Plant Cell. 2008 January; 20(1): 75–87
Guard cells play a critical role in plant growth and development by optimizing gas exchange under variable environments.
Reproductive Biology and Endocrinology 2008, 6:40
Statistical analysis Exploratory data analyses, Kruskal-Wallis test for group comparisons, as well as the Mann-Whitney U test for nonparametric independent two-group comparisons were performed with the program SPSS 14 for Windows (SPSS Inc., Chicago, IL). Differences with P < 0.05 were regarded as statistically significant, P < 0.01 as highly statistically significant. Values of mRNA quantification are given as mean ± standard deviation (SD).
J. Clin. Invest. 118(1): 89-99 (2007)
Stable transfection. The GPC1 antisense construct was prepared by RT-PCR amplification of human placenta cDNA, as described previously (11). Stable transfection of GPC1-AS-1751 into PANC-1 cells was performed by using the lipofectamine method (12), and single clones were isolated after 3-4 weeks. After expansion, cells from each individual clone were screened for expression of GPC1 sense and antisense mRNA by northern blot analysis. Parental PANC-1 cells also were transfected with an empty expression vector carrying the neomycin-resistance gene as a control. Positive clones were routinely grown in selection medium.
Figure 4. Effects of gain of function of Notch ligand杕ediated signals on EPC differentiation. A, Scheme of the insert culture system with 0.4-祄 pores, in which receptors on the surface of the target stem cells in the upper chamber are capable of directly interacting with ligands on the cells in the bottom chamber. In the present study, BM-Lin?/SUP> cells were seeded in the upper chamber, whereas 3T3 stromal cells overexpressing a specific Notch ligand, Jag-1 or Dll-1, were placed in the bottom chamber. B, Reverse-transcription polymerase chain reaction revealed enhanced expression of the specific Notch ligand in the stromal cells stimulated by Jag-1 or Dll-1 signal compared with those transduced with empty vector. C, Reverse-transcription polymerase chain reaction revealed expression of Hes-1 and Hes-5, target effector genes of active Notch, in BM-Lin?/SUP> cells, which were cocultured with 3T3 stromal cells specifically expressing the target Notch ligand gene. D, Fluorescence-activated cell sorting analysis revealed more frequent expression of Flk-1 and CD31 in BM-Lin?/SUP> cells stimulated by Jag-1杕ediated signals, but not Dll-1杕ediated signals, than in those stimulated by empty vector. E, Reverse-transcription polymerase chain reaction to detect expression of typical EPC surface markers and vascular endothelial growth factor in BM-Lin?/SUP> cells stimulated by specific Notch ligand or empty vector. The cellular mRNA level of CD31, Flk-1, and vascular endothelial cadherin (VE-cadherin) was elevated in the Jag-1 group compared with the Dll-1 and empty-vector groups. In contrast, the cellular mRNA level of vascular endothelial growth factor and Flt-1 was similar in all groups. PECAM indicates platelet and endothelial cell adhesion molecule. F, EPC colony-forming assay using BM-KSLs stimulated by specific Notch ligand杕ediated signals revealed significant augmentation of vasculogenic capacity in the Jag-1 group, but not the Dll-1 group, compared with the empty-vector group. CFU indicates colony-forming units. **P<0.01 (n=3 in each group). G, Frequency of apoptotic cells (TUNEL-positive cells) in BM Sca-1+/Lin?/SUP> cells in vitro was significantly lower in EPC-enriched cells stimulated by Jag-1 signal than in those stimulated by Dll-1 signal or empty vector. *P<0.05 (n=3 in each group).
xue, r. et al. investigation of volatile biomarkers in liver cancer blood using solid-phase microextraction and gas chromatography / mass spectrometry. rapid communications in mass spectrometry 1181-1
The emerging field of biomarkers has applications in the diagnosis, staging, prognosis and monitoring of disease progression, as well as in the monitoring of clinical responses to a therapeutic intervention and the development and delivery of personalized treatments to reduce attrition in clinical trials.
Circulation Research. 2004; 94: 46-52
Immunohistochemistry Four weeks after the transplantation, cardiac grafts were cut horizontally into 4 blocks, embedded in OTC compound (Sakura Finetechnical Co, Tokyo, Japan) and kept at ?80°C until staining. Nine slices (3 slices from 3 blocks from the apex) were made for immunostaining with a kit (Histofine SAB-PO kit, Nichirei Co, Tokyo, Japan) for macrophages (MOMA-2) and CD4- and CD8-positive T cells, as previously described.19 Three fields where a coronary artery was recognized in the center were selected from each animal to count the number of macrophages and calculate a percent-positive area in a blind manner at a magnification of ×200.
PLoS One. 2012; 7(3): e33762.
Neuroendocrine small cell cervical carcinoma (SCCC) is an aggressive, rare form of cervical cancer, accounting for less than 3% of all cervical cancers –. SCCC is characterized by a high incidence of early nodal and distant metastases, resulting in a poorer prognosis than other subtypes of cervical cancer –. Previous studies have reported that 60–82% of SCCC patients have lymph-vascular space infiltration or pelvic lymph node metastasis at diagnosis –. Additionally, SCCC exhibits a propensity for rapid distant metastasis via the bloodstream to various sites including the liver, lung, brain, bone, pancreas and lymph nodes, which results in treatment failure in most cases –. Recurrences usually occur within 2 years, and most patients die as a result of early metastasis. It is important to identify the factors responsible for the survival of metastases in order to improve treatment strategies for SCCC. However, due the rarity and the long time period required to enroll a sufficient number of patients, most studies on SCCC are comprised of small series or case reports, making it difficult to determine the optimal therapy.MicroRNAs (miRNAs) are noncoding RNAs 18 to 25 nucleotides in length . The effects of miRNAs are mediated by binding to target mRNAs, to either suppress mRNA translation or degrade miRNA-bound mRNA . Currently, more than several hundred unique mature human miRNAs are known, and many are involved in tumorigenesis, acting either as oncogenes  or tumor suppressors , . Aberrant expression of miRNAs has also been linked to cancer , , suggesting that miRNAs potentially represent prognostic markers, and leading to the use of miRNA profiling for the diagnosis and prognosis of specific cancers. It has been shown that miRNAs are involved in every type of cancer examined to date; however, the expression of miRNAs in SCCC has not been investigated.In this study, miRNA qPCR arrays were performed on 44 SCCC samples, as we hypothesized that investigation of miRNA profiles would provide more information on SCCC, an inadequately understood disease with a poor prognosis.
World J Surg Oncol. 2013; 11: 71.
Quality controlIn the previous study, each subject underwent a clinical examination by two physicians, each of whom had over 3 years of experience working in a breast-surgery department. Each subject completed a questionnaire through interview by one of our trained questionnaire interviewers. At the end of each day, 10% of the questionnaires were randomly inspected to check for completeness, accuracy, and standardization. Any missing data or errors were corrected the following day. After the whole investigation was completed, the questionnaires were entered twice into a computer by two different inputters blinded to the study goals, with the second person correcting any inconsistencies found in the first set of data. Detailed information is available in the previously published paper .
J Biol Chem. 2008 Apr 11;283(15):9986-98.
Tissue Processing, Immunohistochemistry, and Quantitation—Animals were perfused transcardially with a saline solution containing 0.5% sodium nitrate and heparin (1000 units/ml) prior to fixation with 4% paraformaldehyde dissolved in 0.1 m phosphate buffer. Brains were removed and postfixed overnight in buffered 4% paraformaldehyde at 4 °C, incubated in a 30% sucrose solution for 48-72 h at 4 °C until they sank, and sectioned frozen on a sliding microtome in 30-μm-thick coronal sections. Sections were collected and processed for immunohistochemical staining for tyrosine hydroxylase (TH) alone or in combination with the oxidized forms of PRX. The unbiased stereological estimation of the total number of TH-positive cells in the substantia nigra was carried out using the optical fractionator, as previously described in full detail (29). The sampling technique was not affected by tissue volume changes and did not require reference volume determinations. Sampling was done using the Computer-Assisted Stereological Toolbox system, version 2.1.4 (Olympus Denmark A/S, Ballerup, Denmark), using a microscope with a motorized stage run by an IBM-compatible computer. The substantia nigra was delineated at a ×1.25 objective and generated counting areas of 150 × 150 μm. A counting frame (1612 μm2) was placed randomly on the first counting area and moved through all counting areas until the entire delineated area was sampled. Actual counting was carried out using ×100 oil objective. Guard volumes (4 μm from the top and 4-6 μm from the bottom of the section) were excluded from both surfaces to avoid the problem of lost caps, and only the profiles that came into focus within the counting volume (with a depth of 10 μm) were counted. For double immunofluorescent localization of TH and the oxidized forms of PRX, tissue sections were further fixed in 100% methanol for 20 min at -20 °C, permeabilized in 0.2% Triton X-100 in 0.2 m phosphate buffer at room temperature for 1 h, blocked in 3% bovine serum albumin in 0.2 m phosphate buffer at room temperature for 15 min, and then incubated overnight at 4 °C with mouse monoclonal anti-TH (1:2000) and rabbit polyclonal antibody against the oxidized form of PRX (1:200). Tissue sections were rinsed extensively with 0.2 m phosphate buffer and incubated at room temperature for 1 h with a mixture of Alexa Fluor 568-conjugated goat anti-mouse IgG and Alexa Fluor 488-conjugated goat anti-rabbit IgG (1:200; Molecular Probes). Following extensive wash with 0.2 m phosphate buffer, stained tissue sections were examined with an AxioImager D1 fluorescence microscope equipped with a digital image analyzer (Carl Zeiss) or an LSM 510 Meta Laser Scanning Microscope (Carl Zeiss). To assess the number of TH-positive DA neurons or TH-negative cells co-localized with the increased level of the oxidized forms of PRX, manual counts were conducted using 10-15 randomly selected confocal images per condition by an individual who was blind to the treatment conditions. Adjacent tissue sections from each animal were also stained with cresyl violet and subjected to quantitation to validate immunohistochemical determination of nigral neuronal survival.
Am J Gastroenterol. 2013 August; 108(8): 1305–1313.
Radiographic proceduresFor the diagnosis of HCC at study entry, intravenous contrast-enhanced dynamic CT was performed on an outpatient basis using an X-ray CT device with 4, 8, or 16 detector rows (Aquilion 4/16; Toshiba, Tokyo, Japan; LightSpeed Qx/I, LightSpeed Ultra; GE Healthcare, Milwaukee, WI). Images were obtained during the early arterial, late arterial, and equilibrium phases at 28, 40, and 120s after starting the intravenous bolus injection of iopamidol (Iopamiron; Nihon Schering, Osaka, Japan) or iohexol (Omnipaque; Daiichi Sankyo, Tokyo, Japan) at a rate of 2.3–3.3ml/s with a power injector. The total dose of iodine was 0.7g/kg body weight, with an upper limit of 37g iodine. The injection time for the contrast material was 30s. Images were reconstructed with a section thickness of 2.5mm and a reconstruction interval of 1.5mm, and were reviewed by experienced radiologists.CTHA/CTAP was performed on an inpatient basis. First, a 4-Fr modified Shepherd-hook catheter and a 4-Fr hepatic-curve catheter were placed in the celiac artery and superior mesenteric artery, respectively, through bilateral femoral arteries, according to Seldinger's method. Digital subtraction angiography was performed from the celiac artery to evaluate hepatic artery anatomy. A microcatheter was inserted through the 4-Fr catheter and placed in the proper or common hepatic artery for hepatic arteriography.The CTAP catheter was placed in the superior mesenteric artery in all cases. In the case of a replaced or accessory right hepatic artery, the catheter was inserted well beyond the origin of the hepatic artery to prevent contrast medium overflow into the hepatic artery. Less than 30ml of contrast agent, which was diluted to 100mg I/ml, was used before the CTHA/CTAP study. First, CTAP was performed using 90ml nonionic contrast medium diluted to 100mg I/ml, and then CT scanning was performed 30s after the start of the injection at a rate of 3.0ml/s. Multidetector-row CT images were obtained during a single breath hold in a longitudinal direction with collimation of 1mm, table speed of 30mm/s, 120kVp, and 300mAs. CTHA was performed at least 5min after CTAP, using the same parameters. CT scanning was performed at 10 and 45s after the start of contrast medium injection into the microcatheter at a rate of 2.0–2.5ml/s. A total of 30–50ml contrast agent diluted to 100mg I/ml was used. When the liver was perfused by two or more hepatic arteries such as a replaced right hepatic artery, accessory right hepatic artery, or left hepatic artery downstream of the left gastric artery, CTHA was performed from each of the respective arteries. A diagnosis of typical HCC on CTHA/CTAP was defined as a round hypervascular nodule on CTHA with a defect on CTAP, accompanied by corona enhancement during the second phase of CTHA or hypoattenuation during the equilibrium phase of prior dynamic CT ((10,29)).TACE was additionally performed when ≥4 HCC nodules were detected on CTHA/CTAP, as evaluated at the time by the operating radiologist. The procedure used 3.0ml contrast medium, 30mg doxorubicin (Adriacin; Kyowahakko Kirin, Tokyo, Japan), and 3.0ml iodized oil (Lipiodol Ultra-Fluid; Guerbet Japan, Tokyo, Japan). The amounts of contrast medium and iodized oil in this suspension were arbitrarily adjusted according to tumor size. This agent was injected into each feeder of the HCC, followed by infusion of 2-mm-diameter gelatin sponge particles (Gelpart; Nihonkayaku, Tokyo, Japan).CTHA/CTAP images were scrutinized by two experienced radiologists, who made the final diagnosis. The radiologists were not blinded to information regarding the preceding conventional dynamic CT. Preceding intravenous contrast-enhanced dynamic CT was retrospectively reviewed for nodules newly diagnosed by CTHA/CTAP to determine whether the nodules could have been detected on dynamic CT.
Metal ceramic multilayers have come into greater focus due to their promising mechanical, physical and chemical properties, making them practically useful for harsh environments and extreme loading. These composites show improvement in hardness, toughness, wear resistance, thermal resistance, shock resistance and irradiation resistance, to name a few.
Breast cancer is the most common malignancy in women and is the leading cause of death among women world-wide . Despite many advances in the diagnosis and treatment of breast cancer, metastasis remains an insurmountable challenge. Although the molecular mechanisms are not completely understood, it is well established that the formation and growth of new blood vessels is critical for sustained tumor growth and metastasis . Studies have found that benign lesions associated with high vascular density are correlated with an increased risk of developing breast cancer. Traditionally, the mechanism(s) controlling the development of tumor vasculature and perfusion were thought to be endothelial cell-lined vascular networks . Thus, efforts to reduce the growth and spread of breast cancer focused on the mechanism(s) of angiogenesis by which tumors establish a blood supply for survival, growth, and metastasis .
A 51-year-old man with a diagnosis of myelodysplasia and non-Hodgkin's lymphoma underwent an unmatched allogenic bone marrow transplantation and was treated posttransplant with chronic immunosuppressive medication. Eight months following transplantation, he presented with progressive dysarthria, cognitive and visual decline. Evaluation included brain magnetic resonance (MR) imaging demonstrating multifocal areas of increased T2 and FLAIR (fluid attenuated inversion recovery) signals involving the left frontal, parietal, and occipital lobes. The MR lesions demonstrated diffuse increased signal on DWI (diffusion-weighted images) and normal to low signal on ADC (apparent diffusion coefficients). Contrast-enhanced T1 images were unremarkable. Lumbar puncture revealed a mild elevation in cerebrospinal fluid (CSF) protein. CSF PCR assay for viral DNA fragments were negative on two occasions. Serum serology for HIV was negative as well. A brain biopsy was subsequently performed. The clinical and neuroimaging differential diagnoses as well as neuropathologic correlation are presented.,
Reproductive Biology and Endocrinology 2008, 6:40
TLR3 and TLR4 protein is localised to endometrial cells during the menstrual cycle. TLR3 protein staining in healthy late proliferative (LP) tissue was high in luminal and glandular tissue (A, brown precipitate) and lower in LP endometriotic tissue (B). Late secretory (LS) endometrium showed highly expressed TLR3 in the epithelium (C), but weakly in endometriosis (D). Intense staining of TLR4 proteins was shown in mid proliferative (MP) tissue (E). In late proliferative phase of endometriosis, TLR4 proteins were comparably lower (F). TLR4 protein was high in mid secretory (MS) normal endometrium (G), whereas it was decreased in endometriotic MS tissue (H). During the menstrual phase, both TLR3 (I) and TLR4 (J) were highly expressed. Co-immunostaining for TLR4 (green), CD14 (K, red) and CD163 (L, red) demonstrated that TLR4 proteins were expressed by CD14 positive dendritic cells and monocytes (K, yellow) and by CD163 positive macrophages (L, yellow). Localisation of TLR4 to immune cells is marked by a black arrow (J) and by white arrows (K, L). Allhorn et al. Reproductive Biology and Endocrinology 2008 6:40
J. Clin. Invest. 118(1): 89-99 (2007)
Immunohistochemistry of paraffin-embedded and frozen tissues. Paraffin-embedded tumor tissues were sectioned (5 μm thick), mounted on poly-l-lysine-coated glass slides, and allowed to dry overnight at 23°C. These sections were used for detection of PCNA and apoptosis by the ApopTag TUNEL assay, as previously reported (50, 51).