Turning on/off tumor-specific CTL response during progressive tumor growth

Therapeutic vaccinations used to induce CTLs and treat firmly established tumors are generally ineffective. To understand the mechanisms underlying the failure of therapeutic vaccinations, we investigated the fate of tumor-specific CD8+ T cells in tumor-bearing mice with or without vaccinations. Our data demonstrate that tumor-specific CD8+ T cells are activated at the early stage of tumor growth, tumor-specific CTL response reaches a maximal level during progressive tumor growth, and tumor-specific CD8+ T cells lose cytolytic function at the late stage of tumor growth. The early stage therapeutic vaccination induces efficient antitumor activity by amplifying the CTL response, whereas the late-stage therapeutic vaccination is invalid due to tumor-induced dysfunction of CD8+ T cells. However, at the late stage, tumor-specific CD8+ T cells are still present in the periphery. These tumor-specific CD8+ T cells lose cytolytic activity, but retain IFN-gamma secretion function. In contrast to in vitro cultured tumor cells, in vivo growing tumor cells are more resistant to tumor-specific CTL killing, despite an increase of tumor Ag gene expression. Both tumor-induced CD8+ T cell dysfunction at the late stage and immune evasion developed by in vivo growing tumor cells contribute to an eventual inefficacy of therapeutic vaccinations. Our study suggests that it is important to design a vaccination regimen according to the stages of tumor growth and the functional states of tumor-specific CD8+ T cells.     

Cyanidin-3-Glucoside Modulates hsa_circ_0001345/miRNA106b/ATG16L1 Axis Expression as a Potential Protective Mechanism against Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) is the most common form of malignancy in the liver. Autophagy was found to have a significant effect in controlling HCC. Anthocyanins, which are naturally occurring pigments in a variety of fruits and vegetables, have been thoroughly documented to be involved in a variety of bioactive activities and are widely employed for their antioxidant capabilities. Cyanidin-3-glucoside (C3G) extracted from Morus alba L. has promising antioxidant and anti-tumour activities. The current study aims to examine the protective action of C3G against hepatocellular carcinoma through the investigation of the autophagy protein ATG16L1 expression along with its related RNA molecules (hsa_circ_0001345 and miRNA106b) in Wistar rats. In vivo precancerous lesions (PCL) were induced using diethylnitrosamine (DEN) and acetamidofluorene (2-AAF). Rats were treated with C3G (10, 15, and 20 mg/kg; 4 times weekly) for 112 days (16 weeks). Liver function tests, alfa fetoprotein, ATG16L1 expression, hsa_circ_0001345, and miRNA106b differential expression were examined. Liver sections were examined by histological and immunohistochemical approaches. The current study’s findings indicated that C3G administration protects against the negative effects of DEN-2-AAF on liver functions and liver histopathological sections, which nominated C3G as a potential prophylactic agent against HCC.     

Automated classification of atherosclerotic plaque from magnetic resonance images using predictive models

The information contained within multicontrast magnetic resonance images (MRI) promises to improve tissue classification accuracy, once appropriately analyzed. Predictive models capture relationships empirically, from known outcomes thereby combining pattern classification with experience. In this study, we examine the applicability of predictive modeling for atherosclerotic plaque component classification of multicontrast ex vivo MR images using stained, histopathological sections as ground truth. Ten multicontrast images from seven human coronary artery specimens were obtained on a 9.4 T imaging system using multicontrast-weighted fast spin-echo (T1-, proton density-, and T2-weighted) imaging with 39-mum isotropic voxel size. Following initial data transformations, predictive modeling focused on automating the identification of specimen's plaque, lipid, and media. The outputs of these three models were used to calculate statistics such as total plaque burden and the ratio of hard plaque (fibrous tissue) to lipid. Both logistic regression and an artificial neural network model (Relevant Input Processor Network-RIPNet) were used for predictive modeling. When compared against segmentation resulting from cluster analysis, the RIPNet models performed between 25 and 30% better in absolute terms. This translates to a 50% higher true positive rate over given levels of false positives. This work indicates that it is feasible to build an automated system of plaque detection using MRI and data mining.     

Low-density lipoprotein receptor-related protein 8 (LRP8) is upregulated in granulosa cells of bovine dominant follicle: molecular characterization and spatio-temporal expression studies

The low-density lipoprotein (LDL) receptor-related protein 8 (LRP8) is a member of the LDL receptor family that participates in endocytosis and signal transduction. We cloned the full-length bovine LRP8 cDNA in granulosa cells (GC) of the dominant follicle (DF) as well as several LRP8 mRNA splicing variants, including a variant that contains a proline-rich cytoplasmic insert (A759-K817) that is involved in intracellular signaling. Expression of the A759-K817 variant was analyzed in the GC of follicles at different developmental stages: the small follicle (SF; 2-4 mm), the DF at Day 5 (D5) of the estrus cycle, ovulatory follicles (OF) 24 h after hCG injection, and corpora lutea (CL) at D5. RT-PCR analysis showed that expression was predominant in the GC of DF compared to other follicles and CL (P<0.0001), whereas the expression of other related receptors, such as LDLR and VLDLR, did not show differences. Temporal analyses of follicular walls from the OF following hCG treatment revealed a decrease in LRP8 mRNA expression starting 12 h post-hCG treatment (P<0.0001). LRP8 protein was exclusively localized to the GC, with higher levels in the DF than in the SF (P<0.05). RELN mRNA, which encodes an LRP8 ligand, was highly expressed in the theca of the DF as compared to the OF (P<0.004), whereas MAPK8IP1 mRNA, which encodes an LRP8 intracellular interacting partner, is expressed in the GC of the DF. These results demonstrate the differential expression patterns of LRP8, RELN, and MAPK8IP1 mRNAs during final follicular growth and ovulation, and suggest that a RELN/LRP8/MAPK8IP1 paracrine interaction regulates follicular growth.     

Noninvasive detection of macrophages using a nanoparticulate contrast agent for computed tomography

Sudden fibrous cap disruption of 'high-risk' atherosclerotic plaques can trigger the formation of an occlusive thrombus in coronary arteries, causing acute coronary syndromes. High-risk atherosclerotic plaques are characterized by their specific cellular and biological content (in particular, a high density of macrophages), rather than by their impact on the vessel lumen. Early identification of high-risk plaques may be useful for preventing ischemic events. One major hurdle in detecting high-risk atherosclerotic plaques in coronary arteries is the lack of an imaging modality that allows for the identification of atherosclerotic plaque composition with high spatial and temporal resolutions. Here we show that macrophages in atherosclerotic plaques of rabbits can be detected with a clinical X-ray computed tomography (CT) scanner after the intravenous injection of a contrast agent formed of iodinated nanoparticles dispersed with surfactant. This contrast agent may become an important adjunct to the clinical evaluation of coronary arteries with CT.     

Enzymatic characterization of a recombinant isoform hybrid of glutamic acid decarboxylase (rGAD67/65) expressed in yeast

BACKGROUND AND AIMS: Glutamic acid decarboxylase (GAD, EC 4.1.1.15) catalyses the conversion of glutamate to gamma-aminobutyric acid (GABA). The 65 kDa isoform, GAD65 is a potent autoantigen in type 1 diabetes, whereas GAD67 is not. A hybrid cDNA was created by fusing a human cDNA for amino acids 1-101 of GAD67 to a human cDNA for amino acids 96-585 of GAD65; the recombinant (r) protein was expressed in yeast and was shown to have equivalent immunoreactivity to mammalian brain GAD with diabetes sera. We here report on enzymatic and molecular properties of rGAD67/65. METHODS: Studies were performed on enzymatic activity of rGAD67/65 by production of 3H-GABA from 3H-glutamate, enzyme kinetics, binding to the enzyme cofactor pyridoxal phosphate (PLP), stability according to differences in pH, temperature and duration of storage, and antigenic reactivity with various GAD-specific antisera. RESULTS: The properties of rGAD67/65 were compared with published data for mammalian brain GAD (brackets). These included a specific enzyme activity of 22.7 (16.7) nKat, optimal pH for enzymatic activity 7.4 (6.8), K(m) of 1.3 (1.3) mM, efficient non-covalent binding to the cofactor PLP, and high autoantigenic potency. The stability of rGAD67/65 was optimal over 3 months at -80 degrees C, or in lyophilized form at -20 degrees C. CONCLUSIONS: Hybrid rGAD67/65 has enzymatic and other properties similar to those of the mixed isoforms of GAD in preparations from mammalian brain as described elsewhere, in addition to its previously described similar immunoreactivity.     

Glucanolytic Actinomycetes Antagonistic to Phytophthora fragariae var. rubi, the Causal Agent of Raspberry Root Rot

A collection of about 200 actinomycete strains was screened for the ability to grow on fragmented Phytophthora mycelium and to produce metabolites that inhibit Phytophthora growth. Thirteen strains were selected, and all produced (beta)-1,3-, (beta)-1,4-, and (beta)-1,6-glucanases. These enzymes could hydrolyze glucans from Phytophthora cell walls and cause lysis of Phytophthora cells. These enzymes also degraded other glucan substrates, such as cellulose, laminarin, pustulan, and yeast cell walls. Eleven strains significantly reduced the root rot index when inoculated on raspberry plantlets.     

Oral administration with papillomavirus pseudovirus encoding IL-2 fully restores mucosal and systemic immune responses to vaccinations in aged mice

Infectious diseases are one of the major threats for the elderly because their immune system is often compromised, and vaccinations to prevent these infections are not effective. A major defect in their immune system seems to be the inability of T cells to produce IL-2. We used papillomavirus (PV) pseudoviruses (PSVs) as a model vaccine and a gene delivery vector to address how to enhance immune responses to vaccinations. We found that oral immunization with PV PSV induced minimal mucosal and systemic Abs and CTLs specific for the PSVs in aged mice compared with young adult mice. In addition, fewer specific Th cells were generated in the aged mice. When aged mice were immunized with PV PSVs encoding human IL-2, specific Th cells were generated, producing murine IL-2, IL-4, and IFN-gamma. Further, specific Abs and CTLs were induced, resulting in protection against mucosal viral challenge. Thus, this study provided a basis for clinical trials using PV PSVs encoding IL-2 for vaccination of the elderly.