Study On The Occurrence And Development Of Placenta Derived Endothelial Progenitor Cell Initiation Of Infantile Hemangioma

RESEARCH BACKGROUNG Infantile hemangioma(IH)is the most common benign tumor in infants and young children.Neonatal morbidity as high as 10-12%,about 60% of the lesions occurred in the head and neck,severe cases can affect the child's appearance,involvement of organ function,mental health.Without interferometric treatment,most IH patients grow rapidly within one year of age and then slowly degenerate into fibrous adipose tissue during childhood.But about 10% of the IH because of its growing too fast,the area is too large or the growth of special,may lead to disfigurement,deformity,dysfunction and even life-threatening.But the current characteristics of IH proliferation,regression mechanism has not yet elucidated,resulting in its clinical treatment is not satisfactory.At present,the mechanism of the occurrence of IH is mainly focused on the origin of hemangioma endothelial cells.The main hypotheses include genetics and gene mutation theory,placental chorionic endothelial cell theory and stem cell theory,but there is still no theory It is a good idea to elucidate the clinical features of early hyperplasia of IH and spontaneous regression.In the past,IH was initially activated in vascular endothelial cells,but in 2008 Khan et al reported the first report from the IH tissue to extract hemangioma stem cells,confirmed the presence of stem cells in IH.In the previous study,it was found that the patients with hemangioma who had just been delivered were found in the placenta of the placenta,and the endothelial progenitor cells were found in the tissues.The results of HE staining showed that IH and placental hemangioma The morphology of the pathology was similar,while the expression of endothelial progenitor cells was not observed in IH tissues.Vascular endothelial progenitor cells are a kind of stem cells,which can differentiate into vascular endothelial cells,and have the ability of self-renewal,infinite proliferation and multi-directional differentiation,similar to the clinical manifestations of proliferative hemangioma.In summary,this study speculate: vascular sprouting infant hemangiomas,vascular endothelial progenitor cells by placental tissue,mobilization,migration,colonization and settlement in abnormal embryonic fetus certain part,after the baby is born,because the placenta growth factor inhibition produced is released,under the action of estrogen factors,various cytokines,particularly the environmental changes and other signals,selective activation of endothelial progenitor cells,activated endothelial progenitor cells proliferate,which filopodia along angiopoietin Growth factor secretion of adequate local migration,the formation of vascular branches,buds vascular collateral formation of vascular plexus,leading to hemangioma hyperplasia,hemangioma endothelial cell proliferation,rapid blood vessels born(born ~ 1 year old);then a series of signals expression under control of its own path,h EPCs gradually decreased to differentiate into endothelial cells,resting and enters the IH(after 1 year),after changing infant microenvironment in vivo results hemangiomas gradually fibrous connective tissue and adipose tissue replaced,resulting in regression of specific(5 to 10 years);we believe that this push The occurrence of hemangiomas,development,clinical manifestations subside in line with IH,more reasonable explanation.At present,the excessive proliferation of endothelial cells,rapid growth of blood vessels is the biggest feature of IH histopathology.The growth and development of neovascularization is accomplished by a series of vascular growth factors through many signaling pathways.The traditional view that angiogenesis exists only in adults,and angiogenesis is limited to embryos.With the success of h EPCs in adult isolation,it is shown that angiogenesis also occurs in adults.Therefore,angiogenesis,angiogenesis may be the basic process of human angiogenesis.HEPCs are endothelial cells of the precursor cells,morphological can not be h EPCs and other cells to distinguish,so mainly by molecular markers.At present the consensus,CD34,CD133 common mark that is h EPCs. So,a variety of cytokines,the internal environment signal changes is how to regulate the occurrence and development of hemangioma it? At present,in other tumor studies also found that vascular endothelial cells and stromal cells secrete angiogenic factors and angiogenic factors,the common regulation of vascular angiogenesis and vascular formation.Vascular neovascularization is due to vascular endothelial cells in the vascular endothelial growth factor(VEGF)and many other cytokines induced migration and proliferation,leading to large proliferation of blood vessels.VEGF is currently the strongest angiogenic factor found in the neovascularization process of central regulatory factors and vascular endothelial cell-specific mitosis,a variety of tumor animal model studies have confirmed that blocking VEGF signaling pathway can reduce tumor microvessels Density,thereby inhibiting tumor growth.Studies have shown that DLL4 /Notch signaling pathway overexpression,can produce non-functional blood vessels,lumen thinner,blood flow was significantly reduced,leading to tumor ischemia,hypoxia,thereby inhibiting tumor hyperplasia.VEGF can induce DLL4 expression,activation of DLL4 /Notch signaling pathway,suggesting that VEGF on the DLL4/Notch signaling pathway is a positive regulatory role.The activation of this signaling pathway attenuates endothelial-induced angiogenesis in endothelial cells,suggesting that it may feedback to inhibit the functional activity of VEGF signaling pathway.Therefore,only in the vascular and DLL4/Notch signaling pathway under the coordination of hemangioma angiogenesis can be carried out in an orderly manner.OBJECTIVE For further study to confirm the above conjecture,this study intends to,1,in the different stages of IH and placental hemangioma,the detection of CD34 /CD133double-labeled h EPCs expression level,confirmed proliferation and placental hemangioma tissue h EPCs was significantly higher expression,and low expression or low expression of extinction,The results showed that h EPCs were closely related to the development and progression of IH,to confirmed that the development of placental origin h EPCs began to develop IH.2,at different stages of IH,immunohistochemical detection of CD34,CD133,VEGF,DLL4 expression levels,confirmed that VEGF-DLL4 /Notch signaling pathway involved in the regulation of infantile hemangioma angiogenesis.METHODS1.Collection of 50 cases of infants hemangioma in Department of Pathology of the Fifth Affiliated Hospital of Guangzhou Medical University from 2014 to 2017.Including 22 males and 28 females,aged 1 to 2 years old,an average of 1.2 years old,Patients were not given any adjuvant therapy before surgery.All specimens were subjected to routine HE staining,Immunohistochemical SP method for detection of PCNA,according to the Mulliken classification criteria and combined with the expression of PCNA:There were 26 cases of proliferative and 24 cases of degenerative infantile hemangioma in paraffin specimens.Immunohistochemical SP method was used to detect the expression of F?RAg,VEGF,DLL4 and CD34 in infantile hemangioma Proliferative and Hyperplasia period.Quantitative analysis of VEGF and DLL4 expression using the HPIAS-1000 High Definition Color Pathology Report Management System.The measured average optical density,the positive reaction area and the total area of the cells.To provide the basis for the next experiment.2.Collection of sample that born after the discovery of patients with hemangioma in Department of Obstetrics and Gynecology of the First Affiliated Hospital of Guangzhou Medical University from 2014 to 2017.whitin 30 minutes that delivery of the placenta,under aseptic operation,cut the placenta chorion frondosum.Samples were taken from 15 cases after routine HE staining,identification of infants with placental hemangioma in 5cases,including 3 cases of male and female 2 cases,After receiving the consent of the family,5 cases of normal placental tissue were obtained,as a negative control group.Immunohistochemistry and immunofluorescence were used to identify the presence of human endothelial progenitor cells in placental hemangioma and proliferative phase of infantile hemangioma,at the same time discover the expression levels of CD34,CD133 and DLL4.RESULT1.The results of immunohistochemistry showed that the expression of VEGF,DLL4 and CD34 in the proliferative phase was significantly higher than that in the degenerative group and normal skin group(P<0.05),And the difference between the after two groups was not statistically significant(P>0.05).VEGF expression of positive hemangioma MVD was37.6±1.6,VEGF expression of negative hemangioma MVD was 16.4±2.0,the difference between the two very significant.The difference between the two has a very significant significance(t=4.87,P<0.01).Correlation analysis of VEGF and DLL4 positive area ratios showed that VEGF and DLL4 in the proliferative and degenerative IH tissues were positively correlated,r=0.821,P=0.003;DLL4:r=0.830,P=0.003.2.CD34,CD133 and DLL4 were mainly located in the medial villi of placenta,and the positive results showed diffuse distribution of brownish granules,CD34,CD133 and DLL4 were significant.CD34,CD133 and DLL4 positive rate and negative rate,the difference was significant(P <0.01).Immunofluorescence method,CD34 fluorescent staining was green,CD133 was red,double staining was yellow or yellowish green fluorescence,nuclear staining DAPI was blue,to further confirm the above experimental results.CONCLUSIONS1.The expression of VEGF,DLL4 and CD34 in the proliferative phase of infantile hemangioma was high,but low or no expression at the time of extinction,and there are significant differences between proliferative and extinction phase,Suggesting that VEGF-DLL4/Notch signaling pathway involved in the regulation of the development of infantile hemangioma.2.CD34,CD133,DLL4 were high expression in the proliferative phase of infantile hemangioma and placental hemangioma tissue.the expression of endothelial progenitor cells which was mark by CD34 and CD133 staining,at the same time,high expression in the proliferative phase of infantile hemangioma and placental hemangioma tissue.the expression of CD34,CD133,DLL4 and ndothelial progenitor cells low or no expression at the time of extinction,These results suggest that endothelial progenitor cells play an important role in the development of infants hemangioma tissue.