Irreversible electroporation augments ß-glucan induced trained innate immunity for the treatment of pancreatic ductal adenocarcinoma.

BACKGROUND: Pancreatic cancer (PC) is a challenging diagnosis that is yet to benefit from the advancements in immuno-oncologic treatments. Irreversible electroporation (IRE), a non-thermal method of tumor ablation, is used in treatment of select patients with locally-advanced unresectable PC and has potentiated the effect of certain immunotherapies. Yeast-derived particulate ß-glucan induces trained innate immunity and successfully reduces murine PC tumor burden. This study tests the hypothesis that IRE may augment ß-glucan induced trained immunity in the treatment of PC. METHODS: ß-Glucan-trained pancreatic myeloid cells were evaluated ex vivo for trained responses and antitumor function after exposure to ablated and unablated tumor-conditioned media. ß-Glucan and IRE combination therapy was tested in an orthotopic murine PC model in wild-type and Rag-/- mice. Tumor immune phenotypes were assessed by flow cytometry. Effect of oral ß-glucan in the murine pancreas was evaluated and used in combination with IRE to treat PC. The peripheral blood of patients with PC taking oral ß-glucan after IRE was evaluated by mass cytometry. RESULTS: IRE-ablated tumor cells elicited a potent trained response ex vivo and augmented antitumor functionality. In vivo, ß-glucan in combination with IRE reduced local and distant tumor burden prolonging survival in a murine orthotopic PC model. This combination augmented immune cell infiltration to the PC tumor microenvironment and potentiated the trained response from tumor-infiltrating myeloid cells. The antitumor effect of this dual therapy occurred independent of the adaptive immune response. Further, orally administered ß-glucan was identified as an alternative route to induce trained immunity in the murine pancreas and prolonged PC survival in combination with IRE. ß-Glucan in vitro treatment also induced trained immunity in peripheral blood monocytes obtained from patients with treatment-naïve PC. Finally, orally administered ß-glucan was found to significantly alter the innate cell landscape within the peripheral blood of five patients with stage III locally-advanced PC who had undergone IRE. CONCLUSIONS: These data highlight a relevant and novel application of trained immunity within the setting of surgical ablation that may stand to benefit patients with PC.

Biomechanical Evaluation of a Mandibular Spanning Plate Technique Compared to Standard Plating Techniques to Treat Mandibular Symphyseal Fractures.

Purpose. The purpose of this study is to compare the biomechanical behavior of the spanning reconstruction plate compared to standard plating techniques for mandibular symphyseal fractures. Materials and Methods. Twenty-five human mandible replicas were used. Five unaltered synthetic mandibles were used as controls. Four experimental groups of different reconstruction techniques with five in each group were tested. Each synthetic mandible was subjected to a splaying force applied to the mandibular angle by a mechanical testing unit until the construct failed. Peak load and stiffness were recorded. The peak load and stiffness were analyzed using ANOVA and the Tukey test at a confidence level of 95% (P < 0.05). Results. The two parallel plates' group showed statistically significant lower values for peak load and stiffness compared to all other groups. No statistically significant difference was found for peak load and stiffness between the control (C) group, lag screw (LS) group, and the spanning plate (SP1) group. Conclusions. The spanning reconstruction plate technique for fixation of mandibular symphyseal fractures showed similar mechanical behavior to the lag screw technique when subjected to splaying forces between the mandibular gonial angles and may be considered as an alternative technique when increased reconstructive strength is needed.

Human MCS5A1 candidate breast cancer susceptibility gene FBXO10 is induced by cellular stress and correlated with lens epithelium-derived growth factor (LEDGF).

Genetic variation and candidate genes associated with breast cancer susceptibility have been identified. Identifying molecular interactions between associated genetic variation and cellular proteins may help to better understand environmental risk. Human MCS5A1 breast cancer susceptibility associated SNP rs7042509 is located in F-box protein 10 (FBXO10). An orthologous Rattus norvegicus DNA-sequence that contains SNV ss262858675 is located in rat Mcs5a1, which is part of a mammary carcinoma susceptibility locus controlling tumor development in a non-mammary cell-autonomous manner via an immune cell-mediated mechanism. Higher Fbxo10 expression in T cells is associated with Mcs5a increased susceptibility alleles. A common DNA-protein complex bound human and rat sequences containing MCS5A1/Mcs5a1 rs7042509/ss262858675 in electrophoretic mobility shift assays (EMSAs). Lens epithelium-derived growth factor (LEDGF), a stress-response protein, was identified as a candidate to bind both human and rat sequences using DNA-pulldown and mass spectrometry. LEDGF binding was confirmed by LEDGF-antibody EMSA and chromatin immunoprecipitation (ChIP). Ectopic expression of LEDGF/p75 increased luciferase activities of co-transfected reporters containing both human and rat orthologs. Over-expressed LEDGF/p75 increased endogenous FBXO10 mRNA levels in Jurkat cells, a human T-cell line, implying LEDGF may be involved in increasing FBXO10 transcript levels. Oxidative and thermal stress of Jurkat cells increased FBXO10 and LEDGF expression, further supporting a hypothesis that LEDGF binds to a regulatory region of FBXO10 and increases expression during conditions favoring carcinogenesis. We conclude that FBXO10, a candidate breast cancer susceptibility associated gene, is induced by cellular stress and LEDGF may play a role in expression of this gene.

Preparation and characterization of novel elastin-like polypeptide-collagen composites.

Collagen-based biomaterials suffer from poor mechanical properties and rapid degradation. Elastin-like polypeptides (ELPs) possess good biocompatibility and have unique solution properties that allow them to coacervate above a critical temperature. The objective of this research was to prepare a series of freeze dried ELP-collagen composite scaffolds as a proof of concept. Combination of ELP and collagen has the potential to produce composite structures with varying strengths. Four different composite structures were prepared by varying the ratio of ELP to collagen. Increased ELP content in the scaffolds appears to have reduced the residual water content based on Fourier transformed infrared spectroscopy and differential scanning calorimetry. Scanning electron microscopy images of ELP-collagen composites showed a three-dimensional, open porous structure with the formation of characteristic aggregates of ELP. The mechanical testing experiments showed that the elastic modulus, tensile strength, and toughness of ELP-collagen scaffolds were significantly greater than neat collagen scaffolds. The improved mechanical properties were attributed to a homogeneous network structure with additional reinforcement coming from the ELP aggregates. Our study confirms that ELP-collagen composites with superior physical and mechanical properties compared to collagen scaffolds can be produced. Further optimization of design parameters will allow producing ELP-collagen composites for specific biomedical applications.

Rat Mcs1b is concordant to the genome-wide association-identified breast cancer risk locus at human 5q11.2 and MIER3 is a candidate cancer susceptibility gene.

Low-penetrance alleles associated with breast cancer risk have been identified in population-based studies. Most risk loci contain either no or multiple potential candidate genes. Rat mammary carcinoma susceptibility 1b (Mcs1b) is a quantitative trait locus on RN02 that confers decreased susceptibility when Copenhagen (COP)-resistant alleles are introgressed into a Wistar Furth (WF)-susceptible genome. Five WF.COP congenic lines containing COP RN02 segments were compared. One line developed an average of 3.4 ± 2.0 and 5.5 ± 3.6 mammary carcinomas per rat ± SD when females were Mcs1b-resistant homozygous and Mcs1b heterozygous, respectively. These phenotypes were significantly different from susceptible genotype littermates (7.8 ± 3.1 mean mammary carcinomas per rat ± SD, P = 0.0001 and P = 0.0413, respectively). All other congenic lines tested were susceptible. Thus, Mcs1b was narrowed to 1.8 Mb of RN02 between genetic markers ENSRNOSNP2740854 and g2UL2-27. Mammary gland-graft carcinoma susceptibility assays were used to determine that donor (P = 0.0019), but not recipient Mcs1b genotype (P = 0.9381), was associated with ectopic mammary carcinoma outcome. Rat Mcs1b contains sequence orthologous to human 5q11.2, a breast cancer susceptibility locus identified in multiple genome-wide association studies. Human/rat MAP3K1/Map3k1 and mesoderm induction early response (MIER; MIER3)/MIER3 are within these orthologous segments. We identified MIER3 as a candidate Mcs1b gene based on 4.5-fold higher mammary gland levels of MIER3 transcripts in susceptible compared with Mcs1b-resistant females. These data suggest that the human 5q11.2 breast cancer risk allele marked by rs889312 is mammary gland autonomous, and MIER3 is a candidate breast cancer susceptibility gene.

Neovascularization is influenced by androgenic hormones in the tissue implant response.

The objective of this investigation was to demonstrate the effect of androgens on the neovascularization of the fibrous tissue surrounding tricalcium phosphate (TCP) implants. Sixteen animals in four experimental groups (n = 4/group) were implanted with one TCP implant each. Group I animals were implanted with the sham TCP ceramic (Control). Group II animals received a testosterone-loaded ceramic. Group III animals were implanted with a dihydrotestosterone containing bioceramic. Group IV animals received the androstenedione filled bioceramic. At 90 days post-implantation, the fibrous tissue surrounding the implants were evaluated microscopically following staining with routine hematoxylin and eosin (H&E), Masson’s trichrome, and Papanicolaou stains. Using Image Pro (Media Cybernetics, Silver Spring, MD) digital analysis software, data were collected to compare the hormonal effects on the number (per high power field) and size of blood vessels (micrometers, µm) within the fibrous tissue surrounding all four groups. The presence of androgens greatly affected the angiogenic response within the fibrous tissue. All three hormones exhibited less neovascularization compared to the control. Though not as dramatic as androstenedione (3±0), both testosterone (12±1) and dihydrotestosterone (10±1) suppressed the number of blood vessels present in the fibrous tissue capsule compared to control (13±1). However, the circumference of the vessels was much larger for the testosterone (236µm ±8µm) and dihydrotestosterone (256µm±4µm) treated groups compared to the androstenedione (146µm ±7µm) or control (163µm±3µm) groups. The results of this study demonstrate androgens strongly vary in their effect on neovascularization by limiting the number of new vessels developed while contributing to the presence of larger vessels within the fibrous tissue surrounding TCP implants loaded with testosterone and dihydrotestosterone.

Proliferation and morphological transformation of RMK cells exposed to hydroquinine containing ionomers.

Recent research in our laboratories has been directed towards the development of ionomeric polymers and monomers for use in biomedical applications such as adhesives, drug delivery matrices and tissue scaffolds. The chemical Hydroquinone (HQ) aids as a stabilizer and represents a major component in the development of the ionomers. However, hydroquinone in high concentration has the potential to initiate carcinogenic effects on cells. The curing reactions are based on free radical chemistry that require a radical scavenger, ascorbic acid (Asc) to adjust working and setting times and shelf-life stability. The few studies published on HQ have suggested that high dosages of HQ may stimulate apoptosis as well as an increased cellular leakage, however the effect of HQ on the biocompatability is unknown. Therefore the objectives of this study were to measure the functional capacity, cell proliferation and structural integrity of Rhesus monkey kidney epithelial (RMK) cells exposed to ionomer formulations containing 4 different levels of HQ. A total of 90 tubes of RMK (40,000 cells per tube) cells were divided equally into five equal groups. Group I served as a control and group II-V were subjected to ionomers containing 0, 500, 1000, and 2000 ppm HQ. Cell numbers, morphology, cellular and supermatant MDA levels, and total protein analysis were performed. The results suggest: (I) All ionomer groups increased cellular proliferation except for the 2000 ppm HQ group, (II) MDA levels were increased in cells containing 2000 ppm HQ at 24 hours; and 0 ppm at 48 hours. It may be concluded that HQ concentrations over 1000 ppm may adversely affect biocompatability.