Expression of prolactin gene and secretion of prolactin by rat retinal capillary endothelial cells.
OBJECTIVE
Prolactin fragments inhibit blood vessel formation, whereas anti-prolactin antibodies induce angiogenesis in the cornea. Endothelial cells from brain capillaries and the umbilical vein produce prolactin, and this study was undertaken to determine whether retinal capillary endothelial cells could be a source for prolactin in the eye.


METHODS
Primary cultures of rat retinal endothelial cells were investigated for the expression of prolactin mRNA by reverse transcription-polymerase chain reaction (RT-PCR) and Southern blot analysis and by in situ hybridization. The prolactin protein was analyzed by immunocytochemistry, enzyme-linked immunoabsorbent assay, Western blot analysis, and the Nb2-cell bioassay. The effect of prolactin and the 16-kDa prolactin fragment on retinal endothelial cell proliferation was investigated, and the expression of the cloned prolactin receptor was analyzed by RT-PCR and Southern blot analysis.


RESULTS
Retinal endothelial cells expressed prolactin mRNA and full-length 23-kDa prolactin. Prolactin was observed in the cytoplasm of cells and in their conditioned medium at levels 300 times those described in endothelial cells from other vessels and species. Exogenous 16-kDa prolactin inhibited rat retinal endothelial cell proliferation, whereas 23-kDa prolactin was inactive. No evidence was obtained for the expression of the cloned prolactin receptor in these cells, but the prolactin receptor was amplified in whole rat retina.


CONCLUSIONS
Endothelial cells from the microcirculation of rat retina produce and release prolactin. That the cloned prolactin receptor was not expressed in these cells argues against direct autocrine effects of prolactin. Possible paracrine effects are suggested by the expression of the prolactin receptor in retinal tissue.
Ochoa
A
A
Neurobiology Center, National Autonomous University of Mexico, Querétaro, Mexico.
Montes de Oca
P
P
Rivera
J C
JC
Dueñas
Z
Z
Nava
G
G
de La Escalera
G M
GM
Clapp
C
C
D000818:Animals; D015139:Blotting, Southern; D015153:Blotting, Western; D002196:Capillaries / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D019084:Fluorescent Antibody Technique, Indirect; D015870:Gene Expression; D017403:In Situ Hybridization; D011388:Prolactin / Q000096:biosynthesis* / Q000235:genetics* / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D011817:Rabbits; D051381:Rats; D011981:Receptors, Prolactin / Q000378:metabolism; D012171:Retinal Vessels / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D012333:RNA, Messenger; D011981:Receptors, Prolactin; D011388:Prolactin