Background: Mesencephalic astrocyte-derived neurotrophic factor (MANF), a 20 kDa secreted protein, was originally derived from a rat mesencephalic type-1 astrocyte cell line. MANF belongs to a novel evolutionally conserved family of neurotrophic factors along with conserved dopamine neurotrophic factor. In recent years, ever-increasing evidence has shown that both of them play a remarkable protective role against various injuries to neurons in vivo or in vitro. However, the characteristics of MANF expression in the different types of glial cells, especially in astrocytes, remain unclear.
Methods: The model of focal cerebral ischemia was induced by rat middle cerebral artery occlusion. Double-labeled immunofluorescent staining was used to identify the types of neural cells expressing MANF. Primarily cultured glial cells were used to detect the response of glial cells to endoplasmic reticulum stress stimulation. Propidium iodide staining was used to determine dead cells. Reverse transcription PCR and western blotting were used to detect the levels of mRNA and proteins.
Results: We found that MANF was predominantly expressed in neurons in both normal and ischemic cortex. Despite its name, MANF was poorly expressed in glial cells, including astrocytes, in normal brain tissue. However, the expression of MANF was upregulated in the glial cells under focal cerebral ischemia, including the astrocytes. This expression was also induced by several endoplasmic reticulum stress inducers and nutrient deprivation in cultured primary glial cells. The most interesting phenomenon observed in this study was the pattern of MANF expression in the microglia. The expression of MANF was closely associated with the morphology and state of microglia, accompanied by the upregulation of BIP/Grp78.
Conclusions: These results indicate that MANF expression was upregulated in the activated glial cells, which may contribute to the mechanism of ischemia-induced neural injury.