Effective methods for visualizing neurovascular morphology are essential for understanding the normal spinal cord and the morphological alterations associated with diseases. However, ideal techniques for simultaneously imaging neurovascular structure in a broad region of a specimen are still lacking. In this study, we combined Golgi staining with angiography and synchrotron radiation micro-computed tomography (SRμCT) to visualize the 3D neurovascular network in the mouse spinal cord. Using our method, the 3D neurons, nerve fibers, and vasculature in a broad region could be visualized in the same image at cellular resolution without destructive sectioning. Besides, we found that the 3D morphology of neurons, nerve fiber tracts, and vasculature visualized by SRμCT were highly consistent with that visualized using the histological method. Moreover, the 3D neurovascular structure could be quantitatively evaluated by the combined methodology. The method shown here will be useful in fundamental neuroscience studies.
Animals ; Imaging, Three-Dimensional ; Mice ; Neural Networks, Computer ; Spinal Cord/diagnostic imaging* ; Synchrotrons ; X-Ray Microtomography

3D histology is a imaging system for the 3D structural information of cells or tissues. The synchrotron radiation propagation phase contrast micro-CT has been used in 3D imaging methods. However, the simple phase contrast micro-CT did not give sufficient micro-structural information when the specimen contains soft elements, as is the case with many biomedical tissue samples. The purpose of this study is to develop a new technique to enhance the phase contrast effect for soft tissue imaging. Experiments were performed at the imaging beam lines of Pohang Accelerator Laboratory (PAL). The biomedical tissue samples under frozen state was mounted on a computer-controlled precision stage and rotated in 0.18° increments through 180°. An X-ray shadow of a specimen was converted into a visual image on the surface of a CdWO4 scintillator that was magnified using a microscopic objective lens (X5 or X20) before being captured with a digital CCD camera. 3-dimensional volume images of the specimen were obtained by applying a filtered back-projection algorithm to the projection images using a software package OCTOPUS. Surface reconstruction and volume segmentation and rendering were performed were performed using Amira software. In this study, We found that synchrotron phase contrast imaging of frozen tissue samples has higher contrast power for soft tissue than that of non-frozen samples. In conclusion, synchrotron radiation propagation phase contrast cryo-microCT imaging offers a promising tool for non-destructive high resolution 3D histology.
Gyeongsangbuk-do ; Octopodiformes ; Synchrotrons*

BACKGROUND: Synchrotron small-angle X-ray scattering (SAXS) is a very useful technique for experimental study of the nano-structure of the nervous system of animals. The study was designed to evaluate nerve preservation methods for the measurement of SAXS patterns. METHODS: Normal sciatic nerves extracted from male Sprague- Dawley rats were preserved in saline (N = 2), formalin (N = 2) or liquid nitrogen (N = 2) for 1 day, followed by measurement of SAXS patterns. SAXS patterns of normal sciatic nerves (N = 3) extracted just before the initiation of the experiment were used as controls. The study was carried out using the 4C1 beamline at Pohang Accelerator Laboratory in Korea. Incoming X-rays were monochromatized at 11 keV using a double multilayer (WB4C) monochromator with beam size of approximately 0.5 (V) × 0.8 (H) mm2. The exposure time was set at 60 sec, and 8 to 12 images per sample were acquired at a 0.5 mm interval. RESULTS: The periodic peaks of interfibrillar space between collagen fibrils were undetectable. The periodic peaks of the myelin sheath and collagen fibers were weakly detected or undetected in the nerves preserved in normal saline or formalin. The periodic peaks and intensity of the myelin sheath, collagen fibers, and interfibrillar space between collagens in the nerves preserved in liquid nitrogen were comparable to those of nerves in the ex vivo state. CONCLUSIONS: The study results indicated that preservation of nerves in liquid nitrogen is adequate for measurements with SAXS. However, saline and formalin preservation techniques were inadequate for SAXS measurement.
Animals ; Collagen ; Formaldehyde ; Gyeongsangbuk-do ; Humans ; Korea ; Male ; Methods* ; Myelin Sheath ; Nervous System ; Nitrogen ; Rats* ; Sciatic Nerve* ; Synchrotrons*

The shape and structure of granules are controlled by the granulation process, which is one of the main factors to determine the nature of the solid dosage forms. In this article, three kinds of granules of a traditional Chinese medicine for improving appetite and promoting digestion, namely, Jianwei Granules, were prepared using granulation technologies as pendular granulation, high speed stirring granulation, and fluidized bed granulation and the powder properties of them were investigated. Meanwhile, synchrotron radiation X-ray computed micro tomography (SR-µCT) was applied to quantitatively determine the irregular internal structures of the granules. The three-dimensional (3D) structure models were obtained by 3D reconstruction, which were more accurately to characterize the three-dimensional structures of the particles through the quantitative data. The models were also used to quantitatively compare the structural differences of granules prepared by different granulation processes with the same formula, so as to characterize how the production process plays a role in the pharmaceutical behaviors of the granules. To focus on the irregularity of the particle structure, the box counting method was used to calculate the fractal dimensions of the granules. The results showed that the fractal dimension is more sensitive to reflect the minor differences in the structure features than the conventional parameters, and capable to specifically distinct granules in structure. It is proved that the fractal dimension could quantitatively characterize the structural information of irregular granules. It is the first time suggested by our research that the fractal dimension difference (Df,c) between two fractal dimension parameters, namely, the volume matrix fractal dimension and the surface matrix fractal dimension, is a new index to characterize granules with irregular structures and evaluate the effects of production processes on the structures of granules as a new indicator for the granulating process control and optimization.
Drugs, Chinese Herbal ; analysis ; Fractals ; Medicine, Chinese Traditional ; Powders ; Quantitative Structure-Activity Relationship ; Synchrotrons ; Technology, Pharmaceutical ; Tomography, X-Ray Computed

OBJECTIVETo investigate the methods for assessing angiogenesis in experimental liver fibrosis.

METHODSMale C57BL/6 mice were randomly divided into a normal (unmodeled) group and a liver fibrosis model group.The model was established by a 4-week course of 10% CCl4 solution (in olive oil) intraperitoneal injection. Liver vasculature was observed by magnetic resonance (MR),computed tomography (CT),synchrotron radiation X-ray,and von Willibrand factor(vWF) immunofluorescence staining. Liver inflammation and fibrosis were observed by staining with hematoxylin-eosin and Sirius red.The t-test and Pearson's correlation analysis were used for statistical analyses.

RESULTSCompared with the normal group,the model group had more robust inflammation and fibrosis in liver tissue.The liver tissue in the modeled mice showed significant deceases in MR signal intensity on T2WI before and after enhancement (386.67+/-69.04 vs.492.67+/-112.50,t =-2.456, P=0.026).The liver tissue in the modeled mice also showed significantly decreased enhancement CT values (P < 0.01).Synchrotron radiation X-ray imaging showed that the small vessels in the liver tissues of the modeled mice were significantly increased compared to that in the normal mice (P < 0.01).The MR T2W enhanced signal value (439.67+/-104.80, Pearson's r=0.714, P=0.009) and microvascular relative length (676.53+/-122.75, Pearson's r=0.791, P=0.002) were positive correlated with microvessel density (MVD) (14.50+/-5.95),as shown by detection of labeled vWF.The before and after CT enhancement difference (132.60+/-57.02, Pearson's r=-0.612, P=0.034) was negatively correlated with MVD.

CONCLUSIONs MR,CT and synchrotron radiation X-ray imaging can be used for assessing angiogenesis in liver fibrosis,and the findings from each are correlated with the traditional MVD detection method. The two-dimensional imaging of synchrotron radiation X-ray is more intuitive,and has been confirmed as an effective evaluation method for liver angiogenesis.

Animals ; Disease Models, Animal ; Inflammation ; pathology ; Liver Cirrhosis ; pathology ; Liver Function Tests ; Magnetic Resonance Spectroscopy ; Male ; Mice ; Mice, Inbred C57BL ; Neovascularization, Pathologic ; Synchrotrons ; Tomography, X-Ray Computed

Bile Ducts, Intrahepatic ; Dilatation, Pathologic ; Humans ; Synchrotrons

BACKGROUND: To understand the fundamentals of neural tissue injury, experiments on the nano-structured nerve system of animals are essential. This study was designed to reveal the nanostructure changes of an isolated ligatured rat sciatic nerve using the synchrotron small-angle X-ray scattering (SAXS) technique. METHODS: Male Sprague-Dawley rats (weighing approximately 250 grams) were used in this study. The SAXS patterns of 1 week after ligatured nerves (N = 5) and the normal sciatic nerves (N = 5) for the control were acquired after extracted approximately 15 mm before the experiment. Experiments were conducted at the 4C1 beam line at the Pohang Accelerator Laboratory in Korea. The exposure time was 60 sec, and 8 to 12 images per sample were acquired in 0.5 mm intervals, including the regions above, around and below the ligatured position. RESULTS: The periodic peaks of the myelin sheath and the interfibrillar space of collagen completely disappeared at the ligatured position. Farther from the ligatured point, weak and quite different SAXS patterns were observed for the myelin sheath and interfibrillar space. However, the collagen fiber peaks appeared at all positions, although they were weaker near the ligatured position. CONCLUSIONS: The ligature treatment totally destroyed the myelin sheath and interfibrillar space of collagen. In addition, retrograde degeneration developed 2 mm above the ligatured site. The myelin sheath and interfibrillar space of collagen were damaged 6 mm below the ligatured site. However, the collagen fiber structure was not significantly affected by the ligature, indicating a much different structural organization.
Animals ; Collagen ; Gyeongsangbuk-do ; Humans ; Korea ; Ligation ; Male ; Myelin Sheath ; Nanostructures ; Rats* ; Rats, Sprague-Dawley ; Retrograde Degeneration ; Sciatic Nerve* ; Synchrotrons*

BACKGROUND: Direct puncture by a needle is a risk factor for nerve damage. This study was designed to demonstrate nerve damage caused by a needle using the synchrotron small-angle X-ray scattering (SAXS) technique. METHODS: A 15 mm section of rat (Male Spargue-Dawley, about 250 grams) sciatic nerves were involved in this study. The nerve specimen for the experiment (N = 5) was punctured 5 times by a needle (25 G, 100 beveled) under general anesthesia with enflurane. The needle was placed perpendicular to the nerve and the needle bevel was placed parallel to the nerve. The SAXS patterns of the punctured nerves, extracted about 15 min prior to the experiment, were acquired after 1 week. The SAXS patterns of a normal sciatic nerve (N = 5), extracted about 15 min prior to the experiment, were measured in order to provide a comparison. Experiments were carried out at 4C1 beamline at Pohang Accelerator Laboratory in Korea. Incoming X-rays were monochromatized at 11 keV using a double multilayer (WB4C) monochromator; the beam size was around 0.5 (V) x 0.8 (H) mm2. The exposure time was 60 sec, and 8 to 12 images were acquired per sample with a 0.5 mm interval. RESULTS: In the punctured group, the periodic peaks of myelin sheath and collagen fiber were not changed. However, the periodic peaks of interfibrillar distance of collagen were greatly changed. CONCLUSIONS: Direct needle-nerve impalement did not cause damages in myelin sheath and collagen fibers when the needle was placed perpendicular and the needle bevel paralleled to the nerve fiber. This result can imply that the needle slipped into the interfibrillar packing of collagen fibrils.
Anesthesia, General ; Animals ; Collagen ; Enflurane ; Gyeongsangbuk-do ; Korea ; Myelin Sheath ; Needles* ; Nerve Fibers ; Peripheral Nerve Injuries* ; Punctures ; Rats* ; Risk Factors ; Sciatic Nerve* ; Synchrotrons*

BACKGROUND: Pulsed radiofrequency (PRF) may be used in the treatment of patients with some pain syndromes that cannot be controlled by alternative techniques. The objective of the present study is to examine the ultrastructural changes in rat sciatic nerve after PRF, using synchrotron small angle X-ray scattering (SAXS). METHODS: Twenty rats (Male Sprague-Dawley, about 250 grams) were used this study. The PRF is applied to the afferent axons of the sciatic nerves of the rats in ex vivo state, and the ultrastructure of axons were studied after 1 (N = 5), 4 (N = 5), and 6 (N = 5) weeks by SAXS. The control (N = 5) consisted of non-treated sciatic nerve to provide a statistical differential comparison. RESULTS: In the PRF group, the periodic peaks of myelin sheath and collagen fibrils were not changed compared to the control group, in the time progression of 1, 4, and 6 weeks. But the periodic peaks of interfibrillar distance of collagen were greater at 1 and 4 weeks after PRF, comparing to the control group, but it had tendency to return to normal in 6 weeks. CONCLUSIONS: It is suggested that PRF did not induce ultrastructural change of myelin sheath and collagen fiber, but it induced the change of distance between collagen fibrils of the nerve tissue. This change was not caused by thermal injury but by electromagnetic fields and it is reversible.
Animals ; Axons ; Collagen ; Electromagnetic Fields ; Humans ; Myelin Sheath ; Nerve Tissue ; Rats* ; Rats, Sprague-Dawley ; Sciatic Nerve* ; Synchrotrons

The crystal form of solid substance had intrinsic correlation with its three dimensional crystal morphology. Based on the characterization of the three dimensional crystal morphology of clopidogrel bisulfate, this research is to establish a model based on the three dimensional morphological parameters. The granular samples composed of polymorphs of clopidogrel bisulfate and microcrystalline cellulose (MCC) were scanned by synchrotron radiation X-ray microscopic CT technology (SR-microCT) and the three dimensional structural models for which were constructed. Seven groups of three dimensional morphological parameters were calculated. Finally, the mathematical model was established with the multi-layer perception (MLP) artificial neutral network methods to identify and predict the polymorphs of clopidogrel bisulfate. The success rate of the model prediction for the polymorphs of clopidogrel bisulfate was 92.7% and the area under the ROC curve was 96.2%. The polymorphs of drugs could be identified and predicted through the numerical description of the three dimensional morphology. The volume, number of the vertices and the surface area were the major determinants for the identification of the polymorphs of clopidogrel bisulfate.
Crystallization ; Neural Networks (Computer) ; Platelet Aggregation Inhibitors ; chemistry ; ROC Curve ; Radiographic Image Interpretation, Computer-Assisted ; Synchrotrons ; Ticlopidine ; analogs & derivatives ; chemistry ; Tomography, X-Ray Computed