Inflamed In Vitro Retina: Cytotoxic Neuroinflammation and Galectin-3 Expression.
BACKGROUND
Disease progression in retinal neurodegeneration is strongly correlated to immune cell activation, which may have either a neuroprotective or neurotoxic effect. Increased knowledge about the immune response profile and retinal neurodegeneration may lead to candidate targets for treatments. Therefore, we have used the explanted retina as a model to explore the immune response and expression of the immune modulator galectin-3 (Gal-3), induced by the cultivation per se and after additional immune stimulation with lipopolysaccharide (LPS), and how this correlates with retinal neurotoxicity.


METHODS
Post-natal mouse retinas were cultured in a defined medium. One group was stimulated with LPS (100 ng/ml, 24 h). Retinal architecture, apoptotic cell death, and micro- and macroglial activity were studied at the time of cultivation (0 days in vitro (DIV)) and at 3, 4 and 7 DIV using morphological staining, biochemical- and immunohistochemical techniques.


RESULTS
Our results show that sustained activation of macro- and microglia, characterized by no detectable cytokine release and limited expression of Gal-3, is not further inducing apoptosis additional to the axotomy-induced apoptosis in innermost nuclear layer. An elevated immune response was detected after LPS stimulation, as demonstrated primarily by release of immune mediators (i.e. interleukin 2 (IL-2), IL-6, KC/GRO (also known as CLCX1) and tumour necrosis factor-α (TNF-α)), increased numbers of microglia displaying morphologies of late activation stages as well as Gal-3 expression. This was accompanied with increased apoptosis in the two additional nuclear layers, and damage to retinal gross architecture.


CONCLUSIONS
We demonstrate that an immune response characterized by sustained and increased release of cytokines, along with an increase in Gal-3 expression, is accompanied by significant increased neurotoxicity in the explanted retina. Further investigations using the current setting may lead to increased understanding on the mechanisms involved in neuronal loss in retinal neurodegenerations.
Dept. of Biology, Sec. Functional Zoology, Lund University, Lund, Sweden.|Dept. Clinical Sciences in Lund, Div. Ophthalmology, Lund University, Lund, Sweden.
Patrik Maximilian
PM
Bauer
Dept. Clinical Sciences in Lund, Div. Ophthalmology, Lund University, Lund, Sweden.
Marina Castro
MC
Zalis
Dept. Clinical Sciences in Lund, Div. Ophthalmology, Lund University, Lund, Sweden.
Hodan
H
Abdshill
Dept. Experimental Medical Science, Experimental Neuroinflammation Laboratory, Lund University, Lund, Sweden.
Tomas
T
Deierborg
Dept. of Biology, Sec. Functional Zoology, Lund University, Lund, Sweden.
Fredrik
F
Johansson
Dept. Clinical Sciences in Lund, Div. Ophthalmology, Lund University, Lund, Sweden.
Ulrica
U
Englund-Johansson
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002135:Calcium-Binding Proteins / Q000378:metabolism; D053331:Ectodysplasins / Q000378:metabolism; D037502:Galectin 3 / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects*; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D066298:In Vitro Techniques; D007249:Inflammation / Q000139:chemically induced; D007376:Interleukin-2 / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D019394:Ki-67 Antigen / Q000378:metabolism; D008070:Lipopolysaccharides / Q000633:toxicity; D051379:Mice; D008840:Microfilament Proteins / Q000378:metabolism; D017628:Microglia / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D009474:Neurons / Q000187:drug effects*; D012160:Retina / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
C491049:Aif1 protein, mouse; D002135:Calcium-Binding Proteins; D053331:Ectodysplasins; C506019:Eda protein, mouse; D037502:Galectin 3; D005904:Glial Fibrillary Acidic Protein; D007376:Interleukin-2; D015850:Interleukin-6; D019394:Ki-67 Antigen; D008070:Lipopolysaccharides; D008840:Microfilament Proteins; D014409:Tumor Necrosis Factor-alpha; C528900:glial fibrillary astrocytic protein, mouse