| Studies on the Intercellular Communication and Detection of Secretory Proteins of Endothelial Cells of Human Umbilical Blood VesselsAbstractIt is well known that endothelial cells play an important role on the regulation of blood vessels. Blood vessels can be classified into two types according to theirinnervation:theinnervatedbloodvesselsandthenon-innervated blood vessels. The regulating mechanisms of the two kinds of blood vessels are different, and endothelial cells play differential roles on them. However, models of human innervated and non-innervated blood vessels have seldom been reported, and the regulating mechanism of non-innervated blood vessels is not clear yet. In order to establish models of human innervated and non-innervated blood vessels, we investigated the fine structure, ultrastructure and innervation of human inferior mensenteric artery?s branches and human umbilical blood vessels by using HE staining, electron microscopy and immunohistochemistry. Then we explored the regulating mechanism of non-innervated blood vessels by immunohistochemistry, immunofluorescence, RT-PCR, scrape-loading and dye transferring methods.It had been reported that different kinds of vasoactive substances can be secreted by endothelial cells, but the nature of many of them were not completely clear. To better understand the function of endothelium in non-innervated blood vessels, it is necessary to assay the gross secretionof endothel ia1 ce11s. Therefore, in part IIIof' thi s research,we co11ected the serum--free condit ioned medi um of humanumbi 1 ica1 vein endothe1ia1 ce11s (HUVEC), then i so1ated,purified and ana1yzed its componen ts by1 ow--t emperature--vacuum--dry ing me t hod, prot e i n1 ow--t emperature---chromat ograghy, cap i 1 l ary e 1 e c trophore s i s.The main resu1ts are as fo11ows:1. Though human inferior mensenteri c artery's branch andhuman umbi1ica1 artery had equa1 diameter, their finestructure were qui te di fferent. Under e1ec tron 111 i croscope,the endothelia1 ce11s in human umbi1 ica1 vein seenlcd to havemore rough endop1asmi c ret i cu1 um but 1 ess smooth endop1asmi creticu1um than those of the endothe1 ia1 ce11 s in humanumbi 1 ica1 artery and human inferior mensenteric tlrtery ) sb r a n c h.2. Mul t ip1e pepti dergic and monoaminergic nerve fi berssuch as VIP, SP, NPY, 5--HT nerve fibers were tound byimmunohi stochemistry staining in human inferi or mensenter icartery, s branch, whi1e none of these nerve fibers were foundboth in human umbi1 ica1 artery and vein. This suggested thatinferior mensenteric artery could be cons idered as arepresentative mode1 of human innervated b1ood vesse1 s, andumbi1 ica1 b1ood vesse1s cou1d be used ds a reprosentativemode l of human non--i nnervated b1ood vesse1 s.3. Expression of eN0S, V1P, SP, NPY was founC1 in HUVECboth in vivo and in vitro, which suggested endothe l ial ce11scou1d regu1ate non--innervated b1ood vessel s v iaautocr ining/paracr ini ng mu1 t ipl e vasoact i ve subs tances.IV4. Expression of both protein and n1RNA of Cx43 wasdetected in cu1tured HUVEC by immunof1uorescence and RT--PCR.And in scrape--1oading and dye transferring experiment,Lucifer Ye11ow CH, a sma11 mo1ecu1e of f1uorescent dye, wasobserved to transfer from scrape--1oaded ce11s to norma1 ce11 s,which provided an important morpho1ogic evidence to supportdirect communication between HUVEC v ia gap junct i ons.5. In serum--free conditioned medium of ecv304 ce11s (aHUVEC ce1l 1ine), multip1e proteins were detected. Three kindof them had been purified and ana1yzed, their molecu1arweight were 13, 17, 70kD respectively. 0ur study suggestedthat two proteins which weighted 13, 17kD cou1d faci 1 itatethe prol iferation of endothe1 ia1 ce1l, and the other oneweighted 70kD seemed to have no effect on endothe1 ia1 cel 1'sgr ow t h.These resu1ts showed that inferior mensenteri c arterycould be considered as a representative mo... |
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