Three-dimensional reconstruction of tumor microvasculature: simultaneous visualization of multiple components in paraffin-embedded tissue

Three-dimensional (3D) visualization of microscopic structures may provide useful information about the exact 3D configuration, and offers a useful tool to examine the spatial relationship between different components in tissues. A promising field for 3D investigation is the microvascular architecture in normal and pathological tissue, especially because pathological angiogenesis plays a key role in tumor growth and metastasis formation. This paper describes an improved method for 3D reconstruction of microvessels and other microscopic structures in transmitted light microscopy. Serial tissue sections were stained for the endothelial marker CD34 to highlight microvessels and corresponding images were selected and aligned. Alignment of stored images was further improved by automated non-rigid image registration, and automated segmentation of microvessels was performed. Using this technique, 3D reconstructions were produced of the vasculature of the normal brain. Also, to illustrate the complexity of tumor vasculature, 3D reconstructions of two brain tumors were performed: a hemangioblastoma and a glioblastoma multiforme. The possibility of multiple component visualization was shown in a 3D reconstruction of endothelium and pericytes of normal cerebellar cortex and a hemangioblastoma using alternate staining for CD34 and alpha-smooth muscle actin in serial sections, and of a GBM using immunohistochemical double staining. In conclusion, the described 3D reconstruction procedure provides a promising tool for simultaneous visualization of microscopic structures.     

Clinicopathological and prognostic implications of endoglin （CD105） expression in hepatocellular carcinoma and its adjacent non-tumorous liver

AIM: The expression pattern of endoglin (CD105) in hepatocellular carcinoma (HCC) has not been reported so far. We hypothesized that CD105 could differentially highlight a subset of microvessels in HCC, and intratumoral microvessel density (IMVD) by CD105 immunostaining (IMVD-CD105) could provide better prognostic information than IMVD by CD34 immunostaining (IMVD-CD34). METHODS: Paraffin blocks of tumor and adjacent non-tumorous liver tissues from 86 patients who underwent curative resection of HCC were used for this study. Serial sections were stained for CD105 and CD34, respectively, to highlight the microvessels. IMVD was counted according to a standard protocol. RESULTS: In the HCC tissues, CD105 was either negatively or positively stained only in a subset of microvessels. In contrast, CD34 showed positive and more extensive microvessel staining in all cases examined. However, in the adjacent non-tumorous liver sections, CD105 showed a diffuse pattern of microvessel staining in 20 of 86 cases, while CD34 showed negative or only focal staining of the sinusoids around portal area. Correlation with clinicopathological data demonstrated that lower scores of IMVD-CD105 were found in larger sized tumors [mean 41.4/0.74 mm2 (>5 cm tumor) vs 65.9/0.74 mm2 (≤5 cm tumor), P= 0.043] and more aggressive tumors, as indicated by venous infiltration [36.8/0.74 mm2 (present) vs 64.2/0.74 mm2 (absent), P = 0.020], microsatellite nodules [35.1/0.74 mm2 (present) vs 65.9/0.74 mm2 (absent), P= 0.012], and advanced TNM tumor stage [38.8/0.74 mm2 (stage 3 or 4) vs 68.3/0.74 mm2 (stage 1 or 2), P= 0.014]. No prognostic significance was observed when median values were used as cut-off points using either IMVD-CD105 or IMVD-CD34. However, the presence of the diffuse pattern of CD105 expression in the adjacent non-tumorous liver tissues predicted a poorer disease-free survival (median 8.6 vs 21.5 mo, P= 0.026). CONCLUSION: Our data demonstrate that a lower IMVD-CD105 is associated with larger and more aggressive tumors. In this study, IMVD-CD105 did not provide significant prognostic information. However, active angiogenesis as highlighted by diffuse CD105 staining of the microvessels in the adjacent non-tumorous liver tissues is predictive of early recurrence.     

The Use of 3D Contrast-Enhanced CT Reconstructions to Project Images of Vascular Rings and Coarctation of the Aorta

Background: Aortic arch and pulmonary artery anomalies make up a group of vascular structures that have complex three-dimensional (3D) shapes. Tortuosity as well as hypoplasia or atresia of segments of the aortic arch or pulmonary artery makes the conventional two-dimensional (2D) imaging difficult. Methods: Nine patients with native coarctation or recoarctation and 4 patients with a vascular ring had a CT scan as a part of their clinical evaluation. There were 7 males. The mean age was 11.7 years. (range 19 days to 29 years) The mean weight was 22.7 kg (range 3.3–139.0 kg). The dicom data from contrast CT scans were converted by the Amira software package into a 3D image. The areas of interest were selected. The images were then projected in 3D on a standard video monitor and could be rotated 360° in any dimension. Results: Adequate CT scans and 3D reconstructions were obtained in 12 of 13 patients. There were 85–1,044 slices obtained in the adequate studies. We could not reconstruct a 3D image from a patient's CT scan that had only 22 slices. The anatomy defined by 3D was compared to 2D CT imaging and confirmed by cardiac catheterization or direct visualization in the operating room in the 12 patients with adequate 3D reconstructions. In 5 of 12 patients, 3D reconstructions provided valuable spatial information not observed in the conventional 2D scans. Conclusion: We believe that 3D reconstruction of contrast-enhanced CT scans of these complex structures provides additional valuable information that is helpful in the decision-making process.     

Mixed spindle and epithelioid cell type gastrointestinal stromal tumor of the esophagus: a case report

Fewer than 1 % of gastrointestinal stromal tumors (GISTs) are of the esophagus. This report describes a 63-year-old female diagnosed with mixed spindle/epithelioid cell GIST of the esophagus. She was admitted to our hospital with symptoms of nausea and hematemesis. Preoperative imaging showed a huge submucosal tumor in the lower thoracic and abdominal esophagus. Pathologic examination of an endoscopic biopsy sample suggested squamous cell carcinoma. She underwent subtotal esophagectomy and reconstruction with a gastric tube. Postoperative pathological diagnosis revealed a mixed spindle/epithelioid cell type GIST. The tumor measured 8 × 6 cm, with 30–50 mitotic counts per high power field, immunohistochemical positivity for C-kit (CD117) and CD34 and high risk by modified Fletcher classification. Adjuvant chemotherapy with imatinib mesylate was started 3 months after surgery. Preoperative pathological examination, including staining for CD117 and CD34, of biopsy samples of apparently stromal tumors may be required to rule out rare subtypes of GIST.     

上消化道间叶源性肿瘤临床病理特征46例

目的:观察上消化道间叶源性肿瘤(GIMTs)的临床病理及免疫组化特征,探讨其临床诊断及处置策略.方法:46例内镜超声检查拟诊为上消化道间质瘤的病例,采用光镜观察其手术或黏膜切除术(EMR)标本的病理特征,免疫组化检测其CD117,CD34,平滑肌抗体(SMA)和S-100的表达状况,并分析其病理诊断与临床的关系.结果:46例GIMTs中食管肿瘤占24例,其中间质瘤5例(20.8%),平滑肌瘤19例(79.2%);22例胃GIMTs中间质瘤20例(90.9%),平滑肌瘤1例,神经鞘瘤1例.25例上消化道间质瘤中CD117阳性21例(84%)、CD34阳性24例(96%).而平滑肌瘤与神经鞘瘤分别仅表达SMA和S-100.结论:上消化道间叶源性肿瘤光镜下病理特征相似,联合检测CD117,CD34,SMA和S-100等免疫组化标记物可区别上消化道GIMTs.在食管以平滑肌瘤多见;而在胃则以间质瘤多见,平滑肌瘤与神经鞘瘤非常少见.     

Immunohistochemical Expression of PCNA and CD34 in Colorectal Adenomas and Carcinomas Using Specified Automated Cellular Image Analysis System: A Clinicopathologic Study

Background/Aim:

To evaluate the immunohistochemical expression of proliferating cell nuclear antigen (PCNA) and CD34 in colorectal adenomas and carcinomas, and to correlate this expression with different clinicopathologic parameters.

Materials and Methods:

The study was retrospectively designed. A total of 86 tissue samples, including 33 paraffin blocks from patients with colorectal adenomas, 33 paraffin blocks from patients with colorectal adenocarcinomas, and a control group of 20 samples of nontumerous colonic tissue, were included in the study. From each block, 3 sections of 5 ΅m thickness were taken, 1 section was stained with hematoxylin and eosin (H and E) and the other 2 sections were stained immunohistochemically for PCNA and CD34. Scoring of the immunohistochemical staining was performed using a specified automated cellular image analysis system (Digimizer).

Results:

PCNA expression was significantly increased in a sequence of normal mucosa–adenoma–carcinoma. It was significantly higher in adenomas ≥ 1 cm and those with severe dysplasia, and it showed a significant positive correlation with grade and lymph node involvement in colorectal carcinoma. CD34 showed significantly higher expression in carcinoma than adenoma and in adenoma than in the control group. CD34 expression showed a significant correlation with adenomas carrying severe dysplasia and large-sized adenomas (≥1cm). It was significantly correlated with tumor grade, lymphovascular invasion, and lymph node involvement in colorectal carcinoma.

Conclusion:

PCNA plays an important role in colorectal neoplastic progression and can be utilized as ancillary marker for the risk of malignant transformation in colorectal adenomas as it correlates with high grade dysplasia and size. Intratumoral quantification of the mean (A and N) of CD34 in colorectal carcinoma reflects the grade of tumors and can predict lymph node involvement and lymphovascular invasion, to make a useful additional prognostic factor.     

Novel 3D analysis of Claudin-5 reveals significant endothelial heterogeneity among CNS microvessels

Tight junctions (TJs) feature critically in maintaining the integrity of the blood–brain barrier (BBB), and undergo significant disruption during neuroinflammatory diseases. Accordingly, the expression and distribution of CLN-5, a prominent TJ protein in central nervous system (CNS) microvessels and BBB determinant, has been shown to parallel physiological and pathophysiological changes in microvascular function. However, efforts to quantify CLN-5 within the CNS microvasculature in situ, by using conventional two-dimensional immunohistochemical analysis of thin sections, are encumbered by the tortuosity of capillaries and distorted diameters of inflamed venules. Herein, we describe a novel contour-based 3D image visualization and quantification method, employing high-resolution confocal z-stacks from thick immunofluorescently-stained thoraco-lumbar spinal cord cryosections, to analyze CLN-5 along the junctional regions of different-sized CNS microvascular segments. Analysis was performed on spinal cords of both healthy mice, and mice experiencing experimental autoimmune encephalomyelitis (EAE), an animal model of the neuroinflammatory disease multiple sclerosis. Results indicated that, under normal conditions, the density of CLN-5 staining (CLN-5 intensity/ endothelial surface area) was greatest in the capillaries and smaller venules, and least in the larger venules. This heterogeneity in junctional CLN-5 staining was exacerbated during EAE, as spinal venules revealed a significant loss of junctional CLN-5 staining that was associated with focal leukocyte extravasation, while adjacent capillaries exhibited neither CLN-5 loss nor infiltrating leukocytes. However, despite only venules displaying these behaviors, both capillaries and venules evidenced leakage of IgG during disease, further underscoring the heterogeneity of the inflammatory response in CNS microvessels. This method should be readily adaptable to analyzing other junctional proteins of the CNS and peripheral microvasculature, and serve to highlight their role(s) in health and disease.     

Three-dimensional rotational angiography of the left atrium and esophagus—A virtual computed tomography scan in the electrophysiology lab?

BACKGROUND: Three-dimensional (3D) reconstruction of the heart and surrounding structures has been supplementing traditional two-dimensional imaging to guide diagnostic and therapeutic electrophysiologic procedures. Current methods using computed tomography (CT)/magnetic resonance imaging (MRI) reconstruction have certain limitations. OBJECTIVE: We investigated the feasibility of rotational angiography (RA) combined with simultaneous esophagogram to create an intraprocedural 3D reconstruction of the left atrium (LA) and the esophagus. METHODS: Rotational angiography was performed. Contrast was injected via a pigtail catheter positioned in the left or right pulmonary artery to achieve a levophase venous cycle opacification of the ipsilateral pulmonary veins and adjacent LA. Simultaneous administration of oral contrast allowed a 3D reconstruction of the esophagus in the same image. Qualitative and quantitative comparison between the intraprocedural 3D RA and a remote CT scan was performed in 11 consecutive patients undergoing ablation for atrial fibrillation. RESULTS: Adequate visualization of the pulmonary veins, adjacent posterior LA, and esophagus was achieved in 10 patients. Determination of pulmonary transit time to guide the initiation of RA resulted in better-quality imaging. A close correlation between 3D RA and CT was found. Based on close proximity between the LA and esophagus, the ablation procedure was modified in three patients. CONCLUSIONS: Three-dimensional RA of the LA and esophagus is a promising new method allowing intraprocedural 3D reconstruction of these structures comparable in quality to a CT scan. Further studies refining the method are justified because it could eliminate the need for CT/MRI scans before ablation.     

Pancreatic stellate cells regulate blood vessel density in the stroma of pancreatic ductal adenocarcinoma

Background/objectivesThe vascular heterogeneity of pancreatic ductal adenocarcinoma (PDAC) has never been characterised. We analysed the heterogeneous vascular density of human PDAC along with its prognostic correlation.MethodsTissue Microarrays of 87 patients with different pancreatico-biliary pathologies were analysed in an automated manner (Ariol™) after CD31 staining to assess vascular density in juxta-tumoral and panstromal compartments. In vitro and ex vivo assays were carried out to assess the role of PSC.ResultsPDAC has a distinct vascular density and distribution of vessels compared to cholangiocarcinoma. The PDAC juxta-tumoral stroma was hypovascular and the normal adjacent rim was hypervascular compared to the panstromal compartment. These features adversely affected patient prognosis, suggesting a model for spatio-temporal PDAC evolution. Mice aortic rings and 3D organotypic cultures demonstrated pro- and anti-angiogenic signalling from activated PSC and cancer cells respectively. ATRA-induced quiescence suppressed the pro-angiogenic activity of PSC.ConclusionHuman PDAC has variable vascularity at microscopic level suggesting that novel stromal directed therapies would need to be determined by pathological characteristics.     

Assessment of bronchial inflammation using an automated cell recognition system based on colour analysis.

The aim of this study was to develop an automated system of cell recognition based upon colour analysis suitable for microscopic examination of bronchial inflammation. Human bronchi obtained from 17 patients undergoing thoracotomy were embedded in glycolmethacrylate to perform immunohistochemistry with antibodies against: neutrophil elastase, tryptase, chymase, eosinophil cationic protein, CD68, CD3 and immunoglobulin E. The image analysis system calculates three independent criteria (optic density, hue density, hue) combined with morphological parameters to specifically recognize a positive staining. This automated analysis was applied to the study of bronchial inflammation in smokers and nonsmokers in terms of the absolute number of cells and the expression of different markers by a single cell. The use of these criteria enabled the characterization of a positive stain on single (intraclass correlation coefficient (ICC) = 0.88 or serial (ICC = 0.84) sections. Cell counts obtained by the automated system were highly reproducible. Regarding bronchial inflammation, it was found that the number of inflammatory cells was significantly higher in smokers than in nonsmokers, the majority of these cells bearing immunoglobulin E. These results demonstrate that such computerized analysis of colours is a valuable method for quantifying inflammatory cells in bronchial tissue and for analysing the expression of different markers by a single cell.     

Clinical Application of PET/CT Fusion Imaging for Three-Dimensional Myocardial Scar and Left Ventricular Anatomy during Ventricular Tachycardia Ablation

Background: Image integration has the potential to display three-dimensional (3D) scar anatomy and facilitate substrate characterization for ventricular tachycardia (VT) ablation. However, the current generation of clinical mapping systems cannot display 3D left ventricle (LV) anatomy with embedded 3D scar reconstructions or allow display of border zone and high-resolution anatomic scar features.
Objective: This study reports the first clinical experience with a mapping system allowing an integrated display of 3D LV anatomy with detailed 2D/3D scar and border zone reconstruction.
Methods: Ten patients scheduled for VT ablation underwent contrast-enhanced computed tomography (CT) and Rubidium-82 perfusion/F-18 Fluorodeoxyglucose metabolic Positron Emission Tomography (PET) imaging to reconstruct 3D LV and scar anatomy. LV and scar models were co-registered using a 3D mapping system and analyzed with a 17-segment model. Metabolic thresholding was used to reconstruct the 3D border zone. Real-time display of CT images was performed during ablation.
Results: Co-registration (error 4.3 ± 0.7 mm) allowed simultaneous visualization of 3D LV anatomy and embedded scar and guided additional voltage mapping. Segments containing homogenous or partial scar correlated in 94.4% and 85.7% between voltage maps and 3D PET scar reconstructions, respectively. Voltage-defined scar and normal myocardium had relative FDG uptakes of 40 ± 13% and 89 ± 30% (P < 0.05). The 3D border zone correlated best with a 46% metabolic threshold. Real-time display of registered high-resolution CT images allowed the simultaneous characterization of scar-related anatomic changes.
Conclusion: Integration of PET/CT reconstruction allows simultaneous 3D display of myocardial scar and border zone embedded into the LV anatomy as well as the display of detailed scar anatomy. Multimodality imaging may enable a new image-guided approach to substrate-guided VT ablation.     

Preoperative Assessment of Gastric Cancer Vascularity by Flash Echo Imaging

Background: Tumor vascularity as indicated by immunohistochemical staining is a significant prognostic factor in gastric and other cancers. Non-invasive preoperative assessment of the vascularity of gastric cancers has not been possible. We aim to determine the reliability of harmonic flash echo imaging (FEI) for assessment of vascularity of gastric cancers by comparison with CD34 staining of resected specimens. Methods: Twelve patients undergoing surgical resection of advanced gastric cancer were studied. An ultrasound system transmitting ultrasound pulses at 2.3 MHz and receiving them at 4.6 MHz (second harmonic image) was used for harmonic FEI. Approximately 30 s after intravenous injection of ultrasonic contrast medium (SHU 508A, Levovist), second harmonics (4.6 MHz) emitted from microbubbles were obtained to enhance the B-mode images. Using the tumor image showing strongest enhancement in each FEI series, regions of interest were determined to measure mean echo intensity in the tumor. Immunohistochemistry using antibodies against CD34 was carried out in resected specimens. Tumor vascularity was determined by counting stained microvessels. Results: A significant positive correlation was noted between sonographic amplitude determined preoperatively by FEI analysis and number of CD34-stained microvessels in tumor specimens ( r = 0.869, P = 0.004). Conclusion: Vascularity of gastric cancers now can be evaluated non-invasively by harmonic FEI.     

Preoperative assessment of gastric cancer vascularity by flash echo imaging

BACKGROUND: Tumor vascularity as indicated by immunohistochemical staining is a significant prognostic factor in gastric and other cancers. Non-invasive preoperative assessment of the vascularity of gastric cancers has not been possible. We aim to determine the reliability of harmonic flash echo imaging (FEI) for assessment of vascularity of gastric cancers by comparison with CD34 staining of resected specimens. METHODS: Twelve patients undergoing surgical resection of advanced gastric cancer were studied. An ultrasound system transmitting ultrasound pulses at 2.3 MHz and receiving them at 4.6 MHz (second harmonic image) was used for harmonic FEI. Approximately 30 s after intravenous injection of ultrasonic contrast medium (SHU 508A, Levovist), second harmonics (4.6 MHz) emitted from microbubbles were obtained to enhance the B-mode images. Using the tumor image showing strongest enhancement in each FEI series, regions of interest were determined to measure mean echo intensity in the tumor. Immunohistochemistry using antibodies against CD34 was carried out in resected specimens. Tumor vascularity was determined by counting stained microvessels. RESULTS: A significant positive correlation was noted between sonographic amplitude determined preoperatively by FEI analysis and number of CD34-stained microvessels in tumor specimens (r = 0.869, P = 0.004). CONCLUSION: Vascularity of gastric cancers now can be evaluated non-invasively by harmonic FEI.     

Analysis of microvessels in pancreatic cancer: by light microscopy,confocal laser scan microscopy,and electron microscopy

Background/Purpose

The microvessel density (MVD) of most malignant tumors is considered to be strongly related to metastasis and prognosis. Weidner’s “hot spot method” for determining MVD is in general use, but it is possible that cells other than endothelial cells will also be stained. In our previous study, no correlations were observed between MVD determined by the “hot spot method” and prognosis/metastasis. But, using the “lumen method,” we found a correlation with the number of vessel structures only. In the present study, we analyzed the staining of microvessels in pancreatic cancer, using light microscopy, confocal laser scan microscopy (CLSM), and transmission electron microscopy (TEM).

Methods

Microvessel staining of pancreatic cancer with CD34, factor VIII, and CD45 antibodies was examined in consecutive slices by light microscopy. For CLSM, freshly resected specimens were immunostained with factor VIII and fluorescein isothiocynate. For TEM, specimens were fixed with 2.5% glutaraldehyde, treated with 1% osmium tetroxide, and embedded in epoxy resin.

Results

Staining of vessels with CD34 and factor VIII antibodies appeared similar under light microscopy. However, CD34-stained consecutive slices were judged not to reveal vessel structures, and some cells stained with CD45 antibody were similar in appearance to CD34-stained cells. Under CLSM, irregular arrangements of neovascularization, consisting of many branches, were observed, but many positively stained cells not identified as vessels were also seen. Microvessels were distinctly identified under TEM, but the types of individual cells could not be determined.

Conclusions

An integrated, reproducible method for the measurement of MVD is vital. For pancreatic cancer, the “lumen method” is recommended.

     

Bcl-2 and Bcl-x expression in the CD34+ cells of aplastic anaemia patients: relationship with increased apoptosis and upregulation of Fas antigen

Aplastic anaemia (AA) is a syndrome of haemopoietic failure involving increased apoptosis in stem cells. AA CD34+ cells often have upregulated Fas antigen, but this does not explain the increased apoptosis in all patients. To examine whether abnormal expression of the apoptotic modulators Bcl-2 and Bcl-x is involved in increased apoptosis in the CD34+ cells of patients, we examined cells from 19 AA patients and 18 normal controls by triple staining for CD34, Bcl-2 or Bcl-x, together with 7-amino actinomycin D to determine viability or with staining for Fas antigen. We confirmed increased apoptosis of CD34+ cells in patients. All CD34+ cells in patients and controls expressed Bcl-2 and Bcl-x with no significant difference between the groups. In patients, viability of CD34+/Bcl-2hi cells was similar to that of CD34+/Bcl-2lo cells, but CD34+/Bcl-xhi cells were significantly more viable than CD34+/Bcl-xlo cells. CD34+ cells from AA patients expressed upregulated Fas antigen, but this did not correlate with Bcl-2 or Bcl-x expression. These results suggest a more significant role for Bcl-x as an anti-apoptotic regulator in CD34+ cells in AA than Bcl-2. The induction of death by Fas antigen may bypass the anti-apoptotic effect of Bcl-2 and Bcl-x in CD34+ cells in AA.     

Fully automated assessment of inflammatory cell counts and cytokine expression in bronchial tissue

Automated image analysis of bronchial tissue offers the opportunity to quantify stained area and staining intensity in a standardized way to obtain robust estimates of inflammatory cell counts and cytokine expression from multiple large areas of histopathologic sections. We compared fully automated digital image analysis with interactive digital cell counting and semiquantitative scoring of cytokine expression in terms of repeatability and agreement in bronchial biopsies in 52 patients with mild to moderate atopic asthma. Immunohistochemistry with antibodies against CD3, interleukin (IL)-4, IL-5, and interferon-gamma protein was performed on frozen tissue sections, using 3-amino-9-ethylcarbazole as chromogen and hematoxylin as counterstaining. IL-4 and IL-5 messenger RNAs were localized by in situ hybridization without hematoxylin staining. Separation of 3-amino-9-ethylcarbazole and hematoxylin-stained pixels was achieved by linear combination of red- and blue-filtered gray-scale images. Using baseline biopsy specimens, fully automated CD3+ cell counts showed perfect repeatability (r = 1.0) and a strong linear relationship with the interactive procedure (r = 0.98). Automated densitometry showed perfect repeatability (1.0) and a moderate to strong relationship with semiquantitative scoring of protein and messenger RNA expression (r = 0.43-0.89). Relationships between automated and semiquantitative assessments of changes in cytokine expression during 2 years of follow-up were moderate to strong (r = 0.40-0.84). We conclude that fully automated cell counts and automated densitometric analyses in bronchial tissue of patients with asthma are unbiased and help to reduce variability in inflammatory outcomes.     

Predictive value of microvascular density for response to anlotinib in advanced NSCLC

Nonsmall cell lung cancer (NSCLC) is the most common type of lung cancer. This study aimed to categorize the microvessels in advanced NSCLC and determine the relationship between intratumoral microvascular density (MVD) and the efficacy of anlotinib for NSCLC.The clinical data of 68 patients receiving anlotinib as third-line treatment or beyond for advanced NSCLC were retrospectively collected. Microvessels were stained for CD31 and CD34 by using immunohistochemical staining and were classified as undifferentiated (CD31+ CD34−) and differentiated vessels (CD31+ CD34+). The relationship between MVD and anlotinib efficacy and patient prognosis was analyzed.Patients were divided into the high or low MVD groups according to the median MVD of differentiated (9.4 vessels/field) and undifferentiated microvessels (6.5 vessels/field). There were significantly more patients with high undifferentiated-vessel MVD in the disease control group than in the disease progression group (72.7% vs 16.7%, P < .001). Patients with high undifferentiated-vessel MVD had significantly longer median progression-free survival than those with low undifferentiated-vessel MVD (7.1 vs 3.7 months, P < .001).Anlotinib as third- or beyond line therapy is safe and effective for advanced NSCLC. Patients with a higher density of undifferentiated microvessels have better response to anlotinib and longer progression-free survival.     

Image‐Integration of Intraprocedural Rotational Angiography‐Based 3D Reconstructions of Left Atrium and Pulmonary Veins into Electroanatomical Mapping: Accuracy of a Novel Modality in Atrial Fibrillation Ablation

DynaCT Cardiac Integration into Electroanatomical Mapping. Introduction: Exact visualization of complex left atrial (LA) anatomy is crucial for safety and success rates when performing catheter ablation of atrial fibrillation (AF). The aim of our study was to validate the accuracy of integrating rotational angiography‐based 3‐dimensional (3D) reconstructions of LA and pulmonary vein (PV) anatomy into an electroanatomical mapping (EAM) system. Methods and Results: In 38 patients (62 ± 8 years, 25 females) undergoing catheter ablation of paroxysmal (n = 19) or persistent (n = 19) AF, intraprocedural rotational angiography of LA and PVs was performed. The subsequent 3D reconstruction and segmentation of LA and PVs was transferred to the EAM system and registered to the EAM. The distances of all EAM points to corresponding points on the LA syngo® DynaCT Cardiac surface were calculated. Segmentation of LA with clear visualization of adjacent structures was possible in all patients. Also, the integrated segmentation of the LA was used to guide the encirclement of ipsilateral veins, which resulted in PV isolation in all patients. Integration into the 3D mapping system was achieved with a distance error of 2.2 ± 0.4 mm when compared with the EAM surface. Subgroups with paroxysmal and persistent AF showed distance errors of 2.3 ± 0.3 mm and 2.1 ± 0.4 mm, respectively (P = n.s.). Conclusion: Intraprocedural registration of LA and PV anatomy by contrast enhanced rotational angiography was feasible and accurate. There were no differences between patients with paroxysmal or persistent AF. Therefore, integration of rotational angiography‐based reconstructions into 3D EAM systems might be helpful to guide catheter ablation for AF. (J Cardiovasc Electrophysiol, Vol. 21, pp. 278–283, March 2010)     

Slow transit colon constipation is not related to the number of interstitial cells of Cajal

Background and aims Recent studies have demonstrated decreased numbers of interstitial cells of Cajal in patients suffering from severe chronic constipation as measured by c-Kit (CD117) and CD34 immunohistology. In this study, we wished to determine whether there were abnormalities in the number of neurons of the Auerbach's plexus, their CD117 and CD34 immunoreactivity, or the thickness of colon wall sections in patients with refractory slow transit colonic constipation as compared with control subjects.Patients and methods Specimens from 13 patients who had undergone subtotal colectomy for severe chronic constipation refractory to medical treatment were compared with normal controls. Enteric neurons of Auerbach's plexus were counted, and thickness of the circular and longitudinal layer of the muscularis externa as well as total muscularis externa was measured. Quantitative assessment of anti-CD117 and anti-CD34 immunoreactivity was performed using an Automated Cellular Imaging System and expressed as fractional scores.Results Except for a decreased circular muscle layer thickness in the constipated patients, no statistically significant differences were observed between the two groups. In particular, there was no relationship between CD117/CD34 fractional staining score and the duration or severity of disease, despite the selection of highly symptomatic individuals requiring colonic resection.Conclusion Using quantitative immunohistochemistry for CD117/CD34, we could not detect a relationship between fractional CD117/CD34 staining score and chronic constipation as compared to controls.     

Assessment of coronary compensatory enlargement by three-dimensional intravascular ultrasound

Several techniques have been used to demonstrate that human arteries respond to atherosclerosis by increasing their total arterial area to prevent a decrease in blood flow. Three-dimensional reconstructions of coronary arteries can document this compensatory response accurately and specifically. Seven human coronary arteries were reconstructed using intravascular ultrasound and biplane angiography, and vessel geometries were quantified. In all seven vessels, as plaque area increased, overall vessel area increased (R = 0.986, 0.933, 0.984, 0.678, 0.763, 0.963, and 0.830), but luminal cross-sectional area did not significantly decrease. Focal compensatory enlargement was identified in each vessel, and in some cases this response appeared to occur until the vessel was 65% occluded. Luminal enlargement near the proximal ends was attributed to the natural taper of the vessel. The semi-automated, three-dimensional segmentation technique used in this study allows reproducible quantification, as there is no subjective manual tracing involved. Following the intravascular ultrasound transducer in time and space with biplane angiography allows for accurate reconstruction with or without automated pullback devices. Information on the rate of change of vessel measurements is also presented, which, when combined with visualization of accurate 3D geometry, provides a unique assessment of coronary compensatory enlargement. This reconstruction technique can be applied in a clinical environment with no major modification.