Topology,structures, and energy landscapes of human chromosomes

Chromosome conformation capture experiments provide a rich set of data concerning the spatial organization of the genome. We use these data along with a maximum entropy approach to derive a least-biased effective energy landscape for the chromosome. Simulations of the ensemble of chromosome conformations based on the resulting information theoretic landscape not only accurately reproduce experimental contact probabilities, but also provide a picture of chromosome dynamics and topology. The topology of the simulated chromosomes is probed by computing the distribution of their knot invariants. The simulated chromosome structures are largely free of knots. Topologically associating domains are shown to be crucial for establishing these knotless structures. The simulated chromosome conformations exhibit a tendency to form fibril-like structures like those observed via light microscopy. The topologically associating domains of the interphase chromosome exhibit multistability with varying liquid crystalline ordering that may allow discrete unfolding events and the landscape is locally funneled toward “ideal” chromosome structures that represent hierarchical fibrils of fibrils.The genome’s 3D organization is thought to play a crucial role in many biological processes, including gene regulation, DNA replication, and cell differentiation (1). A theoretical framework for picturing the structure and dynamics of chromosomes thus promises significantly to advance our understanding of cell biology. It is a challenge to develop from first principles an energy landscape theory for chromosomes analogous to that for proteins, which has proved rather successful in providing quantitative insights into protein folding thermodynamics, kinetics, and evolution as well as providing protein structure prediction schemes (2–4). The difficulty arises from several factors, the first being the complexity of the team of molecular players that contribute to chromosome organization. Although one chromosome is made of a single DNA molecule, an array of different proteins is involved in its 3D organization (5); of these only a few have been fully characterized. Even at the shortest length scales, the double-stranded DNA is wrapped by core histone proteins into nucleosomes, which can go on to form higher-order structures such as the proposed 30-nm fibers upon stabilization by histone proteins H1 (6). Long-range contacts between genomic loci can also form in the presence of CCCTC-binding factor (CTCF) (7). The second difficulty is that the chromosome is large in molecular terms. It is so large, in fact, that its dynamics can be so slow that the chromosome may be a far from equilibrium structure, unlike most folded proteins (8, 9). To complicate matters further, chromosome organization has been reported to be variable and to depend both on cell type and phase in the cell cycle (10, 11). There are apparently thus many global attractors on any realistic chromosome landscape. An energy landscape theory built at atomistic resolution using physicochemical interactions of known components for chromosome organization seems thus currently out of reach. Here we explore a way to skirt some of the difficulties by using a maximum entropy approach to derive an effective equivalent equilibrium energy landscape for chromosomes by using physical contact frequencies measured in chromosome conformation capture experiments.     

How do I do it: Continuous local anesthetic infusion for children with spinal dysraphism undergoing major reconstruction of the lower urinary tract

Postoperative pain control is a fundamental aspect of contemporary pediatric surgery. While many options for analgesia are available to the general patient population, choices are limited for individuals with spinal dysraphism who undergo major urologic procedures. Continuous infusion of local anesthetics has been shown to improve postoperative pain scores and decreases the need for systemic analgesia. We present our technique for continuous local anesthetic infusion utilizing readily available equipment with limited additional cost.     

变性高效液相色谱分析肺炎支原体基因类型研究

目的 运用变性高效液相色谱（denaturing high-performance liquid chromatography,DHPLC）技术检测肺炎支原体（mycoplasma pneumoniae,MP）基因型,了解儿童MP流行状况.方法 对我院300例患儿鼻咽吸出物进行表型确证试验,采用PCR对标准菌株和MP表型阳性的临床菌株进行PCR扩增,扩增产物经DHPLC分析,通过与标准菌株色谱峰比对进行临床菌株基因分型.结果 300例患儿咽拭子经过24h培养后呈阳性110例.用特异引物扩增MP-129、MP-FH标准株和标本,分别得到2 280 bp和2580bp基因片段.110株临床菌株经DHPLC分析检测出107株是P1-Ⅰ型,均是1b亚型,3株是P1-Ⅱ型,均是2a亚型.结论 运用DHPLC技术检测出本院儿童发生MP感染以P1-Ⅰ、1b亚型为主.     

矩阵针灸联合电温针治疗骶髂关节致密性骨炎42例

目的:观察矩阵针刺联合电温针治疗骶髂关节致密性骨炎(OCI)的临床疗效。方法:将84例患者随机分为观察组44例、对照组40例,对照组口服双氯芬酸钠缓释胶囊治疗,观察组采用矩阵针刺联合电温针治疗。2组均连续治疗2周,观察2组治疗前后视觉模拟评分(VAS)、日本骨科协会评估治疗分数(JOA评分)的变化情况及临床疗效。结果:VAS及JOA评分治疗前后2组组内比较差异有统计学意义(P0.05);治疗后组间比较差异也有统计学意义(P0.05)。优良率观察组为63.64%,对照组为40.00%,2组比较差异有统计学意义(P0.05)。结论:矩阵针刺联合电温针治疗OCI临床疗效显著,并可改善体征及功能活动。     

花芪灵丹丸质控标准研究

目的建立花芪灵丹丸的质量控制标准。方法采用显微鉴别法鉴别花芪灵丹丸中茯苓、牡丹皮、肉桂;采用薄层色谱法鉴别花芪灵丹丸中黄芪、红花、延胡索;采用高效液相色谱法定量检测花芪灵丹丸中丹参酮ⅡA的含量,建立质量控制标准。结果显微鉴别和薄层色谱鉴别效果好,专属性强;含量测定丹参酮ⅡA在0.01592μg~0.796μg范围内,进样量与峰面积积分值呈良好的线性关系,平均加样回收率为101.5%(RSD值3.8%)。结论花芪灵丹丸的质控方法操作简便、结果准确、重现性好,而且能够对花芪灵丹丸进行定性和定量质控,符合中药制剂的质控要求,可用于花芪灵丹丸质量的控制。     

A possible mechanism of low molecular weight protein tyrosine phosphatase (LMW-PTP) activity modulation by glutathione action during human osteoblast differentiation

Objective

Low molecular weight protein tyrosine phosphatases (LMW-PTPs) are a family of enzymes strongly involved in the regulation of cell growth and differentiation. Since there is no information concerning the relationship between osteoblastic differentiation and LMW-PTP expression/activity, we investigated its involvement during human osteoblast-like cells (hFOB 1.19) differentiation. It is known that LMW-PTP is regulated by an elegant redox mechanism, so we also observed how the osteoblastic differentiation affected the reduced glutathione levels.

Design

hFOB 1.19 cells were cultured in DMEM/F12 up to 35 days. The osteoblast phenotype acquisition was monitored by measuring alkaline phosphatase activity and mineralized nodule formation by Von Kossa staining. LMW-PTP activity and expression were measured using the p-nitrophenylphosphate as substrate and Western blotting respectively. Crystal violet assay determined the cell number in each experimental point. Glutathione level was determined by both HPLC and DNTB assays.

Results

LMW-PTP modulation was coincident with the osteoblastic differentiation biomarkers, such as alkaline phosphatase activity and presence of nodules of mineralization in vitro. Likewise LMW-PTP, the reduced glutathione-dependent microenvironment was modulated during osteoblastic differentiation. During this process, LMW-PTP expression/activity, as well as alkaline phosphatase and glutathione increased progressively up to the 21st day (p < 0.001) of culturing, decreasing thereafter.

Conclusions

Our results clearly suggest that LMW-PTP expression/activity was rigorously modulated during osteoblastic differentiation, possibly in response to the redox status of the cells, since it seems to depend on suitable levels of reduced glutathione. In this way, we pointed out LMW-PTP as an important signaling molecule in osteoblast biology and bone formation.     

大鼠下颌骨来源的成骨细胞对甲硝唑和替硝唑的转运

目的：应用SD大鼠下颔骨来源的原代成骨细胞，观察成骨细胞对甲硝唑和替硝唑的转运，探讨人工种植牙给药的影响及可行性。方法：将原代培养的新生鼠成骨细胞（newborn rat’s osteoblast，N）和成年鼠成骨细胞（adult rat’s osteoblast，A）与溶于PBS的药物共同孵育，分别于1，3，5，10min后，弃去细胞外液，收集各时间点细胞悬液，用高效液相色谱法测定细胞内药物量，用考马斯亮蓝法测定细胞蛋白总量。结果：（1）高效液相色谱法可以准确测定2种药物的量。（2）2种成骨细胞均可将甲硝唑和替硝唑转运至细胞内。结论：成骨细胞具有转运硝基眯唑类药物的能力，证实了人工种植牙给药的可行性。     

Effects of low power red laser on induced-dental caries in rats

OBJECTIVE: The purpose of this study was to investigate the effects of low power red laser associated with acidulated phosphate fluoride on the development of induced-dental caries in rats. DESIGN: Dental caries were induced in molars of 40 rats divided into five groups: control group (CG), the teeth were not submitted to any treatment; laser group (LG), teeth were irradiated with a low power red laser (LPRL), power of 30 mW and dose of 5 J/cm(2); fluoride group (FG), teeth were treated with topical acidulated phosphate fluoride (APF) 1.23% applied for 4 min; laser+fluoride group (LFG), teeth were irradiated with LPRL followed by APF; fluoride+laser group (FLG), teeth were treated with APF followed by LPRL. The animals were killed after 48 days, and the first and second molars were extracted to analyze the caries lesion area, microhardness, and calcium and phosphorus ratio. RESULTS: There were no statistical differences among FG, LFG, and FLG regarding to caries area and microhardness, although the caries area were smaller in LFG. Ca/P ratio did not show significant differences among all groups. CONCLUSIONS: Although LPRL before APF application appeared to diminish the caries progression, LPRL did not present any additional benefit compared with acidulated phosphate fluoride on the prevention of induced-dental caries in rats.