Increased synthesis of leukotrienes in the mouse model of diabetic retinopathy.
OBJECTIVE
Evidence suggests that capillary degeneration in early diabetic retinopathy results from chronic inflammation, and leukotrienes have been implicated in this process. The authors investigated the cellular sources of leukotriene biosynthesis in diabetic retinas and the effects of hyperglycemia on leukotriene production.


METHODS
Retinas and bone marrow cells were collected from diabetic and nondiabetic mice. Mouse retinal glial cells and retinal endothelial cells (mRECs) were cultured under nondiabetic and diabetic conditions. Production of leukotriene metabolites was assessed by mass spectrometry, and Western blot analysis was used to quantitate the expression of enzymes and receptors involved in leukotriene synthesis and signaling.


RESULTS
Bone marrow cells from nondiabetic mice expressed 5-lipoxygenase, the enzyme required for the initiation of leukotriene synthesis, and produced leukotriene B(4) (LTB(4)) when stimulated with the calcium ionophore A23187. Notably, LTB(4) synthesis was increased threefold over normal (P < 0.03) in bone marrow cells from diabetic mice. In contrast, retinas from nondiabetic or diabetic mice produced neither leukotrienes nor 5-lipoxygenase mRNA. Despite an inability to initiate leukotriene biosynthesis, the addition of exogenous leukotriene A(4) (LTA(4); the precursor of LTB(4)) to retinas resulted in robust production of LTB(4). Similarly, retinal glial cells synthesized LTB(4) from LTA(4), whereas mRECs produced both LTB(4) and the cysteinyl leukotrienes. Culturing the retinal cells in high-glucose concentrations enhanced leukotriene synthesis and selectively increased expression of the LTB(4) receptor BLT1. Antagonism of the BLT1 receptor inhibited LTB(4)-induced mREC cell death.


CONCLUSIONS
Transcellular delivery of LTA(4) from marrow-derived cells to retinal cells results in the generation of LTB(4) and the death of endothelial cells and, thus, might contribute to chronic inflammation and retinopathy in diabetes.
Talahalli
Ramaprasad
R
Department of Pediatrics, Case Western Reserve University/Rainbow Babies and Children's Hospital, Cleveland, Ohio 44106, USA.
Zarini
Simona
S
Sheibani
Nader
N
Murphy
Robert C
RC
Gubitosi-Klug
Rose A
RA
D000818:Animals; D017209:Apoptosis; D001094:Arachidonate 5-Lipoxygenase / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D001854:Bone Marrow Cells / Q000187:drug effects / Q000378:metabolism; D000001:Calcimycin / Q000494:pharmacology; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D003921:Diabetes Mellitus, Experimental / Q000378:metabolism* / Q000473:pathology; D003930:Diabetic Retinopathy / Q000378:metabolism* / Q000473:pathology; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D005947:Glucose / Q000494:pharmacology; D006943:Hyperglycemia / Q000378:metabolism*; D017572:Leukotriene A4 / Q000494:pharmacology; D007975:Leukotriene B4 / Q000096:biosynthesis*; D013058:Mass Spectrometry; D051379:Mice; D008810:Mice, Inbred C57BL; D009457:Neuroglia / Q000187:drug effects / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D018102:Receptors, Leukotriene B4 / Q000378:metabolism; D012160:Retina / Q000473:pathology; D012171:Retinal Vessels / Q000473:pathology; D020133:Reverse Transcriptase Polymerase Chain Reaction
D017572:Leukotriene A4; C503704:Ltb4r1 protein, mouse; D012333:RNA, Messenger; D018102:Receptors, Leukotriene B4; D007975:Leukotriene B4; D000001:Calcimycin; D001094:Arachidonate 5-Lipoxygenase; D005947:Glucose