A new transgenic reporter line reveals Wnt-dependent Snai2 re-expression and cranial neural crest differentiation in Xenopus

During vertebrate embryogenesis, the cranial neural crest (CNC) forms at the neural plate border and subsequently migrates and differentiates into many types of cells. The transcription factor Snai2, which is induced by canonical Wnt signaling to be expressed in the early CNC, is pivotal for CNC induction and migration in Xenopus. However, snai2 expression is silenced during CNC migration, and its roles at later developmental stages remain unclear. We generated a transgenic X. tropicalis line that expresses enhanced green fluorescent protein (eGFP) driven by the snai2 promoter/enhancer, and observed eGFP expression not only in the pre-migratory and migrating CNC, but also the differentiating CNC. This transgenic line can be used directly to detect deficiencies in CNC development at various stages, including subtle perturbation of CNC differentiation. In situ hybridization and immunohistochemistry confirm that Snai2 is re-expressed in the differentiating CNC. Using a separate transgenic Wnt reporter line, we show that canonical Wnt signaling is also active in the differentiating CNC. Blocking Wnt signaling shortly after CNC migration causes reduced snai2 expression and impaired differentiation of CNC-derived head cartilage structures. These results suggest that Wnt signaling is required for snai2 re-expression and CNC differentiation

Bovine Lhx8, a Germ Cell-SpecificNuclear Factor, Interacts with Figla

LIM homeobox 8 (Lhx8) is a germ cell-specific transcription factor essential for the development of oocytes during early oogenesis. In mice, Lhx8 deficiency causes postnatal oocyte loss and affects the expression of many oocyte-specific genes. The aims of this study were to characterize the bovine Lhx8 gene, determine its mRNA expression during oocyte development and early embryogenesis, and evaluate its interactions with other oocyte-specific transcription factors. The bovine Lhx8 gene encodes a protein of 377 amino acids. A splice variant of Lhx8 (Lhx8_v1) was also identified. The predicted bovine Lhx8 protein contains two LIM domains and one homeobox domain. However, one of the LIM domains in Lhx8_v1 is incomplete due to deletion of 83 amino acids near the N terminus. Both Lhx8 and Lhx8_v1 transcripts were only detected in the gonads but none of the somatic tissues examined. The expression of Lhx8 and Lhx8_v1 appears to be restricted to oocytes as none of the transcripts was detectable in granulosa or theca cells. The maternal Lhx8 transcript is abundant in GV and MII stage oocytes as well as in early embryos but disappear by morula stage. A nuclear localization signal that is required for the import of Lhx8 into nucleus was identified, and Lhx8 is predominantly localized in the nucleus when ectopically expressed in mammalian cells. Finally, a novel interaction between Lhx8 and Figla, another transcription factor essential for oogenesis, was detected. The results provide new information for studying the mechanisms of action for Lhx8 in oocyte development and early embryogenesis

A new Transgenic Reporter Line Reveals Wnt-dependent Snai2 Re-expression and Cranial Neural Crest Differentiation in Xenopus

During vertebrate embryogenesis, the cranial neural crest (CNC) forms at the neural plate border and subsequently migrates and diferentiates into many types of cells. The transcription factor Snai2, which is induced by canonical Wnt signaling to be expressed in the early CNC, is pivotal for CNC induction and migration in Xenopus. However, snai2 expression is silenced during CNC migration, and its roles at later developmental stages remain unclear. We generated a transgenic X. tropicalis line that expresses enhanced green fuorescent protein (eGFP) driven by the snai2 promoter/enhancer, and observed eGFP expression not only in the pre-migratory and migrating CNC, but also the diferentiating CNC. This transgenic line can be used directly to detect defciencies in CNC development at various stages, including subtle perturbation of CNC diferentiation. In situ hybridization and immunohistochemistry confrm that Snai2 is re-expressed in the diferentiating CNC. Using a separate transgenic Wnt reporter line, we show that canonical Wnt signaling is also active in the diferentiating CNC. Blocking Wnt signaling shortly after CNC migration causes reduced snai2 expression and impaired diferentiation of CNC-derived head cartilage structures. These results suggest that Wnt signaling is required for snai2 reexpression and CNC diferentiation

Preferential infection and depletion of Mycobacterium tuberculosis–specific CD4 T cells after HIV-1 infection

HIV-1 preferentially infects M. tuberculosis-specific CD4+ T cells due to their increased production of IL-2

Canonical Wnt Mechanisms in Neural Crest Induction

Canonical Wnt signaling is a pathway that is critical for normal development and the progression of disease. The canonical Wnt signaling pathway was put together carefully by synthesizing decades of research. Over those decades, canonical Wnt signaling was found to be crucial for nearly all aspects of development, but of importance to this thesis, is a key regulatory factor for the development of the highly migratory multipotent stem cells, the neural crest. As our knowledge of the importance of canonical Wnt signaling grew, research is being conducted to further our understanding of how canonical Wnt signaling communicates with other signaling pathways. Over my Ph.D., we have provided new mechanisms for how canonical Wnt signaling cross-talks with Ephrin and Akt signaling. Here, I discuss how EphrinB signaling inhibits canonical Wnt signaling during neural crest induction and in human cell culture. Within that project, we found that the metalloproteinase ADAM19 protects ADAM13 during neural crest induction to inhibit EphrinB signaling, thus activating canonical Wnt signaling. I also discuss how Akt signaling is necessary for neural crest induction to activate canonical Wnt signaling. Within that project we found that the RNA-helicase DDX3 is required for Rac1 expression during neural crest induction to activate Akt signaling and thus activate canonical Wnt signaling. Finally, I discuss the impact of connecting canonical Wnt signaling with Ephrin and Akt signaling in neural crest, development, disease, and potential new discoveries and research

Health-related quality of life in early rheumatoid arthritis: impact of disease and treatment response

OBJECTIVE: To document the burden of early rheumatoid arthritis (RA) on health-related quality of life (HQL) and compare changes in HQL across 2 treatments. STUDY DESIGN: Analysis of HQL scores among patients enrolled in a multicenter, double-blind, randomized control trial of early RA treatment. PATIENTS AND METHODS: A total of 424 patients with early RA were randomized to 1 of 2 treatment groups: etanercept or methotrexate. Patients were treated and followed for 52 weeks. Health-related quality of life was assessed before and throughout treatment using the Medical Outcomes Study Short Form 36 Health Survey (SF-36) and the Health Assessment Questionnaire (HAQ). The HQL burden of RA was established by comparing SF-36 scale scores to general US population norms. The impact of treatment on HQL was determined by comparing scores on both SF-36 and HAQ scales. RESULTS: Before treatment, RA patients showed significant decrements in scores on all SF-36 scales and summary measures in comparison with age- and sex-matched general US population norms, multivariate analysis of variance (MANOVA) F(8,2815) = 204.6, P \u3c .0001. After 52 weeks of treatment, 7 of 8 SF-36 scales and the physical summary measure remained significantly below the general US population norm, MANOVA F(8,2815) = 41.9, P \u3c .0001. Patients randomized to etanercept showed significantly better HQL improvement earlier in treatment than patients randomized to methotrexate on the SF-36 physical summary, MANOVA F(10,4230) = 6.1, P\u3c .0001, the SF-36 arthritis-specific health index, MANOVA F(10,4230) = 8.5, P \u3c .0001, and the HAQ, MANOVA F(10,4230) = 14.7, P \u3c .0001. At 52 weeks, there were no significant differences between treatment groups. CONCLUSIONS: Rheumatoid arthritis places tremendous disease burden on patients\u27 HQL. Successful treatment of early RA improved HQL. Etanercept showed a rapid HQL response

RT-PCR analysis of bovine <i>Lhx8</i> and <i>Lhx8-v1</i> mRNA expression.

<p><b>A</b>: Expression of <i>Lhx8</i> and <i>Lhx8-v1</i> mRNA in bovine tissues. Tissues tested include spleen, stomach, brain, muscle, kidney, liver, heart, intestine, adult ovary, adult testis, fetal testis and fetal ovary. <b>B</b>: Expression of <i>Lhx8</i> and <i>Lhx8-v1</i> mRNA in bovine fetal ovaries from different gestation stages. Fetal ovaries from 90, 95, 100, 150, 160, 200, 210, 230 and 250 day fetuses were analyzed. The ages of fetuses were estimated based on crown-rump length. <b>C</b>: Expression of <i>Lhx8</i> and <i>Lhx8-v1</i> mRNA in oocytes and early embryos. Oocytes and embryos samples used in the analysis include GV- and MII-stage oocytes and 2-cell, 4-cell, 8-cell, 16-cell, and morula- and blastocyst-stage embryos. Bovine <i>RPL19</i> gene was used as a control for RNA quality.</p

Primers used in this study.

<p>Primers used in this study.</p

Multiple sequence alignment of Lhx8 proteins.

<p>Sequence alignment was performed using Clustal Omega (<a href="http://www.ebi.ac.uk/Tools/msa/clustalo/" target="_blank">http://www.ebi.ac.uk/Tools/msa/clustalo/</a>). The functional domains were determined by searching the Pfam database (<a href="http://pfam.xfam.org/search" target="_blank">http://pfam.xfam.org/search</a>). The LIM and Homeobox domains are indicated by green and red boxes, respectively. mLhx8: mouse Lhx8 (NP_034843.2), hLhx8: human Lhx8 (NP_001001933.1), zLhx8: zebrafish Lhx8 (NP_001003980.1), rtLhx8: rainbow trout Lhx8 (unpublished data). bLhx8: bovine Lhx8 (KX898027).</p