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Extracellular fluid flow and chloride content modulate H(+) transport by osteoclasts.

Morethson P - BMC Cell Biol. (2015)

Bottom Line: Bone resorption takes place within the basic multicellular units (BMU), and the surface to be resorbed is isolated from adjacent bone surfaces by a sealing zone between osteoclast membrane and bone matrix, which defines the limits of the resorption lacuna.Considering that the extracellular fluid (ECF) in both BMU and the resorption lacuna can be isolated from its surroundings, I hypothesize that flow and ion composition of the bone ECF in these sites might contribute to the regulation of osteoclast H(+) secretion.The data suggest, for the first time, that ECF flow and Cl(-) content have direct effects on osteoclast H(+) secretion and could be part of a mechanism determining the onset of osteoclast H(+) secretion required for bone resorption.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Biophysics, Institute of Biomedical Sciences, University of São Paulo, São Paulo, SP, Brazil. primoret@icb.usp.br.

ABSTRACT

Background: Bone resorption takes place within the basic multicellular units (BMU), and the surface to be resorbed is isolated from adjacent bone surfaces by a sealing zone between osteoclast membrane and bone matrix, which defines the limits of the resorption lacuna. Considering that the extracellular fluid (ECF) in both BMU and the resorption lacuna can be isolated from its surroundings, I hypothesize that flow and ion composition of the bone ECF in these sites might contribute to the regulation of osteoclast H(+) secretion. To investigate this hypothesis, I evaluated the H(+) secretion properties of individual osteoclasts and osteoclast-like cells (OCL-cells) and investigated whether changes in flow or chloride content of the extracellular solution modify the H(+) secretion properties in vitro.

Results: The results show that 1) osteoclasts are unable to secrete H(+) and regulate intracellular pH (pHi) under continuous flow conditions and exhibit progressive intracellular acidification; 2) the cessation of flow coincides with the onset of H(+) secretion and subsequent progressive intracellular alkalinization of osteoclasts and OCL-cells; 3) osteoclasts exhibit spontaneous rhythmic oscillations of pHi in non-flowing ECF, 4) pHi oscillations are not abolished by concanamycin, NPPB, or removal of extracellular Na(+) or Cl(-); 5) extracellular Cl(-) removal modifies the pattern of oscillations, by diminishing H(+) secretion; 6) pHi oscillations are abolished by continuous flowing of ECF over osteoclasts and OCL-cells.

Conclusions: The data suggest, for the first time, that ECF flow and Cl(-) content have direct effects on osteoclast H(+) secretion and could be part of a mechanism determining the onset of osteoclast H(+) secretion required for bone resorption.

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Related in: MedlinePlus

a. Emitted fluorescence of BCECF-loaded osteoclast-like cells (OCL-cells) individualized in the area defined by the colored circle line by using the MetaFluor 7.1 software (b). pHi of OCL-cells shown in (a). Each line represents one OCL-cell. At the time the flow is stopped (dashed line), pHi become progressively more alkaline, thus indicating H+ secretion since the standard HEPES-buffered solution does not contain HCO3−. c. Transmitted light image of selected osteoclast. d. pHi of the osteoclast shown in c. At the time the flow is stopped (dashed line), pHi become progressively more alkaline, thus indicating H+ secretion since the standard HEPES-buffered solution does not contain HCO3−
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Fig5: a. Emitted fluorescence of BCECF-loaded osteoclast-like cells (OCL-cells) individualized in the area defined by the colored circle line by using the MetaFluor 7.1 software (b). pHi of OCL-cells shown in (a). Each line represents one OCL-cell. At the time the flow is stopped (dashed line), pHi become progressively more alkaline, thus indicating H+ secretion since the standard HEPES-buffered solution does not contain HCO3−. c. Transmitted light image of selected osteoclast. d. pHi of the osteoclast shown in c. At the time the flow is stopped (dashed line), pHi become progressively more alkaline, thus indicating H+ secretion since the standard HEPES-buffered solution does not contain HCO3−

Mentions: In contrast to the results obtained under no flow conditions, with a flow rate of 5 mL/min the cells exhibited progressive intracellular acidification. However, upon the stoppage of extracellular fluid flow H+ secretion commences, resulting alkalinization in osteoclasts (n = 6) and OCL cells (n = 20) (Fig. 5) and the previously described oscillatory pattern resumes (Fig. 6a). Furthermore, reapplying flow—after a period of pHi oscillation—disrupts the rhythmic fluctuations in pHi displayed by the osteoclasts (n = 4) and OCL-cells (n = 20) observed under no flow conditions (Fig. 6b). Despite the fact that primary osteoclasts and OCL-cells may have differences concerning phenotype, the similar effect of extracellular fluid flow observed in both cells indicate that the mechanisms related to the effect of fluid in H+ secretion properties and pHi regulation are preserved in OCL-cells.Fig. 5


Extracellular fluid flow and chloride content modulate H(+) transport by osteoclasts.

Morethson P - BMC Cell Biol. (2015)

a. Emitted fluorescence of BCECF-loaded osteoclast-like cells (OCL-cells) individualized in the area defined by the colored circle line by using the MetaFluor 7.1 software (b). pHi of OCL-cells shown in (a). Each line represents one OCL-cell. At the time the flow is stopped (dashed line), pHi become progressively more alkaline, thus indicating H+ secretion since the standard HEPES-buffered solution does not contain HCO3−. c. Transmitted light image of selected osteoclast. d. pHi of the osteoclast shown in c. At the time the flow is stopped (dashed line), pHi become progressively more alkaline, thus indicating H+ secretion since the standard HEPES-buffered solution does not contain HCO3−
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4536797&req=5

Fig5: a. Emitted fluorescence of BCECF-loaded osteoclast-like cells (OCL-cells) individualized in the area defined by the colored circle line by using the MetaFluor 7.1 software (b). pHi of OCL-cells shown in (a). Each line represents one OCL-cell. At the time the flow is stopped (dashed line), pHi become progressively more alkaline, thus indicating H+ secretion since the standard HEPES-buffered solution does not contain HCO3−. c. Transmitted light image of selected osteoclast. d. pHi of the osteoclast shown in c. At the time the flow is stopped (dashed line), pHi become progressively more alkaline, thus indicating H+ secretion since the standard HEPES-buffered solution does not contain HCO3−
Mentions: In contrast to the results obtained under no flow conditions, with a flow rate of 5 mL/min the cells exhibited progressive intracellular acidification. However, upon the stoppage of extracellular fluid flow H+ secretion commences, resulting alkalinization in osteoclasts (n = 6) and OCL cells (n = 20) (Fig. 5) and the previously described oscillatory pattern resumes (Fig. 6a). Furthermore, reapplying flow—after a period of pHi oscillation—disrupts the rhythmic fluctuations in pHi displayed by the osteoclasts (n = 4) and OCL-cells (n = 20) observed under no flow conditions (Fig. 6b). Despite the fact that primary osteoclasts and OCL-cells may have differences concerning phenotype, the similar effect of extracellular fluid flow observed in both cells indicate that the mechanisms related to the effect of fluid in H+ secretion properties and pHi regulation are preserved in OCL-cells.Fig. 5

Bottom Line: Bone resorption takes place within the basic multicellular units (BMU), and the surface to be resorbed is isolated from adjacent bone surfaces by a sealing zone between osteoclast membrane and bone matrix, which defines the limits of the resorption lacuna.Considering that the extracellular fluid (ECF) in both BMU and the resorption lacuna can be isolated from its surroundings, I hypothesize that flow and ion composition of the bone ECF in these sites might contribute to the regulation of osteoclast H(+) secretion.The data suggest, for the first time, that ECF flow and Cl(-) content have direct effects on osteoclast H(+) secretion and could be part of a mechanism determining the onset of osteoclast H(+) secretion required for bone resorption.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Biophysics, Institute of Biomedical Sciences, University of São Paulo, São Paulo, SP, Brazil. primoret@icb.usp.br.

ABSTRACT

Background: Bone resorption takes place within the basic multicellular units (BMU), and the surface to be resorbed is isolated from adjacent bone surfaces by a sealing zone between osteoclast membrane and bone matrix, which defines the limits of the resorption lacuna. Considering that the extracellular fluid (ECF) in both BMU and the resorption lacuna can be isolated from its surroundings, I hypothesize that flow and ion composition of the bone ECF in these sites might contribute to the regulation of osteoclast H(+) secretion. To investigate this hypothesis, I evaluated the H(+) secretion properties of individual osteoclasts and osteoclast-like cells (OCL-cells) and investigated whether changes in flow or chloride content of the extracellular solution modify the H(+) secretion properties in vitro.

Results: The results show that 1) osteoclasts are unable to secrete H(+) and regulate intracellular pH (pHi) under continuous flow conditions and exhibit progressive intracellular acidification; 2) the cessation of flow coincides with the onset of H(+) secretion and subsequent progressive intracellular alkalinization of osteoclasts and OCL-cells; 3) osteoclasts exhibit spontaneous rhythmic oscillations of pHi in non-flowing ECF, 4) pHi oscillations are not abolished by concanamycin, NPPB, or removal of extracellular Na(+) or Cl(-); 5) extracellular Cl(-) removal modifies the pattern of oscillations, by diminishing H(+) secretion; 6) pHi oscillations are abolished by continuous flowing of ECF over osteoclasts and OCL-cells.

Conclusions: The data suggest, for the first time, that ECF flow and Cl(-) content have direct effects on osteoclast H(+) secretion and could be part of a mechanism determining the onset of osteoclast H(+) secretion required for bone resorption.

Show MeSH
Related in: MedlinePlus