TRIM5α Variations Influence Transduction Efficiency With Lentiviral Vectors in Both Human and Rhesus CD34+ Cells In Vitro and In Vivo

Human immunodeficiency virus type 1 (HIV-1) vectors can transduce human hematopoietic stem cells (HSC), but transduction efficiency varies among individuals. The innate immune factor tripartite motif-containing protein 5α (TRIM5α) plays an important role for restriction of retroviral infection. In this study, we examined whether TRIM5α could account for variations in transduction efficiency using both an established rhesus gene therapy model and human CD34⁺ cell culture. Evaluation of TRIM5α genotypes (Mamu-1, -2, -3, -4, -5, and TrimCyp) in 16 rhesus macaques that were transplanted with transduced CD34⁺ cells showed a significant correlation between TRIM5α Mamu-4 and high gene marking in both lymphocytes and granulocytes 6 months after transplantation. Since significant human TRIM5α coding polymorphisms were not known, we evaluated TRIM5α expression levels in human CD34⁺ cells from 14 donors. Three days after HIV-1 vector transduction, measured transduction efficiency varied significantly among donors and was negatively correlated with TRIM5α expression levels. In summary, transduction efficiency in both rhesus and human CD34⁺ cells was influenced by TRIM5α variations (genotypes and expression levels). Our findings are important for both understanding and mitigating the variability of transduction efficiency for rhesus and human CD34⁺ cells.     

A scoring system for the classification of beta-thalassemia/Hb E disease severity

Beta-thalassemia intermediate patients show a remarkable clinical heterogeneity. We examined the phenotypic diversity of 950 beta-thalassemia/Hb E patients in an attempt to construct a system for classifying disease severity. A novel scoring system based on six independent parameters, hemoglobin level, age at disease presentation, age at receiving first blood transfusion, requirement for transfusion, spleen size, and growth and development, was able to separate patients into three distinctive severity categories: mild, moderate, and severe courses. This system, therefore, can increase the accuracy of studies of genotype-phenotype interactions and facilitate decisions for appropriate patient management.     

High affinity of 4-(4-(dimethylamino)styryl)-N-methylpyridinium transport for assessing organic cation drugs in hepatocellular carcinoma cells

Human organic cation transporter 1 (hOCT1) and human organic cation transporter 3 (hOCT3) are highly expressed in hepatocytes and play important roles in cationic drug absorption, distribution, and elimination. A previous study demonstrated that downregulation of hOCT1 and hOCT3 mRNA was related to hepatocellular carcinoma (HepG2) prognosis and severity. Whether these transporters expressed in HepG2 cells serve for cationic drug delivery has not been investigated. Besides radioactive transport, options for assessing hOCTs in hepatocytes are limited. This study clarified the significant roles of hOCTs in HepG2 by comparing cationic fluorescent 4-(4-(dimethylamino)styryl)-N-methylpyridinium (ASP⁺) with traditional [³H]-1-methyl-4-phenylpyridinium (MPP⁺). The results showed ASP⁺ was preferably transported into HepG2 compared to [³H]-MPP⁺ with high affinity and a high maximal transport rate. Selective transport of ASP⁺ mediated by hOCTs was influenced by extracellular pH, temperature, and membrane depolarization, corresponding to hOCT1 and hOCT3 expressions. Furthermore, transport of cationic drugs, metformin, and paclitaxel in HepG2 cells was blunted by OCT inhibitors, suggesting that hOCT1 and hOCT3 expressed in HepG2 cells exhibit notable impacts on cationic drug actions. The fluorescent ASP⁺-based in vitro model may also provide a rapid and powerful analytical tool for further screening of cationic drug actions and interactions with hOCTs, particularly hOCT1 and hOCT3 in hepatocellular carcinoma.     

Comparison of the effectiveness of oral diazepam and midazolam for the sedation of autistic patients during dental treatment

PURPOSE: This study was undertaken to compare the effectiveness of oral diazepam and midazolam in sedating autistic patients during dental treatment. METHODS: The treatment regimen consisted of nitrous oxide/oxygen inhalation in conjunction with oral administration of either diazepam 0.3 mg/kg or midazolam 0.5 mg/kg in a cross-over design study of 13 subjects aged 5.8 to 14.7 years. A drug was classified as being effective when over 70% of the patients taking the drug were judged as "success" in all 3 behavioral criteria: (1) sleeping; (2) body movement; and (3) crying behaviors. The study was observed by an independent clinician with an intraexaminer reliability of 88%. RESULTS: For sleeping behavior, midazolam was found to be significantly more effective than diazepam as the duration of stimulation increased (P<.05). For the movement and crying behaviors, midazolam also proved to be significantly more effective from the start of treatment through the 35- and 40-min markers, respectively (P<.05). For the remainder of treatment, however, there was no statistically significant difference in these behaviors between the trials (P>.05). Diazepam and midazolam were rated as 77% and 100% successful, according to the overall behavior evaluation criteria (P=.02). CONCLUSIONS: Both diazepam and midazolam were shown to be effective sedative agents, successfully and safely used to sedate autistic patients for dental treatment. Midazolam was significantly more effective than diazepam in those portions of the procedure with increased stimulation.     

Synthesis and characterization of the 7-(4-aminomethyl-1H-1,2,3-triazol-1-yl) analogue of kabiramide C

The 7-(4-aminomethyl-1H-1,2,3-triazol-1-yl) analogue of kabiramide C (5) was synthesized by using the Mitsunobu reaction and 1,3-dipolar cycloaddition. This compound and the intermediate compounds 2 and 4 were shown to bind tightly to G-actin in a 1:1 complex and exhibited the same degree of cytotoxicity as 1. Compound 5 serves as a key intermediate for the synthesis of actin-directed optical probes and drugs.     

A comparison of the effects of clomiphene citrate and the aromatase inhibitor letrozole on superovulation in Asian women with normal ovulatory cycles

Objective. To compare the effect of the aromatase inhibitor letrozole and clomiphene citrate (CC) on superovulation in women with normal ovulation.

Methods. A cross-over randomized study of 22 women with normal ovulation, divided randomly into two equal cohorts, was carried out. Each group of 11 women was randomly allocated to take letrozole or CC for one cycle. After washing out for one cycle, the alternative drug was administered in the subsequent cycle. The number and size of mature follicles, endometrial thickness, and estradiol and progesterone levels were monitored.

Results. The number of mature follicles and estradiol levels on ovulation day were significantly lower in the letrozole group than the CC group (p < 0.05 for both). However, no differences between the two groups in endometrial thickness and pattern were observed. Progesterone levels showed ovulation in all cycles.

Conclusions. The administration of 50 mg CC on days 3–5 was superior to 2.5 mg letrozole for superovulation induction in women with normal ovulation.     

Factors related to community participation by stroke victims six month post-stroke

Participation in the community socially by stroke victims is an optimal outcome post-stroke. We carried out a cohort study to evaluate a model for community participation by Thai stroke victims 6 months post-stroke. Six standardized instruments were used to assess the patient's status 1, 3 and 6 months after stroke. These were the modified Rankin Scale, the National Institute of Health Stroke Scale, the Fugl-Meyer Assessment and the Berg Balance Scale. The performance of activities of daily living and community ambulation were measured using the Barthel Index and walking velocity. Participation in the community was measured by the Stroke Impact Scale. The outcomes demographics and stroke related variables were analyzed using the Generalized Estimating Equations. Of the 98 subjects who completed the follow-up assessment, 72 (86.5%) felt they had more participation in the community 6 months post-stroke. The level of disability, performance of independent activities and length of time receiving physical therapy were associated with the perceived level of participation in the community among stroke victims 6 months post-stroke. To achieve a goal of good participation in the community among stroke victims, health care planning should focus on improving the stroke victim's ability to independently perform daily activities. The average length of physical therapy ranged from 1 to 6 months, at 3 to 8 hours/month. Clinical practice guidelines should be explored to optimize participation in the community.     

Interaction of buspirone and its major metabolites with human organic cation transporters

Buspirone, a cationic drug, is an anxiolytic and antidepressant drug. However, whether buspirone and its metabolites are interacted with organic cationic transporter remains uncertain. In this study, we examined the interaction of buspirone and its major metabolites 1-(2-pyrimidinyl)piperazine (1-PP) and 6-hydroxybuspirone (6′-OH-Bu) with hOCTs using human hepatocellular carcinoma (HepG2), human colorectal adenocarcinoma (Caco-2) cells, and S2 cells expressing OCT1 (S2hOCT1), 2 (S2hOCT2), or 3 (S2hOCT3). Coadministration of buspirone and fluorescent 4-(4-(dimethylamino)styryl)-N-methylpyridinium (ASP⁺) was examined using HepG2 cells, and [³H]-1-methyl-4-phenylpyridinium (MPP⁺) transport was assessed in S2 cell overexpressing hOCTs. The results showed that ASP⁺ transport was suppressed by buspirone with an IC₅₀ of 26.3 ± 2.9 μM without any cytotoxic effects in HepG2 expressing hOCTs cells. Consistently, buspirone strongly inhibited [³H]-MPP⁺ uptake by S2hOCT1, S2hOCT2, and S2hOCT3 cells with an IC_50s of 89.0 ± 1.3 μM, 43.7 ± 7.5 μM, and 20.4 ± 1.0 μM, respectively. Nonetheless, 6′-OH-Bu and 1-PP caused weak or no inhibition on ASP⁺ and [³H]-MPP⁺ transport. These findings suggest the potential interaction of buspirone with organic cation drugs that are handled by hOCT3. However, further clinical relevance is needed to support these findings for preventing drug–drug interaction in patients who take prescribed drugs together with buspirone.