A visible fluorescent nanovaccine based on functional genipin crosslinked ovalbumin protein nanoparticles

Accurate and efficient antigen delivery is crucial for inducing a strong and long-term immune response. A visible protein nanovaccine made from antigen could provide a novel and promising technology for secure and efficient delivery of the antigen with imaging visualization. In this study, a functional nanovaccine based on genipin crosslinked ovalbumin (OVA) fluorescent nanoparticles with chitosan (CS-OVA-NPs) was developed. The nanovaccine can carry abundant antigens by self-crosslinking without additional carriers. The fluorescence imaging technique was applied to monitor and reveal the process of antigen delivery in vivo based on the fluorescence of genipin with a non-invasive and real-time manner. This functional OVA nanovaccine can enhance the uptake of OVA in Dendritic Cells (DCs) and further promote DCs to maturate in vitro. In vivo study further indicated CS-OVA-NPs could trigger antigen-specific immune responses, which demonstrated that this fluorescent nanovaccine provided a novel design approach for accurate and efficient vaccine delivery.     

Pulmonary delivery of polyplexes for combined PAI-1 gene silencing and CXCR4 inhibition to treat lung fibrosis

This report describes the development of polyplexes based on CXCR4-inhibiting poly(ethylenimine) derivative (PEI-C) for pulmonary delivery of siRNA to silence plasminogen activator inhibitor-1 (siPAI-1) as a new combination treatment of pulmonary fibrosis (PF). Safety and delivery efficacy of the PEI-C/siPAI-1 polyplexes was investigated in vitro in primary lung fibroblasts isolated from mice with bleomycin-induced PF. Biodistribution analysis following intratracheal administration of fluorescently labeled polyplexes showed prolonged retention in the lungs. Treatment of mice with bleomycin-induced PF using the PEI-C/siPAI-1 polyplexes resulted in a significant down-regulation of the PAI-1 expression and decreased collagen deposition in the lung. The results of this study provide first evidence of the potential benefits of combined inhibition of CXCR4 and PAI-1 in the pulmonary treatment of PF.     

Nano-topography: Quicksand for cell cycle progression?

The 3-D spatial and mechanical features of nano-topography can create alternative environments, which influence cellular response. In this paper, murine fibroblast cells were grown on surfaces characterized by protruding nanotubes. Cells cultured on such nano-structured surface exhibit stronger cellular adhesion compared to control groups, but despite the fact that stronger adhesion is generally believed to promote cell cycle progression, the time cells spend in G1 phase is doubled. This apparent contradiction is solved by confocal microscopy analysis, which shows that the nano-topography inhibits actin stress fiber formation. In turn, this impairs RhoA activation, which is required to suppress the inhibition of cell cycle progression imposed by p21/p27. This finding suggests that the generation of stress fibers, required to impose the homeostatic intracellular tension, rather than cell adhesion/spreading is the limiting factor for cell cycle progression. Indeed, nano-topography could represent a unique tool to inhibit proliferation in adherent well-spread cells.     

Detection of Memory B Activity Against a Therapeutic Protein in Treatment-Naïve Subjects

Bridging immunoassays commonly used to detect and characterize immunogenicity during biologic development do not provide direct information on the presence or development of a memory anti-drug antibody (ADA) response. In this study, a B cell ELISPOT assay method was used to evaluate pre-existing ADA for anti-TNFR1 domain antibody, GSK1995057, an experimental biologic in treatment naive subjects. This assay utilized a 7-day activation of PBMCs by a combination of GSK1995057 (antigen) and polyclonal stimulator followed by GSK1995057-specific ELISPOT for the enumeration of memory B cells that have differentiated into antibody secreting cells (ASC) in vitro. We demonstrated that GSK1995057-specific ASC were detectable in treatment-naïve subjects with pre-existing ADA; the frequency of drug-specific ASC was low and ranged from 1 to 10 spot forming units (SFU) per million cells. Interestingly, the frequency of drug-specific ASC correlated with the ADA level measured using an in vitro ADA assay. We further confirmed that the ASC originated from CD27⁺ memory B cells, not from CD27^?-naïve B cells. Our data demonstrated the utility of the B cell ELISPOT method in therapeutic protein immunogenicity evaluation, providing a novel way to confirm and characterize the cell population producing pre-existing ADA. This novel application of a B cell ELISPOT assay informs and characterizes immune memory activity regarding incidence and magnitude associated with a pre-existing ADA response.     

<Emphasis Type="Italic">In Vitro</Emphasis>-<Emphasis Type="Italic">In Vivo</Emphasis> Dose Response of Ursolic Acid,Sulforaphane, PEITC,and Curcumin in Cancer Prevention

According to the National Center of Health Statistics, cancer was the culprit of nearly 600,000 deaths in 2016 in the USA. It is by far one of the most heterogeneous diseases to treat. Treatment for metastasized cancers remains a challenge despite modern diagnostics and treatment regimens. For this reason, alternative approaches are needed. Chemoprevention using dietary phytochemicals such as triterpenoids, isothiocyanates, and curcumin in the prevention of initiation and/or progression of cancer poses a promising alternative strategy. However, significant challenges exist in the extrapolation of in vitro cell culture data to in vivo efficacy in animal models and to humans. In this review, the dose at which these phytochemicals elicit a response in vitro and in vivo of a multitude of cellular signaling pathways will be reviewed highlighting Nrf2-mediated antioxidative stress, anti-inflammation, epigenetics, cytoprotection, differentiation, and growth inhibition. The in vitro-in vivo dose response of phytochemicals can vary due, in part, to the cell line/animal model used, the assay system of the biomarker used for the readout, chemical structure of the functional analog of the phytochemical, and the source of compounds used for the treatment study. While the dose response varies across different experimental designs, the chemopreventive efficacy appears to remain and demonstrate the therapeutic potential of triterpenoids, isothiocyanates, and curcumin in cancer prevention and in health in general.     

Nano in nano: Biosynthesized gold and iron nanoclusters cargo neoplastic exosomes for cancer status biomarking

Timely detection is crucial for successful treatment of cancer. The current study describes a new approach that involves utilization of the tumor cell environment for bioimaging with in-situ biosynthesized nanoscale gold and iron probes and subsequent dissemination of Au-Fe nanoclusters from cargo exosomes within the circulatory system. We have isolated the Au-Fe cargo exosomes from the blood of the treated murine models after in situ biosyntheses from their respective pre-ionic solutions (HAuCl₄, FeCl₂), whereas Na₂SeO₃ supplementation added into Au lethal effect. The microarray data of various differentially expressed genes revealed the up-regulated tumor ablation and metal binding genes in SGC-7901 cell lines after treatment with Au-Fe-Se triplet ionic solution. The isolation of Au-Fe nanoclusters cargo exosomes (nano in nano) after secretion from deeply seated tumors may help in early diagnosis and reveal the tumor ablation status during and after the relevant treatment like radio-chemo therapies et al.     

Clinical ultrasound stimulating angiogenesis following drug-release from polymersomes on the ischemic zone for peripheral arterial occlusive disease

Peripheral Arterial Occlusive Disease (PAOD) is an aging disease that affects the quality of life of many people by its intermittent claudication and critical limb ischemia presentations. Traditional treatment and management of PAOD are asking patients to make a life change and medication with antiplatelet, statins and cilostazol, which decrease the possibility of clot formation. Our strategy has employed a magnetic Fe₃O₄-PLGA polymersome to carry the cilostazol into the ischemic area by magnetic attraction following remote-control drug release through low-energy ultrasound exposure. In the animal studies, the cilostazol-loaded Fe₃O₄-PLGA polymersomes were injected and accumulated at ischemic leg through magnetic attraction. Then, using a clinical-use ultrasound machine the leg was irradiated to forward cilostazol release from the accumulated polymersomes. Dramatically, we found an observable result of bloody flux recovery in the leg after 7?days compared to the non-treated leg that showed no evidence of the blood recovery.     

The intraocular staining potential of anthocyanins and their retinal biocompatibility: a preclinical study

Purpose: To perform preclinical studies to determine the efficacy and safety of anthocyanins as stains for the internal limiting membrane (ILM) of the eye.

Materials and methods: Cyanidin (Cya), delphinidin (Del), luteolinidin (Lut), peonidin (Peo) and pelargonidin (Pel) were evaluated. These natural dyes were used to stain the lens capsule and ILM of pig eyes. The effects of these dyes on retinal cell viability was determined using a water-soluble tetrazolium salt assay, and oxidative stress was measured in vitro. Histopathology, in situ TUNEL labelling, transmission electronic microscopy (TEM), and electroretinography (ERG) were performed on rats following the intravitreal and subretinal injection of the neuroprotective dyes.

Results: All anthocyanins stained the lens capsule and ILM of the pigs at a concentration of 1?mg/ml. Del, Lut and Peo were non-toxic and produced survival rates in the ARPE19 and RGC5 cells that were similar to those in control cells. We treated eyes with H₂O₂ and three dyes (Del, Lut, and Peo) to explore the possible neuroprotective effects and observed significantly higher survival rates in the ARPE19 cells treated with Del, Lut or Peo and the RGC5 cells treated with Lut or Peo than those in the control cells. Three dyes were intravitreally and subretinally injected into rats in vivo, and the histology showed mildly disorganized retinal cell layers. TUNEL staining and TEM examinations did not reveal additional toxic effects. Rat ERGs were not altered after intravitreal injections.

Conclusions: This preclinical study, Del, Lut, and Peo show potential as staining agents and warrant further investigation as vital dyes.     

Mutation spectrum of MMACHC in Chinese pediatric patients with cobalamin C disease: A case series and literature review

Cobalamin (cbl) C disease is a rare autosomal recessive inheritance disease, which is the most common cobalamin metabolic disorder. Its clinical phenotype involves multiple systems with varying degrees of severity, where in mild cases can be asymptomatic for many years, whereas severe cases may cause death during the neonatal period. The disease is caused by mutations in the MMACHC gene located on chromosome 1p34.1 that contains 5 exons; among which, exons 1–4 have an 849 bp coding sequence that encodes a protein containing 282 amino acids. Through clinical physical examination and laboratory tests, especially blood and urine screening, we found 28 cblC pediatric patients with clinical manifestations, such as mental retardation, motor development delay, epilepsy, metabolic acidosis, vomiting and diarrhea. By Sanger sequencing, we found homozygous or compound heterozygous mutations of MMACHC in 27 of the patients, and single heterozygous mutation of MMACHC in one of them. The c.609G > A, c.658-660delAAG, c.80A > G and c.482G > A mutations accounted for 43.64% (24/55), 10.91% (6/55), 9.09% (5/55) and 7.27% (4/55) of all the mutations, respectively. This spectrum finding is basically consistent with the previously reported data in Chinese patients. The most common c.609G > A mutation may likely lead to early-onset cblC disease. In previous literature involving a large sample of Caucasian cblC cases, the mutation spectrum of MMACHC gene is almost completely different from that of the Chinese population. The most common mutations in the Caucasian population were c.271dupA, c.394C > T and c.331C > T, which account for 48.05% (542/1128), 13.65% (154/1128) and 7.36% (83/1128) of all the mutant alleles, respectively. The c.271dupA mutation and c.331C > T mutation were mainly associated with early-onset cblC in children less than 1 year old, whilst the c.394C > T mutation was mainly associated with late-onset cblC patients characterised by isolated acute nervous system abnormalities. We also analysed the cause behind the different mutation spectrum of MMACHC gene between the Chinese and Caucasian populations.