Reactive gliosis is a hallmark of many retinal neurodegenerative conditions, including glaucoma. Although a majority of studies to date have concentrated on reactive gliosis in the optic nerve head, very few studies have been initiated to investigate the role of reactive gliosis in the retina. We have previously shown that reactive glial cells synthesize elevated levels of proteases, and these proteases, in turn, promote the death of retinal ganglion cells (RGCs). In this investigation, we have used two glial toxins to inhibit reactive gliosis and have evaluated their effect on protease-mediated death of RGCs. Kainic acid was injected into the vitreous humor of C57BL/6 mice to induce reactive gliosis and death of RGCs. C57BL/6 mice were also treated with glial toxins, alpha-aminoadipic acid (AAA) or Neurostatin, along with KA. Reactive gliosis was assessed by immunostaining of retinal cross sections and retinal flat-mounts with glial fibrillary acidic protein (GFAP) and vimentin antibodies. Apoptotic cell death was assessed by TUNEL assays. Loss of RGCs was determined by immunostaining of flat-mounted retinas with Brn3a antibodies. Proteolytic activities of matrix metalloproteinase-9 (MMP-9), tissue plasminogen activator (tPA), and urokinase plasminogen activator (uPA) were assessed by zymography assays. GFAP-immunoreactivity indicated that KA induced reactive gliosis in both retinal astrocytes and in Muller cells. AAA alone or in combination with KA decreased GFAP and vimentin-immunoreactivity in Mϋller cells, but not in astrocytes. In addition AAA failed to decrease KA-mediated protease levels and apoptotic death of RGCs. In contrast, Neurostatin either alone or in combination with KA, decreased reactive gliosis in both astrocytes and Mϋller cells. Furthermore, Neurostatin decreased protease levels and prevented apoptotic death of RGCs. Our findings, for the first time, indicate that inhibition of reactive gliosis decreases protease levels in the retina, prevents apoptotic death of retinal neurons, and provides substantial neuroprotection.

Activation of ER stress and subsequent loss of stemness of ISCs plays a critical role in BDL-induced systemic inflammation and cholestatic liver injury. Modulation of the ER stress response represents a potential therapeutic strategy for cholestatic liver diseases as well as other inflammatory diseases. (Hepatology 2018;67:1441-1457).

The results show that curcumin attenuates RGC and amacrine cell death despite elevated levels of proteases and raises the possibility that it may be used as a plausible adjuvant therapeutic agent to prevent the loss of these cells in retinal degenerative conditions.

Innate immune cells, such as macrophages (MΦ), play a critical role in the initiation and duration of inflammatory responses directed against foreign pathogens. The elaboration of immune effector by MΦ are coordinated by interactions of microbial stimuli, such as LPS, and critical immuno-modulatory cytokines with receptors present on the MΦ. The interactions mediate a cascade of signal transduction events that ultimately culminate with the activation and nuclear translocation of transcription factors which facilitate appropriate immune responses. Notably, TLR4 stimulation by LPS mediates the activation of NF-κβ , while the immuno-modulatory cytokine IFNγ mediates nuclear translocation of phosphorylated STAT1 dimers. In addition to mediating pro-inflammatory signals, NF-κβ and IFNγ also modulate a critical limiter of pro-inflammatory signals, suppressor of cytokine signaling-1 (SOCS1). Notably the regulation of SOCS1 molecules in response to multiple external stimuli, such as LPS and IFNγ are not well understood. In this study we investigate the mechanisms through time kinetics of SOCS1 upregulation through TLR4 and IFNγ signaling in RAW 264.7 MΦ as it relates to its transcription regulators, NF-κβ and pSTAT1. We found LPS treatment mediated the phosphorylation and subsequent degradation of Iκβ, which is indicative of NF-κβ nuclear translocation, at 0.5–3 hrs, with reaccumulation at 6–12 hrs. LPS-mediated pSTAT1 was detected at 3–6 hrs. In contrast, basal levels of SOCS1 were reduced subsequently to at 0.5–3 hrs, with reaccumulation at 12–48 hrs. While IFNγ stimulated pSTAT1 at 0.5–12 hrs and basal protein levels of SOCS1 were initially reduced further showed enhanced at 6–24 hours.