**4.3. Glycerol transport**

AQP 3 was shown to be expressed in the stratum corneum (SC) at the basal layer of the keratinocytes and plays a role in skin hydration. In AQP-3-deficient mice, SC hydration was significantly reduced due to reduced water content, decreased skin elasticity and wound healing [108]. An important factor which was also attributed to reduced skin hydration in AQP-3-deficient mice is the impaired glycerol transport from the blood to the epidermis through the basal keratinocytes, suggesting the importance of AQP 3 in glycerol transport. Dysregulated expression of AQP 3 has been found in various skin disorders associated with altered epidermal proliferation [109, 110]. In fact, topical or systemic replacement of glycerol prevented skin abnormalities (less hydration and elasticity and impaired barrier function) in the deficient mice [111].

### **4.4. Cell proliferation**

A role for AQP 3 in cell proliferation has been suggested in various cell types. Using corneal epithelial cells, delayed restoration of full-thickness epithelia was seen in AQP-3-deficient mice after scraping. This was confirmed by reduction in proliferating BrdU-positive cells during healing [112]. Reduced keratinocyte cell proliferation was also evident in AQP-3-deficient mice or with siRNA-mediated knockout of AQP 3 in keratinocytes in part through reduction of p38 MAPK activity [113]. Furthermore, the proliferative rate of mouse colonic epithelial cells was significantly reduced in AQP-3-deficient mice, which might explain the enhanced colitis severity in these mice compared to WT mice in the dextran sulfate sodium model of colitis [114].

#### **4.5. Cell adhesion**

AQP 0 is thought to be involved in cell-cell adhesion. It has been found to be expressed in lens fiber cells in the eye and plays a role in maintaining their structure [115]. Loss-of-function mutation of AQP 0 in humans and mice resulted in congenital cataracts [34, 92]. In addition, AQP 4 was shown to mediate weak cell-cell interaction through its short helix in the extracel‐ lular loop [116]. Overexpression of AQP 4 in L-cells (which lack endogenous adhesion molecules) resulted in cell cluster formation, which supports the role of this AQP in intercel‐ lular adhesion.

#### **4.6. Cell migration**

Various AQPs have been shown to be involved in the cell migrative process. AQP 1 is expressed on the leading edge of migrating cultured endothelial cells in association with increased lamellipodia formation. AQP 1 deficiency in cultured endothelial cells results in significant reduction in their migration. Overexpression of AQP 1 or 4 enhanced cell migration along with prominent membrane ruffling at the leading edge [53]. The role of AQP 1 in cell migration was also confirmed using kidney proximal tubule cells where its deficiency reduced cell migration and its overexpression led to enhanced cell migration through the formation of lamella-like membrane protrusions at the cell leading edge [50]. Furthermore, AQP 4 was localized on the leading edge of migrating cultured astroglia cells, and its expression was increased by inducing a small extracellular osmotic gradient. AQP 4 deficiency (by siRNA treatment or cell isolation from AQP-4-deficient mice) resulted in marked reduction in their migratory potential [51, 52]. AQP 3 deficiency in mammalian corneal epithelial cells [51], keratinocytes [113] and fibroblasts [117] also reduced their migrative ability both *in vitro* and *in vivo.*

AQPs enhance cell migration through various mechanisms. They facilitate rapid changes in cell volume and shape, which allows the cells to squeeze through the narrow and irregularly shaped extracellular space; this has been referred to as amoeboidal movement [118]. Also, they increase the local hydrostatic pressure (that push apart adjacent stationary cells), and actin repolymerization, to stabilize cell membrane protrusions at the leading edge which is required for the migratory process [119]. There is some evidence regarding the role of AQP 4 in regulating a complex of intracellular molecules such as alpha-syntrophin involved in mem‐ brane protrusions [120]. Some evidence also suggests a role for AQP 3 in reducing keratinocyte cell migration through reduced p38 MAPK activity [113]; this is generally recognized as an important signaling molecule for cell migration.
