Effect of partial coherence on dimensional measurement sensitivity for DUV scatterfield imaging microscopy.

Optical scatterfield imaging microscopy technique which has the capability of controlling scattered fields in the imaging mode is useful for quantitative nanoscale dimensional metrology that yields precise characterization of nanoscale features for semiconductor device manufacturing process control. To increase the sensitivity in the metrology using this method, it is required to optimize illumination and collection optics that enhance scatterfield signals from the nanoscale targets. Partial coherence of the optical imaging system is used not only for enhancing image quality in the traditional microscopy or lithography but also for increasing the sensitivity of the scatterfield imaging microscopy. This paper presents an empirical investigation of the effect of partial coherence on measurement sensitivity using a deep ultraviolet scatterfield imaging microscope platform that uses a 193 nm excimer laser as a source and a conjugate back focal plane as a unit for controlling partial coherence. Dimensional measurement sensitivity is assessed through analyzing scatterfield images measured at the edge area of periodic multiline structures with nominal linewidths ranging 44-80 nm on a Molybdenum Silicide (MoSi) photomask. Intensities scattered from the targets under the illuminations with various partial coherence factors and two orthogonal polarizations are assessed with respect to sensitivity coefficient. The optimization of partial coherence factor for the target dimension is discussed through the sensitivity coefficient maps.

In Vivo Fluorescence Microendoscopic Monitoring of Stent-Induced Fibroblast Cell Proliferation in an Esophageal Mouse Model.

PURPOSE: To evaluate the feasibility of self-expanding metal stent (SEMS) placement and fluorescence microendoscopic monitoring for determination of fibroblast cell proliferation after stent placement in an esophageal mouse model. MATERIALS AND METHODS: Twenty fibroblast-specific protein (FSP)-1 green fluorescent protein (GFP) transgenic mice were analyzed. Ten mice (Group A) underwent SEMS placement, and fluoroscopic and fluorescence microendoscopic images were obtained biweekly until 8 weeks thereafter. Ten healthy mice (Group B) were used for control esophageal values. RESULTS: SEMS placement was technically successful in all mice. The relative average number of fibroblast GFP cells and the intensities of GFP signals in Group A were significantly higher than in Group B after stent placement. The proliferative cellular response, including granulation tissue, epithelial layer, submucosal fibrosis, and connective tissue, was increased in Group A. FSP-1-positive cells were more prominent in Group A than in Group B. CONCLUSIONS: SEMS placement was feasible and safe in an esophageal mouse model, and proliferative cellular response caused by fibroblast cell proliferation after stent placement was longitudinally monitored using a noninvasive fluorescence microendoscopic technique. The results have implications for the understanding of proliferative cellular response after stent placement in real-life patients and provide initial insights into new clinical therapeutic strategies for restenosis.

Improved efficacy and in vivo cellular properties of human embryonic stem cell derivative in a preclinical model of bladder pain syndrome.

Interstitial cystitis/bladder pain syndrome (IC/BPS) is an intractable disease characterized by severe pelvic pain and urinary frequency. Mesenchymal stem cell (MSC) therapy is a promising approach to treat incurable IC/BPS. Here, we show greater therapeutic efficacy of human embryonic stem cell (hESC)-derived multipotent stem cells (M-MSCs) than adult bone-marrow (BM)-derived counterparts for treating IC/BPS and also monitor long-term safety and in vivo properties of transplanted M-MSCs in living animals. Controlled hESC differentiation and isolation procedures resulted in pure M-MSCs displaying typical MSC behavior. In a hydrochloric-acid instillation-induced IC/BPS animal model, a single local injection of M-MSCs ameliorated bladder symptoms of IC/BPS with superior efficacy compared to BM-derived MSCs in ameliorating bladder voiding function and histological injuries including urothelium denudation, mast-cell infiltration, tissue fibrosis, apoptosis, and visceral hypersensitivity. Little adverse outcomes such as abnormal growth, tumorigenesis, or immune-mediated transplant rejection were observed over 12-months post-injection. Intravital confocal fluorescence imaging tracked the persistence of the transplanted cells over 6-months in living animals. The infused M-MSCs differentiated into multiple cell types and gradually integrated into vascular-like structures. The present study provides the first evidence for improved therapeutic efficacy, long-term safety, and in vivo distribution and cellular properties of hESC derivatives in preclinical models of IC/BPS.

Distinct expression profile of key molecules in crawling-type early gastric carcinoma.

BACKGROUND: Gastric "crawling-type" adenocarcinoma (CRA) is a tumor histologically characterized by irregularly fused glands with low-grade cellular atypia that tends to spread laterally in the mucosa. To date, the expression characteristics of the key molecules involved in CRA, including receptor tyrosine kinases (RTKs), mismatch repair (MMR) proteins, phosphatase and tensin homolog (PTEN), as well as the Epstein-Barr virus (EBV) status, have yet to be uncovered. METHODS: We constructed tissue microarrays of 94 CRAs, 72 conventional-type differentiated adenocarcinomas (CDAs), and 71 intramucosal poorly cohesive adenocarcinomas (PCAs) from early gastric cancers to evaluate and compare the pathological and expression profiles of potential key molecules for molecular classification (EBV; four MMR proteins-MLH1, MSH2, PMS2, and MSH6; three RTKs-HER2, MET, and EGFR; PTEN; and p53). RESULTS: None of the CRAs showed MMR deficiency (0.0 % vs. 5.6 %, CRA vs. CDA, p = 0.036), HER2 overexpression (0.0 % vs. 12.5 %, p = 0.001), or loss of PTEN expression (0.0 % vs. 9.7 %, p = 0.003). Moreover, MET overexpression (4.4 % vs. 19.4 %, p = 0.004), and a mutant p53 pattern (12.4 % vs. 62.5 %, p < 0.001) were significantly less common in CRAs than in CDAs. However, clinicopathological features and all the profile of the molecules of CRAs were close to those of the PCA group. CONCLUSIONS: CRA demonstrated unique clinicopathological characteristics and showed a distinct expression profile of key molecules, which was close to that of a null phenotype. These results support the classification of CRA as a distinct subgroup of gastric adenocarcinoma.

PIK3CA amplification is associated with poor prognosis among patients with curatively resected esophageal squamous cell carcinoma.

To investigate the clinicopathologic characteristics and the prognostic impact of PIK3CA gene amplification in curatively resected esophageal squamous cell carcinoma (ESCC). Using 534 curatively resected ESCCs, the PIK3CA gene copy number was evaluated with fluorescent in situ hybridization. PIK3CA amplification was defined as PIK3CA/centromere 3 ratio is ≥ 2.0 or average number of PIK3CA signals/tumor cell nucleus ≥ 5.0. PIK3CA mutations in exon 9 and 20, encoding the highly conserved helical and kinase domains were assessed by direct sequencing in 388 cases. PIK3CA amplification was detected in 56 (10.5%) cases. PIK3CA amplification was significantly associated with higher T-stage (P=0.026) and pathologic stage (P=0.053). PIK3CA amplification showed a significantly shorter disease free survival (DFS) compared with that of non-amplified group (33.4 vs 63.1 months, P=0.019). After adjusting for gender, tumor location, pathologic stage, histologic grade and adjuvant treatment, PIK3CA amplification was significantly associated with a shorter DFS (adjusted hazard ratio [AHR] 1.53; 95% CI, 1.10-2.17; P=0.02). Though the statistical insignificance, PIK3CA amplification showed tendency of shorter OS (52.1 vs 96.5 moths, P=0.116). PIK3CA mutations were detected in 6 (1.5%) of 388 cases; 5 cases with exon 9 mutations in E545K while one exon 20 mutation in H1047L. PIK3CA amplification is a frequent oncogenic alteration and associated with shorter survival, suggesting its role as a prognostic biomarker in resected ESCC. PIK3CA amplification may represent a promising therapeutic target for ESCC.

Prognostic significance and frequency of EGFR expression and amplification in surgically resected advanced gastric cancer.

OBJECTIVE: The aim of this study is to find the frequency and the role of epidermal growth factor receptor expression as a prognostic biomarker in gastric cancer. METHODS: We evaluated the prognostic value and frequency of epidermal growth factor receptor expression and amplification using immunohistochemistry and silver in situ hybridization in a large cohort of curatively resected gastric cancer. RESULTS: Of the total of 935 cases, 294 (31.4%), 101 (10.8%) and 36 (3.9%) patients showed epidermal growth factor receptor 1+, 2+ and 3+ expression on immunohistochemistry, respectively. Epidermal growth factor receptor-positive (2+/3+) patients more frequently had intestinal type than epidermal growth factor receptor-negative (0/1+) patients (82.5 vs. 44.1%, P < 0.001). After adjusting for sex, age, stage and adjuvant chemotherapy, epidermal growth factor receptor-positive patients had a favorable overall survival outcome compared with epidermal growth factor receptor-negative patients (hazard ratio, 0.734; 95% confidence interval, 0.541-0.997; P = 0.047), especially in Stage III disease (hazard ratio, 0.676; 95% confidence interval, 0.472-0.968; P = 0.033). Among the 393 cases available for  in situ hybridization, the correlation between immunohistochemistry and in situ hybridization was statistically significant (P = 0.001). Thirteen patients with gene amplification (3.3%) did not show different survival outcome with others (P = 0.359). CONCLUSION: Epidermal growth factor receptor positivity was an independent favorable prognostic factor for gastric cancer, especially in Stage III disease.

Epstein-Barr virus positivity, not mismatch repair-deficiency, is a favorable risk factor for lymph node metastasis in submucosa-invasive early gastric cancer.

BACKGROUND: Epstein-Barr virus (EBV)-associated gastric cancer (GC) and microsatellite-instability-high GC are associated with a low prevalence of regional lymph node metastasis (LNM). To evaluate the feasibility of endoscopic treatment of EBV-associated and/or microsatellite-instability-high early GC (EGC), we analyzed the risk factors for LNM using a large series (n = 756) of submucosa-invasive (SM) EGC. METHODS: EBV-encoded RNA in situ hybridization (EBER ISH) and immunohistochemistry for four mismatch repair (MMR) proteins (MLH1, PMS2, MSH2, and MSH6) were performed. The clinicopathologic features and results of EBER ISH and immunohistochemistry were compared according to the LNM status. RESULTS: Among the cases, 146 EGCs (19.3 %) showed LNM. EBV negativity, larger tumor size (greater than 2 cm), deeper level of submucosal invasion, submucosal invasion depth greater than 500 µm, presence of ulceration, and presence of lymphovascular invasion (LVI) were associated with LNM. However, the MMR deficiency was not correlated with LNM. On multivariate regression analysis, larger tumor size (greater than 2 cm; odds ratio 1.6, p = 0.030), deeper level of submucosal invasion (odds ratio 2.9, p = 0.001), LVI (odds ratio 7.4, p < 0.001), and EBV negativity (p = 0.020) were independent risk factors for LNM in SM EGCs. CONCLUSIONS: EBV positivity was a favorable risk factor for LNM in SM EGC. However, MMR deficiency was not associated with the status of LNM. Thus, we suggest that examination with EBER ISH could be considered for endoscopic resected specimens, especially in cases of SM EGC showing no LVI and clear resection margins.

Usefulness of Immunohistochemistry for Microsatellite Instability Screening in Gastric Cancer.

BACKGROUND/AIMS: The usefulness of immunohistochemistry to screen for the microsatellite instability (MSI) phenotype in gastric cancer remains unclear. Moreover, the prognostic value of MSI phenotypes in gastric cancer has been debated. METHODS: The clinicopathologic parameters and survival outcomes of 203 MSI-high (MSI-H) and 261 microsatellite-stable (MSS) advanced gastric cancers (AGCs) were compared. Next, we compared the immunohistochemistry results for hMLH1 and hMSH2 with those of a polymerase chain reaction (PCR)-based method. Kaplan-Meier curves and a Cox proportional hazard regression model were used to conduct survival analyses. RESULTS: The MSI-H AGCs were correlated with older age (p<0.001), female gender (p=0.018), distal location (p<0.001), larger size (p=0.016), and intestinal type (p<0.001). Multivariate analysis revealed that the MSI-H phenotype was an independent favorable factor that was related to overall survival in patients with AGC (p<0.001). Compared with the PCR-based analysis, immunohistochemistry exhibited high sensitivity (91.1%) and specificity (98.5%) in the detection of MSI phenotypes. CONCLUSIONS: MSI-H gastric cancers have distinct clinicopathologic features and better prognoses, which suggests the necessity of MSI analysis in gastric cancer. Immunohistochemistry can be a useful and reliable screening method in the assessment of MSI status in gastric cancer.

Poorly Differentiated Carcinoma Component in Submucosal Layer Should be Considered as an Additional Criterion for Curative Endoscopic Resection of Early Gastric Cancer.

BACKGROUND: Some studies have reported lymph node metastasis (LNM) in early gastric cancer (EGC) cases meeting the expanded criteria for endoscopic resection. Therefore, we investigated whether a minor poorly differentiated carcinoma (PDC) component in the submucosal (SM) layer affects LNM in differentiated EGC. METHODS: We performed surgery in 1096 patients with differentiated SM gastric cancer and compared the clinicopathologic features of node-positive (n = 194) and node-negative (n = 902) differentiated SM cancer, with special reference to the portion of PDC component in the SM layer. RESULTS: When we categorized patients by the proportion of PDC component in the SM layer, we found 840 patients had <5 % and 256 patients had ≥5 % PDC components in the SM layer. The ≥5 % group was significantly associated with younger age, female sex, moderate differentiation, deep SM invasion, lymphovascular invasion (LVI), perineural invasion, and LNM. In multivariate analysis, middle third location, moderate differentiation, SM2 invasion, size >2 cm, LVI, and PDC components in the SM layer were independent risk factors for LNM. When we limited the depth of invasion to SM1, the incidence of LNM was significantly higher in the ≥5 % group. On multivariate analysis, tumor size >2 cm, moderate differentiation, LVI positivity, and ≥5 % PDC components in the SM1 layer were independent risk factors for LNM in SM1 cancer. CONCLUSIONS: The PDC component in the SM layer of differentiated EGC was an independent risk factor of LNM, which might constitute a supplementary criterion in the expanded indications for endoscopic resection in differentiated EGC.

Fibroblast growth factor receptor 1 gene amplification is associated with poor survival in patients with resected esophageal squamous cell carcinoma.

To investigate the frequency and the prognostic impact of fibroblast growth factor receptor 1 (FGFR1) gene amplification in 526 curatively resected esophageal squamous cell carcinoma (ESCC). Using fluorescent in situ hybridization, high amplification was defined by an FGFR1/centromer 8 ratio is ≥ 2.0, or average number of FGFR1 signals/tumor cell nucleus ≥ 6.0, or percentage of tumor cells containing ≥ 15 FGFR1 signals or large cluster in ≥ 10%. Low amplification was defined by ≥ 5 FGFR1 signals in ≥ 50%. FGFR2 and FGFR3 mutations were assessed by direct sequencing in 388 cases and no mutation was detected. High and low amplification were detected in 8.6% and 1.1%, respectively. High FGFR1 amplification had significantly shorter disease-free survival (34.0 vs 158.5 months P=0.019) and overall survival (52.2 vs not reached P=0.022) than low/no amplification group. After adjusting for sex, smoking, stage, histology, and adjuvant treatment, high FGFR1 amplification had a greater risk of recurrence (adjusted hazard ratio [AHR], 1.6; P=0.029) and death (AHR, 1.53; P=0.050). High amplification was significantly higher in current smokers than former and never-smokers (Ptrend<0.001) and increased proportional to smoking dosage. High FGFR1 amplification is a frequent oncogenic alteration and an independent poor prognostic factor in resected ESCC.

Reduced-phase dual-illumination interferometer for measuring large stepped objects.

We present a reduced-phase dual-illumination interferometer (RPDII) that measures the topography of a sample with large step height variation. We experimentally demonstrate the basic principle and the feasibility of this novel single-shot quantitative phase imaging. Two beams of this interferometer illuminate a sample at different incident angles, and two phases of the different incident angles and their phase difference are simultaneously recorded using three spatial frequencies. The relative phase difference between two beams of an RPDII can be controlled by adjusting the angle such that the maximum phase difference is smaller than 2π, and thus there is no phase wrapping ambiguity in the reconstructed phase. One 4f optical system with a transmission grating is used to illuminate the sample with two collimated beams incident at different angles. The feasibility of this technique is demonstrated by measuring the thicknesses of two stepped metal layers with heights of 150 and 660 µm. Although the change in stepped height is more than 1000 times the wavelength of the laser used in our interferometer, the thicknesses of these two metal layers are successfully obtained without the use of an unwrapping algorithm.

Optimum phase shift for quantitative phase microscopy in volume measurement.

Volume measurement of a phase object is one of the most distinctive capabilities of quantitative phase microscopy (QPM). However, the accuracy of a measured volume is limited by the different noises of a measurement system and the finite bandpass filter used in the phase extraction algorithm. In this paper, we analyze the inherent errors in volume measurement with QPM and propose the optimum condition that can minimize these errors. We find that phase information of a sample in the frequency domain nonlinearly oscillates as a function of the phase shift corresponding to the sample and its medium, and that the phase information of a sample inside the bandpass filter can be maximized by a proper phase shift. Through numerical simulations and actual experiments, we demonstrate that the error in phase volume measurement can be effectively reduced by the enhancement of the phase signal inside the bandpass region using an optimum amount of phase, which can be controlled by changing either the medium index or the wavelength of illumination.

Using the Oxford classification of IgA nephropathy to predict long-term outcomes of Henoch-Schönlein purpura nephritis in adults.

Recently, there has been emerging concern that crescents, the main histologic feature of Henoch-Schönlein purpura nephritis, merely reflect active inflammation, and may not be useful in predicting long-term outcomes. We therefore conducted a single-center retrospective study to evaluate whether the new Oxford classification of immunoglobulin A nephropathy can be used to predict long-term outcome in patients with Henoch-Schönlein purpura nephritis. We included 61 biopsy-proven patients with Henoch-Schönlein purpura nephritis between January 1991 and August 2010. In addition to the International Study of Kidney Disease in Children classification, pathologic findings were also evaluated by the Oxford classification. Primary outcomes were defined as either the onset of estimated glomerular filtration rate <60 ml/min per 1.73 m(2) with ≥30% decrease in estimated glomerular filtration rate from baseline or end-stage renal disease. During a median follow-up of 49.3 months, 13 (21%) patients reached the primary end point. A Kaplan-Meier plot showed that renal event-free survival was significantly longer in patients with <50% crescents than in those with crescents in ≥50% of glomeruli (P=0.003). Among the components of the Oxford classification, patients with endocapillary hypercellularity (E1; P=0.016) and tubular atrophy/interstitial fibrosis (T1/T2; P=0.018) had lower renal survival rates than those with E0 and T0. In a multivariate Cox model adjusted for clinical and pathologic factors, E1 (hazard ratio=8.91; 95% confidence interval=1.47-53.88; P=0.017) and T1/T2 (hazard ratio=8.74; 95% confidence interval=1.40-54.38; P=0.020) were independently associated with reaching a primary outcome, whereas the extent of crescentic lesions was not. Our findings suggest that the Oxford classification can be used in predicting long-term outcomes of Henoch-Schönlein purpura nephritis.

Usefulness of Ki-67 (MIB-1) immunostaining in the diagnosis of pulmonary sclerosing hemangiomas.

Pulmonary sclerosing hemangioma (PSH) is an uncommon lung neoplasm with a clinical outcome that is generally benign. However, differentiating PSH from pulmonary carcinoma is sometimes difficult as both lesions share similar histopathologic and immunohistochemical features. In this study, we investigated the usefulness of Ki-67 (MIB-1) immunostaining in the diagnosis of PSH. We compared the staining pattern for Ki-67 (MIB-1) in 29 cases of typical PSH and 79 cases of pulmonary non-small cell carcinoma (NSCLC) using an immunohistochemical method on formalin-fixed paraffin-embedded tissues. In all studied PSH cases, we noted cell membrane and cytoplasmic staining for Ki-67 (MIB-1), but this was not observed in any of the NSCLC cases. The Ki-67 proliferation index was lower in PSH than in the NSCLC cases (mean, 1.1% vs mean, 5.5%; p < 0.001). These findings suggest that cell membrane and cytoplasmic staining for Ki-67 (MIB-1), as well as the Ki-67 proliferation index, may be useful for distinguishing PSH from pulmonary carcinoma.

Diagnostic Accuracy of Cerebrospinal Fluid (CSF) Cytology in Metastatic Tumors: An Analysis of Consecutive CSF Samples.

BACKGROUND: Cerebrospinal fluid (CSF) examination can be used to verify the presence of primary malignancies as well as cases of central nervous system (CNS) metastasis. Because of its importance, there have been several studies concerning the sensitivity of CSF cytology. To determine the practical use and reproducibility of diagnoses based on CSF cytology, we evaluated this test by analyzing cytology results from consecutive CSF samples. METHODS: Between July 2010 and June 2013, 385 CSF cytology samples from 42 patients were collected. The samples were gathered using a ventricular catheter and reservoir. CSF cytology of all patients was examined more than two times with immunocytochemistry for cytokeratin. RESULTS: Primary neoplastic sites and histologic types of patients' metastatic cancer were diverse. The overall sensitivity for detecting malignancy was 41.3%. Even within short-term intervals, diagnoses frequently changed. CONCLUSIONS: Our results were inconsistent, with low sensitivity, when compared to the results of previous studies. However, CSF evaluation can still provide valuable diagnostic and prognostic information because adjuvant treatments are now routinely performed in patients with CNS metastasis. Negative CSF cytology results should not be ignored, and continuous CSF follow-up is essential for following the clinical course of patients with metastatic cancer involving the CNS.

New classification of focal cortical dysplasia: application to practical diagnosis.

BACKGROUND AND PURPOSE: Malformation of cortical development (MCD) is a well-known cause of drug-resistant epilepsy and focal cortical dysplasia (FCD) is the most common neuropathological finding in surgical specimens from drug-resistant epilepsy patients. Palmini's classification proposed in 2004 is now widely used to categorize FCD. Recently, however, Blumcke et al. recommended a new system for classifying FCD in 2011. METHODS: We applied the new classification system in practical diagnosis of a sample of 117 patients who underwent neurosurgical operations due to drug-resistant epilepsy at Severance Hospital in Seoul, Korea. RESULTS: Among 117 cases, a total of 16 cases were shifted to other FCD subtypes under the new classification system. Five cases were reclassified to type IIIa and five cases were categorized as dual pathology. The other six cases were changed within the type I category. CONCLUSIONS: The most remarkable changes in the new classification system are the advent of dual pathology and FCD type III. Thus, it will be very important for pathologists and clinicians to discriminate between these new categories. More large-scale research needs to be conducted to elucidate the clinical influence of the alterations within the classification of type I disease. Although the new FCD classification system has several advantages compared to the former, the correlation with clinical characteristics is not yet clear.