Surgeon, and Institution Characteristics Associated Surgical Preferences in the Pediatric KIDney Stone Care Improvement Network.

OBJECTIVE: To reveal barriers and opportunities to implement evidence for the management of pediatric kidney stone disease, we determined surgeon and institutional factors associated with preferences for the type of surgical intervention for kidney and ureteral stones. METHODS: We conducted a cross-sectional study of urologists participating in the Pediatric KIDney Stone Care Improvement Network (PKIDS) trial. Questionnaires ascertained strengths of urologists' preferences for types of surgery as well as characteristics of participating urologists and institutions. The outcome was the strength of preferences for ureteroscopy, shockwave lithotripsy, and percutaneous nephrolithotomy for four scenarios for which two alternative procedures are recommended by the AUA guidelines: (1) 2 cm kidney stone, (2) 9 mm proximal ureteral stone, (3) 1.5 cm lower pole kidney stone, (4) 1 cm nonlower pole kidney stone. Principal component analysis was performed to identify unique clusters of factors that explain surgical preferences. RESULTS: One hundred forty-eight urologists at 29 sites completed surveys. Stated preferences were highly skewed except for the choice between ureteroscopy and percutaneous nephrolithotomy for a 1.5 cm kidney stone. Shockwave lithotripsy ownership and local practice patterns most frequently associated with the strength of surgeons' preferences for the type of surgery. Principal component analysis revealed that three clusters of stone, patient, and heterogenous characteristics explained 30% of the variance in preferences. CONCLUSION: There is wide variation in the strengths of preferences for surgical interventions supported by current guidelines that are partially explained by surgeon and institutional characteristics. These results reveal opportunities to develop strategies for guidelines that consider real-world drivers of care.

Neural Crest Cells Isolated from the Bone Marrow of Transgenic Mice Express JCV T-Antigen.

JC virus (JCV), a common human polyomavirus, is the etiological agent of the demyelinating disease, progressive multifocal leukoencephalopathy (PML). In addition to its role in PML, studies have demonstrated the transforming ability of the JCV early protein, T-antigen, and its association with some human cancers. JCV infection occurs in childhood and latent virus is thought to be maintained within the bone marrow, which harbors cells of hematopoietic and non-hematopoietic lineages. Here we show that non-hematopoietic mesenchymal stem cells (MSCs) isolated from the bone marrow of JCV T-antigen transgenic mice give rise to JCV T-antigen positive cells when cultured under neural conditions. JCV T-antigen positive cells exhibited neural crest characteristics and demonstrated p75, SOX-10 and nestin positivity. When cultured in conditions typical for mesenchymal cells, a population of T-antigen negative cells, which did not express neural crest markers arose from the MSCs. JCV T-antigen positive cells could be cultured long-term while maintaining their neural crest characteristics. When these cells were induced to differentiate into neural crest derivatives, JCV T-antigen was downregulated in cells differentiating into bone and maintained in glial cells expressing GFAP and S100. We conclude that JCV T-antigen can be stably expressed within a fraction of bone marrow cells differentiating along the neural crest/glial lineage when cultured in vitro. These findings identify a cell population within the bone marrow permissible for JCV early gene expression suggesting the possibility that these cells could support persistent viral infection and thus provide clues toward understanding the role of the bone marrow in JCV latency and reactivation. Further, our data provides an excellent experimental model system for studying the cell-type specificity of JCV T-antigen expression, the role of bone marrow-derived stem cells in the pathogenesis of JCV-related diseases and the opportunities for the use of this model in development of therapeutic strategies.

Bone marrow-derived mesenchymal stem cells undergo JCV T-antigen mediated transformation and generate tumors with neuroectodermal characteristics.

There is now accumulating evidence showing that some tumors may arise from transformed stem cells. In this study we demonstrate that adult bone marrow- derived mesenchymal stem cells (MSCs) undergo neoplastic transformation induced by the human polyomavirus JCV, early protein, T-antigen, and are tumorigenic when transplanted into the flanks of Nude mice as compared to non-transformed MSCs. Histologically, the tumors are heterogeneous with mesenchymal and neural crest characteristics as evidenced by expression of the neural crest markers p75, SOX-10, and S-100, with populations of tumor cells exhibiting characteristics of primitive neuroectodermal cells. In addition, a subset of T-antigen positive tumor cells exhibit a high proliferation index as detected by Ki-67 labeling, and co-express CD133, a marker which is expressed on cancer stem cells. These results show that tumors with neuroectodermal characteristics may arise from transformation of MSCs, a globally accessible adult stem cell with multipotent differentiation capacity. In light of earlier reports on the association of JCV with a broad variety of human tumors, our data suggests that T-antigen transformation of adult stem cells with a multipotent capacity can serve as a possible common origin for some of these cancers, and offers a novel model for oncogenesis.

A population of human brain cells expressing phenotypic markers of more than one lineage can be induced in vitro to differentiate into mesenchymal cells.

Proliferating astrocytic cells from germinal, as well as mature areas of brain parenchyma, have the characteristics of neural stem/progenitor cells and are capable of generating both neurons and glia. We previously reported that primary fetal human brain cells, designated as Normal Human Astrocytes (NHA), expressed, in addition to GFAP, Vimentin and Nestin, low levels of betaIII-Tubulin, an early neuronal marker, and differentiated into neurons and astrocytes in vitro. Here, we showed that primary NHA cells co-express low levels of mesenchymal markers Fibronectin and Collagen-1 in culture. These cells transitioned into mesenchymal-like cells when cultured in adherent conditions in serum containing media. The mesenchymal-like derivatives of these cells were characterized based on their morphological changes, high expression of Vimentin and extracellular matrix (ECM) proteins, Collagen-1 and Fibronectin, and decline of neural markers. When incubated in osteogenic and adipogenic induction media, the mesenchymal-like cells differentiated into osteoblasts and adipocytes. Furthermore, NHA cells express markers of neural crest cells, SOX-10 and p75. These data support the idea of ectoderm-derived mesenchymal lineages. These findings suggest that a population of primitive fetal brain cells with neural/neural crest/mesenchymal phenotype, resembles the remarkable phenotypic plasticity of neural crest cells, and differentiates into adipocytes and osteocytes under the influence of environmental factors.

A population of human brain parenchymal cells express markers of glial, neuronal and early neural cells and differentiate into cells of neuronal and glial lineages.

Glial fibrillary acidic protein (GFAP)-positive cells derived from the neurogenic areas of the brain can be stem/progenitor cells and give rise to new neurons in vitro and in vivo. We report here that a population of GFAP-positive cells derived from fetal human brain parenchyma coexpress markers of early neural and neuronal cells, and have neural progenitor cell characteristics. We used a monolayer culture system to expend and differentiate these cells. During the initial proliferative phase, all cells expressed GFAP, nestin and low levels of betaIII-tubulin. When these cells were cultured in serum and then basic fibroblast growth factor, they generated two distinct progenies: (i) betaIII-tubulin- and nestin-positive cells and (ii) GFAP- and nestin-positive cells. These cells, when subsequently cultured in serum-free media without growth factors, ceased to proliferate and differentiated into two major neural cell classes, neurons and glia. In the cells of neuronal lineage, nestin expression was down-regulated and betaIII-tubulin expression became robust. Cells of glial lineage differentiated by down-regulating nestin expression and up-regulating GFAP expression. These data suggest that populations of parenchymal brain cells, initially expressing both glial and neuronal markers, are capable of differentiating into single neuronal and glial lineages through asymmetric regulation of gene expression in these cells, rather than acquiring markers through differentiation.

Mutational analysis of hSNF5/INI1 and TP53 genes in choroid plexus carcinomas.

We report here the mutational analysis of hSNF5/INI1 and TP53 genes performed on 11 specimens of choroid plexus carcinomas (CPC) in which a large number of abnormalities has been detected by molecular biology techniques. Loss of heterozygosity (LOH) analysis performed on six tumors revealed losses on chromosomes 1, 3, 5, 9, 10, 13, 16, 18, and 22. However, there were no abnormalities on 17p and mutations of the TP53 gene have been observed for two tumors comprising exons 5 and 7, respectively. Exon 4 of hSNF5/INI1 was mutated in one tumor with LOH restricted to the hSNF5/INI1 locus. There was no coexistence of mutations in both analyzed genes. Our analysis confirms the presence of the hSNF5/INI1 mutations and proves involvement of TP53 mutations in sporadic cases of CPC.

Effect of transgene copy number on survival in the G93A SOD1 transgenic mouse model of ALS.

Transgenic mice expressing multiple copies of the G93A mutant form of SOD1 develop motor neuron pathology and clinical symptoms similar to those seen in patients with amyotrophic lateral sclerosis (ALS). The phenotype of these mice is dependent on the number of transgene copies in their genome. Changes in transgene copy number, although rare, can sometimes occur while mating due to intra locus recombination events during meiosis. The objective of this study was to develop a real time quantitative PCR method to determine changes in transgene copy number in these mice and to evaluate the effect of transgene copy number on the phenotype of the G93A SOD1 mouse model of ALS.