Calcium-Signalling in Human Glaucoma Lamina Cribrosa Myofibroblasts

Abstract

Glaucoma is one of the most common causes of treatable visual impairment in the developed world, affecting approximately 64 million people worldwide, some of whom will be bilaterally blind from irreversible optic nerve damage. The optic nerve head is a key site of damage in glaucoma where there is fibrosis of the connective tissue in the lamina cribrosa (LC) extracellular matrix. As a ubiquitous second messenger, calcium (Ca²⁺) can interact with various cellular proteins to regulate multiple physiological processes and contribute to a wide range of diseases, including cancer, fibrosis, and glaucoma. Our research has shown evidence of oxidative stress, mitochondrial dysfunction, an elevated expression of Ca²⁺ entry channels, Ca²⁺-dependent pumps and exchangers, and an abnormal rise in cytosolic Ca²⁺ in human glaucomatous LC fibroblast cells. We have evidence that this increase is dependent on Ca²⁺ entry channels located in the plasma membrane, and its release is from internal stores in the endoplasmic reticulum (ER), as well as from the mitochondria. Here, we summarize some of the molecular Ca²⁺-dependent mechanisms related to this abnormal Ca²⁺-signalling in human glaucoma LC cells, with a view toward identifying potential therapeutic targets for ongoing optic neuropathy.

Keywords: calcium homeostasis; fibrosis; glaucoma; lamina cribrosa.

Figure 2

General concept of Ca²⁺-signalling homeostasis. Stimuli induce both the entry of external Ca²⁺ and the release Ca²⁺ from the internal stores of the ER/SR via IP3R and RYR. In activated cells, Ca²⁺ enters cells through different types of Ca²⁺ channels, including voltage-operated channels (VOC), second messenger-operated channels (SMOC), store-operated channels (SOC), and receptor-operated channels (ROC). We note that VOCs are activated by membrane depolarization and SMOCs are activated by small messenger molecules, such as InsP3. In resting cells, Ca²⁺ is removed from the cell by exchangers and pumps. The NCX and PMCA extrude Ca²⁺ from the cytosol to the extracellular milieu, whereas the ER/SR Ca²⁺ -ATPase (SERCA) pumps pump Ca²⁺ back into the ER. Mitochondria also have an active function during the recovery process in that they sequester Ca²⁺ rapidly through a uniporter, which is then released more slowly back into the cytosol.

Figure 3

Endoplasmic reticulum (ER) stress and unfolded protein response (UPR). The ER’s functions include proper protein synthesis and folding to maintain cellular homeostasis. The disturbance of cellular ATP production or Ca²⁺ concentration affects ER functioning, leading to the excessive accumulation and aggregation of unfolded proteins and generating ER stress, which further activates the UPR. The UPR plays key roles in adaptive responses, feedback control, and cell fates. In an adaptive response, the UPR reduces ER stress and restores ER homeostasis. UPR signalling is inhibited through a negative feedback mechanism.

Figure 1

Examples of phase contrast microscopy images of cultured LC cells from (A) non-glaucomatous and (B) glaucomatous human donors. We note that the glaucomatous LC cell is larger than the non-glaucomatous LC cell.

Figure 4

Ca²⁺ signalling pathways in activated lamina cribrosa fibroblasts in glaucoma. Mechanical and oxidative stress and growth factors (TGFβ) stimulate Ca²⁺ ion channels (L-type, TRPC, Maxi-K⁺) and intracellular Ca²⁺ release from internal stores (ER and mitochondria). This stimulates PLC, which, in turn, activates a variety of signalling pathways, such as RAS/RAF and p38MAPK, as well as PKC p42/44-MAPK, CamK-calcineurin-NFATc, the SERCA pumps, and the PI3K signalling pathways, leading to the activation of Ca²⁺-dependent gene transcription factors (NFATc3 and YAP).