The association between laminin and microglial morphology in vitro

Abstract

Microglia are immune cells in the central nervous system (CNS) that contribute to primary innate immune responses. The morphology of microglia is closely associated with their functional activities. The majority of microglial studies have focused on the ramified or amoeboid morphology; however, bipolar/rod-shaped microglia have recently received much attention. Bipolar/rod-shaped microglia form trains with end-to-end alignment in injured brains and retinae, which is proposed as an important mechanism in CNS repair. We previously established a cell culture model system to enrich bipolar/rod-shaped microglia simply by growing primary microglia on scratched poly-D-lysine (PDL)/laminin-coated surfaces. Here, we investigated the role of laminin in morphological changes of microglia. Bipolar/rod-shaped microglia trains were transiently formed on scratched surfaces without PDL/laminin coating, but the microglia alignment disappeared after 3 days in culture. Amoeboid microglia digested the surrounding laminin, and the gene and protein expression of laminin-cleaving genes Adam9 and Ctss was up-regulated. Interestingly, lipopolysaccharide (LPS)-induced transformation from bipolar/rod-shaped into amoeboid microglia increased the expression of Adam9 and Ctss, and the expression of these genes in LPS-treated amoeboid-enriched cultures remained unchanged. These results indicate a strong association between laminin and morphological transformation of microglia, shedding new light on the role of bipolar/rod-shaped microglia in CNS repair.

Laminins and their receptors in the CNS

“…Microglia are brain-resident immune cells that mediate immune responses in the CNS (Kettenmann et al, 2011). Previous studies show that they exist in two states: a resting state characterized by ramified morphology and an activated state characterized by amoeboid morphology (Laskaris et al, 2016;Tam, Au & Ma, 2016). Recently, bipolar/rod-shaped microglia, which play an important role in CNS repair, have been described (Wierzba-Bobrowicz et al, 2002;Ziebell et al, 2012;Taylor et al, 2014).…”

The Effect of Laminin Surface Modification of Electrospun Silica Nanofiber Substrate on Neuronal Tissue Engineering

“…After 6 days, 12 days and 18 days in 0.010 M PBS, fiber morphology was observed with SEM. After 12 days and 18 days in DMEM, the distribution of the laminin coating on the as-prepared substrates was examined by the immunocytochemistry method with rabbit anti-laminin primary antibody [ 42 ]. First, the substrates for non-specific labeling were blocked with 10% goat serum (Thermo Fisher Scientific).…”

“…PC12 cell viability was determined using a LIVE/DEAD assay kit (Life Technologies, Waltham, MA, USA), which contained Calcein AM and EthD-1 dyes at a concentration of 1 μM. For the determination, PC12 cells were seeded on the as-prepared SNF substrates then assayed after 72 h cultivation, according to the procedure reported previously [ 24 , 42 ]. The dyes diluted in PBS were added to the cultivated cells for 10 mins then 12 images at different positions ( n = 12) were taken for each sample utilizing the fluorescence microscope (Nikon).…”

A Single-Step Surface Modification of Electrospun Silica Nanofibers Using a Silica Binding Protein Fused with an RGD Motif for Enhanced PC12 Cell Growth and Differentiation

“…PC12 cell viability was determined using a Live/Dead assay kit (Life Technologies, Waltham, MA, USA). The PC12 cells were first seeded on the as-prepared SNF substrates then assayed after 72 h of cultivation [ 15 , 34 ]. Calcein AM and Ethidium Homodimer-1 dyes diluted in PBS to a final concentration of 1 μM were added to the cultivated cells for 10 min.…”