Citations

The purpose of this study was to characterize the effect of TIEG1 on the molecular structure of collagen within tail tendon fibers using 3-mo-old female C57BL/6 wild-type (WT) and TIEG1 KO mice. Synchrotron X-ray microdiffraction experiments were carried out on single tendon fibers extracted from the WT and TIEG1 KO dorsal tail tendon. The fibers were scanned in the radial direction, and X-ray patterns were obtained. From these patterns, the meridional direction was analyzed through X-ray intensity profile. In addition, collagen content was investigated using hydroxyproline assays, and qualitative real-time PCR experiments were performed on RNA isolated from fibroblasts to examine specific gene expression changes. The results showed different X-ray diffraction patterns between WT and TIEG1 KO tendon fibers, indicating a disorganization of the collagen structure for the TIEG1 KO compared with WT mice. Furthermore, the analyses of the X-ray intensity profiles exhibited a higher (23 Å) period of collagen for the TIEG1 KO compared with the WT mice. The results of the hydroxyproline assays revealed a significant decrease in the TIEG1 KO compared with WT mice, leading to a decrease in the total amount of collagen present within the TIEG1 KO tendons. Moreover, qualitative real-time PCR results showed differences in the expression profiles of specific genes known to play important roles in tendon fiber development. These data further elucidate the role of TIEG1 on tendon structure and could explain the previous defects in the structure-function relationship found for TIEG1 KO tendon fibers.

Background. A major technical limitation in preclinical cell replacement research is the ability to discriminate between donor and host tissue after transplantation. This problem has been lessened by the availability of transgenic animals that express “reporter” genes, such as green fluorescent protein (GFP). Objective. We determined the usefulness of one such transgenic reporter rat to assess the survival of bone marrow—derived rat mesenchymal stem cells (MSCs) following direct transplantation into the intact adult brain. We also sought to determine if the expression of GFP in the brain affected the survival of the MSCs or the host’s neuroimmune response to the cells. Methods. Rats received intrastriatal injections of sterile transplantation medium, 100 000 normal MSCs, or 100 000 GFP-MSCs and were killed humanely 1, 4, 7, 28, and 42 days posttransplantation for astrocyte and microglial immunohistochemical staining. Results. GFP-MSCs were evident at each examination, although their survival declined over time. Graft volume estimates comparing normal and GFP-MSCs revealed that GFP expression did not adversely affect the survival of the stem cells in the brain. Furthermore, immunostaining for astrocytes and microglia revealed that expression of the reporter protein did not affect the immunogenicity of the stem cells. Conclusions. These data indicate the usefulness of GFP for investigating the survival of MSCs following transplantation to the brain. However, the mechanisms responsible for the poor survival of the stem cells must be elucidated if these cells are to serve cell-based therapies for neurodegenerative disorders.

Daikenchuto (TU-100) is a traditional Japanese (Kampo) medicine used to treat postoperative ileus. TU-100 dose dependently increases gastrointestinal (GI) motility by modulating cholinergic and serotonergic mechanisms in animal studies. The aim of this study was to evaluate the effects of orally administered TU-100 on GI and colonic transit and bowel function in healthy humans. In a randomized, parallel-group, double-blind, placebo-controlled, dose-response study, 60 healthy subjects were randomly assigned to placebo or TU-100 2.5 g or 5 g tid ingested immediately before meals for 5 consecutive days. We measured GI and colonic transit by validated scintigraphy and stool frequency and consistency by daily diaries of bowel function. There were overall treatment effects on colonic filling at 6 h without any significant differences between each dose of TU-100 and placebo. There tended to be overall treatment effects on ascending colon (AC) emptying half-time; the TU-100 (7.5 g/day) treatment significantly accelerated AC emptying compared with placebo. There were numerically higher values of GC24 (which reflect overall colonic transit) with both doses of TU-100, but these changes were not statistically significant. There were no significant overall treatment effects on gastric emptying or stool frequency and consistency. One subject, who received 7.5 g/day of TU-100, had elevated creatine phosphokinase following the study. TU-100 (7.5 g/day) significantly accelerated AC emptying. Further randomized controlled trials in patients with functional constipation or irritable bowel syndrome with constipation are warranted to evaluate the clinical efficacy of TU-100 in these disorders.

Abstract Background: There is strong and consistent evidence that a genetic component contributes to the etiology of chronic lymphocytic leukemia (CLL). A recent genome-wide association study of CLL identified seven genetic variants that increased the risk of CLL within a European population. Methods: We evaluated the association of these variants, or variants in linkage disequilibrium with these variants, with CLL risk in an independent sample of 438 CLL cases and 328 controls. Results: Of these seven single nucleotide polymorphisms (SNP), six had P trend < 0.05 and had estimated odds ratios (OR) that were strikingly comparable to those of the previous study. Associations were seen for rs9378805 [OR, 1.47; 95% confidence intervals (CI), 1.19-1.80; P trend = 0.0003] near IRF4 and rs735665 near GRAMD1B (OR, 1.47; 95% CI, 1.14-1.89; P trend = 0.003). However, no associations (P > 0.05) were found for rs11083846, nor were any found for any SNP in linkage disequilibrium with rs11083846. Conclusions: Our results confirm the previous findings and further support the role of a genetic basis in the etiology of CLL; however, more research is needed to elucidate the causal SNP(s) and the potential manner in which these SNPs or linked SNPs function in CLL pathogenesis. Cancer Epidemiol Biomarkers Prev; 19(4); 1098–102. ©2010 AACR.

Abstract T cell responses are compromised in the elderly. The B7-CD28 family receptors are critical in the regulation of immune responses. We evaluated whether the B7-family and CD28-family receptors were differentially expressed in dendritic cells, macrophages, and CD4+ and CD8+ T cells from young and old mice, which could contribute to the immune dysfunction in the old. Although most of the receptors were equally expressed in all cells, >85% of the old naive CD8+ T cells expressed B7-H1 compared with 25% in the young. Considering that B7-H1 negatively regulates immune responses, we hypothesized that expression of B7-H1 would downregulate the function of old CD8+ T cells. Old CD8+ T cells showed reduced ability to proliferate, but blockade of B7-H1 restored the proliferative capacity of old CD8+ T cells to a level similar to young CD8+ T cells. In vivo blockade of B7-H1 restored antitumor responses against the B7-H1− BM-185–enhanced GFP tumor, such that old animals responded with the same efficiency as young mice. Our data also indicate that old CD8+ T cells express lower levels of TCR compared with young CD8+ T cells. However, following antigenic stimulation in the presence of B7-H1 blockade, the levels of TCR expression were restored in old CD8+ T cells, which correlated with stronger T cell activation. These studies demonstrated that expression of B7-H1 in old CD8+ T cells impairs the proper activation of these cells and that blockade of B7-H1 could be critical to optimally stimulate a CD8 T cell response in the old.

Abstract Tissue injury triggers reparative processes that often involve endothelial progenitor cells (EPCs) recruitment. We hypothesized that atherosclerotic renal artery stenosis (ARAS) activates homing signals that would be detectable in both the kidney and EPCs, and attenuated on renal repair using selective cell-based therapy. Pigs were treated with intrarenal autologous EPC after 6 weeks of ARAS. Four weeks later, expression of homing-related signals in EPC and kidney, single kidney function, microvascular (MV) density, and morphology were compared with untreated ARAS and normal control pigs (n = 7 each). Compared with normal EPC, EPC from ARAS pigs showed increased stromal cell-derived factor (SDF)-1, angiopoietin-1, Tie-2, and c-kit expression, but downregulation of erythropoietin (EPO) and its receptor. The ARAS kidney released the c-kit-ligand stem cell factor, uric acid, and EPO, and upregulated integrin β2, suggesting activation of corresponding homing signaling. However, angiopoietin-1 and SDF-1/CXCR4 were not elevated. Administration of EPC into the stenotic kidney restored angiogenic activity, improved MV density, renal hemodynamics and function, decreased fibrosis and oxidative stress, and attenuated endogenous injury signals. The ARAS kidney releases specific homing signals corresponding to cognate receptors expressed by EPC. EPC show plasticity for organ-specific recruitment strategies, which are upregulated in early atherosclerosis. EPC are renoprotective as they attenuated renal dysfunction and damage in chronic ARAS, and consequently decreased the injury signals. Importantly, manipulation of homing signals may potentially allow therapeutic opportunities to increase endogenous EPC recruitment.

AbstractExperience with new‐born screening (NBS) for disorders of fatty‐acid oxidation (FAOD) is now becoming available from an increasing number of programs worldwide. The spectrum of FAOD differs widely between ethnic groups. Incidence calculations from reports from Australia, Germany, and the USA of a total of 5,256,999 newborns give a combined incidence of all FAOD of approximately 1:9,300. However, it appears to be much lower in Asians. Consequently, a significant prevalence and evidence for a clear benefit of NBS is proven for medium‐chain acyl‐CoA dehydrogenase deficiency (MCAD) only in countries with a high percentage of Caucasians, with very‐long‐chain acyl‐CoA dehydrogenase deficiency (VLCAD) and long‐chain 3‐hydroxy acyl‐CoA dehydrogenase deficiency (LCHAD) being additional candidates. The long‐term benefit for many disorders has still to be evaluated and will require international collaboration, especially for the rarest disorders. Short‐chain acyl‐CoA dehydrogenase deficiency (SCAD) [as well as Systemic carnitine transporter deficiency (CTD) and dienoyl‐CoA reductase deficiency (DE‐RED)] are conditions of uncertain clinical significance, but most FAOD have a spectrum of clinical presentations (healthy–death). Confirmatory diagnostic procedures should be agreed upon to ensure international comparability of results and evidence‐based modifications. The case of short‐chain acyl‐CoA dehydrogenase deficiency (SCAD) deficiency shows that even inclusion of conditions without a clearly known natural course may prove useful with respect to gain of knowledge and consecutive exclusion of a biochemical abnormality without clinical significance, although this line of argument implies the existence of structured follow‐up programs and bears ethical controversies. As a final conclusion, the accumulated evidence suggests all FAOD should to be included into tandem mass spectrometry (MS/MS)‐based NBS programs provided sufficient laboratory performance is guaranteed.

Abstract Epidermal growth factor receptor (EGFR) vIII is a mutated EGFR that is frequently overexpressed in glioblastomas and implicated in response to receptor tyrosine kinase inhibitors. In this study, we investigate the effect of ZD6474 (ZACTIMA, vandetanib), a dual inhibitor for vascular endothelial growth factor receptor 2 and EGFR on growth and angiogenesis of gliomas expressing EGFRvIII. We used two glioma xenograft models, U87MG cells overexpressing EGFRvIII and short-term cultured primary glioma GBM8 cells with EGFRvIII. ZD6474 inhibited tumor growth and angiogenesis and induced cell apoptosis in various brain gliomas. Moreover, significant inhibition of EGFRvIII-expressing U87MG and GBM8 gliomas was observed compared with their controls. Magnetic resonance imaging analysis using the apparent diffusion coefficient and three-dimensional T2*weighed measurements validated ZD6474 inhibition on tumor growth and angiogenesis in EGFRvIII-expressing GBM8 gliomas. Mechanistically, ZD6474 shows better inhibition of cell growth and survival of U87MG/EGFRvIII, GBM6, and GBM8 cells that express EGFRvIII than U87MG or GBM14 cells that have nondetectable EGFRvIII through attenuation of activated phosphorylation of signal transducer and activator of transcription 3, Akt, and Bcl-XL expression. Albeit in lesser extent, ZD6474 also displays suppressions of U87MG/EGFR and GBM12 cells that overexpress wild-type EGFR. Additionally, ZD6474 inhibits activation of extracellular signal-regulated kinase 1/2 in both types of cells, and expression of a constitutively active phosphoinositide 3-kinases partially rescued ZD6474 inhibition in U87MG/EGFRvIII cells. Taken together, these data show that ZD6474 significantly inhibited growth and angiogenesis of gliomas expressing EGFRvIII by specifically blocking EGFRvIII-activated signaling mediators, suggesting a potential application of ZD6474 in treatments for glioblastomas that overexpress EGFRvIII. Mol Cancer Ther; 9(4); 929–41. ©2010 AACR.