Determination Of Microvessel Density Research Articles

Loss of Gucy1a3 causes poor post-stroke recovery by reducing angiogenesis via the HIF-1α/VEGFA signaling pathway in mice

ObjectivesIschemic stroke is a common and debilitating disease that can cause permanent neurological damage. Gucy1a3, which encodes the α1 subunit of soluble guanylyl cyclase, has been reported to be associated with functional recovery after ischemic stroke. However, the mechanism is still not well understood. In the present study, we investigated the effects of Gucy1a3 on (i) post-stroke recovery; (ii) vascular endothelial growth factor A (VEGFA) and hypoxia inducible factor 1 alpha (HIF-1α) expression; and (iii) angiogenesis after ischemic stroke. Materials and methodsWild-type and Gucy1a3 knockout C57BL/6J male mice were respectively used to establish the models of permanent middle cerebral artery occlusion (pMCAO). Neurological deficit scores were evaluated at 24 h and 96 h after pMCAO. Cerebral infarct volume was measured by 2,3,5-triphenyltetrazolium chloride (TTC) staining. For determining microvessel density, immunohistochemical analysis was performed with CD31. The expression of VEGFA and HIF-1α was detected by western blotting. ResultsOur results suggest that loss of Gucy1a3 increased the infarct volume and aggravated neurological deficits after pMCAO. In addition, the Gucy1a3 knockout brains exhibited significantly lower microvessel densities and VEGFA and HIF-1α expression levels than the wild-type brains at 96 h post-pMCAO. ConclusionsOur study indicates that GUCY1A3 might be involved in angiogenesis after ischemic stroke. Further investigation of GUCY1A3 will provide a new therapeutic target for stroke.

Determination of Microvessel Density by CD34 Immunoreactivity in Female Breast Cancer and its Relation with Grading and Staging

Background: Angiogenesis is a basic process that enables neoplasm to thrive. Microvessel density (MVD) evaluation is an accepted parameter for assessing the angiogenesis process within a tumour. The research was focused to evaluate angiogenesis by using CD34 immunomarker in invasive breast cancer and to correlate the microvessel density with grades and stages. Objective: To determine microvessel density by CD34 immunoreactivity in female breast cancer and its relation with grading and staging. Methods: This cross-sectional descriptive type of study was conducted in the Department of Pathology, Rajshahi Medical College. Forty-five untreated cases of breast cancer were included in this study between the period of July 2019 to June 2021 and paraffin embedded sections were obtained from representative mastectomy specimen. The sections were stained with hematoxylin and eosin stain followed by evaluation of angiogenesis by using CD34 antibody. Results: In this study, most of the patients (71.1%) belonged to age < 50 years where the mean age was found 47.5 + 12.4 years. In 45 cases, tumour sizes ranged from 1-8 cm. 86.7% (39 cases) ranges from 0-5 cm, where only (6 cases) 13.3% were more than 5 cm. About 30 cases (66.7%) lymph nodes were positive for metastasis. According to histopathological grade 51.1% belonged to grade III followed by 28.9% grade II and 20% grade I. Histological stage showed most of the cases was stage II (48.9 %) followed by stage I (26.7 %) then stage III (24.4 %). Upon statistical analysis, a significant relation was obtained between MVD with increasing histologic grades and stages. Conclusion: As a new prognostic marker it would be of great value in diagnosis and in identifying patients at high risk of tumor recurrence more accurately. By this way breast cancer patients might be benefited from adjuvant therapies.

Prognostic and predictive significance of VEGF, CD31, and Ang-1 in patients with metastatic clear cell renal cell carcinoma treated with first-line sunitinib.

The most common type of renal cell carcinoma (RCC) is clear cell renal cell carcinoma (ccRCC), which has a high metastatic potential. Even though the International Metastatic RCC Database Consortium (IMDC) risk model is conventionally utilized for selection and stratification of patients with metastatic RCC (mRCC), there remains an unmet demand for novel prognostic and predictive markers. The goal of this study was to analyze the expression of Vascular endothelial growth factor (VEGF), Cluster of Differentiation 31 (CD31) to determine microvessel density, and Angiopoietin-1 (Ang-1) in primary kidney tumors, as well as their predictive and prognostic value in patients with metastatic ccRCC (mccRCC) who were treated with first-line sunitinib. The study included 35 mccRCC patients who were treated with first-line sunitinib in period between 2009 and 2019. Immunofluorescence was used to examine biomarker expression in tissue specimens of the primary tumor and surrounding normal kidney tissue. Median disease-free survival (DFS) was longer in patients with negative and low tumor VEGF score than in patients with medium tumor VEGF score (p=0.02). Those with low tumor CD31 expression had a longer median DFS than patients with high tumor CD31 expression (p=0.019). There was no correlation between Ang-1 expression and DFS. The expression of biomarkers in normal kidney tissue was significantly lower than in tumor tissue (p<0.001). In conclusion, higher VEGF scores and greater CD31 expression were associated with longer DFS, but neither of these biomarkers correlated with progression-free survival or overall survival.

Immunohistochemical and immunofluorescence expression profile of lymphatic endothelial cell markers in oral cancer.

Lymphangiogenesis makes an important contribution to the tumour microenvironment (TME), but little is known about this in oral squamous cell carcinoma (OSCC). Archival formalin-fixed paraffin-embedded specimens (28 OSCC, 10 inflamed and 6 normal oral mucosa controls) were processed using immunohistochemistry (IHC) with antibodies against lymphatic markers D2-40 (podoplanin), LYVE-1, VEGFR3 and Prox1. After the endothelial cells had been highlighted by the various markers for lymphatic endothelium, the positive stained cells and vessels were identified and counted in a systematic manner to determine microvessel density. Double-labelling immunofluorescence (DLIF) was used to investigate the specificity of D2-40 and LYVE-1 to lymphatic endothelial cells (LECs) as opposed to blood ECs. There was higher D2-40 and Prox1 lymphatic vessel density (P=.001) in the OSCC group when compared with both control groups. Some malignant keratinocytes expressed lymphatic markers, as did a much smaller number of epithelial cells in the control groups. DLIF showed that no vessels co-expressed D2-40/CD34 or LYVE/CD34. Some D2/40+ LVs were LYVE- . D2-40 was the most specific LEC marker in OSCC tissues. These results establish that the OSCC TME contains significantly more lymphatic vessels expressing D2-40 and Prox1 than the control groups, which may play a role in facilitating lymphatic invasion and metastases.

Endothelial Cells of a Normal Liver and with Hepatocellular Carcinoma

The article presents a systematic review of the literature data on the role of endothelial cells and features of vascularization in the development of hepatocellular carcinoma. The capillaries of the normal liver are represented by sinusoids, which are characterized by the presence of specific fenestrations in endothelial cells and the absence of a basal membrane under the endothelium. The article shows that, as a rule, the development of hepatocellular carcinoma occurs upon chronic liver lesions, and it is a multistage process of the progression of tissue and cellular atypism, as well as changes in the vascularization of tumor tissue. The vascular network that forms in the tumor tissue is characterized by structural and functional atypism. The development and progression of liver carcinoma is accompanied by changes in the structure and metabolism of endothelial cells in the tumor node, as well as chromosomal aberrations with impaired gene expression and growth factors. The processes of angiogenesis and vascularization are based on the morphological determination of microvessel density on immunohistochemical specimens. The clinical diagnosis of hepatocellular carcinoma is based on the detection of radiation characteristics of the tumor, including those due to changes in vascularization of the tumor tissue. An increase in the malignancy grade of hepatocellular carcinoma is accompanied by a decrease in blood flow through the portal vein system and, accordingly, an increase in arterial blood flow. It is noted that knowledge of the processes of tumor angiogenesis is required to develop antitumor, targeted, antiangiogenic drugs.

Microvessels in tumor tissue in patients with central nervous system tumors

Angiogenesis plays an essential role in the development of a tumor. The most preferred histological evaluation method of this process is the determination of microvessel density (MVD) in the biopsy. Numerous studies report the possibility of using various MRI techniques for imaging the network of neoplastic vessels and determining the angiogenesis in them, as well as the subsequent analysis of the obtained images. In this study, a classic MRI imaging with intravenous contrast agent administration was used, and in order to ensure objectification and repeatability, and to eliminate human error that may occur in classic MRI image analysis, an automatic evaluation algorithm was used to determine the degree of angiogenesis, which was then presented in the form of a statistical pixels analysis. The study involved 48 adult patients of both sexes, treated surgically at the Department of Neurosurgery, Nicolaus Copernicus University Collegium Medicum in Bydgoszcz, due to a solid brain tumor. The highest mean density of MVD microvessels in tumor tissue was observed in high-grade gliomas. The high level of MVD expression in tumors of the central nervous system showed a positive correlation with the high degree of tumor malignancy. Based on the conducted research, it was concluded that the determination of the level of MVD expression in the tumor tissue is an additional prognostic factor in neoplasms of the central nervous system. The comparison of the results obtained with the tests performed routinely translates into a more accurate selection of treatment and assessment of the prognosis for survival.

MicroRNA-16 Inhibits Glioblastoma Growth in Orthotopic Model by Targeting Cyclin D1 and WIP1.

IntroductionTo examine the molecular mechanism by which miRNA-16 (miR-16) suppresses glioblastoma in vitro and in vivo.MethodsGene expression of miR-16 in normal brain tissues and human glioma cell lines was examined. To characterize the functional role of miR-16 in vitro, miR-16 was ectopically expressed in U87 cells by lentiviral transduction. Expression of miR-16 downstream targets cyclin D1 and Bcl-2 in U87 was studied using Western blotting. Cell proliferation and clonogenic property were examined using CCK-8 and clone formation assay, respectively. Migration and invasiveness of U87 was studied using wound-healing assay and transwell assay, respectively. In vivo tumorigenic properties of the miR-16-transduced U87 cells were examined in an orthotopic xenograft model. Immunohistochemistry was performed to examine cyclin D1, WIP1 and CD31 expressions.ResultsExpression of miR-16 was reduced in glioblastoma cell lines compared to normal human brain tissues. Ectopic miR-16 expression reduced cyclin D1 and Bcl-2 in U87 cells. miR-16 also induced apoptosis, reduced cell proliferation and clone formation. Furthermore, miR-16 suppressed U87 migration in wound-healing assay and invasion across transwell membrane in vitro. In an orthotopic tumor model, overexpression of miR-16 inhibited tumor growth in vivo was accompanied with reduction in cyclin D1 and WIP1 expression in the xenografts. CD31 expression in miR-16-overexpressed xenografts was also decreased. The determined microvessel density of the miR-16 overexpression group was significantly lower than those groups treated with vehicle and empty vector.DiscussionMicroRNA-16 exhibits inhibitory effects of glioblastoma. MicroRNA-16 and its downstream targets could be potential therapeutic targets for treatment of glioblastoma.

Nasal polyps in eosinophilic granulomatosis with polyangiitis: Structured histopathology and CD105 expression.

Distinguishing the prodromal nasal polyposis of eosinophilic granulomatosis with polyangiitis (EGPA) from chronic rhinosinusitis with nasal polyps (CRSwNP) is a challenge for rhinologists and rheumatologists. It has recently been reported that angiogenesis and CD105 expressed on vascular endothelial cells could have a role in the pathogenesis and development of nasal polyps. This exploratory study examined the structured histopathology of nasal polyps in patients with EGPA and CRSwNP, comparing CD105 expression in their nasal tissue with that of a control group with no chronic sinonasal inflammation. A structured histopathological study was performed on surgical specimens of nasal tissue from 32 adults (13 with EGPA, 14 with CRSwNP, 5 controls), considering CD105 as a marker to determine microvessel density (MVD). The mean eosinophil count was higher in EGPA patients with tissue inflammation (p=.002), and in CRSwNP patients with sub-epithelial edema (p=.009). Neutrophil infiltration was significantly associated with severe tissue inflammation in EGPA patients (p=.04), but with the absence of fibrosis in CRSwNP patients (p=.04). In the EGPA group, CD105-MVD correlated with tissue eosinophil count (p=.05). Mean CD105-MVD was significantly higher in EGPA patients with mucosal ulceration (p=.004). In the CRSwNP group, a CD105-MVD correlated positively and significantly with tissue eosinophil count (p=.01). Alongside the known abundance of eosinophils, other cells might contribute to inflammatory processes. Neutrophils may amplify inflammation, eosinophil recruitment and tissue damage. CD105 expression in CRSwNP and EGPA nasal polyps supports the hypothesized involvement of angiogenesis in the pathogenesis and development of nasal polyps.

Endorectal power Doppler ultrasonography is a reliable method for evaluation of rectal cancer angiogenesis.

We aimed to assess the preoperative rectal cancer angiogenesis with Endorectal Power Doppler Ultrasonography by using the Power Doppler Vascularity Index (PDVI) calculated by imaging analysis software, and to compare it with the microvessel density (MVD) in surgical specimens PATIENTS AND METHODS: This study included 110 patients (39 females; mean age 61.5 years) with rectal cancer. Immunohistochemical staining of surgical specimens with anti-CD-31 antibody was used for MVD evaluation. The PDVI of each tumor was calculated using Endorectal Power Doppler with computer-assisted quantification of colour pixels. Mean MVD - 163 ± 69 microvessels/mm2 (50-328) was used as a cutoff point, differentiating two groups of tumors with high (> 160 mm2) and low (≤ 160 mm2) angiogenic activity. Mean PDVI of 8.9 ± 6.0% (0-27.3) was used as a cutoff point, dividing two groups of tumors with high (> 8%) and low (≤ 8%) PDVI. The MVD and the PDVI showed a good positive correlation (r = 0.438, p = 0.002). Patients with low PDVI had 25 months longer overall survival (p < 0.05) than patients with high PDVI. Patients with low MVD had 36 months longer survival (p < 0.05). Endorectal Power Doppler Ultrasonography is a reliable and noninvasive method for assessment of the extent of rectal cancer angiogenesis. Tumor angiogenesis assessed by the PDVI correlated with histological MVD determination and could predict survival rates. Endorectal Power Doppler examination is a useful and reproducible method for in vivo preoperative quantitative assessment of tumor vascularization.

Abstract B095: Antitumor activity of lenvatinib mesilate (LEN) via angiogenesis inhibition and tumor FGF signaling pathway in the human hepatocellular carcinoma models

Abstract Background: LEN is an oral multi-targeted tyrosine kinase inhibitor that selectively inhibits vascular endothelial growth factor receptor (VEGFR)1-3, fibroblast growth factor receptor (FGFR)1-4, platelet-derived growth factor receptor α, RET, and KIT. LEN was noninferior to sorafenib (SOR) in overall survival (OS) in a phase 3 study in patients with unresectable hepatocellular carcinoma (HCC) (medians: LEN, 13.6 vs SOR, 12.3 months; hazard ratio: 0.92; 95% confidence interval, 0.79-1.06) and statistically superior to SOR in progression-free survival (PFS), time to progression (TTP), and objective response rate (ORR). We previously reported that LEN showed antitumor activity in multiple HCC xenograft models including patient-derived xenograft (PDx) models. Here, we investigated the mode of action of LEN in preclinical human HCC models to examine antitumor angiogenesis activity in vivo and the effects on FGF signaling pathways both in vitro and in vivo in the preclinical HCC models. Methods: Antiproliferative activity of LEN and SOR was examined using human HCC cells (Hep 3B2.1-7, HuH-7, SNU398, and PLC/PRF/5) in vitro, and antitumor and anti-angiogenesis activity were examined using these HCC cell lines and PDx models. The effect on the FGF signaling pathway was evaluated by determining the phosphorylation of FRS2, a direct substrate of FGFRs, with immunoblotting analysis in both cultured cells and tumor xenografts. Anti-angiogenesis activity was evaluated by determining microvessel density (MVD) within tumor xenografts by immunohistochemical analysis using anti-CD31 antibody. Result: LEN decreased the phosphorylation of FRS2 in FGF19-overexpressing Hep3B2.1-7 and HuH-7 cells at 0.03 to 3 μmol/L in a concentration-dependent manner. LEN showed selective antiproliferative activity against these cells with half-maximal inhibitory concentration (IC50) values of 0.23 and 0.42 μmol/L, respectively, and those concentrations were consistent with ones for inhibitions of the phosphorylation of FRS2. In tumor xenografts, LEN also decreased the phosphorylation of FRS2 by the single administration of LEN between 3 and 30 mg/kg. SNU398 cells are known to have a relatively weak dependence on the FGF signaling pathway without FGF19 expression. LEN inhibited in vitro proliferation of SNU398 cells, and the phosphorylation of FRS2 both in vitro and in vivo at higher doses than in Hep 3B2.1-7 and HuH-7 cells. SOR did not selectively inhibit in vitro proliferation or the phosphorylation of FRS2 in HCC cells with FGF activation in vitro and in vivo. In HCC PDx and PLC/PRF/5 xenograft models, LEN significantly decreased the MVD in a dose-dependent manner between 3 and 30 mg/kg, and it was correlated with tumor growth inhibition in each models. SOR showed clear anti-angiogenesis activity at 30 mg/kg, but not at 10 mg/kg. Conclusion: These results suggest that inhibition of the FGF signaling pathway via FGFR with LEN affects proliferation of HCC cells with activation of the FGF signaling pathway, and the potent anti-angiogenic activity underlies the antitumor activity of LEN in preclinical HCC xenograft models. Citation Format: Taisuke Hoshi, Masahiro Matsuki, Yuji Yamamoto, Yukinori Minoshima, Junji Matsui, Yasuhiro Funahashi. Antitumor activity of lenvatinib mesilate (LEN) via angiogenesis inhibition and tumor FGF signaling pathway in the human hepatocellular carcinoma models [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr B095.

Iodine Concentration in Spectral CT: Assessment of Prognostic Determinants in Patients With Gastric Adenocarcinoma.

The purpose of this study was to use virtual monochromatic spectral CT to investigate the usefulness of iodine concentration (IC) and its correlation with clinicopathologically determined prognostic factors in gastric adenocarcinoma. From June 2012 to March 2015, 34 patients with gastric adenocarcinoma underwent arterial and portal venous phase spectral CT. The ICs in the arterial and portal venous phases were calculated and then normalized with the aorta as normalized IC (NIC). The surgical specimen was evaluated with CD34 staining to determine microvessel density (MVD). The correlation between imaging results and clinicopathologic findings was investigated for histologic grading, lymph node metastasis, serosal involvement, distant metastasis, pathologic TNM stage, and MVD. The mean arterial phase NIC value of tumors was 0.12 ± 0.03, portal venous phase NIC value was 0.39 ± 0.06, and MVD was 26.94 ± 7.87 vessels per high-power field (×400). Both arterial phase and portal venous phase NIC values were significantly higher in poorly differentiated gastric adenocarcinomas (p = 0.005) than in moderately differentiated tumors (p = 0.013). There was no significant correlation between NIC and serosal involvement or distant metastasis. There was significant correlation between the NIC and MVD in gastric adenocarcinoma (arterial phase NIC, p = 0.013; portal venous phase NIC, p = 0.001). However, neither the arterial nor the portal venous phase NIC of gastric adenocarcinoma had a significant relation to lymphatic metastasis or pathologic TNM stage. There was a significant difference between the high and low MVD groups with respect to portal venous phase NIC (p = 0.045). NIC can serve as a useful predictor of angiogenesis and degree of differentiation of moderately and poorly differentiated gastric adenocarcinomas.

Role of Small Interfering RNA Silencing Protein Kinase C-α Gene on the Occurrence of Ultrafiltration Failure in Peritoneal Dialysis Rats.

This study aims to explore the effect of PKC-α gene silencing on the occurrence of ultrafiltration failure (UFF) in peritoneal dialysis (PD) rats. Forty-eight male SD rats were collected to establish 5/6 renal resection uremic and uremic PD rats models. Rats were assigned into control, sham operation, uremia, PD-2 W (peritoneal dialysis for 2 weeks), PD-4 W (peritoneal dialysis for 4 weeks), negative control (NC) (peritoneal dialysis for 4 weeks, and injected 0.1 mg/kg blank plasmid into abdominal cavity) and PKC-α siRNA (peritoneal dialysis for 4 weeks, and injected 0.1 mg/kg PKC-α siRNA into abdominal cavity) groups. CD34 staining was performed to determine microvessel density (MVD) for peritoneal tissues. The mRNA and protein expression of PKC-α in peritoneal tissue were detected by qRT-PCR and Western blot. Compared with the control group, MVD, the mRNA and protein expression of PKC-α were significantly increased in rats of the uremia, PD-2 W, PD-4 W, NC, and PKC-α siRNA groups. Compared with the uremia group, MVD, the mRNA and protein expression of PKC-α were increased, the changes observed in the PD-4 W and NC groups were better obvious than in the PD-2 W group. In comparison with the PD-4 W and NC groups, MVD, the mRNA and protein expression of PKC-α in rats were decreased in the PKC-α siRNA group. PKC-α gene has a high expression in uremic PD rats, and PKC-α gene silencing is able to increase UF while decrease MVD and glucose transport in peritoneal tissues thus reversing UFF in PD rats. J. Cell. Biochem. 118: 4607-4616, 2017. © 2017 Wiley Periodicals, Inc.

MP22-07 CORRELATION OF CAD PEAK LESION ENHANCEMENT WITH QUANTITATIVE TUMOR ANGIOGENESIS TO NON-INVASIVELY ASSESS FURHMAN GRADES I-IV IN PATIENTS WITH CLEAR CELL RENAL CELL CARCINOMA

You have accessJournal of UrologyKidney Cancer: Epidemiology & Evaluation/Staging I1 Apr 2017MP22-07 CORRELATION OF CAD PEAK LESION ENHANCEMENT WITH QUANTITATIVE TUMOR ANGIOGENESIS TO NON-INVASIVELY ASSESS FURHMAN GRADES I-IV IN PATIENTS WITH CLEAR CELL RENAL CELL CARCINOMA Heidi Coy, Jonathan Young, Michael Douek, Matthew Brown, Anthony Sisk, James Sayre, and Steven Raman Heidi CoyHeidi Coy More articles by this author , Jonathan YoungJonathan Young More articles by this author , Michael DouekMichael Douek More articles by this author , Matthew BrownMatthew Brown More articles by this author , Anthony SiskAnthony Sisk More articles by this author , James SayreJames Sayre More articles by this author , and Steven RamanSteven Raman More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.661AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES To assess if Computer Aided Detection (CAD) of peak lesion attenuation discriminates among Fuhrman Grades I-IV and correlates with an increase in tumor angiogenesis in clear cell RCC (ccRCC) on four-phase MDCT. METHODS We reviewed a cohort of patients with ccRCC and preoperative multiphasic multidetector CT imaged with a 4-phase renal mass protocol (unenhanced, corticomedullary (C), nephrographic (N), and excretory (E)). A whole lesion 3D contour was obtained in all phases with proprietary software. The CAD algorithm determined a 0.5cm diameter region of peak enhancement ≤300HU within the 3D lesion contour. For assessment of quantitative angiogenesis, immunohistochemical staining for CD34 to determine microvessel density (MVD) was performed. T-tests were used to compare peak multiphasic enhancement and microvessel density among Fuhrman grades I-IV. P values less than 0.05 were considered to be significant. RESULTS 107 patients (71(64%) men and 40(35%) women) with 111 unique ccRCC lesions (16 (14%) Fuhrman grade I, 64 (58%) Fuhrman grade II, 23 (21%) Fuhrman Grade III, 8 (7%) Fuhrman grade IV) were analyzed. In the C phase, CAD peak lesion enhancement discriminated grade I from II (150 HU vs. 185 HU, p=0.006), I from III (150 HU vs. 178 HU, p=0.054), I from IV (150 HU vs 229HU, p<0.001). This directly correlated with an increase in quantitative angiogenesis (MVD): I from II (2134 mm2 vs. 4710 mm2, p=0.004), I from III (2134 mm2 vs. 5162 mm, p=0.001), I from IV (2134 mm2 vs 6076 mm2, p=0.057). CONCLUSIONS CAD peak lesion enhancement discriminates Fuhrman grades 1-IV on multiphasic CT and correlates with an increase in tumor angiogenesis. This may be helpful to triage patients to active surveillance, interventional therapy or, if validated, may be useful to monitor results with anti-angiogenic therapy. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e259 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Heidi Coy More articles by this author Jonathan Young More articles by this author Michael Douek More articles by this author Matthew Brown More articles by this author Anthony Sisk More articles by this author James Sayre More articles by this author Steven Raman More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Genetic Features of Aflatoxin-Associated Hepatocellular Carcinoma

Dietary exposure to aflatoxin is an important risk factor for hepatocellular carcinoma (HCC). However, little is known about the genomic features and mutations of aflatoxin-associated HCCs compared with HCCs not associated with aflatoxin exposure. We investigated the genetic features of aflatoxin-associated HCC that can be used to differentiate them from HCCs not associated with this carcinogen. We obtained HCC tumor tissues and matched non-tumor liver tissues from 49 patients, collected from 1990 through 2016, at the Qidong Liver Cancer Hospital Institute in China-a high-risk region for aflatoxin exposure (38.2% of food samples test positive for aflatoxin contamination). Somatic variants were identified using GATK Best Practices Pipeline. We validated part of the mutations from whole-genome sequencing and whole-exome sequencing by Sanger sequencing. We also analyzed genomes of 1072 HCCs, obtained from 5 datasets from China, the United States, France, and Japan. Mutations in 49 aflatoxin-associated HCCs and 1072 HCCs from other regions were analyzed using the Wellcome Trust Sanger Institute mutational signatures framework with non-negative matrix factorization. The mutation landscape and mutational signatures from the aflatoxin-associated HCC and HCC samples from general population were compared. We identified genetic features of aflatoxin-associated HCC, and used these to identify aflatoxin-associated HCCs in datasets from other regions. Tumor samples were analyzed by immunohistochemistry to determine microvessel density and levels of CD34 and CD274 (PD-L1). Aflatoxin-associated HCCs frequently contained C>A transversions, the sequence motif GCN, and strand bias. In addition to previously reported mutations in TP53, we found frequent mutations in the adhesion G protein-coupled receptor B1 gene (ADGRB1), which were associated with increased capillary density of tumor tissue. Aflatoxin-associated HCC tissues contained high-level potential mutation-associated neoantigens, and many infiltrating lymphocytes and tumors cells that expressed PD-L1, compared to HCCs not associated with aflatoxin. Of the HCCs from China, 9.8% contained the aflatoxin-associated genetic features, whereas 0.4%-3.5% of HCCs from other regions contained these genetic features. We identified specific genetic and mutation features of HCCs associated with aflatoxin exposure, including mutations in ADGRB1, compared to HCCs from general populations. We associated these mutations with increased vascularization and expression of PD-L1 in HCC tissues. These findings might be used to identify patients with HCC due to aflatoxin exposure, and select therapies.

Treatment of novel IL17A inhibitor in glioblastoma implementing 3rd generation co-culture cell line and patient-derived tumor model

Despite many treatment options, cancer remains a growing problem and has become the second leading cause of death globally. Here, we present fluorescence molecular tomography (FMT) data regarding the reversion of third generation co-cultured U87+DBTRG and patient-derived GBM tumor model after treatment with novel IL17A inhibitor named FLVM and FLVZ (organic derivatives of caffeic acid). FMT was used to determine tumor angiogenesis volume (assessment of number of blood vessel; the expression of angiogenic factors CD34 and other angiogenic cancer bio-markers) in U87+DBTRG and patient-derived gliomas. Immunohistochemistry was used to determine microvessel density [CD34], and cell proliferation [Ki67]. Western blot was used to assess the interleukin 17A [IL17A], vascular endothelial growth factor [VEGF] and hypoxia-inducible factor-1α [HIF-1α]. Antibody array was used to assess the cancer bio-markers in co-cultured U87+DBTRG gliomas. Animal survival was found to be significantly increased (P<0.0001) after FLVM treatment compared with control-IL17A. After FMT detection, FLVM, administered orally, was found to decrease tumor growth (P<0.0001). FLVM and FLVZ administration resulted in significant decreases in tumor hypoxia [HIF-1α (P<0.05)], angiogenesis [CD34 (P<0.05)], VEGF, IL17A and cell proliferation [Ki67 (P<0.05)] and caused a significant increase of Bax, caspase and FasL (P<0.05), compared with untreated animals. Additionally, Leptin, LPL (P<0.01), FFA (P<0.05) and adipogenesis were downregulated and no additive toxicity was found in mice except calorie-restriction like effect. Use of FLVM can be considered as a novel inhibitor of IL17A for the treatment of human gliomas.

Danshensu, a major water-soluble component of Salvia miltiorrhiza, enhances the radioresponse for Lewis Lung Carcinoma xenografts in mice.

The molecule 3-(3,4-dihydroxyphenyl)-2-hydroxypropanoic acid (danshensu), a herbal preparation used in traditional Chinese medicine, has been found to possess potential antitumor and anti-angiogenesis effects. The aim of the present study was to investigate the efficacy of the combination of radiation therapy (RT) with danshensu in the treatment of Lewis lung carcinoma (LLC) xenografts, whilst exploring and evaluating the mechanism involved. In total, 8-week old female C57BL/6J mice were randomly assigned into 3 groups to receive: RT, RT + cisplatin and RT + danshensu, respectively, when LLC reached 100–150 mm3. Each group was divided into 7 subgroups according to the different irradiation doses that were administered. Tumor growth curves were created and the sensitization enhancement ratios of the drugs were calculated. The experiment was then repeated, and the 4 groups of tumor-bearing mice were treated with natural saline, danshensu, RT + danshensu and RT, respectively. The mice were sacrificed on day 7, and tumor tissue and blood were collected to determine microvessel density, the expression of proangiogenic factors, and the levels of blood thromboxane B2 and 6-keto-prostaglandin-F1α. Tumor hypoxia was also detected using in vivo fluorescence imaging. With respect to LLC xenografts, treatment with danshensu + RT significantly enhanced the effects of tumor growth inhibition (P<0.05). Furthermore, tumor vasculature was remodeled and microcirculation was improved, which significantly reduced tumor hypoxia (P<0.05). The present study demonstrated that danshensu significantly enhanced the radioresponse of LLC xenografts in mice. The mechanism involved may be associated with the alleviation of tumor cell hypoxia following treatment with danshensu + RT, caused by the improvement of tumor microcirculation and the remodeling of tumor vasculature.

Reconsideration of the clinical and histopathological significance of angiogenesis in prostate cancer: Usefulness and limitations of microvessel density measurement.

Angiogenesis plays important roles in tumor growth and cancer cell dissemination in almost all cancers. In prostate cancer, there is general agreement that increased angiogenesis is an important factor in determining tumor development and prognosis in these patients. Microvessel density is recognized as a useful marker for evaluating the angiogenic status of cancer tissues. Many investigators have reported that microvessel density is significantly associated with pathological features and outcomes in prostate cancer patients; however, some researchers have expressed opposing opinions. As the reason for such discrepancy, previous reports have suggested differences in the methodologies of measuring microvessel density in cancer tissues. In the present review, we focus on the variation in such methods, including the selected area and the method used for (semi)quantification. In particular, we discuss the relationship between malignancy potential, tumor progression, and survival and differences in the antibodies used for detection of endothelial cells in detail. We briefly compare the pathological significance and prognostic roles of microvessel density measured using von Willebrand factor, CD31, CD34, and CD105. Based on these analyses, the advantages and limitations of microvessel density measurements in prostate cancer tissues are clarified. Improved "real" and "specific" markers of cancer-related angiogenesis are necessary for better predictions of prognoses and for discussion of treatment strategies for patients with prostate cancer. However, establishment of a satisfactory cancer-related endothelial marker could take a long time. Therefore, knowledge regarding the pathological significance of microvessel density - based on understanding of the advantages and limitations of microvessel density determination methods - is important.

Effects of acitretin combined with clarithromycin on tumor growth and angiogenesis in human oral epidermoid carcinoma xenografts in nude mice

Objective To evaluate the effects of acitretin combined with clarithromycin on tumor growth in human oral epidermoid carcinoma xenografts in nude mice, and to investigate their antitumor mechanisms. Methods A cell line of human oral epidermoid carcinoma was subcutaneously inoculated into 31 Balb/c nude mice to establish a xenograft model of human skin tumor. Then, the nude mice were randomly classified into 6 groups according to a double blind protocol: control group（n = 6）remaining untreated, placebo group（n = 5）treated with wheat flour, acitretin group（n = 5）treated with acitretin 7.2 mg/kg per day, clarithromycin group（n = 5）treated with clarithromycin 100 mg/kg per day, acitretin + placebo group（n = 5）treated with both acitretin（7.2 mg/kg per day）and wheat flour, and acitretin + clarithromycin group（n = 5）treated with acitretin（7.2 mg/kg per day）and clarithromycin 100 mg/kg per day. All the drugs were intragastrically administrated once daily. After three weeks of treatment, mice were sacrificed and xenografts were removed. Then, the size and weight of xenografts were measured, and pathological analysis was conducted. Real time-PCR was performed to quantify the mRNA expressions of vascular endothelial growth factor（VEGF）and nuclear factor（NF）-κB, and immunohistochemistry was carried out to observe the expression of VEGF as well as to determine microvessel density（MVD）and Ki-67 proliferation index. By using the software SPSS 19.0, analysis of variance was performed for comparison of measurement data, and least significant difference（LSD）test for paired comparisons. Results Both the size and weight of xenografts in the acitretin + clarithromycin group were significantly lower than those in the other groups（all P < 0.05）. Real-time fluorescence-based PCR revealed weaker mRNA expressions of VEGF and NF-κB in the acitretin + clarithromycin group compared with the control group, clarithromycin group and acitretin group（all P < 0.05）. As immunohistochemistry showed, the acitretin + clarithromycin group displayed a decrease in the expression rate（all P < 0.01）and staining intensity of VEGF, MVD（all P < 0.01）with a sparse distribution of microvessels, Ki-67 proliferation index（all P < 0.05）and proliferative activity of tumor cells compared with the control group, clarithromycin group and acitretin group. Conclusion Acitretin combined with clarithromycin can synergistically inhibit the growth of human oral epidermoid carcinoma xenografts in nude mice, downregulate VEGF expression, and suppress angiogenesis and tumor proliferation. Key words: Xenograft model antitumor assays; Acitretin; Clarithromycin; Carcinoma, squamous cell; Vascular endothelial growth factors; NF-kappa B; Ki-67

The Role of Bevacizumab in the Management of Head and Neck Squamous Cell Carcinoma Patients

Angiogenesis is a crucial step in tumor growth and metastasis. Clinical association of tumor vascularity with tumor aggressiveness has been clearly demonstrated in a wide variety of tumor types including head and neck squamous cell carcinoma (HNSCC) [1]. Thus, determination of microvessel density in tumor tissue can be useful in estimation of prognosis. Inhibition of angiogenesis can repress the growth rate of tumor cells, and also induce apoptosis due to reduced nutrition and oxygen supply to the tumors. Vascular endothelial growth factor (VEGF), which is a major element for many angiogenic processes, binds VEGF receptor (VEGFR) to stimulate endothelial cell proliferation and migration [2,3]. Thus, determination of microvessel density in tumor tissue can be useful as a predictive factor for the effectiveness of bevacizumab in chemoradiotherapeutic strategies.

Clinicopathological significance of ASC amino acid transporter-2 expression in pancreatic ductal carcinoma.

ASC amino acid transporter-2 (ASCT2) is highly expressed in cancer cells. However, the clinicopathological significance of ASCT2 expression in pancreatic cancer remains unclear. The aim of this study was to investigate the clinical significance of ASCT2 expression in pancreatic cancer. Ninety-seven patients with surgically resected pancreatic ductal adenocarcinoma were evaluated. Tumour sections were stained by immunohistochemistry for ASCT2, Ki67, CD34 (to determine microvessel density), phospho-AKT (p-AKT) and phospho-mammalian target of rapamycin (p-mTOR) expression. ASCT2 was expressed in 54% (52/97) of tumours. Statistically significant differences in patient age, T stage, N stage, lymphatic permeation, vascular invasion, Ki67, and CD34 and p-mTOR expression were observed between tumours with and without ASCT2 expression. Multivariate analysis confirmed that vascular invasion, ASCT2 expression and Ki67 expression were independent predictive factors for a poorer prognosis. ASCT2 expression plays an important role in tumour cell growth, and is a promising pathological marker for predicting a worse outcome in pancreatic cancer.