It has been reported that an antioxidant-rich, blueberry-supplemented rat diet may retard brain aging in the rat. The present study determined whether such supplementation could prevent impaired object recognition memory and elevated levels of the oxidative stress-responsive protein, nuclear factor-kappa B (NF-kappaB) in aged Fischer-344 rats. Twelve aged rats had been fed a 2% blueberry supplemented diet for 4 months prior to testing. Eleven aged rats and twelve young rats had been fed a control diet. The rats were tested for object recognition memory on the visual paired comparison task. With a 1-h delay between training and testing, aged control diet rats performed no better than chance. Young rats and aged blueberry diet rats performed similarly and significantly better than the aged control diet group. Levels of NF-kappaB in five brain regions of the above subjects were determined by western blotting assays. In four regions, aged control diet rats had significantly higher average NF-kappaB levels than young animals on the control diet. In four regions, aged blueberry diet rats had significantly lower levels of NF-kappaB than aged control diet rats. Normalized NF-kappaB levels (averaged across regions and in several individual regions) correlated negatively and significantly with the object memory scores.

Current efforts to develop Zika virus (ZIKV) subunit vaccines have been focused on pre-membrane (prM) and envelope (E) proteins, but the role of NS1 in ZIKV-specific immune response and protection is poorly understood. Here, we develop an attenuated recombinant vesicular stomatitis virus (rVSV)-based vaccine expressing ZIKV prM-E-NS1 as a polyprotein. This vectored vaccine candidate is attenuated in mice, where a single immunization induces ZIKV-specific antibody and T cell immune responses that provide protection against ZIKV challenge. Co-expression of prM, E, and NS1 induces significantly higher levels of Th2 and Th17 cytokine responses than prM-E. In addition, NS1 alone is capable of conferring partial protection against ZIKV infection in mice even though it does not induce neutralizing antibodies. These results demonstrate that attenuated rVSV co-expressing prM, E, and NS1 is a promising vaccine candidate for protection against ZIKV infection and highlights an important role for NS1 in ZIKV-specific cellular immune responses.

The diagnosis of vascular neoplasms is often facilitated by the use of immunohistochemical markers such as factor VIII-related antigen, CD31, and CD34. However, the relative sensitivity and specificity of these markers have not been compared in cat vascular neoplasms. In this study, these 3 immunohistochemical markers were evaluated in 61 endothelial neoplasms (50 hemangiosarcomas and 11 hemangiomas) in 59 cats. All neoplasms were labeled by all 3 markers. CD34 had the highest average immunolabeling intensity in neoplastic endothelial cells. CD31 had the lowest average background labeling, followed by CD34 and factor VIII-related antigen, respectively. CD34 expression was also examined in 130 nonvascular neoplasms of cats; 14 of 62 epithelial neoplasms, 39 of 43 mesenchymal neoplasms, 8 of 23 leukocytic neoplasms, and 2 of 2 melanomas were positive. Given the broad expression of CD34 in mesenchymal neoplasms, this marker has limited diagnostic relevance for vascular neoplasms of cats.

CD8؉ T cell responses are critical to the control of replication and reactivation associated with gammaherpesvirus infection. Type I interferons (IFNs) have been shown to have direct and indirect roles in supporting CD8؉ T cell development and function during viral infection; however, the role of type I interferons during latent viral infection has not been examined. Mice deficient in type I IFN signaling (IFNAR1 ؊/؊ mice) have high levels of reactivation during infection with murine gammaherpesvirus 68 (MHV68), a murine gammaherpesvirus model for Epstein-Barr virus. We hypothesized that type I IFNs function to enhance the anti-gammaherpesvirus CD8 ؉ T cell response. To test this, IFNAR1 ؊/؊ mice were infected with MHV68 and the CD8 ؉ T cell response was analyzed. In the absence of type I IFN signaling, there was a marked increase in short-lived effector CD8 ؉ T cells, and MHV68-specific CD8 ؉ T cells had upregulated expression of PD-1 and reduced tumor necrosis factor alpha (TNF-␣), gamma IFN (IFN-␥), and interleukin-2 (IL-2) production. Suppressing MHV68 replication early in infection using the antiviral cidofovir rescued CD8 ؉ T cell cytokine production and reduced PD-1 expression. However, suppressing high levels of reactivation in IFNAR1؊/؊ mice failed to improve CD8 ؉ T cell cytokine production during latency. T cell-specific abrogation of type I IFN signaling showed that the effects of type I IFNs on the CD8 ؉ T cell response during MHV68 infection are independent of direct type I IFN signaling on T cells. Our findings support a model in which type I IFNs likely suppress MHV68 replication, thus limiting viral antigen and facilitating an effective gammaherpesvirus-directed CD8 ؉ T cell response. IMPORTANCE The murine gammaherpesvirus MHV68 has both genetic and biologic homology to the human gammaherpesvirus Epstein-Barr virus (EBV), which infects over 90% of humans. Latent EBV infection and reactivation are associated with various life-threatening diseases and malignancies. Host suppression of gammaherpesvirus latency and reactivation requires both CD8 T he gammaherpesvirus-directed CD8ϩ T cell response is critical to the control of replication and reactivation associated with Epstein-Barr virus (EBV) infection, and individuals with either genetic or acquired immunodeficiencies are highly susceptible to EBV-associated diseases (1-3). Adoptive transfer of EBVspecific CD8 ϩ T cells has been successfully utilized to treat EBVassociated lymphoproliferative disease (4, 5). In addition, CD8 ϩ T cells prevent in vivo tumor outgrowth of B cell cancer lines immortalized by murine gammaherpesvirus 68 (MHV68), a wellcharacterized virus model for EBV (6). Thus, CD8ϩ T cells can suppress gammaherpesvirus-associated malignancies. The promise of immunotherapy and vaccine development relies on our understanding of factors that promote a highly effective gammaherpesvirus-directed CD8 ϩ T cell response. CD8 ϩ T cells responding to their cognate antigen require three signals for survival and differentiation: antige...

Objective: To determine the diagnostic accuracy of optical coherence tomography (OCT) to assess surgical margins of canine soft tissue sarcoma (STS) and determine the influence of observer specialty and training. Study design: Blinded clinical prospective study. Animals: Twenty-five dogs undergoing surgical excision of STS. Methods: In vivo and ex vivo surgical margins were imaged with OCT after tumor resection. Representative images and videos were used to generate a training presentation and data sets. These were completed by 16 observers of four specialties (surgery, radiology, pathology, and OCT researchers). Images and videos from data sets were classified as cancerous or noncancerous. Results: The overall sensitivity and specificity were 88.2% and 92.8%, respectively, for in vivo tissues and 82.5% and 93.3%, respectively, for ex vivo

The diagnostic evaluation of nonalcoholic fatty liver disease (NAFLD) relies on the initial detection of hepatomegaly or elevated serum aminotransferases by the primary care provider. In this investigation, the aptitude of the primary care provider to identify children with hepatomegaly and initiate an evaluation for NAFLD is determined. The physical examination findings and requests for diagnostic testing of 18 primary care physicians (and pediatric gastroenterologists) on 11 obese school-aged children, including a subset of children with hepatomegaly and NAFLD, were analyzed. In children with NAFLD, clinicians detected hepatomegaly in 1.4% of encounters and requested serum liver chemistries in 12.5% of encounters. Hepatomegaly is detected poorly in obese children by primary care physicians and thereby increases the likelihood of a delayed or omitted evaluation for NAFLD.

Human metapneumovirus (hMPV) is a major causative agent of upper-and lower-respiratory-tract infections in infants, the elderly, and immunocompromised individuals worldwide. Like all pneumoviruses, hMPV encodes the zinc binding protein M2-1, which plays important regulatory roles in RNA synthesis. The M2-1 protein is phosphorylated, but the specific role(s) of the phosphorylation in viral replication and pathogenesis remains unknown. In this study, we found that hMPV M2-1 is phosphorylated at amino acid residues S57 and S60. Subsequent mutagenesis found that phosphorylation is not essential for zinc binding activity and oligomerization, whereas inhibition of zinc binding activity abolished the phosphorylation and oligomerization of the M2-1 protein. Using a reverse genetics system, recombinant hMPVs (rhMPVs) lacking either one or both phosphorylation sites in the M2-1 protein were recovered. These recombinant viruses had a significant decrease in both genomic RNA replication and mRNA transcription. In addition, these recombinant viruses were highly attenuated in cell culture and cotton rats. Importantly, rhMPVs lacking phosphorylation in the M2-1 protein triggered high levels of neutralizing antibody and provided complete protection against challenge with wild-type hMPV. Collectively, these data demonstrated that phosphorylation of the M2-1 protein upregulates hMPV RNA synthesis, replication, and pathogenesis in vivo. IMPORTANCEThe pneumoviruses include many important human and animal pathogens, such as human respiratory syncytial virus (hRSV), hMPV, bovine RSV, and avian metapneumovirus (aMPV). Among these viruses, hRSV and hMPV are the leading causes of acute respiratory tract infection in infants and children. Currently, there is no antiviral or vaccine to combat these diseases. All known pneumoviruses encode a zinc binding protein, M2-1, which is a transcriptional antitermination factor. In this work, we found that phosphorylation of M2-1 is essential for virus replication and pathogenesis in vivo. Recombinant hMPVs lacking phosphorylation in M2-1 exhibited limited replication in the upper and lower respiratory tract and triggered strong protective immunity in cotton rats. This work highlights the important role of M2-1 phosphorylation in viral replication and that inhibition of M2-1 phosphorylation may serve as a novel approach to develop live attenuated vaccines as well as antiviral drugs for pneumoviruses. Human metapneumovirus (hMPV) is one of the leading causes of upper and lower respiratory tract infection in humans, particularly in infants, young children, the elderly, and immunocompromised individuals. The virus was first identified in children suffering from respiratory illness in the Netherlands in 2001 (1). Subsequent epidemiological studies suggested that 5 to 15% of all respiratory tract infections in infants and young children were caused by hMPV, a proportion second only to that of human respiratory syncytial virus (hRSV) (2-5). hMPV belongs to the Metapneumovirus genus within the Pneu...

Histopathologic surgical margin assessment in veterinary patients is an imprecise science with assessment limited to a small proportion of the surgical margin due to time and finances. Incomplete excision of canine mast cell tumours (MCTs) alters treatment recommendations and prognosis. Optical coherence tomography (OCT) is a novel imaging modality that has been reported in a single veterinary study for surgical margin assessment. Twenty‐five dogs with 34 MCTs were enrolled in a prospective pilot‐study to assess the imaging characteristics of canine MCTs with OCT and to evaluate the feasibility and utility of OCT‐guided histopathology. All dogs underwent routine surgical excision of MCTs. OCT imaging was used to assess the entire surgical margin prior to placement in formalin. Either normal areas or areas suspected of incomplete MCT excision were inked. Standard histopathologic sectioning and tangential sectioning of inked areas were performed and compared to OCT results. OCT identified MCT near the surgical margin in 10 of 26 specimens (38.4%). Four specimens suspicious for incomplete margins on OCT had incomplete MCT excision that was missed on standard histopathologic sectioning. Six specimens had OCT‐guided sections taken as suspicious, which did not show MCT on histopathology. OCT‐guided pathology sections were able to detect incompletely excised MCT near the surgical margin with a sensitivity of 90% and specificity of 56.2% in this preliminary study. OCT imaging shows promise for guiding pathologists to areas of interest to improve the diagnostic accuracy of surgical margin assessment in excised canine MCTs.