We report solution structural studies on 9--16-kilobase (kb) fragments of the 30-nm chromatin fiber isolated from calf thymus nuclei. Samples were stabilized by dimethylsuberimidate cross-linking in 100 mM salt concentration to ensure retention of a compact conformation. Electron microscopy, sedimentation diffusion, light scattering, and gel electrophoresis were used to characterize materials which were fractionated by size by utilizing sucrose gradient sedimentation. Measurements reported include the translational frictional coefficient as determined by quasielastic light scattering and the rotational frictional coefficient as deduced from transient electric dichroism. These frictional properties were combined to yield 33 +/- 3 nm for the diameter of the fiber and a length of 1.5 +/- 0.1 nm per nucleosome. Assuming a superhelix pitch of 11 nm, we calculate 7.5 +/- 0.5 nucleosomes per superhelical turn. The 30-nm fiber was found to reach saturation of electric field orientation at about 10--13 kV/cm and to lack a detectable permanent dipole moment, implying no polarity of the fiber. The limiting reduced dichroism rho was found to be +0.06, intermediate between the values expected if the nucleosomal disk diameters were parallel (rho expected = -3/8) or perpendicular (rho expected +3/4) to the fiber axis. This result implies an average angle of 51 degrees between the fiber axis and the local DNA (nucleosomal) superhelix axis and rules out many of the simple models which have been proposed for the detailed structure of the 30-nm fiber.

Relative to nonreplicating DNA in mature simian virus 40 (SV40) chromosomes, newly synthesized DNA in replicating SV40 chromosomes was found to be hypersensitive to the nonspecific endonucleases, micrococcal nuclease (MNase), DNase I, and DNase II. Nascent DNA, pulse labeled in either intact cells or nuclear extracts supplemented with cytosol, was digested about 5-fold faster and about 25% more extensively than uniformly labeled DNA in mature viral chromosomes. Pulse-chase experiments in vitro revealed a time-dependent chromatin maturation process that involved two distinct steps: (i) conversion of prenucleosomal DNA (PN-DNA) into immature nucleosomal oligomers and (ii) maturation of newly assembled chromatin into a structure with increased nuclease resistance. PN-DNA was hypersensitive to MNase, releasing short DNA fragments which were subsequently solubilized by the nuclease. However, when the nascent PN-DNA was specifically removed by digestion of replicating viral chromosomes with Escherichia coli exonuclease III (3'-5') and phage T7 exonuclease (5'-3'), subsequent digestion of the remaining chromatin with MNase revealed the same degree of hypersensitivity observed prior to exonuclease treatment. Furthermore, newly assembled nucleosomal oligomers, isolated after a brief MNase digestion of replicating viral chromosomes, were also hypersensitive to MNase relative to oligomers isolated from mature chromosomes. Hybridization analysis of the DNA in these immature oligomers revealed that it originated from both sides of replication forks. Inhibition of DNA polymerase alpha by aphidicolin inhibited conversion of PN-DNA into nucleosomes but did not inhibit loss of nucleosomal hypersensitivity to MNase. In contrast, components in the soluble fraction of the subcellular system ("cytosol") were required for both DNA replication and chromatin maturation. Analysis of the nucleoprotein products from a MNase digestion of replicating and mature SV40 chromosomes failed to detect a change in nucleosome structure that corresponded to the loss of nuclease hypersensitivity. However, the results presented demonstrate that both PN-DNA and newly assembled immature chromatin, present on both arms of SV40 replication forks, contribute to the commonly observed hypersensitivity of newly replicated chromatin to endonucleases.

SynopsisWe report electric-dichroism and electron-microscopic studies of chromatin fibers fixed by protein-protein crosslinking at salt concentrations ranging from 10 to 100 mM. The results confirm a progressive disorganization of the fiber as the salt concentration is lowered. The positive dichroism and large polarizability anisotropy characteristic of the 300-A diameter fiber found in 100 mM salt are replaced by negative dichroism and smaller effective polarizability anisotropy or dipole moment for samples fixed at lower salt concentration. We interpret the results in terms of segmental, field-induced orientation of the disorganized structure which is present in low salt concentrations. We also observed a field-induced absorbance decrease in chromatin fibers fixed at salt concentration at and below 100 mM. All three optical effects, namely overall orientation of the high-salt fixed fiber, segmental orientation of the low-salt fixed fiber, and field-induced absorbance decrease, occur on roughly the same time scale, 20-100 ps for 50 nucleosome polynucleosomes. The polarizability anisotropy of fibers fixed in 100 mM salt was found to be proportional to the length of the fragment and to the reciprocal square root of the conductivity of the solution used for electric-dichroism measurements. Addition of Mg2+ to the measurement buffer affected the dichroism amplitude of samples fixed below 100 mM salt but not those fixed at 100 mM salt. The results reinforce the need for caution in interpreting electric-dichroism measurements on chromatin fibers because of possible field-induced distortion effects.

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ObjectiveChronic subdural hematoma drainage is one of the most common procedures performed in neurosurgical practice. Not only burr hole drainage but also small craniotomy (diameter 3–5 cm) is frequently used neurosurgical treatment of chronic subdural hematomas. We assessed to compare the postoperative recurrence rates between burr hole drainage versus small craniotomy with closed-system drainage for chronic subdural hematomas.MethodsFrom January 2016 to December 2018, 75 patients who were treated with burr hole drainage and small craniotomy with closed system drainage for the symptomatic chronic subdural hematoma were enrolled. Pre and postoperative computed tomography (CT) were used for radiologic evaluation. The choice of procedure was decided by preoperative CT images.Results60 patients out of 75 patients underwent burr hole drainage, whereas 15 patients underwent small craniotomy. The overall postoperative recurrence rate was 16%. The recurrence occurred in 8 patients out of 60 patients in burr hole drainage group (13.3%) and 7 patients out of 15 patients in small craniotomy group (46.7%). The number of days of hospitalization was 10.3 days in burr hole drainage group and 15.7 days in small craniotomy group.ConclusionBurr hole drainage would be sufficient to evacuate chronic subdural hematoma with lower recurrence rate, but small craniotomy was also needed in some cases such as hematoma has solid portion or multiple septum.