Compact laboratory-based X-ray microscope enabling nondestructive 3D structure acquisition of mouse nephron with high speed and better user accessibility.

X-ray microscopes adopting computed tomography enable nondestructive 3D visualization of biological specimens at micron-level resolution without conventional 2D serial sectioning that is a destructive/laborious method and is routinely used for analyzing renal biopsy in clinical diagnosis of kidney diseases. Here we applied a compact commercial system of laboratory-based X-ray microscope to observe a resin-embedded osmium-stained 1-mm strip of a mouse kidney piece as a model of renal biopsy, toward a more efficient diagnosis of kidney diseases. A reconstructed computed tomography image from several hours of data collection using CCD detector allowed us to unambiguously segment a single nephron connected to a renal corpuscle, which was consistent with previous reports using serial sectioning. Histogram analysis on the segmented nephron confirmed that the proximal and distal tubules were distinguishable on the basis of their X-ray opacities. A 3D rendering model of the segmented nephron visualized a convoluted structure of renal tubules neighboring the renal corpuscle and a branched structure of efferent arterioles. Furthermore, another data collection using scientific complementary metal-oxide semiconductor detector with a much shorter data acquisition time of 15 min provided similar results from the same samples. These results suggest a potential application of the compact laboratory-based X-ray microscope to analyze mouse renal biopsy.

Nondestructive cellular-level 3D observation of mouse kidney using laboratory-based X-ray microscopy with paraffin-mediated contrast enhancement.

For three-dimensional observation of unstained bio-specimens using X-ray microscopy with computed tomography (CT), one main problem has been low contrast in X-ray absorption. Here we introduce paraffin-mediated contrast enhancement to visualize biopsy samples of mouse kidney using a laboratory-based X-tray microscope. Unlike conventional heavy-atom staining, paraffin-mediated contrast enhancement uses solid paraffin as a negative contrast medium to replace water in the sample. The medium replacement from water to paraffin effectively lowers the absorption of low-energy X-rays by the medium, which eventually enhances the absorption contrast between the medium and tissue. In this work, paraffin-mediated contrast enhancement with 8 keV laboratory X-rays was used to visualize cylindrical renal biopsies with diameters of about 0.5 mm. As a result, reconstructed CT images from 19.4 h of data collection achieved cellular-level resolutions in all directions, which provided 3D structures of renal corpuscles from a normal mouse and from a disease model mouse. These two structures with and without disease allowed a volumetric analysis showing substantial volume differences in glomerular subregions. Notably, this nondestructive method presents CT opacities reflecting elemental composition and density of unstained tissues, thereby allowing more unbiased interpretation on their biological structures.

Real-space modeling for complex structures based on small-angle X-ray scattering.

A three-dimensional real-space model has been created for hierarchical materials by matching observed and simulated small-angle X-ray scattering patterns. The simulation is performed by arranging the positions of small primary particles and constructing an aggregate structure in a finite-sized cell. In order to avoid the effect of the finite size of the cell, the cell size is extended to infinity by introducing an asymptotic form of the long-range correlations among the primary particles. As a result, simulations for small-angle X-ray scattering patterns can be performed correctly in the low-wavenumber regime (<0.1 nm-1), allowing the model to handle hundred-nanometre-scale structures composed of primary particles of a few nanometres in size. An aerogel structure was determined using this model, resulting in an excellent match with the experimental scattering pattern. The resultant three-dimensional model can generate cross-sectional images similar to those obtained by transmission electron microscopy, and the calculated pore-size distribution is in accord with that derived from the gas adsorption method.

Complete Ring Artifacts Reduction Procedure for Lab-Based X-ray Nano CT Systems.

In this article, we introduce a new ring artifacts reduction procedure that combines several ideas from existing methods into one complex and robust approach with a goal to overcome their individual weaknesses and limitations. The procedure differentiates two types of ring artifacts according to their cause and character in computed tomography (CT) data. Each type is then addressed separately in the sinogram domain. The novel iterative schemes based on relative total variations (RTV) were integrated to detect the artifacts. The correction process uses the image inpainting, and the intensity deviations smoothing method. The procedure was implemented in scope of lab-based X-ray nano CT with detection systems based on charge-coupled device (CCD) and scientific complementary metal-oxide-semiconductor (sCMOS) technologies. The procedure was then further tested and optimized on the simulated data and the real CT data of selected samples with different compositions. The performance of the procedure was quantitatively evaluated in terms of the artifacts' detection accuracy, the comparison with existing methods, and the ability to preserve spatial resolution. The results show a high efficiency of ring removal and the preservation of the original sample's structure.

Visualization of internal 3D structure of small live seed on germination by laboratory-based X-ray microscopy with phase contrast computed tomography.

BACKGROUND: The visualization of internal 3D-structure of tissues at micron resolutions without staining by contrast reagents is desirable in plant researches, and it can be achieved by an X-ray computed tomography (CT) with a phase-retrieval technique. Recently, a laboratory-based X-ray microscope adopting the phase contrast CT was developed as a powerful tool for the observation of weakly absorbing biological samples. Here we report the observation of unstained pansy seeds using the laboratory-based X-ray phase-contrast CT. RESULTS: A live pansy seed within 2 mm in size was simply mounted inside a plastic tube and irradiated by in-house X-rays to collect projection images using a laboratory-based X-ray microscope. The phase-retrieval technique was applied to enhance contrasts in the projection images. In addition to a dry seed, wet seeds on germination with the poorer contrasts were tried. The phase-retrieved tomograms from both the dry and the wet seeds revealed a cellular level of spatial resolutions that were enough to resolve cells in the seeds, and provided enough contrasts to delineate the boundary of embryos manually. The manual segmentation allowed a 3D rendering of embryos at three different stages in the germination, which visualized an overall morphological change of the embryo upon germination as well as a spatial arrangement of cells inside the embryo. CONCLUSIONS: Our results confirmed an availability of the laboratory-based X-ray phase-contrast CT for a 3D-structural study on the development of small seeds. The present method may provide a unique way to observe live plant tissues at micron resolutions without structural perturbations due to the sample preparation.

[Long-Term Survival Due to Combination Therapy in a Patient with Locally Advanced Body-Tail Pancreatic Cancer].

A 49-year-old man was referred to a neighboring hospital with a chief complaint of abdominal pain.He was diagnosed with locally advanced body-tail pancreatic cancer that had invaded the celiac artery and SMA.He came to our department after undergoing radiotherapy, 2.5 Gy×22 Fr, and chemotherapy with gemcitabine(GEM)and S-1.The same chemotherapy was continued for 15 months until DIC occurred.He was subsequently treated with GEM only and then S-1 only in sequence for 6 years.We decided to stop the chemotherapy because the original lesion had been stable for a long time.After 1 month, a hard nodule appeared in the subcutaneous layer of the navel.Although resection was performed and he received chemotherapy, he died after surviving a total of 7 years and 10 months.This case is important when considering whether to discontinue chemotherapy with a stable long-term pancreatic cancer patient.

Hard x-ray phase contrast imaging using a tabletop Talbot-Lau interferometer with multiline embedded x-ray targets.

We demonstrate hard x-ray phase contrast imaging (XPCI) using a tabletop Talbot-Lau interferometer in which the x-ray source and source grating are replaced with an x-ray source with multiline metal targets embedded in a diamond substrate. This source realizes an array of linear x-ray sources of a few micrometers width without fabrication difficulty because of the shallow penetration depth of electrons irradiated to the metal targets. This enhances the coherence of x rays from each linear source and allows XPCI within 45 cm source-detector distance under 1.2 W input power for 8 keV x rays.

[A case of port site recurrence after thoracoscopic resection for lung metastasis of cecal cancer].

We experienced a case of port site recurrence after thoracoscopic resection for lung metastasis of cecal cancer. The patient was a 57-year-old woman who underwent right hemicolectomy at the age of 51 following a diagnosis of cecal carcinoma. She underwent video-assisted thoracic surgery for pulmonary metastasis 2 years after the first surgery. She underwent local resection for a retroperitoneal pelvic wall recurrence 3.5 years after the first surgery. Chest wall port site recurrence occurred 5.5 years after the first surgery, and she underwent partial resection of the left lung and chest wall. Subsequent treatment has been performed with adjuvant chemotherapy, and she is healthy with no evidence of recurrence 7 years and 11 months after the initial surgery. In this case, a good prognosis was obtained by frequent local resection and adjuvant chemotherapy for metachronous multiple metastases.

[A case of gastric endocrine cell carcinoma that responded to chemotherapy with CPT-11+CDDP].

A 57-year-old male patient was referred to our department with a diagnosis of #3 lymph node recurrence of early gastric cancer after treatment of endoscopic submucosal dissection (ESD). The pathological diagnosis of the ESD specimen was neuroendocrine cell carcinoma of the stomach with positive immunohistochemical staining of chromogranin A. The diameter of the tumor was 10 mm and the depth of invasion was pSM2. Distal partial gastrectomy with standard lymph node dissection (D2) was performed. The pathological findings were negative for malignancy in the resected stomach and positive in 2 of the #3 lymph nodes. Adjuvant chemotherapy of S-1 was administered, but a recurrence in the paraaortic lymph nodes was revealed by follow up X-ray computed tomography (X-CT) 3 months later. The case was considered as a S-1 failure, and the chemotherapy was changed to the irinotecan(CPT-11) +cisplatin(CDDP). A clinical complete response (CR) was obtained after two courses and maintained for up to twenty months.

[A case of drug resistant recurrent gastrointestinal stromal tumor treated surgically by resection, partial hepatectomy and diaphragmectomy].

This 69-year-old man underwent a partial jejunectomy for gastrointestinal stromal tumor(GIST) at the age of 60, and subsequently hepatectomy of segment 5, 6, 7 for liver metastasis of GIST a year later. An irregular mass close to the cutting stump of the liver, and a mass that showed enhanced-effect at segment 4 was discovered 28 months after hepatectomy. In order to treat this second recurrence, we administered imatinib and sunitinib sequentially. The tumor subsequently became drug-resistant, so we removed it surgically together with the liver and a portion of right diaphragm, and a tumor in segment 4. The patient shows no recurrent sign 4 months after surgery. This case suggests that surgical resection should be considered for partially drug resistant GIST.

High resolution grazing-incidence in-plane x-ray diffraction for measuring the strain of a Si thin layer.

We have measured the strain of a thin Si layer deposited on a SiGe layer using a high resolution x-ray diffraction system. The Si layer was deposited on the SiGe layer in order to introduce a tensile strain to the Si layer. To measure the in-plane lattice constant accurately, we have employed so-called grazing-incidence in-plane diffraction. For this measurement, we have made a new five-axis x-ray goniometer which has four ordinal circles (ω, 2θ, χ, φ) plus a counter-χ-axis for selecting the exit angle of the diffracted x-rays. In grazing-incidence geometry, an incident x-ray is focused on the sample surface in order to obtain good diffraction intensity even though the layer thickness is less than 5 nm. Because diffracted x-rays are detected through analyzer crystals, the diffraction angle can be determined with an accuracy of ± 0.0003°. This indicates that the strain sensitivity is about 10( - 5) when we measure in-plane Si 220 diffraction. Use of x-ray diffraction could be the best standard metrology method for determining strain in thin layers. Furthermore, we have demonstrated that incident/exit angle selected in-plane diffraction is very useful for height/depth selective strain determination.

[A case of AGC with pCR after preoperative chemotherapy including S-1 plus cisplatin].

A 79-year-old man complaining of epigastralgia was examined and diagnosed with advanced gastric cancer (UML, Type 5, Ant-Less-Gre, cT4a, cN1, cH0, cP1, cStage IV). A poor prognosis was predicted, but we tried preoperative chemotherapy hoping for a down-staging of the tumor. We chose a regimen of S-1 plus cisplatin as follows: S-1 (60 mg/m2) was administered orally for 3 weeks followed 2 weeks of rest, and cisplatin (50 mg/m2) was administered by intravenous drip on day 8. After three cycles of treatment, diagnostic laparoscopic examination revealed a suspected serosal invasion of the main tumor, but peritoneal dissemination was not seen, and abdominal washing cytology was negative. After the fourth cycle of treatment, total gastrectomy with lymph node dissection (D1+No. 7, 8a, 9, R0) was performed. Histological examination of the resected specimens revealed no residual cancer cells in the primary lesion or regional lymph nodes, resulting in a diagnosis of complete response to chemotherapy according to the Japanese Classification of Gastric Carcinoma. The postoperative course was uneventful, and he has been fine as an outpatient.

[A case report of pathologically complete response with chemo-radiation for rectal cancer].

A 60-year-old female presented to a local hospital with a chief complaint of melena. Since she was found to have a neoplastic lesion in the rectum on endoscopy, she was referred to our hospital. Barium-enema examination and endoscopy showed a 5-cm type-2 lesion proximal to the dentate line, which was diagnosed by biopsy as a moderately differentiated adenocarcinoma. Pelvic CT and MRI showed lower rectal wall thickening with pararectal lymphadenopathy, and FDP-PET showed increased uptake in these sites. Under a diagnosis of cT3, cN2, M0, Stage III b cancer, she started with preoperative chemoradiotherapy, consisting of 50 Gy in 25 fractions and oral UFT (400 mg)/Uzel (75 mg). After therapy, the tumor showed only a 2-cm ulceration, and FDG-PET revealed no abnormal uptake. Under a diagnosis of cT1, cN0, M0, Stage I cancer, super-low anterior resection of the rectum with D2 lymphadenectomy was performed. Histopathological examination showed that cancer cells forming a glandular structure had completely disappeared, leaving a fibrous scar, consistent with a Grade 3 (pCR).

[A case of retroperitoneal dedifferentiated liposarcoma successfully treated with IFM and CDDP].

A 53-year-old man was evaluated for a chief complaint of abdominal bloating. Physical examination revealed an abdominal distention and ascites, and CT showed multiple large retroperitoneal masses. The patient was diagnosed with retroperitoneal liposarcoma. Surgery was performed including the tumor, small bowel, and sigmoid resection, and an artificial anus was constructed. Multiple tumors in the peritoneum were noted. Large dark red tumors that were hemorrhagic were resected, but the yellowish tumors were unresectable. On histopathology, the dark red lesions showed dedifferentiated liposarcoma, and the yellowish lesions showed well-differentiated liposarcoma. One month postoperatively, peritoneal dissemination increased including nodular infiltration of the artificial anus and multiple hepatic metastases. Despite VAC chemotherapy (VCR 1.5 mg, ACD 0.5 mg, CPA 900 mg), progressive disease (PD) was noted. As second-line chemotherapy, weekly IFM (2 g)+CDDP (30 mg) was given. Shrinkage of the tumor infiltrates in the artificial anus, decreased abdominal bloating, and improved QOL were observed. A partial response (PR) against peritoneal dissemination was achieved. However, hepatic metastases increased, and the patient died 6 months after surgery. This case suggests that IFM+CDDP may be useful in dedifferentiated liposarcoma.

[A case of anal canal carcinoma with systemic lymph node metastases successfully treated by bevacizumab+mFOLFOX6 therapy].

We describe our experience with a patient who had unresectable anal signet-ring cell carcinoma with extensive metastases to the lymph nodes, lungs, and bones. The patient was treated with bevacizumab (Bev)+mFOLFOX6 and achieved complete response (CR). He was a man in his fifties, who visited a local doctor with the chief complaints of swelling in the axillary and inguinal regions. Signet-ring cell carcinoma was diagnosed by examination of a biopsy specimen from the inguinal lymph nodes. A search for the primary tumor was performed, and anal canal carcinoma with pagetoid spread was detected in the perianal region. Positron emission tomography-computed tomography (PET-CT) showed an accumulation in lymph nodes throughout the body, as well as in the lungs and the bones. Bev+mFOLFOX6 therapy was initiated. After completion of 4 courses, the lymph nodes were no longer palpable. PET-CT scanning showed no accumulation. During the 8th course, tumor markers decreased to the normal range, and CR was diagnosed. When 13 courses had been completed, the patient experienced grade 3 numbness of the hands and feet, so his treatment was changed to Bev+FOLFIRI therapy. In conclusion, Bev+mFOLFOX6 therapy achieved 6 months of CR in our patient who had anal signet-ring cell carcinoma with systemic metastases, which seemed likely to have a very poor prognosis.

[A case of IL-6 producing malignant mesothelioma of abdomen with thrombocytosis].

A 77-year-old man was admitted to our hospital with abdominal pain and ascites. He had an occupational history of working with asbestos. Abdominal CT showed multiple nodular lesions with enhancement by contrast medium in the cavity. Platelet counts, CRP and serum IL-6 level were increased. Biopsied materials obtained by laparoscopy showed oval cells with rich cytoplasm growing in an epithelial pattern. To clarify the characteristics of the cells, immunohistochemistry was performed. Calretinin and CK5/6 were positive, and CEA, S-100 protein, c-kit and CD34 were negative, result in confirmation of a diagnosis of malignant mesothelioma. Because IL-6, IL-6 receptor and VEGF were expressed markedly, the patient received chemotherapy for IL-6 suppression. During the treatment, thrombocytosis imploved satisfactorily.

Adjuvant immunochemotherapy with protein-bound polysaccharide K for colon cancer in relation to oncogenic beta-catenin activation.

PURPOSE: Protein-bound polysaccharide K is an immunotherapeutic agent that promotes apoptosis by inhibiting nuclear factor-kappaB activation in cancer cells. We previously showed that oncogenic beta-catenin activates nuclear factor-kappaB and inhibits apoptosis by up-regulating beta-transducin repeat-containing protein. We investigated whether the activation state of beta-catenin in the primary tumor is associated with differences in survival rates of patients with colon cancer undergoing immunochemotherapy with 5-fluorouracil plus polysaccharide K vs. chemotherapy with 5-fluorouracil alone. METHODS: We assessed the activation states of beta-catenin and nuclear factor-kappaB in the primary tumors of 202 colon cancer patients, and analyzed the data in terms of the clinicopathologic characteristics and survival of patients undergoing the two forms of adjuvant therapy. RESULTS: We found two distinct patterns of nuclear accumulation of activated beta-catenin in the tumor cells: diffuse nuclear accumulation in 89 cases (44 percent) and selective nuclear accumulation at the tumor invasion front in 18 cases (9 percent). Nuclear factor-kappaB activation was found in 64 cases (32 percent). In patients with diffuse nuclear accumulation-type beta-catenin activation, immunochemotherapy significantly improved recurrence-free survival, cancer death survival, and overall survival rates compared with patients receiving chemotherapy alone. No survival benefit was found in cases with nuclear accumulation at the tumor invasion front-type beta-catenin activation or no activation. Similarly, immunochemotherapy favored the survival of patients with nuclear factor-kappaB activation. Multivariate analysis established the TNM stage and administration of polysaccharide K as independent prognostic factors in the patients with diffuse nuclear accumulation-type beta-catenin activation. CONCLUSIONS: The presence of diffuse nuclear accumulation-type beta-catenin activation identifies patients with colon cancer who respond better to immunotherapy with polysaccharide K.

[Combination chemotherapy of TS-1, docetaxel and CDDP produces a remarkable response in a patient with advanced esophageal cancer].

We report a patient with advanced esophageal cancer who achieved a complete response to combination chemotherapy of TS-1, docetaxel and CDDP. A 74-year-old man was admitted to our hospital for advanced esophageal cancer with a complaint of dysphagia. He received chemotherapy, consisting of TS-1 100 mg/body, docetaxel 35 mg/m(2), and CDDP 10 mg/m(2), every 3 weeks. TS-1 was administered for 14 days followed by 7 days rest; docetaxel and CDDP were administered by intravenous infusion at day one and day 8 after beginning TS-1. After three cycles of chemotherapy, his dysphagia disappeared, and endoscopic examination of the primary esophageal tumor showed a complete response. Endoscopic examination with biopsy confirmed the disappearance of the esophageal cancer. No severe side effects were observed during this chemotherapy. Combination chemotherapy of TS-1, docetaxel, and CDDP can thus be effective for advanced esophageal cancer.

Pilot study of low-dose, divided maximum tolerated dose of CPT-11 in 21 consecutive patients with metastatic colorectal or gastric cancer.

PURPOSE: We devised a new treatment regimen, delivering a frequent low dose of CPT-11, calculated by dividing the maximum tolerated dose (MTD) to reduce its toxicity without impairing its efficacy. METHODS: CPI-11, 25 mg/m2, determined by dividing the MTD dose per month by 12, was given on days 1, 2, and 3 of every week, to 21 consecutive patients; 12 with metastatic colon cancer and 9 with metastatic gastric cancers. RESULTS: The total delivered dose of CPI-11 per patient was more than 1,000 mg in 17 (80.1%) of the 21 patients. Grade 3 marrow depression developed in 3 (14.3%) patients, and although nausea, vomiting, alopecia, and diarrhea developed in some patients, these side effects were all categorized as grade 2 or milder. The antitumor effect was evaluated in 18 patients with measurable lesions, who had received CPI-11 according to our regimen for at least 3 weeks. Of these 18 patients, 10, 7, and 1, respectively, had a found to have partial response, no change, or progression of disease, demonstrating a 55.6% efficacy rate [colon 6/10 (60.0%) and stomach 4/8 (50.0%)]. Moreover, time to progression (TTP) was greater than 90 days in 12 (75.0%) of these 18 patients. CONCLUSION: These results show that our low-dose, divided MTD of CPI-11 regimen is a promising method of reducing toxicity and strengthening the antitumor effect, justifying further large-scale comparative clinical studies to verify this potential.