Dataset Information

Improved Determination of Subnuclear Position Enabled by Three-Dimensional Membrane Reconstruction.

ABSTRACT: Many aspects of chromatin biology are influenced by the nuclear compartment in which a locus resides, from transcriptional regulation to DNA repair. Further, the dynamic and variable localization of a particular locus across cell populations and over time makes analysis of a large number of cells critical. As a consequence, robust and automatable methods to measure the position of individual loci within the nuclear volume in populations of cells are necessary to support quantitative analysis of nuclear position. Here, we describe a three-dimensional membrane reconstruction approach that uses fluorescently tagged nuclear envelope or endoplasmic reticulum membrane marker proteins to precisely map the nuclear volume. This approach is robust to a variety of nuclear shapes, providing greater biological accuracy than alternative methods that enforce nuclear circularity, while also describing nuclear position in all three dimensions. By combining this method with established approaches to reconstruct the position of diffraction-limited chromatin markers-in this case, lac Operator arrays bound by lacI-GFP-the distribution of loci positions within the nuclear volume with respect to the nuclear periphery can be quantitatively obtained. This stand-alone image analysis pipeline should be of broad practical utility for individuals interested in various aspects of chromatin biology, while also providing, to our knowledge, a new conceptual framework for investigators who study organelle shape.

SUBMITTER: Zhao Y

PROVIDER: S-EPMC4945324 | biostudies-literature | 2016 Jul

REPOSITORIES: biostudies-literature

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Publications

Improved Determination of Subnuclear Position Enabled by Three-Dimensional Membrane Reconstruction.

Zhao Yao Y Schreiner Sarah M SM Koo Peter K PK Colombi Paolo P King Megan C MC Mochrie Simon G J SG

Biophysical journal 20160701 1

Many aspects of chromatin biology are influenced by the nuclear compartment in which a locus resides, from transcriptional regulation to DNA repair. Further, the dynamic and variable localization of a particular locus across cell populations and over time makes analysis of a large number of cells critical. As a consequence, robust and automatable methods to measure the position of individual loci within the nuclear volume in populations of cells are necessary to support quantitative analysis of ...[more]

PMID: 27410730

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Project description:Background/objectivesIt is important to restore the tibiofemoral relationship as well as the anterior knee laxity for more successful anterior cruciate ligament (ACL) reconstruction, since a residual abnormality in the tibiofemoral relationship would lead an abnormal stress on the articular cartilages/menisci and consequently increase the risk of osteoarthritis in the future. This study aimed to sequentially clarify the three-dimensional tibiofemoral relationship before and after anatomic anterior cruciate ligament (ACL) reconstruction under an anterior tibial load with a gravity-assisted radiographic technique in the prone position.MethodsFifteen patients with unilateral ACL injury participated in the study. Anatomic triple-bundle ACL reconstruction was performed using semitendinosus tendon autografts. During the computed tomography scans that were performed preoperatively, and those performed at 3 weeks and at 6 months postoperatively, the patients lay in the prone position with the knee flexed at 15°, wherein the calf weight could exert an anterior drawer force on the tibia due to gravity. Three-dimensional the tibial position relative to the femur were evaluated for each time point, followed by calculation of side-to-side differences in the parameters between the ACL-deficient/ACL-reconstructed knees and the contralateral intact knees. Seven healthy volunteers were enrolled in the control group and the side-to-side differences (right minus left) in these parameters were calculated.ResultsThe tibia in the ACL-deficient knee was located anteriorly by 3.5 ± 1.1 mm and rotated internally by 2.4° ± 2.3°; these values were significantly larger than the corresponding values of -0.2 ± 1.5 mm and 0.1° ± 2.2° in the control group. However, at 3 weeks postoperatively, the tibia in the ACL-reconstructed knee was over-constrained as compared to that in the control group; it was located posteriorly by 2.5 ± 1.4 mm and rotated externally by 3.4° ± 3.4°. At 6 months postoperatively, no significant difference was observed in the tibial displacements/rotations between the patient and control groups. The side-to-side difference in the anterior knee laxity at the manual maximum anterior load was 0.1 ± 1.2 mm at 6 months postoperatively, with a significant improvement over the preoperative value of 7.4 ± 2.5 mm.ConclusionsAnatomic ACL reconstruction could restore not only the normal anterior knee laxity, but also the normal tibiofemoral relationship even under an anterior tibial load.

Dataset Information

Improved Determination of Subnuclear Position Enabled by Three-Dimensional Membrane Reconstruction.

Publications

Improved Determination of Subnuclear Position Enabled by Three-Dimensional Membrane Reconstruction.

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