Glutathione determines chronic myeloid leukemia vulnerability to an inhibitor of CMPK and TMPK.

Bcr-Abl transformation leads to chronic myeloid leukemia (CML). The acquirement of T315I mutation causes tyrosine kinase inhibitors (TKI) resistance. This study develops a compound, JMF4073, inhibiting thymidylate (TMP) and cytidylate (CMP) kinases, aiming for a new therapy against TKI-resistant CML. In vitro and in vivo treatment of JMF4073 eliminates WT-Bcr-Abl-32D CML cells. However, T315I-Bcr-Abl-32D cells are less vulnerable to JMF4073. Evidence is presented that ATF4-mediated upregulation of GSH causes T315I-Bcr-Abl-32D cells to be less sensitive to JMF4073. Reducing GSH biosynthesis generates replication stress in T315I-Bcr-Abl-32D cells that require dTTP/dCTP synthesis for survival, thus enabling JMF4073 susceptibility. It further shows that the levels of ATF4 and GSH in several human CML blast-crisis cell lines are inversely correlated with JMF4073 sensitivity, and the combinatory treatment of JMF4073 with GSH reducing agent leads to synthetic lethality in these CML blast-crisis lines. Altogether, the investigation indicates an alternative option in CML therapy.

DNMT3b protects centromere integrity by restricting R-loop-mediated DNA damage.

This study used DNA methyltransferase 3b (DNMT3b) knockout cells and the functional loss of DNMT3b mutation in immunodeficiency-centromeric instability-facial anomalies syndrome (ICF) cells to understand how DNMT3b dysfunction causes genome instability. We demonstrated that R-loops contribute to DNA damages in DNMT3b knockout and ICF cells. More prominent DNA damage signal in DNMT3b knockout cells was due to the loss of DNMT3b expression and the acquirement of p53 mutation. Genome-wide ChIP-sequencing mapped DNA damage sites at satellite repetitive DNA sequences including (peri-)centromere regions. However, the steady-state levels of (peri-)centromeric R-loops were reduced in DNMT3b knockout and ICF cells. Our analysis indicates that XPG and XPF endonucleases-mediated cleavages remove (peri-)centromeric R-loops to generate DNA beaks, causing chromosome instability. DNMT3b dysfunctions clearly increase R-loops susceptibility to the cleavage process. Finally, we showed that DNA double-strand breaks (DSBs) in centromere are probably repaired by error-prone end-joining pathway in ICF cells. Thus, DNMT3 dysfunctions undermine the integrity of centromere by R-loop-mediated DNA damages and repair.

A Convenient and Sensitive Method for Deoxynucleoside Triphosphate Quantification by the Combination of Rolling Circle Amplification and Quantitative Polymerase Chain Reaction.

Measurement of four dNTP pools is important for investigating metabolism, genome stability, and drug action. In this report, we developed a two-step method for quantitating dNTPs by the combination of rolling circle amplification (RCA) and quantitative polymerase chain reaction (qPCR). We used CircLigase to generate a single-strand DNA in circular monomeric configuration, which was then used for the first step of RCA reaction that contained three dNTPs in excess for quantification of one dNTP at limiting levels. The second step is the amplification of RCA products by qPCR, in which one primer was designed to be completely annealed with the polymeric ssDNA product but not the monomeric template DNA. Using 1 amol of the template in the assay, each dNTP from 0.02 to 2.5 pmol gave a linearity with r2 > 0.99, and the quantification was not affected by the presence of rNTPs. We further found that the preparation of biological samples for the RCA reaction required methanol and chloroform extraction. The method was so sensitive that 1 × 104 cells were sufficient for dNTP quantification with the results similar to those determined by a radio-isotope method using 2 × 105 cells. Thus, the RCA/qPCR method is convenient, cost-effective, and highly sensitive for dNTP quantification.

Systematic review of apraxia treatments to improve occupational performance outcomes.

The objective was to assess effectiveness of apraxia treatments using a systematic review. In contrast to previous reviews, each study was rated as to its applicability to occupational therapy practice and its focus on occupational performance using the FAME rating system (defined by four categories: Feasibility, Appropriateness, Meaningfulness, Effectiveness). This systematic review included eight studies: four randomized controlled trials (level 1 evidence) and four pre-post designs (level 3 evidence). Three treatment approaches were reported: errorless learning with training of details; gesture training; and strategy training. FAME scores ranged from A to C. All studies reported significant treatment effects, but only one demonstrated an impact on observed occupational performance that transferred from clinic to home.

Participation of young adults with high-functioning autism in Taiwan: a pilot study.

This pilot study aimed to investigate the activity participation of young adults with high-functioning autism (HFA) living in Taiwan. Eleven young adults with HFA, their caring adults, and 11 matched typically developing youth were recruited across Taiwan. The Adolescent and Young Adult Participation Sort-Taiwanese version (AYAPS-T) was administered to all three groups to compare the activity participation. In addition, youth with HFA identified activities in which they desired to participate and barriers hindering their participation. The results of this study suggest that youth with HFA had lower participation rates in activities across different domains than their typically developing peers. Youth with HFA were able to identify the activities they desired to do and the barriers hindering their participation. No significant differences in participation were found between the results reported by the caring adults and youth with HFA. Occupational therapy practitioners may work on eliminating the personal and environmental barriers that impede participation as youth with HFA transition out of secondary school.

The development of Adolescent and Young Adult Participation Sort--Taiwanese version.

The purpose of this pilot research was to translate the Adolescent and Young Adult Participation Sort into Taiwanese (AYAPS-T), an assessment tool measuring the activity participation and the self-identified barriers of youth transitioning into adulthood. The study included five phases: translation, cultural adaptation, taking photographs, field testing for content validity and pilot testing of test-retest reliability. A literature review, expert review and translation with back-translation were conducted for the initial activities. The activities were then edited on the basis of the feedback of 23 Taiwanese youth for content validity. Test-retest reliability (intraclass correlation coefficient = 0.91) of the AYAPS-T reported by 22 Taiwanese youth was high. With the activities designed to meet the developmental needs of young adults, the methodology of sorting photographs, the barrier list to identify possible obstacles and some psychometric properties established, the AYAPS-T serves as a reliable and valid tool to identify engagement of young adults in their occupations. Because of the nature of a pilot study, results are limited by a small sample size and limited evidence of psychometric properties. Psychometric properties such as interrater reliability and internal consistency as well as construct validity and concurrent validity need to be tested on a larger sample size.

Regulation of adult human mesenchymal stem cells into osteogenic and chondrogenic lineages by different bioreactor systems.

The aim of this study was to examine the feasibility of expanding and regulating mesenchymal stem cells (MSCs) from isolated adult human bone marrow mononuclear cells, seeded on gelatin-hyaluronic acid biomatrices, and then to quantitatively compare the gene expression in three different culture systems. Individual and interactive effects of model system parameters on construct structure, function, and molecular properties were evaluated. The results showed that these adult human MSCs even at old age not only expressed primitive mesenchymal cell markers but also maintained a high level of colony-forming efficiency and were capable of differentiating into osteoblasts, chondrocytes, and adipocytes upon appropriate inductions. After 21 days of culture, we found that the osteoblastic and chondrocytic lineage gene expression were earlier and higher expressed in spinner flask bioreactor culture group when compared with the static culture and rotating wall vessel reactor culture. The osteogenic lineage proteins type I collagen, alkaline phosphatase, and osteocalcin were strongly stained in histological sections of spinner flask bioreactor culture, whereas these were less detected in the other two groups, especially in rotating wall vessel reactor culture. As for the markers associated with the chondrogenic lineage differentiation proteins, type II collagen was apparently expressed in spinner flask culture group, while the expression of proteoglycans (aggreacan, decorin) in three culture conditions took the lead of each other. We conclude that the spinner flask bioreactor with appropriate induction medium reported in this study may be used to rapidly expand adult MSCs and is likely to possess better induction results toward osteoblastic and chondrocytic lineages.