lower-energy VM imaging. The authors recommend routine reconstruction of VM images at 50 keV when using dual-energy CT to exploit the increased contrast properties. For reduction of beam-hardening artifacts, VM imaging at 120 keV is useful for the initial assessment.
•A CRISPR/Cas9 system has been developed for the methylotrophic yeast P. pastoris.•Several parts for expression (promoters for CAS9 and gRNAs, CAS9 CDSs…) needed optimization.•Only 6/95 constructs ...designed achieved high efficiency genome targeting.•Targeting rates close to 100% and multiplexing were demonstrated for several genes.•This report may serve as a guideline to implement CRISPR/Cas9 in other eukaryotes.
The methylotrophic yeast Pichia pastoris (Komagataella phaffii) is one of the most commonly used expression systems for heterologous protein production. However the recombination machinery in P. pastoris is less effective in contrast to Saccharomyces cerevisiae, where efficient homologous recombination naturally facilitates genetic modifications. The lack of simple and efficient methods for gene disruption and specifically integrating cassettes has remained a bottleneck for strain engineering in P. pastoris. Therefore tools and methods for targeted genome modifications are of great interest.
Here we report the establishment of CRISPR/Cas9 technologies for P. pastoris and demonstrate targeting efficiencies approaching 100%. However there appeared to be a narrow window of optimal conditions required for efficient CRISPR/Cas9 function for this host. We systematically tested combinations of various codon optimized DNA sequences of CAS9, different gRNA sequences, RNA Polymerase III and RNA Polymerase II promoters in combination with ribozymes for the expression of the gRNAs and RNA Polymerase II promoters for the expression of CAS9. Only 6 out of 95 constructs were functional for efficient genome editing.
We used this optimized CRISPR/Cas9 system for gene disruption studies, to introduce multiplexed gene deletions and to test the targeted integration of homologous DNA cassettes. This system allows rapid, marker-less genome engineering in P. pastoris enabling unprecedented strain and metabolic engineering applications.
Colorectal cancer (CRC) is one of the most common tumor entities worldwide and a common cause of cancer-associated death. Colorectal cancer liver metastases (CRLM) thereby constitute a severe ...life-limiting factor. The therapy of CRLM presents a major challenge and surgical resection as well as systemic chemotherapy remain the first-line treatment options. Over the years several locoregional, vascular- and image-based treatments offered by interventional radiologists have emerged when conventional therapies fail, or metastases recurrence occurs. Among such options is the conventional/traditional transarterial chemoembolization (cTACE) by local injection of a combination of chemotherapeutic- and embolic-agents. A similar treatment is the more recent irinotecan-loaded drug-eluting beads TACE (DEBIRI-TACE), which are administered using the same approach. Numerous studies have shown that these different types of chemoembolization can be applied in different clinical settings safely. Furthermore, such treatments can also be combined with other local or systemic therapies. Unfortunately, due to the incoherent patient populations of studies investigating TACE in CRLM, critics state that the definite evidence supporting positive patient outcomes is still lacking. In the following article we review studies on conventional and DEBIRI-TACE. Although highly dependent on the clinical setting, prior therapies and generally the study population, cTACE and DEBIRI-TACE show comparable results. We present the most representative studies on the different chemoembolization procedures and compare the results. Although there is compelling evidence for both approaches, further studies are necessary to determine which patients profit most from these therapies. In conclusion, we determine TACE to be a viable option in CRLM in different clinical settings. Nevertheless, a multidisciplinary approach is desired to offer patients the best possible care.
Graphical abstract Highlights ► The application of Pichia pastoris for biopharmaceutical production is described. ► Synthetic biology approaches and perspectives to improve production are reviewed. ► ...Glycoengineering efforts to produce humanized, uniform glycoproteins are covered. ► The design and application of synthetic promoter variants are highlighted. ► The molecular toolbox available for synthetic biology in P. pastoris is discussed.
The tumor microenvironment plays an important role in modulating tumor progression. Earlier, we showed that S100A8/A9 proteins secreted by myeloid-derived suppressor cells (MDSC) present within ...tumors and metastatic sites promote an autocrine pathway for accumulation of MDSC. In a mouse model of colitis-associated colon cancer, we also showed that S100A8/A9-positive cells accumulate in all regions of dysplasia and adenoma. Here we present evidence that S100A8/A9 interact with RAGE and carboxylated glycans on colon tumor cells and promote activation of MAPK and NF-κB signaling pathways. Comparison of gene expression profiles of S100A8/A9-activated colon tumor cells versus unactivated cells led us to identify a small cohort of genes upregulated in activated cells, including Cxcl1, Ccl5 and Ccl7, Slc39a10, Lcn2, Zc3h12a, Enpp2, and other genes, whose products promote leukocyte recruitment, angiogenesis, tumor migration, wound healing, and formation of premetastatic niches in distal metastatic organs. Consistent with this observation, in murine colon tumor models we found that chemokines were upregulated in tumors, and elevated in sera of tumor-bearing wild-type mice. Mice lacking S100A9 showed significantly reduced tumor incidence, growth and metastasis, reduced chemokine levels, and reduced infiltration of CD11b(+)Gr1(+) cells within tumors and premetastatic organs. Studies using bone marrow chimeric mice revealed that S100A8/A9 expression on myeloid cells is essential for development of colon tumors. Our results thus reveal a novel role for myeloid-derived S100A8/A9 in activating specific downstream genes associated with tumorigenesis and in promoting tumor growth and metastasis.
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