Intensity-modulated radiotherapy with planned Gamma Knife radiosurgery boost for head and neck cancer with extensive disease in proximity to critical structures.

BACKGROUND: To describe intensity-modulated radiotherapy (IMRT) with Gamma Knife Radiosurgery (GKRS) boost for locally advanced head and neck cancer (HNC) with disease near dose-limiting structures. METHODS: Patients with HNC treated with IMRT/GKRS as part of a combined modality approach between 2011 and 2021 were reviewed. Local control, overall survival and disease-specific survival were estimated using the Kaplan Meier method. RESULTS: Twenty patients were included. Nineteen patients had T3-4 tumors. Median follow-up was 26.3 months. GKRS site control was 95%. Two patients progressed at the treated primary site, one patient failed at the edge of the GKRS treatment volume, with no perineural or intracranial failure. 2-year OS was 94.7% (95% CI: 85.2%-100%). Concurrent chemotherapy was given in nine patients (45%). One patient (5%) received induction/concurrent chemotherapy. Brain radionecrosis occurred in three patients, one of which was biopsy-proven. CONCLUSIONS: IMRT plus GKRS boost results in excellent disease control near critical structures with minimal toxicity.

Clinical outcomes of dose-escalated re-irradiation in patients with recurrent high-grade glioma.

Background: Re-irradiation for recurrent gliomas is a controversial treatment option with no clear standard dose or concurrent systemic therapy. Methods: This series represents a single-institution retrospective review of patients treated with re-irradiation for recurrent high-grade glioma. After 2012, patients were commonly offered concurrent bevacizumab as a cytoprotective agent against radiation necrosis. Kaplan-Meier method was used to estimate overall survival and progression-free survival. Cox proportional hazards regression was used to identify factors associated with overall survival and progression-free survival. Results: Between 2001 and 2021, 52 patients underwent re-irradiation for a diagnosis of recurrent high-grade glioma. 36 patients (69.2%) had a histologic diagnosis of glioblastoma at the time of re-irradiation. The median BED10 (biological equivalent dose 10 Gy) of re-irradiation was 53.1 Gy. Twenty-one patients (40.4%) received concurrent bevacizumab with re-irradiation. Median survival for the entire cohort and for glioblastoma at the time of recurrence patients was 6.7 months and 6.0 months, respectively. For patients with glioblastoma at the time of recurrence, completing re-irradiation (HR 0.03, P < .001), use of concurrent bevacizumab (HR 0.3, P = .009), and the BED10 (HR 0.9, P = .005) were predictive of overall survival. Nine patients developed grade 3-5 toxicity; of these, 2 received concurrent bevacizumab and 7 did not (P = .15). Conclusion: High dose re-irradiation with concurrent bevacizumab is feasible in patients with recurrent gliomas. Concurrent bevacizumab and increasing radiation dose may improve survival in patients with recurrent glioblastoma.

Intermodel comparison for the radiological assessment of the Zapadnoe and Tessenderlo case studies with implications for selection of remediation strategy.

Risk assessment provides a key input for determining the need for and extent of remedial actions necessary for sites contaminated with naturally occurring radioactive material or nuclear legacy sites. The choice of a modelling approach for risk assessment, and the corresponding toolsets should fit the assessment context, taking account of the complexity, and be clearly related to the questions to be addressed in the decision-making process. One of the objectives of Working Group 1 of IAEA Modelling and Data for Radiological Impact Assessments II (MODARIA II) Programme is to perform intermodel comparisons for case studies of selected sites, in particular, to help illustrate the applicability of different models and approaches as inputs to decision-making processes. This intercomparison exercise, which included the analysis of potential consequences on the management strategy for contaminated sites, has been performed for two sites: The former uranium mill tailings facility at Zapadnoe, Ukraine, and the phosphate processing facility at Tessenderlo, Belgium. Several models and computer codes have been used for one or both of these cases: AMBER, GoldSim, NORM And LegacY Site Assessment, Preliminary Remediation Goals (PRG)-dose compliance concentration calculator, and RESRAD-OFFSITE. The assessments explore the implications of using differing assessment frameworks and assumptions, as well as alternative modelling tools, on model outputs and as input for corresponding decisions on remediation strategy. This paper reviews both similarities and differences in the results of assessments performed using these different models. It discusses how different approaches can complement one another to help build confidence in the evidence base underpinning decisions. It also discusses the appropriateness of the different modelling approaches in a given assessment context. In one of the case studies in particular (Tessenderlo case study), the remediation strategy is essentially driven by the contamination of the site with heavy metals, such as cadmium. This has significant consequences on the choice of the most adequate approaches and scenarios for assessing the radiological risk and balancing their relative importance with other impacts. The development of a holistic approach to risk assessment is, therefore, highlighted.

Characterization and implementation of a miniature X-ray system for live cell microscopy.

The study of radiation effects on biological tissues is a diverse field of research with direct applications to improve human health, in particular in the contexts of radiation therapy and space exploration. Understanding the DNA damage response following radiation exposure, which is a key determinant for mutagenesis, requires reproducible methods for delivering known doses of ionizing radiation (IR) in a controlled environment. Multiple IR sources, including research X-ray and gamma-ray irradiators are routinely used in basic and translational research with cell and animal models. These systems are however not ideal when a high temporal resolution is needed, for example to study early DNA damage responses with live cell microscopy. Here, we characterize the dose rate and beam properties of a commercial, miniature, affordable, and versatile X-ray source (Mini-X). We describe how to use Mini-X on the stage of a fluorescence microscope to deliver high IR dose rates (up to 29 Gy/min) or lower dose rates (≤ 0.1 Gy/min) in live cell imaging experiments. This article provides a blueprint for radiation biology applications with high temporal resolution, with a step-by-step guide to implement a miniature X-ray system on an imaging platform, and the information needed to characterize the system.

Preoperative Optimization of Promotility in Robotic Prostatectomy and Minimally Invasive Kidney Surgery.

INTRODUCTION: Patients are routinely discharged postoperative day 1 following minimally invasive surgery (MIS) for prostate cancer and kidney cancer. Delays in discharge are often related to gastrointestinal symptoms such as nausea, abdominal pain and vomiting; however, the role of baseline constipation in these symptoms and resultant delays in discharge is unclear. We conducted a prospective observational study to describe the incidence of baseline constipation among patients undergoing MIS prostate and kidney surgery, and its relationship to length of stay (LOS). METHODS: Consenting adult patients undergoing MIS procedures for kidney and prostate cancer completed constipation symptom questionnaires perioperatively. Clinicopathological data were collected prospectively. Delay in discharge, defined as LOS >2 days, was the primary outcome. Patients were stratified by the primary outcome and preoperative Patient Assessment of Constipation Symptoms (PAC-SYM) scores were compared. RESULTS: A total of 97 patients enrolled, of whom 29 underwent radical nephrectomy, 34 underwent robotic partial nephrectomy and 34 underwent robotic prostatectomy. Constipation symptoms were reported in 67/97 patients (69%). A total of 17/97 patients (18%) had a delay in discharge. Patients who discharged on time had a median PAC-SYM score of 2 (IQR 2-9) compared to 4 (IQR 0-7.5) for those with a delay (p=0.021). Patients who had a delay with gastrointestinal symptoms had a median PAC-SYM score of 5 (IQR 1.5-11.5, p=0.032). CONCLUSIONS: Seven out of 10 patients undergoing routine MIS procedures report constipation symptoms, which may represent a target for preoperative interventions to reduce LOS after surgery.

A revolution in biochemistry and molecular biology education informed by basic research to meet the demands of 21st century career paths.

The National Science Foundation estimates that 80% of the jobs available during the next decade will require math and science skills, dictating that programs in biochemistry and molecular biology must be transformative and use new pedagogical approaches and experiential learning for careers in industry, research, education, engineering, health-care professions, and other interdisciplinary fields. These efforts require an environment that values the individual student and integrates recent advances from the primary literature in the discipline, experimentally directed research, data collection and analysis, and scientific writing. Current trends shaping these efforts must include critical thinking, experimental testing, computational modeling, and inferential logic. In essence, modern biochemistry and molecular biology education must be informed by, and integrated with, cutting-edge research. This environment relies on sustained research support, commitment to providing the requisite mentoring, access to instrumentation, and state-of-the-art facilities. The academic environment must establish a culture of excellence and faculty engagement, leading to innovation in the classroom and laboratory. These efforts must not lose sight of the importance of multidimensional programs that enrich science literacy in all facets of the population, students and teachers in K-12 schools, nonbiochemistry and molecular biology students, and other stakeholders. As biochemistry and molecular biology educators, we have an obligation to provide students with the skills that allow them to be innovative and self-reliant. The next generation of biochemistry and molecular biology students must be taught proficiencies in scientific and technological literacy, the importance of the scientific discourse, and skills required for problem solvers of the 21st century.

Deletion of fatty acid transport protein 2 (FATP2) in the mouse liver changes the metabolic landscape by increasing the expression of PPARα-regulated genes.

Fatty acid transport protein 2 (FATP2) is highly expressed in the liver, small intestine, and kidney, where it functions in both the transport of exogenous long-chain fatty acids and the activation of very-long-chain fatty acids. Here, using a murine model, we investigated the phenotypic impacts of deleting FATP2, followed by a transcriptomic analysis using unbiased RNA-Seq to identify concomitant changes in the liver transcriptome. WT and FATP2-null (Fatp2-/-) mice (5 weeks) were maintained on a standard chow diet for 6 weeks. The Fatp2-/- mice had reduced weight gain, lowered serum triglyceride, and increased serum cholesterol levels and attenuated dietary fatty acid absorption. Transcriptomic analysis of the liver revealed 258 differentially expressed genes in male Fatp2-/- mice and a total of 91 in female Fatp2-/- mice. These genes mapped to the following gene ontology categories: fatty acid degradation, peroxisome biogenesis, fatty acid synthesis, and retinol and arachidonic acid metabolism. Targeted RT-quantitative PCR verified the altered expression of selected genes. Of note, most of the genes with increased expression were known to be regulated by peroxisome proliferator-activated receptor α (PPARα), suggesting that FATP2 activity is linked to a PPARα-specific proximal ligand. Targeted metabolomic experiments in the Fatp2-/- liver revealed increases of total C16:0, C16:1, and C18:1 fatty acids; increases in lipoxin A4 and prostaglandin J2; and a decrease in 20-hydroxyeicosatetraenoic acid. We conclude that the expression of FATP2 in the liver broadly affects the metabolic landscape through PPARα, indicating that FATP2 provides an important role in liver lipid metabolism through its transport or activation activities.

Remodeling of Chlamydomonas Metabolism Using Synthetic Inducers Results in Lipid Storage during Growth.

Microalgae accumulate lipids during stress such as that of nutrient deprivation, concomitant with cessation of growth and depletion of chloroplasts. By contrast, certain small chemical compounds selected by high-throughput screening in Chlamydomonas reinhardtii can induce lipid accumulation during growth, maintaining biomass. Comprehensive pathway analyses using proteomics, transcriptomics, and metabolomics data were acquired from Chlamydomonas cells grown in the presence of one of two structurally distinct lipid activators. WD10784 stimulates both starch and lipid accumulation, whereas WD30030-treated cells accumulate only lipids. The differences in starch accumulation are largely due to differential effects of the two compounds on substrate levels that feed into starch synthesis and on genes encoding starch metabolic enzymes. The compounds had differential effects on photosynthesis, respiration, and oxidative stress pathways. Cells treated with WD10784 showed slowed growth over time and reduced abundance of photosynthetic proteins, decreased respiration, and increased oxidative stress proteins, glutathione, and reactive oxygen species specific to this compound. Both compounds maintained central carbon and nitrogen metabolism, including the tricarboxylic acid cycle, glycolysis, respiration, and the Calvin-Benson-Bassham cycle. There were few changes in proteins and transcripts related to fatty acid biosynthesis, whereas proteins and transcripts for triglyceride production were elevated, suggesting that lipid synthesis is largely driven by substrate availability. This study reports that the compound WD30030 and, to a lesser extent WD10784, increases lipid and lipid droplet synthesis and storage without restricting growth or biomass accumulation by mechanisms that are substantially different from nutrient deprivation.

Treatment Outcomes and Dose Rate Effects Following Gamma Knife Stereotactic Radiosurgery for Vestibular Schwannomas.

BACKGROUND: Gamma Knife radiosurgery (GKRS; Elekta AB) remains a well-established treatment modality for vestibular schwannomas. Despite highly effective tumor control, further research is needed toward optimizing long-term functional outcomes. Whereas dose-rate effects may impact post-treatment toxicities given tissue dose-response relationships, potential effects remain largely unexplored. OBJECTIVE: To evaluate treatment outcomes and potential dose-rate effects following definitive GKRS for vestibular schwannomas. METHODS: We retrospectively reviewed 419 patients treated at our institution between 1998 and 2015, characterizing baseline demographics, pretreatment symptoms, and GKRS parameters. The cohort was divided into 2 dose-rate groups based on the median value (2.675 Gy/min). Outcomes included clinical tumor control, radiographic progression-free survival, serviceable hearing preservation, hearing loss, and facial nerve dysfunction (FND). Prognostic factors were assessed using Cox regression. RESULTS: The study cohort included 227 patients with available follow-up. Following GKRS 2-yr and 4-yr clinical tumor control rates were 98% (95% CI: 95.6%-100%) and 96% (95% CI: 91.4%-99.6%), respectively. Among 177 patients with available radiographic follow-up, 2-yr and 4-yr radiographic progression-free survival rates were 97% (95% CI: 94.0%-100.0%) and 88% (95% CI: 81.2%-95.0%). The serviceable hearing preservation rate was 72.2% among patients with baseline Gardner-Robertson class I/II hearing and post-treatment audiological evaluations. Most patients experienced effective relief from prior headaches (94.7%), tinnitus (83.7%), balance issues (62.7%), FND (90.0%), and trigeminal nerve dysfunction (79.2%), but not hearing loss (1.0%). Whereas GKRS provided effective tumor control independently of dose rate, GKRS patients exposed to lower dose rates experienced significantly better freedom from post-treatment hearing loss and FND (P = .044). CONCLUSION: Whereas GKRS provides excellent tumor control and effective symptomatic relief for vestibular schwannomas, dose-rate effects may impact post-treatment functional outcomes. Further research remains warranted.

Changes in lipid classes of Drosophila melanogaster in response to selection for three stress traits.

Laboratory selection on environmental stress traits is an evolutionary approach that is informative in the context of understanding stress adaptation. Here we characterize changes in a lipidome of Drosophila melanogaster in lines selected for increased heat (elevated heat knockdown refractoriness), cold (decreased time to recover from chill-coma) and desiccation survival. Selection for desiccation resistance resulted in changes in multiple lipid classes used to characterize a lipidome. This included a decrease in triacylglycerols (TAGs) which is relevant to interpretation of storage lipid levels in previous D. melanogaster desiccation survival selection experiments. Chill-coma recovery rate selection was expected to show extensive changes in lipid classes, but only phosphatidic acids exhibited significant change. Selection for increased heat knockdown resistance resulted in a substantial change in the abundance of a class of lipids (diacylglycerols) which could play a role in mediating the heat shock response or result in an increase in neutral lipid mobilization.

Fatty acid transport protein 2 reprograms neutrophils in cancer.

Polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) are pathologically activated neutrophils that are crucial for the regulation of immune responses in cancer. These cells contribute to the failure of cancer therapies and are associated with poor clinical outcomes. Despite recent advances in the understanding of PMN-MDSC biology, the mechanisms responsible for the pathological activation of neutrophils are not well defined, and this limits the selective targeting of these cells. Here we report that mouse and human PMN-MDSCs exclusively upregulate fatty acid transport protein 2 (FATP2). Overexpression of FATP2 in PMN-MDSCs was controlled by granulocyte-macrophage colony-stimulating factor, through the activation of the STAT5 transcription factor. Deletion of FATP2 abrogated the suppressive activity of PMN-MDSCs. The main mechanism of FATP2-mediated suppressive activity involved the uptake of arachidonic acid and the synthesis of prostaglandin E2. The selective pharmacological inhibition of FATP2 abrogated the activity of PMN-MDSCs and substantially delayed tumour progression. In combination with checkpoint inhibitors, FATP2 inhibition blocked tumour progression in mice. Thus, FATP2 mediates the acquisition of immunosuppressive activity by PMN-MDSCs and represents a target to inhibit the functions of PMN-MDSCs selectively and to improve the efficiency of cancer therapy.

Experimental studies on zeta potential of flocculants for harvesting of algae.

An experimental study was performed to evaluate the comparative efficiency of bio-flocculant (waste egg shell), laboratory available calcium carbonate (LACC) and alum (Al2 (SO4)3) for harvesting of unicellular microalga, Chlorella pyrenoidosa. The influence of pH on zeta potential (ζ) was also studied to explain the chemistry of flocculation process. The maximum harvesting efficiency (99%) was obtained with alum with deformities in algal cell surfaces. Waste egg-shell material is developed as a low-cost bio-flocculant for harvesting of Chlorella pyrenoidosa using 100 mg egg-shell bio-flocculant/L and 100 mg LACC/L, zeta potential analysis was completed to further understand the chemistry of harvesting efficiency over the different ranges of pH (2.0, 4.0, 6.0, 8.0, and 10.0). The optimized range for harvesting efficiency (HE) of pH is 4.0-8.0 for both flocculants. Maximal harvesting efficiency was achieved at pH 4.0 (99%) and pH 8.0 (95%) with bio-flocculant and LACC respectively. Hence, bio-flocculant based harvesting method is found as the best way to dewatering the algal biomass from aqueous medium with entire and intact algal cell surface with environment friendly and cost-effective approach.

An investigation of clinical treatment field delivery verification using cherenkov imaging: IMRT positioning shifts and field matching.

PURPOSE: Cherenkov light emission has been shown to correlate with ionizing radiation dose delivery in solid tissue. An important clinical application of Cherenkov light is the real-time verification of radiation treatment delivery in vivo. To test the feasibility of treatment field verification, Cherenkov light images were acquired concurrent with radiation beam delivery to standard and anthropomorphic phantoms. Specifically, we tested two clinical treatment scenarios: (a) Observation of field overlaps or gaps in matched 3D fields and (b) Patient positioning shifts during intensity modulated radiation therapy (IMRT) field delivery. Further development of this technique would allow real-time detection of treatment delivery errors on the order of millimeters so that patient safety and treatment quality can be improved. METHODS: Cherenkov light emission was captured using a PI-MAX4 intensified charge coupled device (ICCD) system (Princeton Instruments). All radiation delivery was performed using a Varian Trilogy linear accelerator (linac) operated at 6 MV or 18 MV for photon and 6 MeV or 16 MeV for electron studies. Field matching studies were conducted with photon and electron beams at gantry angles of 0°, 15°, and 45°. For each modality and gantry angle, a total of three data sets were acquired. Overlap and gap distances of 0, 2, 5, and 10 mm were tested and delivered to solid phantom material of 30 × 30 × 5 cm3 . Phantom materials used were white plastic water and brown solid water. Tests were additionally performed on an anthropomorphic phantom with an irregular surface. Positioning shift studies were performed using IMRT fields delivered to a thoracic anthropomorphic phantom. For thoracic phantom measurements, the camera was placed laterally to observe the entire right side of the phantom. Fields were delivered with known translational patient positioning shifts in four directions. Changes in the Cherenkov fluence were evaluated through the generation of difference maps from unshifted Cherenkov images. All images were evaluated using ImageJ, Python, and MATLAB software packages. RESULTS: For matched fields, Cherenkov images were able to quantitate matched field separations with discrepancies between 2 and 4 mm, depending on gantry angle and beam energy or modality. For all photon and electron beams delivered at a gantry angle of 0°, image analysis indicated average discrepancies of less than 2 mm for all field gaps and overlaps, with 83% of matched fields exhibiting discrepancies less than 1 mm. Beams delivered obliquely to the phantom surface exhibited average discrepancies as high as 4 mm for electron beams delivered at large oblique angles. Finally, for IMRT field delivery, vertical and lateral patient positioning shifts of 2 mm were detected in some cases, indicating the potential detectability threshold of using this technique alone. CONCLUSIONS: Our study indicates that Cherenkov imaging can be used to support and bolster current treatment delivery verification techniques, improving our ability to recognize and rectify millimeter-scale delivery and positioning errors.

Immune regulation and anti-cancer activity by lipid inflammatory mediators.

Rodent and clinical studies have documented that myeloid cell infiltration of tumors is associated with poor outcomes, neutrophilia and lymphocytopenia. This contrasts with increased lymphocyte infiltration of tumors, which is correlated with improved outcomes. Lifestyle parameters, such as obesity and diets with high levels of saturated fat and/or omega (ω)-6 polyunsaturated fatty acids (PUFAs), can influence these inflammatory parameters, including an increase in extramedullary myelopoiesis (EMM). While tumor secretion of growth factors (GFs) and chemokines regulate tumor-immune-cell crosstalk, lifestyle choices also contribute to inflammation, abnormal pathology and leukocyte infiltration of tumors. A relationship between obesity and high-fat diets (notably saturated fats in Western diets) and inflammation, tumor incidence, metastasis and poor outcomes is generally accepted. However, the mechanisms of dietary promotion of an inflammatory microenvironment and targeted drugs to inhibit the clinical sequelae are poorly understood. Thus, modifications of obesity and dietary fat may provide preventative or therapeutic approaches to control tumor-associated inflammation and disease progression. Currently, the majority of basic and clinical research does not differentiate between obesity and fatty acid consumption as mediators of inflammatory and neoplastic processes. In this review, we discuss the relationships between dietary PUFAs, inflammation and neoplasia and experimental strategies to improve our understanding of these relationships. We conclude that dietary composition, notably the ratio of ω-3 vs ω-6 PUFA regulates tumor growth and the frequency and sites of metastasis that together, impact overall survival (OS) in mice.

A novel model to correlate hydrogel spacer placement, perirectal space creation, and rectum dosimetry in prostate stereotactic body radiotherapy.

BACKGROUND: The SpaceOAR hydrogel is employed to limit rectal radiation dose during prostate radiotherapy. We identified a novel parameter - the product of angle θ and hydrogel volume - to quantify hydrogel placement. This parameter predicted rectum dosimetry and acute rectal toxicity in prostate cancer patients treated with stereotactic body radiotherapy to 36.25 Gy in 5 fractions. METHODS: Twenty men with low- and intermediate-risk prostate cancer underwent hydrogel placement from 2015 to 2017. Hydrogel symmetry was assessed on the CT simulation scan in 3 axial slices (midgland, 1 cm above midgland, 1 cm below midgland). Two novel parameters quantifying hydrogel placement - hydrogel volume and angle θ formed by the prostate, hydrogel, and rectum - were measured, and the normalized product of θ and hydrogel volume calculated. These were then correlated with perirectal distance, rectum maximum 1-3 cc point doses (rDmax 1-3 cc), and rectum volumes receiving 80-95% of the prescription dose (rV80-95%). Acute rectal toxicity was recorded per RTOG criteria. RESULTS: In 50% of patients, hydrogel placement was symmetric bilaterally to within 1 cm of midline in all three CT simulation scan axial slices. Lateral hydrogel asymmetry < 2 cm in any one axial slice did not affect rectum dosimetry, but absence of hydrogel in the inferior axial slice resulted in a mean increase of 171 cGy in the rDmax 1 cc (p < 0.005). The perirectal distance measured at prostate midgland, midline (mean 9.1 ± 4.3 mm) correlated strongly with rV95 (R2 0.6, p < 0.001). The mean hydrogel volume and θ were 10.3 ± 4.5 cc and 70 ± 49°, respectively. Perirectal distance, rV95 and rDmax 1 cc correlated with hydrogel angle θ (p < 0.01), and yet more strongly with the novel metric θ*hydrogel volume (p < 0.001). With a median follow up of 14 months, no rectal toxicity >grade 2 was observed. Low grade rectal toxicity was observed in a third of men and resolved within 1 month of SBRT. Men who had these symptoms had higher rDmax 1 cc and smaller θ*hydrogel volume measurements. CONCLUSIONS: Optimal hydrogel placement occurs at prostate midgland, midline. The novel parameter θ*hydrogel volume describes a large proportion of rectum dosimetric benefit derived from hydrogel placement, and can be used to assess the learning curve phenomenon for hydrogel placement.

Long-chain omega-3 polyunsaturated fatty acids decrease mammary tumor growth, multiorgan metastasis and enhance survival.

Epidemiological studies show a reduced risk of breast cancer (BC) in women consuming high levels of long-chain (LC) omega-3 (ω-3) fatty acids (FAs) compared with women who consumed low levels. However, the regulatory and mechanistic roles of dietary ω-6 and LC-ω-3 FAs on tumor progression, metastasis and survival are poorly understood. Female BALB/c mice (10-week old) were pair-fed with a diet containing ω-3 or an isocaloric, isolipidic ω-6 diet for 16 weeks prior to the orthotopic implantation of 4T1 mammary tumor cells. Major outcomes studied included: mammary tumor growth, survival analysis, and metastases analyses in multiple organs including pulmonary, hepatic, bone, cardiac, renal, ovarian, and contralateral MG (CMG). The dietary regulation of the tumor microenvironment was evaluated in mice autopsied on day-35 post tumor injection. In mice fed the ω-3 containing diet, there was a significant delay in tumor initiation and prolonged survival relative to the ω-6 diet-fed group. The tumor size on day 35 post tumor injection in the ω-3 group was 50% smaller and the frequencies of pulmonary and bone metastases were significantly lower relative to the ω-6 group. Similarly, the incidence/frequencies and/or size of cardiac, renal, ovarian metastases were significantly lower in mice fed the ω-3 diet. The analyses of the tumor microenvironment showed that tumors in the ω-3 group had significantly lower numbers of proliferating tumor cells (Ki67+)/high power field (HPF), and higher numbers of apoptotic tumor cells (TUNEL+)/HPF, lower neo-vascularization (CD31+ vessels/HPF), infiltration by neutrophil elastase+ cells, and macrophages (F4/80+) relative to the tumors from the ω-6 group. Further, in tumors from the ω-3 diet-fed mice, T-cell infiltration was 102% higher resulting in a neutrophil to T-lymphocyte ratio (NLR) that was 76% lower (p < 0.05). Direct correlations were observed between NLR with tumor size and T-cell infiltration with the number of apoptotic tumor cells. qRT-PCR analysis revealed that tumor IL10 mRNA levels were significantly higher (six-fold) in the tumors from mice fed the ω-3 diet and inversely correlated with the tumor size. Our data suggest that dietary LC-ω-3FAs modulates the mammary tumor microenvironment slowing tumor growth, and reducing metastases to both common and less preferential organs resulting in prolonged survival. The surrogate analyses undertaken support a mechanism of action by dietary LC-ω-3FAs that includes, but is not limited to decreased infiltration by myeloid cells (neutrophils and macrophages), an increase in CD3+ lymphocyte infiltration and IL10 associated anti-inflammatory activity.

Induction of oil accumulation by heat stress is metabolically distinct from N stress in the green microalgae Coccomyxa subellipsoidea C169.

Algae are often promoted as feedstock organisms to produce a sustainable petroleum fossil fuel alternative. However, to induce lipid accumulation most often requires a severe stress that is difficult to induce in large batch cultures. The objective of this study is to analyze and mathematically model heat stress on growth, chlorophyll content, triacylglyceride, and starch synthesis in algae. We initially screened 30 algal species for the most pronounced induction of lipid droplets from heat stress using confocal microscopy and mass spectroscopy techniques. One species, Coccomyxa subellipsoidea C169, was selected and subjected to further biochemical analyses using a jacketed bioreactor amended with 1% CO2 at 25°C, 30°C, 32°C, 33°C, 34°C, 35°C, and 36°C. Lipid and starch accumulation was less extreme than N stress. Growth was reduced above 25°C, but heat stress induced lipid droplet synthesis was negatively correlated with growth only past a demonstrated threshold temperature above 32°C. The optimal temperature for lipid accumulation was 35°C, which led to 6% of dry weight triglyceride content and a 72% reduction from optimal growth after 5 days. Fatty acid influx rates into triglycerides and 15N labeling of amino acids and proteins indicate that heat stress is mechanistically distinct from N stress. Thus, this study lends support to a novel hypothesis that lipid droplet triglycerides result from a redistribution of carbon flux as fatty acids to neutral storage lipids over membrane or other lipids.