Vaccination against neoantigens induced in cross-priming cDC1 in vivo.

The conventional type 1 dendritic cells (cDC1) play a pivotal role in protective immunity against pathogens and cancer. However, their low frequency in the blood and tissues limits their use in immune therapy. We have recently described a method to vaccinate against neoantigens that are induced in tumor cells by targeted delivery of a TAP siRNA to dendritic cells using a TLR9 binding CpG oligonucleotide. Since TLR9 is also expressed in immune suppressive myeloid populations TLR9 targeting could reduce the effectiveness of this approach. Here, we describe a modular multivalent antibody platform to target the TAP siRNA to resident Clec9a expressing cDC1 and show that it leads to selective and sustained TAP downregulation in cDC1 and inhibits tumor growth in mice more effectively than CpG targeted siRNA. To induce DC maturation an agonistic CD40 antibody was administered to the siRNA treated mice. To obviate the need for a second drug formulation and reduce the risk of toxicity, we exploited the multivalent nature of this targeting platform to co-deliver the TAP siRNA and a DC maturation agent, a CpG containing oligonucleotide, to cDC1 in vivo and show that it was more effective than Clec9a targeting of TAP siRNA in combination with CD40 antibody. This study describes a way to manipulate the function of cDC1 cells in vivo using a broadly applicable antibody-based targeting platform to deliver multiple biological agents to specific cells in vivo to potentiate (immune) therapy and to probe the biology of specific cell types in their natural settings.

High waterborne Mg does not attenuate the toxic effects of Fe, Mn, and Ba on Na+ regulation of Amazonian armored catfish tamoatá (Hoplosternum litoralle).

Formation water (FoW) is a by-product from oil and gas production and usually has high concentrations of soluble salts and metals. Calcium (Ca) and magnesium (Mg) have been shown to reduce the toxicity of metals to aquatic animals, and previous study showed that high waterborne Ca exerts mild effect against disturbances on Na+ regulation in Amazonian armored catfish tamoatá (Hoplosternum littorale) acutely exposed to high Fe, Mn, and Ba levels. Here, we hypothesized that high Mg levels might also reduce the toxic effects of these metals on Na+ regulation of tamoatá. The exposure to 5% FoW promoted an increase in Na+ uptake and a rapid accumulation of Na+ in all tissues analyzed (kidney

Radiation-Induced Enhancement of Antitumor T-cell Immunity by VEGF-Targeted 4-1BB Costimulation.

Radiotherapy can elicit systemic immune control of local tumors and distant nonirradiated tumor lesions, known as the abscopal effect. Although this effect is enhanced using checkpoint blockade or costimulatory antibodies, objective responses remain suboptimal. As radiotherapy can induce secretion of VEGF and other stress products in the tumor microenvironment, we hypothesized that targeting immunomodulatory drugs to such products will not only reduce toxicity but also broaden the scope of tumor-targeted immunotherapy. Using an oligonucleotide aptamer platform, we show that radiation-induced VEGF-targeted 4-1BB costimulation potentiated both local tumor control and abscopal responses with equal or greater efficiency than 4-1BB, CTLA-4, or PD1 antibodies alone. Although 4-1BB and CTLA-4 antibodies elicited organ-wide inflammatory responses and tissue damage, VEGF-targeted 4-1BB costimulation produced no observable toxicity. These findings suggest that radiation-induced tumor-targeted immunotherapy can improve the therapeutic index and extend the reach of immunomodulatory agents. Cancer Res; 77(6); 1310-21. ©2017 AACR.

4-1BB Aptamer-Based Immunomodulation Enhances the Therapeutic Index of Radiation Therapy in Murine Tumor Models.

PURPOSE: To report a novel strategy using oligonucleotide aptamers to 4-1BB as an alternate method for costimulation, and show that combinatorial therapy with radiation improves the therapeutic ratio over equivalent monoclonal antibodies. METHODS AND MATERIALS: Subcutaneous 4T1 (mouse mammary carcinoma) tumors were established (approximately 100 mm(3)), and a radiation therapy (RT) dose/fractionation schedule that optimally synergizes with 4-1BB monoclonal antibody (mAb) was identified. Comparable tumor control and animal survival was observed when either 4-1BB antibody or aptamer were combined with RT using models of breast cancer and melanoma (4T1 and B16-F10). Off-target CD8(+) T-cell toxicity was evaluated by quantification of CD8(+) T cells in livers and spleens of treated animals. RESULTS: When combined with 4-1BB mAb, significant differences in tumor control were observed by varying RT dose and fractionation schedules. Optimal synergy between RT and 4-1BB mAb was observed at 5 Gy × 6. Testing 4-1BB mAb and aptamer independently using the optimal RT (5 Gy × 6 for 4T1/Balb/c and 12 Gy × 1 for B16/C57BL6J mouse models) revealed equivalent tumor control using 4-1BB aptamer and 4-1BB mAb. 4-1BB mAb, but not 4-1BB aptamer-treated animals, exhibited increased lymphocytic liver infiltrates and increased splenic and liver CD8(+) T cells. CONCLUSIONS: Radiation therapy synergizes with 4-1BB mAb, and this effect is dependent on RT dose and fractionation. Tumor control by 4-1BB aptamer is equivalent to 4-1BB mAb when combined with optimal RT dose, without eliciting off-target liver and spleen CD8(+) expansion. 4-1BB aptamer-based costimulation affords a comparable and less toxic strategy to augment RT-mediated tumor control.

Sodium fluxes in tamoatá, Hoplosternum litoralle, exposed to formation water from Urucu Reserve (Amazon, Brazil).

Formation water (produce water or oil field brine) from oil and gas production usually has high concentrations of soluble salts and metals. The objective of this study was to examine the effect of formation water from Urucu Reserve, Amazon, on whole-body uptake and internal distribution of newly accumulated Na+ in juvenile tamoatá, Hoplosternum litoralle. Groups of fish were submitted to nine treatments for 3 h in 400-ml chambers: control (well water), 5% formation water, and well water with respective concentrations of 5% formation water of Ca2+, Fe, Mn, Ba2+, Fe+Ca2+, Mn+Ca2+, and Ba+Ca2+ added. Specimens of tamoatá exposed to 5% formation water presented a very high Na+ influx, probably due to the high Na+ levels in this water. Waterborne Fe and Mn stimulated Na+ influx, but Fe increased Na+ efflux, causing Na+ loss. Waterborne Mn, on the other hand, decreased Na+ efflux, reducing Na+ loss by this species. Waterborne Ca2+ also affected Na+ influx but had no significant effect on net Na+ fluxes. These results demonstrated that spilling of formation water in ion-poor Amazon rivers would dramatically disrupt osmoregulatory balance of tamoatá and probably other Amazon fish species, impairing their survival and reduce biodiversity.

Quantification of carbon nanotube induced adhesion of osteoblast on hydroxyapatite using nano-scratch technique.

This paper explores the nano-scratch technique for measuring the adhesion strength of a single osteoblast cell on a hydroxyapatite (HA) surface reinforced with carbon nanotubes (CNTs). This technique efficiently separates out the contribution of the environment (culture medium and substrate) from the measured adhesion force of the cell, which is a major limitation of the existing techniques. Nano-scratches were performed on plasma sprayed hydroxyapatite (HA) and HA-CNT coatings to quantify the adhesion of the osteoblast. The presence of CNTs in HA coating promotes an increase in the adhesion of osteoblasts. The adhesion force and energy of an osteoblast on a HA-CNT surface are 17 ± 2 µN/cell and 78 ± 14 pJ/cell respectively, as compared to 11 ± 2 µN/cell and 45 ± 10 pJ/cell on a HA surface after 1 day of incubation. The adhesion force and energy of the osteoblasts increase on both the surfaces with culture periods of up to 5 days. This increase is more pronounced for osteoblasts cultured on HA-CNT. Staining of actin filaments revealed a higher spreading and attachment of osteoblasts on a surface containing CNTs. The affinity of CNTs to conjugate with integrin and other proteins is responsible for the enhanced attachment of osteoblasts. Our results suggest that the addition of CNTs to surfaces used in medical applications may be beneficial when stronger adhesion of osteoblasts is desired.

Wear behavior and in vitro cytotoxicity of wear debris generated from hydroxyapatite-carbon nanotube composite coating.

This work evaluates the effect of carbon nanotube (CNT) addition to plasma-sprayed hydroxyapatite (HA) coating on its tribological behavior, biocompatibility of the coating, and cytotoxicity of CNT-containing wear debris. Biological response of the CNT-containing wear debris is critical for osteoblasts, the bone-forming cells, and macrophages, the cells that clear up wear debris from blood stream. The addition of 4 wt % CNTs to HA coating reduces the volume of wear debris generation by 80% because of the improved elastic modulus and fracture toughness. CNT reinforcement has a pronounced effect on the particle size in the wear debris and subsequent biological response. There was a slight increase in the numbers and viability of osteoblasts grown on HA-CNT compared with HA alone. The cytotoxic effect of HA and HA-CNT debris to macrophages and osteoblasts was similar, demonstrating that loose CNT does not pose a problem to these cells.

Boron nitride nanotube reinforced hydroxyapatite composite: mechanical and tribological performance and in-vitro biocompatibility to osteoblasts.

This study proposes boron nitride nanotube (BNNT) reinforced hydroxyapatite (HA) as a novel composite material for orthopedic implant applications. The spark plasma sintered (SPS) composite structure shows higher density compared to HA. Minimal lattice mismatch between HA and BNNT leads to coherent bonding and strong interface. HA-4 wt% BNNT composite offers excellent mechanical properties-120% increment in elastic modulus, 129% higher hardness and 86% more fracture toughness, as compared to HA. Improvements in the hardness and fracture toughness are related to grain refinement and crack bridging by BNNTs. HA-BNNT composite also shows 75% improvement in the wear resistance. The wear morphology suggests localized plastic deformation supported by the sliding of outer walls of BNNT. Osteoblast proliferation and cell viability show no adverse effect of BNNT addition. HA-BNNT composite is, thus, envisioned as a potential material for stronger orthopedic implants.

Protective effect of high alkalinity against the deleterious effects of chronic waterborne cadmium exposure on the detection of alarm cues by juvenile silver catfish (Rhamdia quelen).

The objective of the present study was to analyze the effect of chronic cadmium (Cd) exposure at two alkalinity levels (63 and 92 mg l(-1) CaCO(3)) on the antipredatory behavior of juvenile silver catfish (Rhamdia quelen) exposed to conspecific skin extract and predator odor. At an alkalinity of 63 mg l(-1) CaCO(3), 30 days of exposure to either 4.5 or 8.0 microg l(-1) Cd impaired the catfish's antipredatory response to alarm cues. However, silver catfish exposed to 4.5 microg l(-1) Cd at an alkalinity of 92 mg l(-1) CaCO(3) responded to skin extract and predator odor. In catfish exposed to 8.0 microg l(-1) Cd at the same alkalinity, only the number of feeding bites decreased, and this occurred only for specimens exposed to predator odor. Our results show that higher alkalinity protected against the deleterious effects of Cd on alarm cue detection but only in the larvae exposed to the lowest waterborne Cd level.

Toxicity of cadmium for silver catfish Rhamdia quelen (Heptapteridae) embryos and larvae at different alkalinities.

The present study evaluated the effect of waterborne Cd in eggs and larvae of silver catfish, Rhamdia quelen, at two alkalinity levels (63 and 92 mg . L(-1) CaCO(3)): 0.5 (control), 4.5, 8, and 18 microg . L(-1) Cd. The fertilization rate was similar in all treatments, but the number of eggs with irregular surface was significantly higher in those exposed to 18 microg . L(-1) Cd and 63 mg . L(-1) CaCO(3). Three days after hatching survival of larvae exposed to 4.5 and 8 microg . L(-1) Cd and 63 mg . L(-1) CaCO(3) was significantly lower than that of control larvae, and 21 days after hatching survival decreased with the increase in waterborne Cd levels and total cadmium accumulation in the larvae. These significant differences and relationships were not observed in larvae maintained at 92 mg . L(-1) CaCO(3). Furthermore, in the lower alkalinity, a higher incidence of barbels and spinal column deformities was observed in the highest waterborne Cd concentration. At the end of the experimental period, 21 days after hatching, larval survival rate, length, weight, head height, membranous layer thickness, and biomass decreased with the increase in waterborne Cd levels in the lower alkalinity. However, these relationships were not observed in larvae maintained at 92 mg . L(-1) CaCO(3). The percentage specific growth rate was lower in larvae exposed to the highest waterborne Cd concentration at 63 mg . L(-1) CaCO(3). These results suggest that alkalinity of 92 mg . L(-1) CaCO(3) reduces waterborne Cd toxicity in silver catfish larvae.

A mathematical model for growth in weight of silver catfish (Rhamdia quelen) (Heptapteridae, Siluriformes, Teleostei)

The use of a mathematical model applied to biological science helps to predict the specific data. Based on biological data (weight and age) of silver catfish, Rhamdia quelen, a mathematical model was elaborated based on a nonlinear difference equation to demonstrate the relationship between age and growth in weight. Silver catfish growth was described following the Beverton-Holt model Pt+1 = (r Pt) / (1+ a Pt ), where r > 0 is the maximum growth rate and a > 0 is a constant of growth inhibition. The solution of this equation is Pt= 1 /{[1/P0 - a / (r-1)] 1/r t + a/ (r-1)}, were P0 is the initial weight of the fish. Through this model it was observed that the female reaches the theoretical maximum weight approximately at the age of 18 years and the male at the age of 12 years in a natural environment.

A formulação de modelos matemáticos aplicado às ciências biológicas auxilia na previsão de dados específicos. Fundamentado em dados biológicos (peso e idade) de jundiá, Rhamdia quelen, elaborou-se um modelo matemático com base em equações a diferenças não lineares para demonstrar a relação entre idade e crescimento em peso. O crescimento do jundiá foi descrito segundo o modelo de Beverton-Holt Pt+1 = (r Pt) / (1+ a Pt), onde r > 0 é a taxa de crescimento máxima e a > 0 é uma constante de inibição do crescimento. A solução dessa equação é Pt= 1 / {[1/P0 - a / (r-1)] 1/r t + a/ (r-1)}, onde P0 é o peso inicial do peixe. Por esse modelo foi observado que fêmeas alcançam o peso máximo aproximadamente aos 18 anos e os machos aos 12 anos, em ambiente natural.