In vivo Mn-enhanced MRI for early tumor detection and growth rate analysis in a mouse medulloblastoma model.

Mouse models have increased our understanding of the pathogenesis of medulloblastoma (MB), the most common malignant pediatric brain tumor that often forms in the cerebellum. A major goal of ongoing research is to better understand the early stages of tumorigenesis and to establish the genetic and environmental changes that underlie MB initiation and growth. However, studies of MB progression in mouse models are difficult due to the heterogeneity of tumor onset times and growth patterns and the lack of clinical symptoms at early stages. Magnetic resonance imaging (MRI) is critical for noninvasive, longitudinal, three-dimensional (3D) brain tumor imaging in the clinic but is limited in resolution and sensitivity for imaging early MBs in mice. In this study, high-resolution (100 µm in 2 hours) and high-throughput (150 µm in 15 minutes) manganese-enhanced MRI (MEMRI) protocols were optimized for early detection and monitoring of MBs in a Patched-1 (Ptch1) conditional knockout (CKO) model. The high tissue contrast obtained with MEMRI revealed detailed cerebellar morphology and enabled detection of MBs over a wide range of stages including pretumoral lesions as early as 2 to 3 weeks postnatal with volumes close to 0.1 mm(3). Furthermore, longitudinal MEMRI allowed noninvasive monitoring of tumors and demonstrated that lesions within and between individuals have different tumorigenic potentials. 3D volumetric studies allowed quantitative analysis of MB tumor morphology and growth rates in individual Ptch1-CKO mice. These results show that MEMRI provides a powerful method for early in vivo detection and longitudinal imaging of MB progression in the mouse brain.

Expression and purification of the membrane enzyme selenoprotein K.

Selenoprotein K (SelK) is a membrane protein residing in the endoplasmic reticulum. The function of SelK is mostly unknown; however, it has been shown to participate in anti-oxidant defense, calcium regulation and in the endoplasmic reticulum associated protein degradation (ERAD) pathway. In order to study the function of SelK and the role of selenocysteine in catalysis, we have tested heterologous expression of human SelK in E. coli. Consequently, we have developed an over-expression strategy that exploits the maltose binding protein as a fusion partner to stabilize and solubilize SelK. The fusion partner can be cleaved from SelK in the presence of a variety of detergents compatible with structural characterization and the protein purified to homogeneity. SelK acquires a helical secondary structure in detergent micelles, even though it was predicted to be an intrinsically disordered protein due to its high percentage of polar residues. The same strategy was successfully applied to preparation of SelK binding partner - selenoprotein S (SelS). Hence, this heterologous expression and purification strategy can be applied to other members of the membrane enzyme family to which SelK belongs.

Hedgehog inhibition as an anti-cancer strategy.

Dysregulated Hedgehog (Hh) signaling has been implicated in a growing number of human cancers. Although first identified as an important developmental signaling pathway crucial for cellular proliferation, differentiation, and migration during organogenesis in invertebrates, these fundamental processes have been co-opted in human cancers. Initial evidence for the Hh pathway in tumor biology comes from mutations of signaling pathway components in a hereditary cancer syndrome that typically results in basal-cell carcinoma and medulloblastoma. Subsequent analysis revealed that Hh pathway mutations are found in sporadic tumors as well as activated Hh signaling in several epithelial cancers independent of Hh pathway mutation status. Further, recent evidence has demonstrated paracrine Hh signaling within stromal cells of the tumor microenvironment with implications for drug delivery. Several Hh antagonists targeting the Hh receptor, Smoothened (SMO), have been developed and show efficacy in preclinical studies and early-stage clinical trials in humans. However, major issues with these small molecule compounds include rapid acquired resistance, potential developmental toxicities secondary to use in children, and limited efficacy in cancers driven by Hh signaling downstream of the SMO receptor.

Evaluation of changes in trabecular alveolar bone during growth using conventional panoramic radiographs.

OBJECTIVE: To assess changes of the alveolar trabecular bone during growth using panoramic radiographs and to detect possible differences in trabecular bone patterns when comparing individuals of various ages and genders. MATERIALS AND METHODS: Conventional panoramic radiographs of 18 young (eight females, 10 males) and 21 adult (12 females, nine males) subjects were taken at 2 years (T1) and 10 years (T2) after the end of orthodontic treatment. At T1, mean ages were 15.6 ± 0.9 years and 31.3 ± 9.7 years in the young and the adult groups, respectively. A three-scale visual analysis was used to evaluate bilaterally the alveolar bone trabeculation in the interdental spaces, from the distal side of the first mandibular premolar to the mesial side of the second lower molar. An analysis of variance (ANOVA), associated with t-tests whenever significance was found, was used to appraise the role of the age, the extent of the follow-up period and the gender on trabecular bone structure. RESULTS: The adult group had a denser alveolar bone trabeculation, compared to the young group. This was also observed in the 8 years follow-up recordings among the adults, but no statistically significant differences were found in the growing individuals. No gender discrepancy was detected. CONCLUSIONS: From puberty to the middle age adulthood, denser alveolar bone trabeculation in the mandible seems to be related to the age. No differences were found between male and female subjects in the sample.

Assessment of trabecular pattern on periapical and panoramic radiographs: a pilot study.

OBJECTIVE: This methodological study aimed to determine whether the mandibular trabecular bone assessment from panoramic radiographs, using a visual index, corresponds to the evaluation obtained from periapical radiographs. MATERIAL AND METHODS: A panoramic radiograph and corresponding periapical radiographs of the region of the lower premolars and molars were collected from each of 32 patients (mean age 18.5 +/- 5.5 years). Two calibrated observers assessed randomly the interdental sites between the first molar and second premolar and between the two premolars on all the radiographs using a visual index. Evaluations were repeated with an interval of 60 days. The results of the repeated evaluations were used to assess intra- and inter-observer agreements, employing Kappa statistics. Spearman's correlation was used to determine the association between assessments of panoramic and periapical radiographs. RESULTS: In total, 79 interdental sites were evaluated on the panoramic and periapical radiographs. The visual analysis of periapical radiographs revealed intra-observer agreements of 0.88 for observer 1 and 0.93 for observer 2, and an inter-observer agreement of 0.82. The intra-observer agreement for panoramic radiographs was 0.79 and 0.83 for observers 1 and 2, respectively, and the inter-observer agreement was 0.79. A substantial correlation was found between periapical and panoramic radiographs (rho = 0.737, p = 0.001). CONCLUSIONS: Although panoramic radiographs are less reliable than periapical radiographs, they can be used for assessment of the trabecular bone pattern with the aid of a visual index. Training on the method is recommended to obtain results with a high reproducibility.

Presentation of osteitis and osteomyelitis pubis as acute abdominal pain.

Osteitis pubis is the most common inflammatory condition of the pubic symphysis and may present as acute abdominal, pelvic, or groin pain. Osteomyelitis pubis can occur concurrently and spontaneously with osteitis pubis. Primary care physicians should consider these conditions in patients presenting with abdominal and pelvic pain. A thorough history, including type of physical activity, and a focused physical examination will be useful, and imaging modalities may be helpful. A biopsy and culture of the pubic symphysis will usually confirm the diagnosis. Treatment for osteitis pubis generally involves rest and anti-inflammatory medications. Failure with this conservative treatment should alert the physician to the possibility of osteomyelitis, which needs treatment with antibiotics. Prognosis for recovery is excellent with definitive diagnosis and treatment.

Development of the deep cerebellar nuclei: transcription factors and cell migration from the rhombic lip.

The deep cerebellar nuclei (DCN) are the main output centers of the cerebellum, but little is known about their development. Using transcription factors as cell type-specific markers, we found that DCN neurons in mice are produced in the rhombic lip and migrate rostrally in a subpial stream to the nuclear transitory zone (NTZ). The rhombic lip-derived cells express transcription factors Pax6, Tbr2, and Tbr1 sequentially as they enter the NTZ. A subset of rhombic lip-derived cells also express reelin, a key regulator of Purkinje cell migrations. In organotypic slice cultures, the rhombic lip was necessary and sufficient to produce cells that migrate in the subpial stream, enter the NTZ, and express Pax6, Tbr2, Tbr1, and reelin. In later stages of development, the subpial stream is replaced by the external granular layer, and the NTZ organizes into distinct DCN nuclei. Tbr1 expression persists to adulthood in a subset of medial DCN projection neurons. In reeler mutant mice, which have a severe cerebellar malformation, rhombic lip-derived cells migrated to the NTZ, despite reelin deficiency. Studies in Tbr1 mutant mice suggested that Tbr1 plays a role in DCN morphogenesis but is not required for reelin expression, glutamatergic differentiation, or the initial formation of efferent axon pathways. Our findings reveal underlying similarities in the transcriptional programs for glutamatergic neuron production in the DCN and the cerebral cortex, and they support a model of cerebellar neurogenesis in which glutamatergic and GABAergic neurons are produced from separate progenitor compartments.

Pax6, Tbr2, and Tbr1 are expressed sequentially by radial glia, intermediate progenitor cells, and postmitotic neurons in developing neocortex.

The developing neocortex contains two types of progenitor cells for glutamatergic, pyramidal-projection neurons. The first type, radial glia, produce neurons and glia, divide at the ventricular surface, and express Pax6, a homeodomain transcription factor. The second type, intermediate progenitor cells, are derived from radial glia, produce only neurons, and divide away from the ventricular surface. Here we show that the transition from radial glia to intermediate progenitor cell is associated with upregulation of Tbr2, a T-domain transcription factor, and downregulation of Pax6. Accordingly, Tbr2 expression in progenitor compartments (the subventricular zone and ventricular zone) rises and falls with cortical plate neurogenesis. The subsequent transition from intermediate progenitor cell to postmitotic neuron is marked by downregulation of Tbr2 and upregulation of Tbr1, another T-domain transcription factor. These findings delineate the transcription factor sequence Pax6 --> Tbr2 --> Tbr1 in the differentiation of radial glia --> intermediate progenitor cell --> postmitotic projection neuron. This transcription factor sequence is modified in preplate neurons, in which Tbr2 is transiently coexpressed with Tbr1, and in the direct differentiation pathway from radial glia --> postmitotic projection neuron, in which Tbr2 is expressed briefly or not at all.