Diagnostic Value of a Combined Serum α-Hydroxybutyrate Dehydrogenase, Carcinoembryonic Antigen and Glycoantigen 125 Test for Early-Stage Breast Cancer.

Objective: To investigate the diagnostic value of the combined detection of α-hydroxybutyrate dehydrogenase (α-HBDH), carcinoembryonic antigen (CEA) and cancer antigen 125 (CA125) in early-stage breast cancer (ESBC). Methods: This was a retrospective analysis of 169 patients with ESBC, 138 patients with benign breast disease (BBD) and 200 normal healthy controls (NHCs). The levels of serum α-HBDH, CEA and CA125 in the two groups were detected. The receiver operating characteristic (ROC) curve and area under the curve (AUC) were used to analyse the diagnostic value of the above indicators alone and in combination for ESBC. Results: The levels of α-HBDH, CEA and CA125 in the ESBC group were significantly higher than those in the BBD and NHC groups ([118.18 ± 11.19 vs 91.24 ± 9.17 vs 89.38 ± 9.01, F = 6.189, p = 0.004], [2.39 ± 1.12 vs 1.48 ± 0.76 vs 1.58 ± 0.58, F = 5.362, p = 0.017] and [14.44 ± 6.78 vs 11.19 ± 3.17 vs 7.18 ± 4.71, F = 8.912, p = 0.001], respectively). In the ESBC group, the positive rate of combined detection was higher than that of single detection (96.12% vs 72.64% vs 53.67% vs 42.41%, X2 = 27.174, p < 0.05). ROC curve analysis showed that serum α-HBDH, CEA, CA125 alone and combined detection in the diagnosis of ESBC. The sensitivity was 48.1%, 63.6%, 44.2% and 54.5%, the specificity was 75.4%, 75.4%, 86.0% and 91.2% and the AUC was 0.654, 0.715, 0.636 and 0.772, respectively. The diagnostic value of combined detection was the highest. Conclusion: The levels of serum α-HBDH, CEA and CA125 in ESBC are high, and the combined detection of the three has a high diagnostic value for ESBC.

The Comprehensive Steroidome in Complete TSPO/PBR Knockout Mice under Basal Conditions.

The 18 kDa translocator protein (TSPO/PBR) is a multifunctional evolutionary highly conserved outer mitochondrial membrane protein. Decades of research has reported an obligatory role of TSPO/PBR in both mitochondrial cholesterol transport and, thus, steroid production. However, the strict dependency of steroidogenesis on TSPO/PBR has remained controversial. The aim of this study was to provide insight into the steroid profile in complete C57BL/6-Tspotm1GuWu(GuwiyangWurra)-knockout male mice (TSPO-KO) under basal conditions. The steroidome in the brain, adrenal glands, testes and plasma was measured by gas chromatography coupled to tandem mass spectrometry (GC-MS/MS). We found that steroids present in wild-type (WT) mice were also detected in TSPO-KO mice, including pregnenolone (PREG), progestogens, mineralo-glucocorticosteroids and androgens. The concentrations of PREG and most metabolites were similar between genotypes, except a significant decrease in the levels of the 5α-reduced metabolites of progesterone (PROG) in adrenal glands and plasma and of the 5α-reduced metabolites of corticosterone (B) in plasma in TSPO-KO compared to WT animals, suggesting other regulatory functions for the TSPO/PBR. The expression levels of the voltage-dependent anion-selective channel (VDAC-1), CYP11A1 and 5α-reductase were not significantly different between both groups. Thus, the complete deletion of the tspo gene in male mice does not impair de novo steroidogenesis in vivo.

The cancer-testis antigen a-kinase anchor protein 3 facilitates breast cancer progression via activation of the PTEN/PI3K/AKT/mTOR signaling.

The cancer-testis antigen A-kinase anchor protein 3 (AKAP3) has been shown to have a strong association with breast cancer (BC). However, its role in BC progression received scant attention. We aimed to explore the prognostic implication of aberrant AKAP3 expression for a better knowledge of BC progression and improved treatment. AKAP3 expression was quantitated using tissue microarrays and immunohistochemistry (IHC). Cell viability, invasion, migration, apoptosis, and expressions of PTEN/PI3K/AKT/mTOR signaling components were assessed in AKAP3-overexpressed or si-AKAP3-transfected BC cells. Finally, elevated AKAP3 expression was observed in BC versus paracancerous tissues. BC patients with high AKAP3 expression showed a worse prognosis than low expression patients (P < 0.0001). AKAP3 overexpressions fueled cell growth, proliferation, migration, and invasion in HCC1937 and MDA-MB-468 BC cell lines, alongside increased expressions of PI3K/AKT/mTOR signaling components and PTEN suppression. These effects were pronouncedly reversed, together with elevated apoptosis, in cells transfected with si-AKAP3. Therefore, AKAP3 is upregulated in BC and promotes BC cell growth, invasion, and migration via PTEN/PI3K/AKT/mTOR signaling activation. It may serve as a prognosis indicator for BC survival.

Long-term diazepam treatment enhances microglial spine engulfment and impairs cognitive performance via the mitochondrial 18 kDa translocator protein (TSPO).

Benzodiazepines are widely administered drugs to treat anxiety and insomnia. In addition to tolerance development and abuse liability, their chronic use may cause cognitive impairment and increase the risk for dementia. However, the mechanism by which benzodiazepines might contribute to persistent cognitive decline remains unknown. Here we report that diazepam, a widely prescribed benzodiazepine, impairs the structural plasticity of dendritic spines, causing cognitive impairment in mice. Diazepam induces these deficits via the mitochondrial 18 kDa translocator protein (TSPO), rather than classical γ-aminobutyric acid type A receptors, which alters microglial morphology, and phagocytosis of synaptic material. Collectively, our findings demonstrate a mechanism by which TSPO ligands alter synaptic plasticity and, as a consequence, cause cognitive impairment.

Microgravity × Radiation: A Space Mechanobiology Approach Toward Cardiovascular Function and Disease.

In recent years, there has been an increasing interest in space exploration, supported by the accelerated technological advancements in the field. This has led to a new potential environment that humans could be exposed to in the very near future, and therefore an increasing request to evaluate the impact this may have on our body, including health risks associated with this endeavor. A critical component in regulating the human pathophysiology is represented by the cardiovascular system, which may be heavily affected in these extreme environments of microgravity and radiation. This mini review aims to identify the impact of microgravity and radiation on the cardiovascular system. Being able to understand the effect that comes with deep space explorations, including that of microgravity and space radiation, may also allow us to get a deeper understanding of the heart and ultimately our own basic physiological processes. This information may unlock new factors to consider with space exploration whilst simultaneously increasing our knowledge of the cardiovascular system and potentially associated diseases.

Mitochondrial Translocator Protein (TSPO) Expression in the Brain After Whole Body Gamma Irradiation.

The brain's early response to low dose ionizing radiation, as may be encountered during diagnostic procedures and space exploration, is not yet fully characterized. In the brain parenchyma, the mitochondrial translocator protein (TSPO) is constitutively expressed at low levels by endothelial cells, and can therefore be used to assess the integrity of the brain's vasculature. At the same time, the inducible expression of TSPO in activated microglia, the brain's intrinsic immune cells, is a regularly observed early indicator of subtle or incipient brain pathology. Here, we explored the use of TSPO as a biomarker of brain tissue injury following whole body irradiation. Post-radiation responses were measured in C57BL/6 wild type (Tspo +/+) and TSPO knockout (Tspo -/-) mice 48 h after single whole body gamma irradiations with low doses 0, 0.01, and 0.1 Gy and a high dose of 2 Gy. Additionally, post-radiation responses of primary microglial cell cultures were measured at 1, 4, 24, and 48 h at an irradiation dose range of 0 Gy-2 Gy. TSPO mRNA and protein expression in the brain showed a decreased trend after 0.01 Gy relative to sham-irradiated controls, but remained unchanged after higher doses. Immunohistochemistry confirmed subtle decreases in TSPO expression after 0.01 Gy in vascular endothelial cells of the hippocampal region and in ependymal cells, with no detectable changes following higher doses. Cytokine concentrations in plasma after whole body irradiation showed differential changes in IL-6 and IL-10 with some variations between Tspo-/- and Tspo +/+ animals. The in vitro measurements of TSPO in primary microglial cell cultures showed a significant reduction 1 h after low dose irradiation (0.01 Gy). In summary, acute low and high doses of gamma irradiation up to 2 Gy reduced TSPO expression in the brain's vascular compartment without de novo induction of TSPO expression in parenchymal microglia, while TSPO expression in directly irradiated, isolated, and thus highly activated microglia, too, was reduced after low dose irradiation. The potential link between TSPO, its role in mitochondrial energy metabolism and the selective radiation sensitivity, notably of cells with constitutive TSPO expression such as vascular endothelial cells, merits further exploration.

Control of Neuroinflammation through Radiation-Induced Microglial Changes.

Microglia, the innate immune cells of the central nervous system, play a pivotal role in the modulation of neuroinflammation. Neuroinflammation has been implicated in many diseases of the CNS, including Alzheimer's disease and Parkinson's disease. It is well documented that microglial activation, initiated by a variety of stressors, can trigger a potentially destructive neuroinflammatory response via the release of pro-inflammatory molecules, and reactive oxygen and nitrogen species. However, the potential anti-inflammatory and neuroprotective effects that microglia are also thought to exhibit have been under-investigated. The application of ionising radiation at different doses and dose schedules may reveal novel methods for the control of microglial response to stressors, potentially highlighting avenues for treatment of neuroinflammation associated CNS disorders, such as Alzheimer's disease and Parkinson's disease. There remains a need to characterise the response of microglia to radiation, particularly low dose ionising radiation.

Steric Hindrance- and Work Function-Promoted High Performance for Electrochemical CO Methanation on Antisite Defects of MoS2 and WS2.

CO methanation from electrochemical CO reduction reaction (CORR) is significant for sustainable environment and energy, but electrocatalysts with excellent selectivity and activity are still lacking. Selectivity is sensitive to the structure of active sites, and activity can be tailored by work function. Moreover, intrinsic active sites usually possess relatively high concentration compared to artificial ones. Here, antisite defects MoS2 and WS2 , intrinsic atomic defects of MoS2 and WS2 with a transition metal atom substituting a S2 column, were investigated for CORR by density functional theory calculations. The steric hindrance from the special bowl structure of MoS2 and WS2 ensured good selectivity towards CO methanation. Coordination environment variation of the active sites, the under-coordinated Mo or W atoms, effectively lowered the work function, making MoS2 and WS2 highly active for CO methanation with the required potential of -0.47 and -0.49 V vs. reversible hydrogen electrode, respectively. Moreover, high concentration of active sites and minimal structural deformation during the catalytic process of MoS2 and WS2 enhanced their attraction for future commercial application.

Effects of dietary Enteromorpha powder on reproduction-related hormones and genes during the late laying period of Zi geese.

OBJECTIVE: The aim of this study was to investigate the effects of Enteromorpha powder supplementation on reproduction-related hormones and genes in the late laying period of Zi geese. METHODS: A total of 312 (1-year-old) Zi geese with similar laying rate were randomly divided into 2 groups with 6 replicates each, each with 21 female geese and 5 male geese. The control group was fed with a basal diet and the test group was fed with a diet containing 3% Enteromorpha powder. The trial period lasted for 7 weeks. RESULTS: Our results showed that the laying rate was improved in the test group at each week of trial (p<0.01), and the levels of estradiol in serum and prolactin in ovary were increased compared with the control group (p<0.05). CONCLUSION: Based on above results, Enteromorpha powder supplementation at 3% could promote reproductive performance during the late laying period of Zi geese.

Author Correction: Selective, high-contrast detection of syngeneic glioblastoma in vivo.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Selective, high-contrast detection of syngeneic glioblastoma in vivo.

Glioblastoma is a highly malignant, largely therapy-resistant brain tumour. Deep infiltration of brain tissue by neoplastic cells represents the key problem of diffuse glioma. Much current research focuses on the molecular makeup of the visible tumour mass rather than the cellular interactions in the surrounding brain tissue infiltrated by the invasive glioma cells that cause the tumour's ultimately lethal outcome. Diagnostic neuroimaging that enables the direct in vivo observation of the tumour infiltration zone and the local host tissue responses at a preclinical stage are important for the development of more effective glioma treatments. Here, we report an animal model that allows high-contrast imaging of wild-type glioma cells by positron emission tomography (PET) using [18 F]PBR111, a selective radioligand for the mitochondrial 18 kDa Translocator Protein (TSPO), in the Tspo-/- mouse strain (C57BL/6-Tspotm1GuMu(GuwiyangWurra)). The high selectivity of [18 F]PBR111 for the TSPO combined with the exclusive expression of TSPO in glioma cells infiltrating into null-background host tissue free of any TSPO expression, makes it possible, for the first time, to unequivocally and with uniquely high biological contrast identify peri-tumoral glioma cell invasion at preclinical stages in vivo. Comparison of the in vivo imaging signal from wild-type glioma cells in a null background with the signal in a wild-type host tissue, where the tumour induces the expected TSPO expression in the host's glial cells, illustrates the substantial extent of the peritumoral host response to the growing tumour. The syngeneic tumour (TSPO+/+) in null background (TSPO-/-) model is thus well suited to study the interaction of the tumour front with the peri-tumoral tissue, and the experimental evaluation of new therapeutic approaches targeting the invasive behaviour of glioblastoma.

The Translocator Protein (TSPO) in Mitochondrial Bioenergetics and Immune Processes.

The translocator protein (TSPO) is an outer mitochondrial membrane protein that is widely used as a biomarker of neuroinflammation, being markedly upregulated in activated microglia in a range of brain pathologies. Despite its extensive use as a target in molecular imaging studies, the exact cellular functions of this protein remain in question. The long-held view that TSPO plays a fundamental role in the translocation of cholesterol through the mitochondrial membranes, and thus, steroidogenesis, has been disputed by several groups with the advent of TSPO knockout mouse models. Instead, much evidence is emerging that TSPO plays a fundamental role in cellular bioenergetics and associated mitochondrial functions, also part of a greater role in the innate immune processes of microglia. In this review, we examine the more direct experimental literature surrounding the immunomodulatory effects of TSPO. We also review studies which highlight a more central role for TSPO in mitochondrial processes, from energy metabolism, to the propagation of inflammatory responses through reactive oxygen species (ROS) modulation. In this way, we highlight a paradigm shift in approaches to TSPO functioning.

C-type lectin-like receptor 2 and zonulin are associated with mild cognitive impairment and Alzheimer's disease.

OBJECTIVE: Increased permeability and changes in gut microbiota contributed to the pathogenesis of Alzheimer's disease (AD). Zonulin is a key modulator that regulates intestinal barrier function. Peripheral platelet alterations have been involved in AD pathology. C-type lectin-like receptor 2 (CLEC-2) is a receptor on the platelet surface for activation. The purpose of this study was to determine zonulin and CLEC-2 levels in mild cognitive impairment (MCI) and AD, and investigate the relationship between zonulin and CLEC-2. METHODS: In this study, CLEC-2 and zonulin levels were measured using ELISA assay in 110 AD patients, 110 MCI patients, and 110 non-demented control subjects. RESULTS: Increased CLEC-2 and zonulin levels were observed in MCI and AD patients. Furthermore, AD patients had higher CLEC-2 and zonulin levels compared with MCI patients. In addition, CLEC-2 levels were positively correlated with zonulin levels, after adjusting confounding factors (r = .592, P < .001). Multivariate analysis revealed that increased CLEC-2 and zonulin levels were significantly associated with reduced Mini-Mental State Examination (MMSE) score. CONCLUSIONS: C-type lectin-like receptor 2 is correlated with zonulin after adjusting confounding covariates. Moreover, increased CLEC-2 and zonulin are the significant factors for reduced MMSE score in MCI and AD. Further studies are needed.

Adaptive in vivo device for theranostics of inflammation: Real-time monitoring of interferon-γ and aspirin.

Cytokines mediate and control immune and inflammatory responses. Complex interactions exist among cytokines, inflammation, and the innate and adaptive immune responses in maintaining homeostasis, health, and well-being. On-demand, local delivery of anti-inflammatory drugs to target tissues provides an approach for more effective drug dosing while reducing the adverse effects of systemic drug delivery. This work demonstrates a proof-of-concept theranostic approach for inflammation based on analyte-kissing induced signaling, whereby a drug (in this report, aspirin) can be released upon the detection of a target level of a proinflammatory cytokine (i.e., interferon-γ (IFN-γ)) in real time. The structure-switching aptamer-based biosensor described here is capable of quantitatively and dynamically detecting IFN-γ both in vitro and in vivo with a sensitivity of 10 pg mL-1. Moreover, the released aspirin triggered by the immunoregulatory cytokine IFN-γ is able to inhibit inflammation in a rat model, and the release of aspirin can be quantitatively controlled. The data reported here provide a new and promising strategy for the in vivo detection of proinflammatory cytokines and the subsequent therapeutic delivery of anti-inflammatory molecules. This universal theranostic platform is expected to have great potential for patient-specific personalized medicine. STATEMENT OF SIGNIFICANCE: We developed an adaptive in vivo sensing device whereby a drug, aspirin, can be released upon the detection of a proinflammatory cytokine, interferon-γ (IFN-γ), in real time with a sensitivity of 10 pg mL-1. Moreover, the aspirin triggered by IFN-γ depressed inflammation in the rat model and was delivered indirectly through blood and cerebrospinal fluid or directly to the inflammation tissue or organ without adverse gastrointestinal effects observed in the liver and kidney. We envision that, for the first time, patients with chronic inflammatory disease can receive the right intervention and treatment at the right time. Additionally, this technology may empower patients to monitor their personalized health and disease management program, allowing real-time diagnostics, disease monitoring, and precise and effective treatments.

Microfluidic Actuation via 3D-Printed Molds toward Multiplex Biosensing of Cell Apoptosis.

Multiplexed analysis of biochemical analytes such as proteins, enzymes, and immune products using a microfluidic device has the potential to cut assay time, reduce sample volume, realize high-throughput, and decrease experimental error without compromising sensitivity. Despite these huge benefits, the need for expensive specialized equipment and the complex photolithography fabrication process for the multiplexed devices have, to date, prevented widespread adoption of microfluidic systems. Here, we present a simple method to fabricate a new microfluidic-based multiplexed biosensing device by taking advantage of 3D-printing. The device is an integration of normally closed (NC) microfluidic valving units which offer superior operational flexibility by using PDMS membrane (E ∼ 1-2 MPa) and require minimized energy input (1-5 kPa). To systematically engineer the device, we first report on the geometrical and operational analysis of a single 3D-printed valving unit. Based on the characterization, we introduce a full prototype multiplexed chip comprising several microfluidic valves. The prototype offers-for the first time in a 3D-printed microfluidic device-the capability of on-demand performce of both a sequential and a parallel biochemical assay. As a proof of concept, our device has been used to simultaneously measure the apoptotic activity of 5 different members of the caspase protease enzyme family. In summary, the 3D-printed valving system showcased in this study overcomes traditional bottlenecks of microfabrication, enabling a new class of sophisticated liquid manipulation required in performing multiplexed sensing for biochemical assays.

Turn-On Fluorescence Aptasensor on Magnetic Nanobeads for Aflatoxin M1 Detection Based on an Exonuclease III-Assisted Signal Amplification Strategy.

In order to satisfy the need for sensitive detection of Aflatoxin M1 (AFM1), we constructed a simple and signal-on fluorescence aptasensor based on an autocatalytic Exonuclease III (Exo III)-assisted signal amplification strategy. In this sensor, the DNA hybridization on magnetic nanobeads could be triggered by the target AFM1, resulting in the release of a single-stranded DNA to induce an Exo III-assisted signal amplification, in which numerous G-quadruplex structures would be produced and then associated with the fluorescent dye to generate significantly amplified fluorescence signals resulting in the increased sensitivity. Under the optimized conditions, this aptasensor was able to detect AFM1 with a practical detection limit of 9.73 ng kg-1 in milk samples. Furthermore, the prepared sensor was successfully used for detection of AFM1 in the commercially available milk samples with the recovery percentages ranging from 80.13% to 108.67%. Also, the sensor performance was evaluated by the commercial immunoassay kit with satisfactory results.

DNA physical properties outperform sequence compositional information in classifying nucleosome-enriched and -depleted regions.

The nucleosome is the fundamental structural unit of eukaryotic chromatin and plays an essential role in the epigenetic regulation of cellular processes, such as DNA replication, recombination, and transcription. Hence, it is important to identify nucleosome positions in the genome. Our previous model based on DNA deformation energy, in which a set of DNA physical descriptors was used, performed well in predicting nucleosome dyad positions and occupancy. In this study, we established a machine-learning model for predicting nucleosome occupancy in order to further verify the physical descriptors. Results showed that (1) our model outperformed several other sequence compositional information-based models, indicating a stronger dependence of nucleosome positioning on DNA physical properties; (2) nucleosome-enriched and -depleted regions have distinct features in terms of DNA physical descriptors like sequence-dependent flexibility and equilibrium structure parameters; (3) gene transcription start sites and termination sites can be well characterized with the distribution patterns of the physical descriptors, indicating the regulatory role of DNA physical properties in gene transcription. In addition, we developed a web server for the model, which is freely accessible at http://lin-group.cn/server/iNuc-force/.

In vivo [64Cu]CuCl2 PET imaging reveals activity of Dextran-Catechin on tumor copper homeostasis.

Given the strong clinical evidence that copper levels are significantly elevated in a wide spectrum of tumors, copper homeostasis is considered as an emerging target for anticancer drug design. Monitoring copper levels in vivo is therefore of paramount importance when assessing the efficacy of copper-targeting drugs. Herein, we investigated the activity of the copper-targeting compound Dextran-Catechin by developing a [64Cu]CuCl2 PET imaging protocol to monitor its effect on copper homeostasis in tumors. Methods: Protein expression of copper transporter 1 (CTR1) in tissue microarrays representing 90 neuroblastoma patient tumors was assessed by immunohistochemistry. Western blotting analysis was used to study the effect of Dextran-Catechin on the expression of CTR1 in neuroblastoma cell lines and in tumors. A preclinical human neuroblastoma xenograft model was used to study anticancer activity of Dextran-Catechin in vivo and its effect on tumor copper homeostasis. PET imaging with [64Cu]CuCl2 was performed in such preclinical neuroblastoma model to monitor alteration of copper levels in tumors during treatment. Results: CTR1 protein was found to be highly expressed in patient neuroblastoma tumors by immunohistochemistry. Treatment of neuroblastoma cell lines with Dextran-Catechin resulted in decreased levels of glutathione and in downregulation of CTR1 expression, which caused a significant decrease of intracellular copper. No changes in CTR1 expression was observed in normal human astrocytes after Dextran-Catechin treatment. In vivo studies and PET imaging analysis using the neuroblastoma preclinical model revealed elevated [64Cu]CuCl2 retention in the tumor mass. Following treatment with Dextran-Catechin, there was a significant reduction in radioactive uptake, as well as reduced tumor growth. Ex vivo analysis of tumors collected from Dextran-Catechin treated mice confirmed the reduced levels of CTR1. Interestingly, copper levels in blood were not affected by treatment, demonstrating potential tumor specificity of Dextran-Catechin activity. Conclusion: Dextran-Catechin mediates its activity by lowering CTR1 and intracellular copper levels in tumors. This finding further reveals a potential therapeutic strategy for targeting copper-dependent cancers and presents a novel PET imaging method to assess patient response to copper-targeting anticancer treatments.

Cellular Sources and Regional Variations in the Expression of the Neuroinflammatory Marker Translocator Protein (TSPO) in the Normal Brain.

The inducible expression of the mitochondrial translocator protein 18 kDa (TSPO) by activated microglia is a prominent, regular feature of acute and chronic-progressive brain pathology. This expression is also the rationale for the continual development of new TSPO binding molecules for the diagnosis of "neuroinflammation" by molecular imaging. However, there is in the normal brain an ill-defined, low-level constitutive expression of TSPO. Taking advantage of healthy TSPO knockout mouse brain tissue to validate TSPO antibody specificity, this study uses immunohistochemistry to determine the regional distribution and cellular sources of TSPO in the normal mouse brain. Fluorescence microscopy revealed punctate TSPO immunostaining in vascular endothelial cells throughout the brain. In the olfactory nerve layers and glomeruli of the olfactory bulb, choroid plexus and ependymal layers, we confirm constitutive TSPO expression levels similar to peripheral organs, while some low TSPO expression is present in regions of known neurogenesis, as well as cerebellar Purkinje cells. The distributed-sparse expression of TSPO in endothelial mitochondria throughout the normal brain can be expected to give rise to a low baseline signal in TSPO molecular imaging studies. Finally, our study emphasises the need for valid and methodologically robust verification of the selectivity of TSPO ligands through the use of TSPO knockout tissues.

BAMLET kills chemotherapy-resistant mesothelioma cells, holding oleic acid in an activated cytotoxic state.

Malignant pleural mesothelioma is an aggressive cancer with poor prognosis. Here we have investigated in vitro efficacy of BAMLET and BLAGLET complexes (anti-cancer complexes consisting of oleic acid and bovine α-lactalbumin or ß-lactoglobulin respectively) in killing mesothelioma cells, determined BAMLET and BLAGLET structures, and investigated possible biological mechanisms. We performed cell viability assays on 16 mesothelioma cell lines. BAMLET and BLAGLET having increasing oleic acid content inhibited human and rat mesothelioma cell line proliferation at decreasing doses. Most of the non-cancer primary human fibroblasts were more resistant to BAMLET than were human mesothelioma cells. BAMLET showed similar cytotoxicity to cisplatin-resistant, pemetrexed-resistant, vinorelbine-resistant, and parental rat mesothelioma cells, indicating the BAMLET anti-cancer mechanism may be different to drugs currently used to treat mesothelioma. Cisplatin, pemetrexed, gemcitabine, vinorelbine, and BAMLET, did not demonstrate a therapeutic window for mesothelioma compared with immortalised non-cancer mesothelial cells. We demonstrated by quantitative PCR that ATP synthase is downregulated in mesothelioma cells in response to regular dosing with BAMLET. We sought structural insight for BAMLET and BLAGLET activity by performing small angle X-ray scattering, circular dichroism, and scanning electron microscopy. Our results indicate the structural mechanism by which BAMLET and BLAGLET achieve increased cytotoxicity by holding increasing amounts of oleic acid in an active cytotoxic state encapsulated in increasingly unfolded protein. Our structural studies revealed similarity in the molecular structure of the protein components of these two complexes and in their encapsulation of the fatty acid, and differences in the microscopic structure and structural stability. BAMLET forms rounded aggregates and BLAGLET forms long fibre-like aggregates whose aggregation is more stable than that of BAMLET due to intermolecular disulphide bonds. The results reported here indicate that BAMLET and BLAGLET may be effective second-line treatment options for mesothelioma.