Quantitative analysis of vascular endothelial growth factor, microvascular density and their clinicopathologic features in human hepatocellular carcinoma

BACKGROUND: Angiogenesis is known to be essential to the survival, growth, invasion, and metastasis of tumor cells. Vascular endothelial growth factor (VEGF) are an important angiogenic factor regulating tumor angiogenesis, but its significance and tumor pathologic features are un- clear in hepatocellular carcinoma (HCC). In the present study, we analyzed expression of tissue VEGF, alteration of microvascular density (MVD) in microvessel angiogenesis, development and metastasis of HCC, and level of serum VEGF in differential diagnosis of benign and malignant liv- er diseases. METHODS: Tumor specimens were prospectively collected from HCC patients undergoing resection. Total RNAs were extracted and the expression levels were detected from different parts of HCC tissues. The cellular distributions of VEGF and MVD of liver tumors and their paracancerous and distal cancerous tissues were investigated by streptavi- din peroxidase (S-P) immunohistochemistry, respectively. The VEGF levels of circulating blood and hepatoma tissues were measured by enzyme-linked immunosorbent assay. RESULTS: The incidence of VEGF expression was 63.9% in HCCs (23/36 cases), 78.3% in non-encapsulated HCCs (18/23), and 90.9% in HCCs with extrahepatic metastasis (10/11), respectively. The VEGF expression was tightly correlated with MVD (P <0.01). The MVD in HCC with metastasis, low differentiation or non-encapsulation was significantly higher than that in HCC with intact capsule, high differentiation, or no metastasis. No significant diffe- rence was found between VEGF, MVD, tumor size, and hepatitis virus infection. The level of total RNA in HCC tis- sues was significantly lower but the VEGF level significantly higher than those in paracancerous or distal cancerous ones (P<0.01). The abnormal expression levels of VEGF in sera of HCC patients were directly correlated with the me- tastasis and recurrence of tumors. CONCLUSION: The high expression of VEGF and abnor- mality of tissue MVD are useful predictors for vascular inva- sion and metastasis of liver tumors.

Correlation of Vascular Endothelial Growth Factor and CD105-Microvascular Density in Primary Pterygium

The relationship between the expression of vascular endothelial growth factor（VEGF） and microvascular density（MVD） marked by CD105（CD105-MVD）,and that between CD105-MVD and the clinicopathological characteristics of primary pterygium were investigated.The streptavidin-biotin complex（SABC） immunohistochemical staining in paraffin-embedded tissues was used to detect the expression of VEGF in 23 cases of primary pterygia and 7 normal conjunctival specimens.The antibody against CD105 was used to display vascular endothelial cells,and MVD was examined by counting the CD105-positive vascular endothelial cells.The correlations of VEGF and CD105-MVD,and those of CD105-MVD and clinicopathological data were analyzed by using SPSS 12.0.The expression of VEGF was significantly increased in epithelia（P=0.000）,endothelia and stroma cells（P=0.005） in primary pterygia as compared with normal conjunctivae.The CD105-MVD in pterygia（mean 19.22±6.68） was higher than that in normal conjunctivae（mean 4.00±2.15,P=0.000）.MVD in pterygia was significantly associated with the Tan classification（P=0.000） and the VEGF expression level in the stroma（P=0.020）,but not with sex（P=0.61）,age（P=0.150） or the VEGF expression level in the epithelia（P=0.518）.Our results suggest that over-expression of VEGF and high CD105-MVD in primary pterygium may contribute to the progression by increasing angiogenesis and growth of primary pterygium.

Correlation between expression of two transforming growth factor-beta 1 receptors and microvascular density in a rat model of cerebral ischemia and reperfusion injury

The effects of transforming growth factor-β1 （TGF-β1） are currently controversial. Whether TGF-β1 promotes or inhibits revascularization under different conditions remains poorly understood. Based on previous studies, the current experiment established rat models of cerebral ischemia and reperfusion injury （IRI）, and demonstrated that pathological and functional damage was also increased after IRI. The most serious damage was observed at 3 days after reperfusion, at which time microvascular density fell to its lowest level. Soon afterwards, microvascular density increased, new collateral circulation was gradually established at 4 to 7 days after reperfusion, and pathological damage and neurological deficits were improved. TGF-β1, activin receptor-like kinase 5 （ALK5） mRNA and protein expression levels increased gradually over time. In contrast, ALK1 mRNA and protein expression decreased over the same period. A significant negative correlation was detected between microvascular density and expression of the ALK5 gene transcript. There was no correlation between microvascular density and ALK1 gene transcriptional expression following cerebral IRI in a rat model. These findings suggest that ALK5, rather than ALK1, is the critical receptor in the TGF-β1 signal pathways after cerebral IRI.

Close relationship between mediators of inflammation and pancreatic cancer:Our experience

In this editorial,we focus specifically on the mechanisms by which pancreatic inflammation affects pancreatic cancer.Cancer of the pancreas remains one of the deadliest cancer types.The highest incidence and mortality rates of pancreatic cancer are found in developed countries.Trends of pancreatic cancer incidence and mortality vary considerably worldwide.A better understanding of the etiology and identification of the risk factors is essential for the primary prevention of this disease.Pancreatic tumors are characterized by a complex microenvironment that orchestrates metabolic alterations and supports a milieu of interactions among various cell types within this niche.In this editorial,we highlight the foundational studies that have driven our understanding of these processes.In our experimental center,we have carefully studied the mechanisms of that link pancreatic inflammation and pancreatic cancer.We focused on the role of mast cells(MCs).MCs contain pro-angiogenic factors,including tryptase,that are associated with increased angiogenesis in various tumors.In this editorial,we address the role of MCs in angiogenesis in both pancreatic ductal adenocarcinoma tissue and adjacent normal tissue.The assessment includes the density of c-Kit receptor-positive MCs,the density of tryptase-positive MCs,the area of tryptasepositive MCs,and angiogenesis in terms of microvascularization density.

Microvessel changes in the gerbil hippocampus after cerebral ischemia and reperfusion by Buyang Huanwu decoction pretreatment

Buyang Huanwu decoction （BYHWD） is a classic recipe for the prevention and treatment of ischemic cerebrovascular disease. Gerbils were pretreated with BYHWD, and then subjected to cerebral ischemia and reperfusion. Microvascular changes were determined with laser Doppler monitoring, tannic acid-ferric chloride mordant, and electron microscopy. Results showed that BYHWD pretreatment could enhance the function of hippocampal microvessels, prevent injury, and increase microvasular density and microvasular area density. Thus, these results suggest that BYHWD pretreatment could prevent microvascular occlusion, enhance the capacity of microvascular reperfusion, increase cerebral blood flow, and inhibit neuronal damage, and may be an effective therapy against brain ischemic injury.

Relationship between RGS5 expression and differentiation and angiogenesis of gastric carcinoma

AIM:To explore the regulator of G-protein signaling 5 (RGS5) expression in gastric carcinoma and its association with differentiation and microvascular density (MVD).METHODS:Expression of RGS5 and CD34 were examined in 76 cases of gastric carcinoma,including 22 cases with lymph node metastasis and 54 cases without lymph node metastasis determined by immunohistochemistry (IHC).MVD was assessed using CD34 monoclonal antibody.The presence of RGS5 and CD34 was analyzed by IHC using the Envision technique.RESULTS:The RGS5 expression in gastric carcinoma was positively correlated with the differentiation of the tumor (r=0.345,P < 0.001),but not related with age,gender,tumor size,clinical stage and lymph node metastasis (P > 0.05).The average MVD in the group with lymph node metastasis was significantly higher than that in the group without lymph node metastasis (P < 0.05).RGS5 expression was negatively correlated with the average MVD (P < 0.05).CONCLUSION:RGS5 expression level in gastric carcinoma is associated with the differentiation and MVD of the tumor,and may be used as an important parameter for determining the prognosis of gastric carcinoma patients.

Expression of Nerve Growth Factor and Hypoxia Inducible Factor-1α and Its Correlation with Angiogenesis in Non-Small Cell Lung Cancer

Summary： In order to investigate the expression of nerve growth factor （NGF） and hypoxia inducible factor-1α （HIF-1α） and its correlation with angiogenesis in non-small cell lung cancer （NSCLC）, paraffin-embedded tissue blocks from 20 patients with NSCLC were examined. Twenty corresponding para-cancerous lung tissue specimens were obtained to serve as a control. The expression of NGF, HIF-1α, and vascular endothelial growth factor （VEGF） in the NSCLC tissues was detected by using immunohistochemistry. The microvascular density （MVD） was determined by CD31 staining. The resuits showed that the expression levels ofNGF, HIF-1α and VEGF in the NSCLC tissues were remarkably higher than those in the para-cancerous lung tissues （P〈0.05）. There was significant difference in the MVD between the NSCLC tissues （9.19±1.43） and para-cancerous lung tissues （2.23±1.19） （P〈0.05）. There were positive correlations between NGF and VEGF, between HIF-1α and VEGF, and between NGF and HIF-1α in NSCLC tissues, with the spearman correlation coefficient being 0.588, 0.519 and 0.588, respectively. In NSCLC tissues, the MVD had a positive correlation with the three factors （P〈0.05）. Theses results suggest that NGF and HIF-1α are synergically involved in the angiogenesis of NSCLC.

Inhibitory Effect of Endostar in Combination with Radiotherapy in a Mouse Model of Human CNE2 Nasopharyngeal Carcinoma

The inhibitory effects of Endostar in combination with radiotherapy in BALB/c nude mice model of human CNE2 nasopharyngeal carcinoma and the mechanism were investigated.In nude mice model of CNE2 nasopharyngeal carcinoma,the inhibitory rate and the sensitizing enhancement ratio（E/O） were calculated according to the tumor volumes in different groups.The expression of microvascular density（MVD） in tumor tissues was examined by using immunohistochemistry staining.The transcription of VEGF gene was detected by using RT-PCR.The inhibitory rate in Endostar＋radiotherapy group was higher than in other groups.In Endostar＋radiotherapy group,the tumor volume was significantly decreased and the E/O ratio was 2.335,suggesting that Endostar could be a radiosensitizer.The expression of MVD of tumor tissues in Endostar＋radiotherapy group was reduced significantly.The expression of the MVD in treatment groups was significantly different from that in control group（P〈0.05）.Compared to other groups,VEGF mRNA expression in Endostar＋radiotherapy group was decreased remarkably.Endostar in combination with radiotherapy significantly inhibited the growth of CNE2 tumor.The combination therapy decreased the expression of VEGF,and inhibited tumor angiogenesis and proliferation.When combined with radiotherapy,Endostar acted as a radiosensitizer.

Basic Fibroblast Growth Factor and Fibroblast Growth Factor Receptor-1 in Human Meningiomas

The expression of basic fibroblast growth factor (bFGF) and fibroblast growth factor receptor-1 (FGFR-1) in human meningiomas and the relationships between their expression and the tumors' histological features and angiogenesis were investigated by means of immunohistochemical technique. The expression of bFGF and FGFR-1 was detected by antibody of bFGF or FGFR-1. The tumors' angiogenesis was evaluated by microvascular density (MVD) and, which was observed by use of CD34-antibody immunohistochemically. The results showed that there were varied degrees of the expression of bFGF and FGFR-1 proteins in meningiomas. The expression was correlated with the tumors' histological characters and angiogenesis. It was concluded that bFGF and FGFR-1 might play important roles in meningiomas' angiogenesis and proliferation. The expression positive rate of bFGF and FGFR-1 may provide an indication of evaluating the histological and malignant degree of the tumor.

Interrogating the interplay of angiogenesis and immunity in metastatic colorectal cancer

Colon cancer is the third most common malignancy and the fifth most frequent cause of death from neoplastic disease worldwide.At the time of diagnosis,more than 20%of patients already have metastatic disease.In the last 20 years,the natural course of the disease has changed due to major changes in the management of metastatic disease such as the advent of novel surgical and local therapy approaches as well as the introduction of novel chemotherapy drugs and targeted agents such as anti-epidermal growth factor receptor,anti-BRAF and antiangiogenics.Angiogenesis is a complex biological process of new vessel formation from existing ones and is an integral component of tumor progression supporting cancer cells to grow,proliferate and metastasize.Many molecules are involved in this proangiogenic process,such as vascular endothelial growth factor and its receptors on endothelial cells.A well-standardized methodology that is applied to assess angiogenesis in the tumor microenvironment is microvascular density by using immunohistochemistry with antibodies against endothelial CD31,CD34 and CD105 antigens.Even smaller molecules,such as the micro-RNAs,which are small non-coding RNAs,are being studied for their usefulness as surrogate biomarkers of angiogenesis and prognosis.In this review,we will discuss recent advances regarding the investigation of angiogenesis,the crosstalk between elements of the immune microenvironment and angiogenesis and how a disorganized tumor vessel network affects the trafficking of CD8+T cells in the tumor bed.Furthermore,we will present recent data from clinical trials that combine antiangiogenic therapies with immune checkpoint inhibitors in colorectal cancer.

Lymphatic metastasis is related to the epithelialmesenchymal transition and expressions of VEGF, MMP-9, and COX-2 in breast cancer

The invasion and metastasis of breast cancer are supposed to involve several stages in which epithelial-mesenchymal transition(EMT)is regarded as the mechan-istic basis for the behavior of cancer cells.A series of factors related to EMT are apparently involved in such process.The current study aimed to investigate the contributions of EMT and related factors in lymph node metastasis of breast cancer.The expressions of E-cadherin(E-Cad),N-cadherin(N-Cad),vascular endothelial cell growth factor(VEGF),matrix metalloproteinase-9(MMP-9),cyclooxygenase-2(COX-2),and CD34 were examined in 74 cases of breast cancer,including 39 cases with lymph node metastasis and 35 cases without lymph node metastasis by immunohistochemistry.Multivariable Cox proportional hazards model was used to analyze the patients’prognosis.The expressions of N-Cad,VEGF,MMP-9,and COX-2 in cases with lymph node metastasis were significantly higher than those without lymph node metastasis(P<0.05),while the E-Cad level was inversely related to status of lymph node metastasis(P<0.05).The metastasis rate of lymph node in the cases with EMT(lower E-Cad expression and higher N-Cad expression)was 78.3%,while that without EMT(higher E-Cad expression and lower N-Cad expression)was 11.1%.There was a statistical difference in the expression of COX-2 protein between histological grade I and grade II or III,respectively(P<0.05).In the cases with higher grade,the expression of E-Cad was decreased,while that of N-Cad was increased.Higher microvascular density(MVD)was also found to be significantly associated with lymphatic metastasis(P<0.05),and the cases with higher MVD had shorter survival time.This study indicates that EMT and expressions of VEGF,MMP-9 and COX-2,and MVD value are strongly correlated with lymph node metastasis in breast cancer.

Angiogenesis in acute myeloid leukemia

Angiogenesis is a word that refers to new blood vessel formation,and this process is of fundamental importance for physiological development and tissue homeostasis,as well as the genesis of several diseases,including tumors.Thus,studies carried out in the last years have shown that angiogenesis is essential for the growth of many solid tumors.Angiogenesis is also important for the growth of many hematological malignancies,including acute myeloid leukemia(AML).Endothelial cells are essential constituents of the bone marrow vascular niches,structures essential for the survival and maintenance of normal hematopoietic stem/progenitor cells.Bone marrow endothelial cells play an essential role in leukemia development and there is growing evidence that a targeting of both leukemic and endothelial cells of the leukemic vascular niche may improve the efficacy of antileukemic therapies.Bone marrow angiogenesis is frequently increased in AML,is morphologically evidenced as increased microvascular density,and is typically associated with some AML subtypes.The molecular mechanisms underlying the increased angiogenesis in some AML subtypes have been defined.In conclusion,a better understanding of angiogenesis as well as the fundamental interactions between bone marrow endothelial cells and leukemic stem cells may contribute to improve antileukemia treatments.

A review of functional slit lamp biomicroscopy

Functional slit lamp biomicroscopy(FSLB)is a novel device which consists of a traditional slit-lamp and a digital camera.It can quantitatively assess vessel diameter,blood flow velocity,and blood flow rate and can create noninvasive microvascular perfusion maps(nMPMs).At present,FSLB is mainly used in contact lens(CL)and dry eye disease(DED)studies to advance our understanding of ocular surface microcirculation.FSLB-derived blood flow and vessel density measures are significantly altered in CL wearers and DED patients compared to normal people.These subtle changes in the ocular surface microcirculation may contribute to the monitoring of potential diseases of the body and provide a new way to diagnose dry eye disease.Therefore,this may also indicate that FSLB can be more widely applied in the study of other diseases to reveal the relationship between changes in ocular surface microcirculation and systemic diseases.The purpose of this paper is to summarize the functions of FSLB and the related studies especially in CL and DED.