Evaluation of GABA Production and Probiotic Activities of Enterococcus faecium BS5

Probiotics and Antimicrobial Proteins - Gamma-aminobutyric acid (GABA) is a principal inhibitory neurotransmitter in the central nervous system and is produced by irreversible decarboxylation of...     

Monolayer formation of human osteoblastic cells on vertically aligned multiwalled carbon nanotube scaffolds

Monolayer formation of SaOS‐2 (human osteoblast‐like cells) was observed on VACNT (vertically aligned multiwalled carbon nanotubes) scaffolds without purification or functionalization. The VACNT were produced by a microwave plasma chemical vapour deposition on titanium surfaces with nickel or iron as catalyst. Cell viability and morphology studies were evaluated by LDH (lactate dehydrogenase) release assay and SEM (scanning electron microscopy), respectively. The non‐toxicity and the flat spreading with monolayer formation of the SaOs‐2 on VACNT scaffolds surface indicate that they can be used for biomedical applications.     

Targeted Training of Ultrasonic Vocalizations in Aged and Parkinsonian Rats

Voice deficits are a common complication of both Parkinson disease (PD) and aging; they can significantly diminish quality of life by impacting communication abilities. ^{1, 2} Targeted training (speech/voice therapy) can improve specific voice deficits,^{3, 4} although the underlying mechanisms of behavioral interventions are not well understood. Systematic investigation of voice deficits and therapy should consider many factors that are difficult to control in humans, such as age, home environment, age post-onset of disease, severity of disease, and medications. The method presented here uses an animal model of vocalization that allows for systematic study of how underlying sensorimotor mechanisms change with targeted voice training. The ultrasonic recording and analysis procedures outlined in this protocol are applicable to any investigation of rodent ultrasonic vocalizations.The ultrasonic vocalizations of rodents are emerging as a valuable model to investigate the neural substrates of behavior.^5-8 Both rodent and human vocalizations carry semiotic value and are produced by modifying an egressive airflow with a laryngeal constriction.^{9, 10} Thus, rodent vocalizations may be a useful model to study voice deficits in a sensorimotor context. Further, rat models allow us to study the neurobiological underpinnings of recovery from deficits with targeted training.To model PD we use Long-Evans rats (Charles River Laboratories International, Inc.) and induce parkinsonism by a unilateral infusion of 7 μg of 6-hydroxydopamine (6-OHDA) into the medial forebrain bundle which causes moderate to severe degeneration of presynaptic striatal neurons (for details see Ciucci, 2010).^{11, 12} For our aging model we use the Fischer 344/Brown Norway F1 (National Institute on Aging).Our primary method for eliciting vocalizations is to expose sexually-experienced male rats to sexually receptive female rats. When the male becomes interested in the female, the female is removed and the male continues to vocalize. By rewarding complex vocalizations with food or water, both the number of complex vocalizations and the rate of vocalizations can be increased (Figure 1).An ultrasonic microphone mounted above the male''s home cage records the vocalizations. Recording begins after the female rat is removed to isolate the male calls. Vocalizations can be viewed in real time for training or recorded and analyzed offline. By recording and acoustically analyzing vocalizations before and after vocal training, the effects of disease and restoration of normal function with training can be assessed. This model also allows us to relate the observed behavioral (vocal) improvements to changes in the brain and neuromuscular system.     

Immunophenotyping of human HT29 colon cancer cell primary tumours and their metastases in severe combined immunodeficient mice

The immunophenotype of HT29 human colon cancer cells implanted into severe combined immunodeficient mice was assessed in primary tumours and their metastases in the lungs using an indirect immunohistochemical method. After primary tumours were surgically removed, the metastases were given time to develop, thus paralleling the clinical situation. While vimentin was negative in both primary and secondary tumours, E-cadherin was present as membrane-bound labelling in the primary tumours only. Whereas the markers p53, MIB1, PCNA and CEA were consistently positive in both primary and metastatic tumours, CD44 variant 6 and CA125 were negative in metastases but positive in the primary tumours. There was a significant increase in the percentage of cells labelled for p53 in the primary tumours compared with the metastases. For the proliferation markers, there was no significant difference in labelling between primary tumours and metastases for MIB1. Of the cytokeratins examined, CK 20 gave the strongest and most consistent reaction in both primary and secondary tumours. The results indicate that, for certain immunohistochemical markers, results are the same in both primary tumours and metastases. Hence, in these cases, antigens that are expressed on the primary tumour as well as on the metastases can serve as target molecules for immunologically based forms of treatment of metastases. This revised version was published online in November 2006 with corrections to the Cover Date.     

Comparisons of the molecular evolutionary process at rbcL and ndhF in the grass family (Poaceae)

We examine rate heterogeneity among evolutionary lineages of the grass family at two plasmid loci, ndhF and rbcL, and we introduce a method to determine whether patterns of rate heterogeneity are correlated between loci. We show both that rates of synonymous evolution are heterogeneous among grass lineages and that are heterogeneity is correlated between loci at synonymous sites. At nonsynonymous sites, the pattern of rate heterogeneity is not correlated between loci, primarily due to an aberrant pattern of rate heterogeneity at nonsynonymous sites of rbcL. We compare patterns of synonymous rate heterogeneity to predictors based on the generation time effect and the speciation rate hypotheses. Although there is some evidence for generation time effects, neither generation time effects nor speciation rates appear to be sufficient to explain patterns of rate heterogeneity in the grass plastid sequences.      

Fully Automated RNAscope In Situ Hybridization Assays for Formalin‐Fixed Paraffin‐Embedded Cells and Tissues

Biomarkers such as DNA, RNA, and protein are powerful tools in clinical diagnostics and therapeutic development for many diseases. Identifying RNA expression at the single cell level within the morphological context by RNA in situ hybridization provides a great deal of information on gene expression changes over conventional techniques that analyze bulk tissue, yet widespread use of this technique in the clinical setting has been hampered by the dearth of automated RNA ISH assays. Here we present an automated version of the RNA ISH technology RNAscope that is adaptable to multiple automation platforms. The automated RNAscope assay yields a high signal‐to‐noise ratio with little to no background staining and results comparable to the manual assay. In addition, the automated duplex RNAscope assay was able to detect two biomarkers simultaneously. Lastly, assay consistency and reproducibility were confirmed by quantification of TATA‐box binding protein (TBP) mRNA signals across multiple lots and multiple experiments. Taken together, the data presented in this study demonstrate that the automated RNAscope technology is a high performance RNA ISH assay with broad applicability in biomarker research and diagnostic assay development. J. Cell. Biochem. 117: 2201–2208, 2016. © 2016 The Authors. Journal of Cellular Biochemistry Published by Wiley Periodicals, Inc.