Commissioning and initial experience with a commercial software for in vivo volumetric dosimetry

Introduction and purposeDosimetry Check (DC) (Math Resolutions) is a commercial EPID-based dosimetry software, which allows performing pre-treatment and transit dosimetry. DC provides an independent verification of the treatment, being potentially of great interest due to the high benefits of the in vivo volumetric dosimetry, which guarantee the treatment delivery and anatomy constancy. The aim of this work is to study the differences in dose between DC and the Treatment Planning System (TPS) to establish an accuracy level of the system.Material and methodsDC v.3.8 was used along with Varian Clinac iX accelerator equipped with EPID aS1000 and Eclipse v.10.0 with AAA and Acuros XB calculation algorithms. The DC evaluated version is based on a pencil beam calculation algorithm. Various plans were generated over several homogeneous and heterogeneous phantoms. Isocentre point doses and gamma analysis were evaluated.ResultsTotal dose differences at the isocentre between DC and TPS for the studied plans are less than 2%, but single field contributions achieve greater values. In the presence of heterogeneities, the discrepancies can reach up to 15%. In transit mode, DC does not consider properly the couch attenuation, especially when there is an air gap between phantom and couch.ConclusionsThe possibility of this in vivo evaluation and the potentiality of this new system have a very positive impact on improving patient QA. But improvements are required in both calculation algorithm and integration with the record and verify system.     

Genetically encoded sensors for measuring histamine release both in vitro and in vivo

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A sensitive GRAB sensor for detecting extracellular ATP in vitro and in vivo

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Preconditioning influences mesenchymal stem cell properties in vitro and in vivo

Various diseases and toxic factors easily impair cellular and organic functions in mammals. Organ transplantation is used to rescue organ function, but is limited by scarce resources. Mesenchymal stem cell (MSC)‐based therapy carries promising potential in regenerative medicine because of the self‐renewal and multilineage potency of MSCs; however, MSCs may lose biological functions after isolation and cultivation for a long time in vitro. Moreover, after they are injected in vivo and migrate into the damaged tissues or organs, they encounter a harsh environment coupled with death signals due to the inadequate tensegrity structure between the cells and matrix. Preconditioning, genetic modification and optimization of MSC culture conditions are key strategies to improve MSC functions in vitro and in vivo, and all of these procedures will contribute to improving MSC transplantation efficacy in tissue engineering and regenerative medicine. Preconditioning with various physical, chemical and biological factors is possible to preserve the stemness of MSCs for further application in studies and clinical tests. In this review, we mainly focus on preconditioning and the corresponding mechanisms for improving MSC activities in vitro and in vivo; we provide a glimpse into the promotion of MSC‐based cell therapy development for regenerative medicine. As a promising consequence, MSC transplantation can be applied for the treatment of some terminal diseases and can prolong the survival time of patients in the near future.     

Activity-based bioluminescence probes for in vivo sensing applications

Bioluminescence imaging is a highly sensitive technique commonly used for various in vivo applications. Recent efforts to expand the utility of this modality have led to the development of a suite of activity-based sensing (ABS) probes for bioluminescence imaging by ‘caging’ of luciferin and its structural analogs. The ability to selectively detect a given biomarker has presented researchers with many exciting opportunities to study both health and disease states in animal models. Here, we highlight recent (2021–2023) bioluminescence-based ABS probes with an emphasis on probe design and in vivo validation experiments.     

Cholesteryl hemiazelate identified in CVD patients causes in vitro and in vivo inflammation

Oxidation of PUFAs in LDLs trapped in the arterial intima plays a critical role in atherosclerosis. Though there have been many studies on the atherogenicity of oxidized derivatives of PUFA-esters of cholesterol, the effects of cholesteryl hemiesters (ChEs), the oxidation end products of these esters, have not been studied. Through lipidomics analyses, we identified and quantified two ChE types in the plasma of CVD patients and identified four ChE types in human endarterectomy specimens. Cholesteryl hemiazelate (ChA), the ChE of azelaic acid (n-nonane-1,9-dioic acid), was the most prevalent ChE identified in both cases. Importantly, human monocytes, monocyte-derived macrophages, and neutrophils exhibit inflammatory features when exposed to subtoxic concentrations of ChA in vitro. ChA increases the secretion of proinflammatory cytokines such as interleukin-1β and interleukin-6 and modulates the surface-marker profile of monocytes and monocyte-derived macrophage. In vivo, when zebrafish larvae were fed with a ChA-enriched diet, they exhibited neutrophil and macrophage accumulation in the vasculature in a caspase 1- and cathepsin B-dependent manner. ChA also triggered lipid accumulation at the bifurcation sites of the vasculature of the zebrafish larvae and negatively impacted their life expectancy. We conclude that ChA behaves as an endogenous damage-associated molecular pattern with inflammatory and proatherogenic properties.     

Oncostatin M inhibits differentiation of rat stem Leydig cells in vivo and in vitro

Oncostatin M (OSM) is a pleiotropic cytokine within the interleukin six family of cytokines, which regulate cell growth and differentiation in a wide variety of biological systems. However, its action and underlying mechanisms on stem Leydig cell development are unclear. The objective of the present study was to investigate whether OSM affects the proliferation and differentiation of rat stem Leydig cells. We used a Leydig cell regeneration model in rat testis and a unique seminiferous tubule culture system after ethane dimethane sulfonate (EDS) treatment to assess the ability of OSM in the regulation of proliferation and differentiation of rat stem Leydig cells. Intratesticular injection of OSM (10 and 100 ng/testis) from post‐EDS day 14 to 28 blocked the regeneration of Leydig cells by reducing serum testosterone levels without affecting serum luteinizing hormone and follicle‐stimulating hormone levels. It also decreased the levels of Leydig cell‐specific mRNAs (Lhcgr, Star, Cyp11a1, Hsd3b1, Cyp17a1 and Hsd11b1) and their proteins by the RNA‐Seq and Western blotting analysis. OSM had no effect on the proliferative capacity of Leydig cells in vivo. In the seminiferous tubule culture system, OSM (0.1, 1, 10 and 100 ng/mL) inhibited the differentiation of stem Leydig cells by reducing medium testosterone levels and downregulating the expression of Leydig cell‐specific genes (Lhcgr, Star, Cyp11a1, Hsd3b1, Cyp17a1 and Hsd11b1) and their proteins. OSM‐mediated action was reversed by S3I‐201 (a STAT3 antagonist) or filgotinib (a JAK1 inhibitor). These data suggest that OSM is an inhibitory factor of rat stem Leydig cell development.     

Mobile genetic elements: in silico,in vitro,in vivo

Mobile genetic elements (MGEs), also called transposable elements (TEs), represent universal components of most genomes and are intimately involved in nearly all aspects of genome organization, function and evolution. However, there is currently a gap between the fast pace of TE discovery in silico, driven by the exponential growth of comparative genomic studies, and a limited number of experimental models amenable to more traditional in vitro and in vivo studies of structural, mechanistic and regulatory properties of diverse MGEs. Experimental and computational scientists came together to bridge this gap at a recent conference, ‘Mobile Genetic Elements: in silico, in vitro, in vivo’, held at the Marine Biological Laboratory (MBL) in Woods Hole, MA, USA.     

Proscillaridin A induces apoptosis and inhibits the metastasis of osteosarcoma in vitro and in vivo

The side effects of chemotherapy, drug resistance, and tumor metastasis hinder the development of treatment for osteosarcoma, leading to poor prognosis of patients with the disease. Proscillaridin A, a kind of cardiac glycoside, has been proven to have anti-proliferative properties in many malignant tumors, but the efficacy of the drug in treating osteosarcoma is unclear. In the present study, we assessed the effects of Proscillaridin A on osteosarcoma and investigated its underlying action mechanism. The cell cytotoxicity assay showed that Proscillaridin A significantly inhibited the proliferation of 143B cells in a dose- and time-dependent manner. Also, flow cytometry and invasion assay revealed that Proscillaridin A induced apoptosis and reduced 143B cell motility. Western blotting and PCR were used to detect the expressions of Bcl-xl and MMP2 and showed that mRNA/protein expression levels decreased significantly in Proscillaridin A-treated osteosarcoma cells. Using a mouse xenograft model, we found that Proscillaridin A treatment significantly inhibited tumor growth and lung metastasis in vivo and decreased the expression levels of Bcl-xl and MMP2. No noticeable side effect was observed in the liver, kidney, and hematological functions. Conclusively, Proscillaridin A suppressed proliferation, induced apoptosis, and inhibited 143B cell metastasis in vitro and in vivo, and these effects could be mediated by downregulating the expressions of Bcl-xl and MMP2.     

Neurons require glucose uptake and glycolysis in vivo

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Gut bacterial nutrient preferences quantified in vivo

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HAM: A new epitope-tag for in vivo protein labeling

The ability to metabolically label proteins with ³⁵S-methionine is critical for the analysis of protein synthesis and turnover. Despite the importance of this approach, however, efficient labeling of proteins in vivo is often limited by a low number of available methionine residues, or by deleterious side-effects associated with protein overexpression. To overcome these limitations, we have created a methionine-rich variant of the widely used HA tag, called HAM, for use with ectopically expressed proteins. Here we describe the development of a series of vectors, and corresponding antisera, for the expression and detection of HAM-tagged proteins in mammalian cells. We show that the HAM tag dramatically improves the sensitivity of ³⁵S-methionine labeling, and permits the analysis of Myc oncoprotein turnover even when HAM-tagged Myc is expressed at levels comparable to that of the endogenous protein. Because of the improved sensitivity provided by the HAM tag, the vectors and antisera described here should be useful for the analysis of protein synthesis and destruction at physiological levels of protein expression.     

In silico,in vitro,and in vivo machine learning in synthetic biology and metabolic engineering

Among the main learning methods reviewed in this study and used in synthetic biology and metabolic engineering are supervised learning, reinforcement and active learning, and in vitro or in vivo learning.In the context of biosynthesis, supervised machine learning is being exploited to predict biological sequence activities, predict structures and engineer sequences, and optimize culture conditions.Active and reinforcement learning methods use training sets acquired through an iterative process generally involving experimental measurements. They are applied to design, engineer, and optimize metabolic pathways and bioprocesses.The nascent but promising developments with in vitro and in vivo learning comprise molecular circuits performing simple tasks such as pattern recognition and classification.     

Betulinic acid induces apoptosis and suppresses metastasis in hepatocellular carcinoma cell lines in vitro and in vivo

Hepatocellular carcinoma (HCC) is a high incidence and mortality malignant tumour globally. Betulinic acid (BA) is a pentacyclic triterpenoid with potential pro‐apoptotic activities which widely found in many plants. In this study, we determined the effects of BA on proliferation, apoptosis, invasion, and metastasis in HCC cell lines and on tumour growth and pulmonary metastasis in mice. The results suggested that BA could inhibit cell viability and proliferation of HCC cell lines including HepG2, LM3, and MHCC97H. In addition, BA induced apoptosis of HepG2 cells characterised condensed nuclei and nuclear fragmentation. Moreover, western blot analysis showed that BA‐induced apoptosis associated with increasing of pro‐apoptotic protein Bax and cleaved caspase‐3 and decreasing of anti‐apoptotic protein Bcl‐2. Meanwhile, BA also reduced the reactive oxygen species (ROS) level. Furthermore, BA also significantly inhibited HCC growth in vivo and blocked pulmonary metastasis of HCC by regulating the metastasis‐related proteins including MMP‐2, MMP‐9, and TIMP2 without obvious toxicity. In all, the present study suggested that BA might be a promising anti‐HCC drug candidate by inhibiting proliferation, inducing apoptosis, and blocking metastasis.     

Long-term ex vivo and in vivo monitoring of tumor progression by using dual luciferases

We propose a new concept of tumor progression monitoring using dual luciferases in living animals to reduce stress for small animals and the cost of luciferin. The secreted Cypridina luciferase (CLuc) was used as an ex vivo indicator to continuously monitor tumor progression. On the other hand, the non-secreted firefly luciferase was used as an in vivo indicator to analyze the spatial distribution of the tumor at suitable time points indicated by CLuc. Thus, the new monitoring systems that use dual luciferases are available, allowing long-term bioluminescence imaging under minimal stress for the experimental animals.