• Molecules and Cells
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• v.37 no.5
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• pp.418-425
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• 2014

Extracellular hypotonic stress can affect cellular function. Whether and how hypotonicity affects immune cell function remains to be elucidated. Macrophages are immune cells that play key roles in adaptive and innate in immune reactions. The purpose of this study was to investigate the role and underlying mechanism of hypotonic stress in the function of bone marrow-derived macrophages (BMDMs). Hypotonic stress increased endocytic activity in BMDMs, but there was no significant change in the expression of CD80, CD86, and MHC class II molecules, nor in the secretion of TNF-${\alpha}$ or IL-10 by BMDMs. Furthermore, the enhanced endocytic activity of BMDMs triggered by hypotonic stress was significantly inhibited by chloride channel-3 (ClC-3) siRNA. Our findings suggest that hypotonic stress can induce endocytosis in BMDMs and that ClC-3 plays a central role in the endocytic process.

• Korean Journal of Veterinary Research
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• v.36 no.2
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• pp.305-312
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• 1996

Cell volume regulatory mechanisms are usually disclosed by exposure of cell to anisotonic media. If a cell is suddenly exposed to hypotonic media, it swells initially like an osmometer but within minutes regains its original cell volume. This behavior has been labelled as regulatory cell volume decrease(RVD). RVD is believed to result from the loss of permeable ions through the membrane. In this study, we examined hypotonically induced changes in the membrance currents involved in RVD by using whole cell voltage clamp technique in the unfertilized hamster egg. At -40mV of the holding potential, the stationary current was maintained in the hamster egg exposed to isotonic solution composed of, mainly, 115mM NaCl and 40mM mannitol. Hypotonic solution was prepared by removing mannitol. Therefore, the concentrations of $Na^+$ and $Cl^-$ in this hypotonic media were the same as those in the isotonic solution. Following 30 to 60 sec after applying the hypotonic media to the egg, the inward current was evoked. This inward current was eliminated by $100{\mu}M$ 4-acetamido-4'-isothiocyanostil-bene-2,2'-disulfonic acid(SITS), an anion channel blocker, leaving the small outward current component. Further addition of 2mM $Ba^{2+}$, a broad $K^+$ channel blocker, completely abolished the small outward current left even in the presence of SITS during hypotonic stress. These results suggest that $K^+$ and $Cl^-$ move out of cells, resulting in RVD. To test the involvement of $Na^+$ in RVD, 20mM Na-isethionate was substituted for mannitol in isotonic media(135mM $Na^+$) and Na-isethionate (20mM) was freed the hypotonic solution. Only $Cl^-$ concentration in both isotonic and hypotonic media was kept constant at 115mM, whereas concentration of $Na^+$ was lowered in hypotonic solution to 115mM from 135mM in isotonic solution. Hypotonic medium induced the outward current in the egg equilibrated isotonically. This current was reduced by $100{\mu}M$ SITS but was augmented by 2 mM $Ba^{2+}$. In terms of RVD, these results imply that $Cl^-$ efflux is coupled with $K^+$, maybe for electroneutrality during hypotonic stress and/or with $Na^+$ via unknown transport mechanism(s). From the overall results, the hypotonic stress facilitates the movement of $Cl^-$ and $K^+$ out of the hamster egg to regain cellular volume with electroneutrality. If there exist a difference in $[Na^+]_0$ between isotonic and hypotonic solution, another transport mechanism concerned with $Na^+$ may, at least partly, participate in regulatory volume decrease.

• International Journal of Vascular Biomedical Engineering
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• v.4 no.2
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• pp.7-12
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• 2006

The present study investigated the effects of the osmotic environment on the rheological properties of erythrocytes and their suspensions. In an iso-osmotic medium, erythrocytes forming a biconcave discocyte under resting conditions, exhibited high deformability. In a low-osmotic medium, the deformability of erythrocytes, which swelled and exhibited a spherical shape, significantly decreased at a high shear stress and the high-shear viscosity of the cell suspension was slightly higher than that of normal blood. Hyper-osmotic stress, however, which caused to form echinocytes, decreased cell deformability but exhibited smaller viscosity in low shear rates than iso-osmotic blood viscosity. These results showed a close relation with the aggregability measurements, in that hypertonic blood showed lower aggregability than the hypotonic and isotonic RBC suspensions. These findings indicate that the physicochemical environment has a strong influence on the rheological properties of the erythrocyte and its suspensions.

• The Korean Journal of Physiology and Pharmacology
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• v.19 no.3
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• pp.249-255
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• 2015

Wnk kinase maintains cell volume, regulating various transporters such as sodium-chloride cotransporter, potassium-chloride cotransporter, and sodium-potassium-chloride cotransporter 1 (NKCC1) through the phosphorylation of oxidative stress responsive kinase 1 (OSR1) and STE20/SPS1-related proline/alanine-rich kinase (SPAK). However, the activating mechanism of Wnk kinase in specific tissues and specific conditions is broadly unclear. In the present study, we used a human salivary gland (HSG) cell line as a model and showed that $Ca^{2+}$ may have a role in regulating Wnk kinase in the HSG cell line. Through this study, we found that the HSG cell line expressed molecules participating in the WNK-OSR1-NKCC pathway, such as Wnk1, Wnk4, OSR1, SPAK, and NKCC1. The HSG cell line showed an intracellular $Ca^{2+}$ concentration ($[Ca^{2+}]_i$) increase in response to hypotonic stimulation, and the response was synchronized with the phosphorylation of OSR1. Interestingly, when we inhibited the hypotonically induced $[Ca^{2+}]_i$ increase with nonspecific $Ca^{2+}$ channel blockers such as 2-aminoethoxydiphenyl borate, gadolinium, and lanthanum, the phosphorylated OSR1 level was also diminished. Moreover, a cyclopiazonic acid-induced passive $[Ca^{2+}]_i$ elevation was evoked by the phosphorylation of OSR1, and the amount of phosphorylated OSR1 decreased when the cells were treated with BAPTA, a $Ca^{2+}$ chelator. Finally, through that process, NKCC1 activity also decreased to maintain the cell volume in the HSG cell line. These results indicate that $Ca^{2+}$ may regulate the WNK-OSR1 pathway and NKCC1 activity in the HSG cell line. This is the first demonstration that indicates upstream $Ca^{2+}$ regulation of the WNK-OSR1 pathway in intact cells.