Citations

Abstract Human erythropoiesis is a complex multistep process that involves the differentiation of early erythroid progenitors to mature erythrocytes. Here we show that it is feasible to differentiate and mature human embryonic stem cells (hESCs) into functional oxygen-carrying erythrocytes on a large scale (1010-1011 cells/6-well plate hESCs). We also show for the first time that the oxygen equilibrium curves of the hESC-derived cells are comparable with normal red blood cells and respond to changes in pH and 2,3-diphosphoglyerate. Although these cells mainly expressed fetal and embryonic globins, they also possessed the capacity to express the adult β-globin chain on further maturation in vitro. Polymerase chain reaction and globin chain specific immunofluorescent analysis showed that the cells increased expression of β-globin (from 0% to > 16%) after in vitro culture. Importantly, the cells underwent multiple maturation events, including a progressive decrease in size, increase in glycophorin A expression, and chromatin and nuclear condensation. This process resulted in extrusion of the pycnotic nuclei in up to more than 60% of the cells generating red blood cells with a diameter of approximately 6 to 8 μm. The results show that it is feasible to differentiate and mature hESCs into functional oxygen-carrying erythrocytes on a large scale.

ABSTRACT The accumulation of filamentous a‐synuclein (α‐S) is associated with Parkinson's disease. It remains controversial as to the mode (antiparallel or parallel) of α‐S self‐assembly and whether an exact alignment of the central hydrophobic region is essential. In the present study, we performed in vitro assembly using α‐S with or without the attachment of artificial leucine zippers (Zips) capable of forming either parallel or antiparallel coiled coils and included a spacer in one derivative. Results showed that Zips accelerate filament assembly in both the parallel and antiparallel fashions, that a precise alignment of the central hydrophobic region is not essential, and that the antiparallel pairs displayed the highest thioflavin T signals. More importantly, cells expressing Zip‐fused α‐S, but not α‐S alone, formed α‐S immunopositive and thioflavin S‐positive inclusions in 7 days. The results suggest that α‐S can assemble in both parallel and antiparallel modes but have a higher tendency to assemble in the latter mode and that cells overexpressing Zip‐fused α‐S may be used to screen α‐S assembly inhibitors due to enhanced ability to form inclusions.—Jiang, P., Ko, L., Jansen, K. R., Golde, T. E., Yen, S.‐H. Using leucine zipper to facilitate α‐synuclein assembly. FASEB J. 22, 3165–3174 (2008)

Abstract Purpose: MicroRNA (miRNA) is a new class of small, noncoding RNA. The purpose of this study was to determine if miRNAs are differentially expressed in hepatocellular carcinoma (HCC). Experimental Design: More than 200 precursor and mature miRNAs were profiled by real-time PCR in 43 and 28 pairs of HCC and adjacent benign liver, respectively, and in normal liver specimens. Results: Several miRNAs including miR-199a, miR-21, and miR-301 were differentially expressed in the tumor compared with adjacent benign liver. A large number of mature and precursor miRNAs were up-regulated in the adjacent benign liver specimens that were both cirrhotic and hepatitis-positive compared with the uninfected, noncirrhotic specimens (P < 0.01). Interestingly, all of the miRNAs in this comparison had increased expression and none were decreased. The expression of 95 randomly selected mRNAs was not significantly altered in the cirrhotic and hepatitis-positive specimens, suggesting a preferential increase in the transcription of miRNA. Comparing the miRNA expression in the HCC tumors with patient's survival time revealed two groups of patients; those with predominantly lower miRNA expression and poor survival and those with predominantly higher miRNA expression and good survival (P < 0.05). A set of 19 miRNAs significantly correlated with disease outcome. A number of biological processes including cell division, mitosis, and G1-S transition were predicted to be targets of the 19 miRNAs in this group. Conclusion: We show that a global increase in the transcription of miRNA genes occurs in cirrhotic and hepatitis-positive livers and that miRNA expression may prognosticate disease outcome in HCC.

Abstract Using ABQ diagnosis, the sensitivity to detect VF of densitometric versus radiographic assessment in 755 postmenopausal women was 71-81% and specificity was 97%. Misdiagnosis was influenced by image quality and was more common for mild deformities. Introduction: Using densitometric vertebral fracture assessment (VFA), prevalent fractures are identified when vertebral height appears reduced by ≥20%. However, this approach does not discriminate between osteoporotic vertebral fracture (VF) and nonosteoporotic deformity, which increases the false-positive rate. Algorithm-based qualitative diagnosis (ABQ) focuses on vertebral endplate fracture to exclude these deformities but has not been applied in VFA. We wished to determine whether densitometric image quality is adequate for ABQ assessment. Our aims were to (1) calculate agreement between VFA and radiography using ABQ to identify prevalent VF and (2) identify the primary reasons for any discordant diagnosis. Methods: Radiographic and densitometric spine images for postmenopausal women at low risk (LR; n = 459) and high risk (HR; n = 298) of VF were assessed using ABQ. Agreement between imaging modalities for VF diagnosis was assessed by κ statistics using ABQ radiographic readings as the gold standard. Results: The prevalence of VF was 11-29% (radiography) and 9-26% (VFA) in the LR and HR groups, respectively. Agreement between imaging modalities was good or very good (κ = 0.62-0.81 in the LR and HR populations). The sensitivity to detect women with VF by VFA was 71% and 84% in the LR and HR populations, respectively, and specificity was 97%. Fifty-two (77%) and 60 (61%) of vertebrae misclassified by VFA in the LR and HR populations were mild fractures and 37 (54%) and 62 (63%) were wedge fractures. One third of fractures missed by VFA were related to poor or unreadable image quality (n = 27 and 28 vertebrae in the LR and HR populations, respectively). Conclusions: There was good agreement between VFA and radiography using ABQ to identify prevalent VF in women at LR or HR of osteoporotic VF. Vertebrae misclassified by VFA were primarily mild fractures or deformities, and two thirds of all fractures missed by VFA were related to poor or unreadable image quality.

We present an efficient feature selection algorithm for computer aided detection (CAD) computed tomographic (CT) colonography. The algorithm (1) determines an appropriate piecewise linear network (PLN) model by cross validation, (2) applies the orthonormal least square (OLS) procedure to the PLN model utilizing a Modified Schmidt procedure, and (3) uses a floating search algorithm to select features that minimize the output variance. The undesirable "nesting effect" is prevented by the floating search approach, and the piecewise linear OLS procedure makes this algorithm very computationally efficient because the Modified Schmidt procedure only requires one data pass during the whole searching process. The selected features are compared to those obtained by other methods, through cross validation with support vector machines (SVMs).

Abstract The use of histone deacetylase (HDAC) inhibitors has shown promise for a variety of malignancies. In this investigation, we define the activity of this class of inhibitors in combination with traditional cytotoxic chemotherapy in endometrial cancer cells. Significant reductions in growth were observed in Ark2 and KLE endometrial cancer cells following treatment with paclitaxel, doxorubicin, carboplatin, or the HDAC inhibitor trichostatin A (TSA). However, only combined treatment with TSA/paclitaxel caused synergistic inhibition of cell growth. This combination also resulted in significant changes in cell morphology. Using cell cycle analysis, nuclear staining, and Western blot analysis for poly(ADP-ribose) polymerase and caspase-9 degradation products, TSA/paclitaxel showed the most dramatic activation of the apoptotic cascade. These effects were also observed when the HDAC inhibitors HDAC inhibitor-1 or oxamflatin were substituted for TSA. The anticancer properties of paclitaxel are known to result in part from inhibition of microtubule depolymerization, which results in apoptosis. We show that TSA administration also stabilizes microtubules via α-tubulin acetylation. Furthermore, using Western blot and immunohistochemical analysis, treatment with TSA/paclitaxel led to a significant increase in acetylated tubulin and microtubule stabilization. These effects were confirmed in a mouse xenograft model. Moreover, TSA/paclitaxel resulted in a 50% reduction in tumor weight compared with either agent alone. This study provides in vivo evidence of nonhistone protein acetylation as one possible mechanism by which HDAC inhibitors reduce cancer growth. The TSA/paclitaxel combination seems to hold promise for the treatment of serous endometrial carcinoma and other malignancies with limited sensitivity to paclitaxel. [Mol Cancer Ther 2006;5(11):2767–76]

Purpose Palliation of bone pain can be achieved in men with androgen-independent prostate cancer treated with docetaxel and estramustine (DE) or mitoxantrone and prednisone (MP). While Southwest Oncology Group trial 99-16 demonstrated a survival improvement of DE over MP, the study also was designed to compare the palliation of disease-related symptoms. Methods Pain palliation and global quality of life (QOL) were the two primary patient-reported outcomes. Pain was measured with the Present Pain Intensity scale of the McGill Pain Questionnaire-Short Form. The European Organisation for Research and Treatment of Cancer Core Quality of Life Questionnaire C30 (QLQ-C30) and its Prostate Cancer Module (PR25) measured QOL and symptom status. Pain and analgesic use were measured at random assignment, every cycle for eight cycles, and 1 year from random assignment; the QLQ-C30 and the PR25 were administered at random assignment, before cycle four (week 10) and cycle eight (month 6) and at 1 year. In addition to the primary intent-to-treat, missing at random analysis, sensitivity analyses were performed to assess robustness of global QOL conclusions under alternative informative missing data assumptions. Results Six hundred seventy four eligible patients received DE (n = 338) or MP (n = 336). In an intention-to-treat analysis, median overall survival was 17.5 months for the DE arm and 15.6 months for the MP arm (P = .02). There were no statistically significant differences in pain palliation between the treatment arms. The sensitivity analyses showed a consistent lack of statistically significant global QOL differences for the two arms. Conclusion DE had superior clinical efficacy (overall survival, time-to-progression, and prostate-specific antigen declines) with similar global QOL and pain palliation in the MP arm.