A Cohort Study of the Milk Microbiota of Healthy and Inflamed Bovine Mammary Glands From Dryoff Through 150 Days in Milk

The objective of this longitudinal cohort study was to describe the milk microbiota of dairy cow mammary glands based on inflammation status before and after the dry period. Individual mammary quarters were assigned to cohorts based on culture results and somatic cell count (SCC) at dryoff and twice in the first 2 weeks post-calving. Mammary glands that were microbiologically negative and had low SCC (< 100,000 cells/mL) at all 3 sampling periods were classified as Healthy (n = 80). Microbiologically negative mammary glands that had SCC ≥150,000 cells/mL at dryoff and the first post-calving sample were classified as Chronic Culture-Negative Inflammation (CHRON; n = 17). Quarters that did not have both culture-negative milk and SCC ≥ 150,000 cells/mL at dryoff but were culture-negative with SCC ≥ 150,000 at both post-calving sampling periods were classified as Culture-Negative New Inflammation (NEWINF; n = 6). Mammary glands with bacterial growth and SCC ≥ 150,000 cells/mL at all 3 periods were classified as Positive (POS; n = 3). Milk samples were collected from all enrolled quarters until 150 days in milk and subjected to microbiota analysis. Milk samples underwent total DNA extraction, a 40-cycle PCR to amplify the V4 region of the bacterial 16S rRNA gene, and next-generation sequencing. Healthy quarters had the lowest rate of PCR and sequencing success (53, 67, 83, and 67% for Healthy, CHRON, NEWINF, and POS, respectively). Chao richness was greatest in milk collected from Healthy quarters and Shannon diversity was greater in milk from Healthy and CHRON quarters than in milk collected from glands in the NEWINF or POS cohorts. Regardless of cohort, season was associated with both richness and diversity, but stage of lactation was not. The most prevalent OTUs included typical gut- and skin-associated bacteria such as those in the phylum Bacteroidetes and the genera Enhydrobacter and Corynebacterium. The increased sequencing success in quarters with worse health outcomes, combined with the lack of bacterial growth in most samples and the high PCR cycle number required for amplification of bacterial DNA, suggests that the milk microbiota of culture-negative, healthy mammary glands is less abundant than that of culture-negative glands with a history of inflammation

High mobility group box 1 (HMGB1) acts as an “alarmin” to promote acute myeloid leukaemia progression

High mobility group box 1 (HMGB1) is a non-histone protein localised in the cell nucleus, where it interacts with DNA and promotes nuclear transcription events. HMGB1 levels are elevated during acute myeloid leukaemia (AML) progression followed by participation of this protein in triggering signalling events in target cells as a pro-inflammatory stimulus. This mechanism was hypothesised to be employed as a survival pathway by malignant blood cells and our aims were therefore to test this hypothesis experimentally. Here we report that HMGB1 triggers the release of tumour necrosis factor alpha (TNF-?) by primary human AML cells. TNF-? induces interleukin 1 beta (IL-1?) production by healthy leukocytes, leading to IL-1?-induced secretion of stem cell factor (SCF) by competent cells (for example endothelial cells). These results were verified in mouse bone marrow and primary human AML blood plasma samples. In addition, HMGB1 was found to induce secretion of angiogenic vascular endothelial growth factor (VEGF) and this process was dependent on the immune receptor Tim-3. We therefore conclude that HMGB1 is critical for AML progression as a ligand of Tim-3 and other immune receptors thus supporting survival/proliferation of AML cells and possibly the process of angiogenesis

Integrating treatment and education for mood disorders: an adolescent case report

This case study illustrates one successful outcome of an intensive, outpatient, treatment project for adolescents with mood disorders. An 18-year-old female with symptoms across several DSM-IV Axis I classifications, including a depressive disorder, and her parents participated in a year-long, multimodal intervention that included mood-focused psychoeducation and coaching designed to impact on her, her family, school, and community systems. Self-report, clinician-driven, and ecologically valid measures were used to assess treatment effects on psychiatric symptoms and psychosocial functioning. Results on the Child and Adolescent Functional Assessment Scale demonstrated considerable gains in the following areas: Home, school/work, social behavior, self-harm, thinking/communication, and substance use. During the intervention, she went from failing several of her classes to graduating from high school. In addition, she made the Honours\u27 List in her first semester at a local community college. A discussion of intervention pluses and pitfalls specific to the case highlight the necessity to influence the various spheres of the young person\u27s life

COLOX: a new blood-based test for colorectal cancer (CRC)screening

BACKGROUND: The objective is to develop a cost-effective, reliable and non invasive screening test able to detect early CRCs and adenomas. This is done on a nucleic acids multigene assay performed on peripheral blood mononuclear cells (PBMCs). METHODS: A colonoscopy-controlled study was conducted on 179 subjects. 92 subjects (21 CRC, 30 adenoma >1 cm and 41 controls) were used as training set to generate a signature. Other 48 subjects kept blinded (controls, CRC and polyps) were used as a test set. To determine organ and disease specificity 38 subjects were used: 24 with inflammatory bowel disease (IBD),14 with other cancers (OC). Blood samples were taken and PBMCs were purified. After the RNA extraction, multiplex RT-qPCR was applied on 92 different candidate biomarkers. After different univariate and multivariate analysis 60 biomarkers with significant p-values (<0.01) were selected. 2 distinct biomarker signatures are used to separate patients without lesion from those with CRC or with adenoma, named COLOX CRC and COLOX POL. COLOX performances were validated using random resampling method, bootstrap. RESULTS: COLOX CRC and POL tests successfully separate patients without lesions from those with CRC (Se 67%, Sp 93%, AUC 0.87), and from those with adenoma > 1cm (Se 63%, Sp 83%, AUC 0.77). 6/24 patients in the IBD group and 1/14 patients in the OC group have a positive COLOX CRC. CONCLUSION: The two COLOX tests demonstrated a high Se and Sp to detect the presence of CRCs and adenomas > 1 cm. A prospective, multicenter, pivotal study is underway in order to confirm these promising results in a larger cohort

Study design.

<p>In a first screening phase performed on the OpenArray system, 670 genes were profiled on 93 samples. Out of these, 163 genes were selected and further tested in phase 2 on additional 51 samples. The final dataset included 144 samples profiled with 163 genes. A 29-gene panel was compiled based on highest power to discriminate AP/CRC from controls by univariate and multivariate analysis. Finally, the 29-gene panel was validated with a LightCycler480 platform, commonly used in clinical laboratories.</p

Validation of the 29-gene panel on the LightCycler480 qPCR platform.

<p>Scatter plots comparing analyses performed for each gene on the datasets generated on the LC480 and OpenArray platforms. The following variables have been used: <b>A.</b> Mean normalized expression values (ΔCp and ΔCt) (R²: 0.933), <b>B.</b> Mean standard deviations (SD) relative to each target gene measured, <b>C.</b> Gene expression fold changes between the CRC and the control group (linear absolute values), <b>D.</b> p-values from statistical testing between the CRC and the control group (log transformed). Lines represent a p-value<0.05. Gene names have been overlapped to the graphs.</p

Functional analysis of the 29-gene panel.

<p>The table reports the most significantly represented biological functions within the gene panel. P-values measure the likelihood that the association between a set of biomarkers and a given Ingenuity Pathway Analysis (IPA) functional category is random. The p-value is calculated using the right-tailed Fisher Exact Test. The number of genes associated with a specific function is reported in the last column.</p><p>Functional analysis of the 29-gene panel.</p

Receiving Operating Characteristics (ROC) analysis.

<p><b>A.</b> Summary of the false and true positive rates of the 29-gene panel in classifying CRC cases. <b>B.</b> Summary of the false and true positive rates of the 29-gene panel in classifying AP cases. Analyses were performed using 500 bootstrap validations. The boxplots represent the distribution of the 500 bootstraps. The black line represents the average values over 500 bootstraps for clinical specificity and sensitivity.</p

Proangiogenic Factor PlGF Programs CD11b+ Myelomonocytes in Breast Cancer during Differentiation of Their Hematopoietic Progenitors.

Tumor-mobilized bone marrow-derived CD11b(+) myeloid cells promote tumor angiogenesis, but how and when these cells acquire proangiogenic properties is not fully elucidated. Here, we show that CD11b(+) myelomonocytic cells develop proangiogenic properties during their differentiation from CD34(+) hematopoietic progenitors and that placenta growth factor (PlGF) is critical in promoting this education. Cultures of human CD34(+) progenitors supplemented with conditioned medium from breast cancer cell lines or PlGF, but not from nontumorigenic breast epithelial lines, generate CD11b(+) cells capable of inducing endothelial cell sprouting in vitro and angiogenesis in vivo. An anti-Flt-1 mAb or soluble Flt-1 abolished the generation of proangiogenic activity during differentiation from progenitor cells. Moreover, inhibition of metalloproteinase activity, but not VEGF, during the endothelial sprouting assay blocked sprouting induced by these proangiogenic CD11b(+) myelomonocytes. In a mouse model of breast cancer, circulating CD11b(+) cells were proangiogenic in the sprouting assays. Silencing of PlGF in tumor cells prevented the generation of proangiogenic activity in circulating CD11b(+) cells, inhibited tumor blood flow, and slowed tumor growth. Peripheral blood of breast cancer patients at diagnosis, but not of healthy individuals, contained elevated levels of PlGF and circulating proangiogenic CD11b(+) myelomonocytes. Taken together, our results show that cancer cells can program proangiogenic activity in CD11b(+) myelomonocytes during differentiation of their progenitor cells in a PlGF-dependent manner. These findings impact breast cancer biology, detection, and treatment. Cancer Res; 71(11); 3781-91. ©2011 AACR