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Kupffer cells promote lead nitrate-induced hepatocyte apoptosis via oxidative stress.

Pagliara P, Carlà EC, Caforio S, Chionna A, Massa S, Abbro L, Dini L - Comp Hepatol (2003)

Bottom Line: RESULTS: The in vivo hepatic apoptosis, induced by Pb(NO3)2 was prevented by a pre-treatment with gadolinium chloride (GdCl3), a Kupffer cells toxicant, that suppresses Kupffer cell activity and reduces to a half the apoptotic rate.In addition, in vivo Pb(NO3)2 administration deprives hepatocytes of reduced glutathione, whereas the loss of this important oxidation-preventing agent is considerably mitigated or abolished by pre-treatment with GdCl3.CONCLUSION: Pb(NO3)2 has, at most, a direct necrotic (but not apoptogenic) effect on hepatocytes and HepG2 cells, giving a clue about the regulatory role of Kupffer cells in the induction of liver apoptosis after a single Pb(NO3)2 injection without pre-treatment with GdCl3, probably via secreting soluble factors that trigger oxidative stress in target cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Science and Biological and Environmental Technologies, University of Lecce, via per Monteroni, Lecce, Italy. luciana.dini@unile.it

ABSTRACT
BACKGROUND: Apoptosis and its modulation are crucial factors for the maintenance of liver health, allowing hepatocytes to die without provoking a potential harmful inflammatory response through a tightly controlled and regulated process. Since Kupffer cells play a key role in the maintenance of liver function, the aim of this study was to verify whether Kupffer cells are involved in the induction of liver apoptosis after i.v. injection of Pb(NO3)2 likely by secretion mechanisms. RESULTS: The in vivo hepatic apoptosis, induced by Pb(NO3)2 was prevented by a pre-treatment with gadolinium chloride (GdCl3), a Kupffer cells toxicant, that suppresses Kupffer cell activity and reduces to a half the apoptotic rate. In addition, in vivo Pb(NO3)2 administration deprives hepatocytes of reduced glutathione, whereas the loss of this important oxidation-preventing agent is considerably mitigated or abolished by pre-treatment with GdCl3. However, incubation of isolated hepatocytes and Kupffer cells and HepG2 cells with Pb(NO3)2 for 24 hours induced necrotic but not apoptotic cells. Apoptosis of hepatocytes and HepG2 cells was observed only after the addition of conditioned medium obtained from Kupffer cells cultured for 24 hours with Pb(NO3)2, thus indicating the secretion of soluble mediators of apoptosis by Kupffer cells. Apoptosis in the HepG2 cells was observed upon 24-hours incubation of HepG2 cells with 1 mM buthionine sulfoximine, a glutathione depleting agent, thus showing that there is an oxidative apoptogenic pathway in HepG2 cells. CONCLUSION: Pb(NO3)2 has, at most, a direct necrotic (but not apoptogenic) effect on hepatocytes and HepG2 cells, giving a clue about the regulatory role of Kupffer cells in the induction of liver apoptosis after a single Pb(NO3)2 injection without pre-treatment with GdCl3, probably via secreting soluble factors that trigger oxidative stress in target cells.

No MeSH data available.


Related in: MedlinePlus

Nomarsky light microscopy of rat hepatocyte cultures. Cells were incubated with Pb(NO3)2 (10 mM) or with conditioned medium (CM) from Kupffer cells incubated for 24 hours with Pb(NO3)2 (a) control hepatocytes (b) 24 hours incubation with Pb(NO3)2. Apoptotic hepatocytes are not present. (c) Apoptotic cells (arrows) are present when incubated with CM from Kupffer cells incubated for 24 hours with Pb(NO3)2. (d-e) Hepatocytes pre-incubated with 1 mM BSO for 24 hours, and subsequently incubated for 24 hours (d) or 48 hours (e) with Pb(NO3)2. Deprivation of GHS leads to apoptosis also upon incubation with Pb(NO3)2. Arrows indicate apoptotic hepatocytes that are characterized by shrinkage in volume, round shape, very condensed cytoplasm and dark nucleus. Bar = 10 μm.
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Figure 3: Nomarsky light microscopy of rat hepatocyte cultures. Cells were incubated with Pb(NO3)2 (10 mM) or with conditioned medium (CM) from Kupffer cells incubated for 24 hours with Pb(NO3)2 (a) control hepatocytes (b) 24 hours incubation with Pb(NO3)2. Apoptotic hepatocytes are not present. (c) Apoptotic cells (arrows) are present when incubated with CM from Kupffer cells incubated for 24 hours with Pb(NO3)2. (d-e) Hepatocytes pre-incubated with 1 mM BSO for 24 hours, and subsequently incubated for 24 hours (d) or 48 hours (e) with Pb(NO3)2. Deprivation of GHS leads to apoptosis also upon incubation with Pb(NO3)2. Arrows indicate apoptotic hepatocytes that are characterized by shrinkage in volume, round shape, very condensed cytoplasm and dark nucleus. Bar = 10 μm.

Mentions: Necrosis and apoptosis were evaluated at fixed time intervals (6, 8, 16, 24, 48 hours) by light microscopy in cultures of hepatocytes and Kupffer cells isolated from normal rats and incubated up to 48 hours with 10 mM Pb(NO3)2 (Table 5). More than 95% of Kupffer cells died within 24 hours of incubation with Pb(NO3)2, and only cell debris were found in the culture wells at 48 hours. Within 24 hours of culture in the presence of Pb(NO3)2 some necrotic hepatocytes were observed, whereas almost all hepatocytes were necrotic at 48 hours. Similar results were found for hepatocytes incubated with KNO3. 80% necrotic hepatocytes were observed at 24 hours of incubation with Pb(C2H3O2)2. Apoptosis was never detected after incubation with Pb(NO3)2, Pb(C2H3O2)2 or KNO3 for up to 48 hours, but it was observed only after incubation with conditioned medium (CM) of Kupffer cells cultured for 24 hours in presence of Pb(NO3)2; values ranged from about 35% at 24 hours to about 48% at 48 hours of incubation (Fig. 3 and Table 5). Necrosis was never observed in such condition. Figure 3 shows many apoptotic hepatocytes detaching from culture dishes.


Kupffer cells promote lead nitrate-induced hepatocyte apoptosis via oxidative stress.

Pagliara P, Carlà EC, Caforio S, Chionna A, Massa S, Abbro L, Dini L - Comp Hepatol (2003)

Nomarsky light microscopy of rat hepatocyte cultures. Cells were incubated with Pb(NO3)2 (10 mM) or with conditioned medium (CM) from Kupffer cells incubated for 24 hours with Pb(NO3)2 (a) control hepatocytes (b) 24 hours incubation with Pb(NO3)2. Apoptotic hepatocytes are not present. (c) Apoptotic cells (arrows) are present when incubated with CM from Kupffer cells incubated for 24 hours with Pb(NO3)2. (d-e) Hepatocytes pre-incubated with 1 mM BSO for 24 hours, and subsequently incubated for 24 hours (d) or 48 hours (e) with Pb(NO3)2. Deprivation of GHS leads to apoptosis also upon incubation with Pb(NO3)2. Arrows indicate apoptotic hepatocytes that are characterized by shrinkage in volume, round shape, very condensed cytoplasm and dark nucleus. Bar = 10 μm.
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Related In: Results  -  Collection

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Figure 3: Nomarsky light microscopy of rat hepatocyte cultures. Cells were incubated with Pb(NO3)2 (10 mM) or with conditioned medium (CM) from Kupffer cells incubated for 24 hours with Pb(NO3)2 (a) control hepatocytes (b) 24 hours incubation with Pb(NO3)2. Apoptotic hepatocytes are not present. (c) Apoptotic cells (arrows) are present when incubated with CM from Kupffer cells incubated for 24 hours with Pb(NO3)2. (d-e) Hepatocytes pre-incubated with 1 mM BSO for 24 hours, and subsequently incubated for 24 hours (d) or 48 hours (e) with Pb(NO3)2. Deprivation of GHS leads to apoptosis also upon incubation with Pb(NO3)2. Arrows indicate apoptotic hepatocytes that are characterized by shrinkage in volume, round shape, very condensed cytoplasm and dark nucleus. Bar = 10 μm.
Mentions: Necrosis and apoptosis were evaluated at fixed time intervals (6, 8, 16, 24, 48 hours) by light microscopy in cultures of hepatocytes and Kupffer cells isolated from normal rats and incubated up to 48 hours with 10 mM Pb(NO3)2 (Table 5). More than 95% of Kupffer cells died within 24 hours of incubation with Pb(NO3)2, and only cell debris were found in the culture wells at 48 hours. Within 24 hours of culture in the presence of Pb(NO3)2 some necrotic hepatocytes were observed, whereas almost all hepatocytes were necrotic at 48 hours. Similar results were found for hepatocytes incubated with KNO3. 80% necrotic hepatocytes were observed at 24 hours of incubation with Pb(C2H3O2)2. Apoptosis was never detected after incubation with Pb(NO3)2, Pb(C2H3O2)2 or KNO3 for up to 48 hours, but it was observed only after incubation with conditioned medium (CM) of Kupffer cells cultured for 24 hours in presence of Pb(NO3)2; values ranged from about 35% at 24 hours to about 48% at 48 hours of incubation (Fig. 3 and Table 5). Necrosis was never observed in such condition. Figure 3 shows many apoptotic hepatocytes detaching from culture dishes.

Bottom Line: RESULTS: The in vivo hepatic apoptosis, induced by Pb(NO3)2 was prevented by a pre-treatment with gadolinium chloride (GdCl3), a Kupffer cells toxicant, that suppresses Kupffer cell activity and reduces to a half the apoptotic rate.In addition, in vivo Pb(NO3)2 administration deprives hepatocytes of reduced glutathione, whereas the loss of this important oxidation-preventing agent is considerably mitigated or abolished by pre-treatment with GdCl3.CONCLUSION: Pb(NO3)2 has, at most, a direct necrotic (but not apoptogenic) effect on hepatocytes and HepG2 cells, giving a clue about the regulatory role of Kupffer cells in the induction of liver apoptosis after a single Pb(NO3)2 injection without pre-treatment with GdCl3, probably via secreting soluble factors that trigger oxidative stress in target cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Science and Biological and Environmental Technologies, University of Lecce, via per Monteroni, Lecce, Italy. luciana.dini@unile.it

ABSTRACT
BACKGROUND: Apoptosis and its modulation are crucial factors for the maintenance of liver health, allowing hepatocytes to die without provoking a potential harmful inflammatory response through a tightly controlled and regulated process. Since Kupffer cells play a key role in the maintenance of liver function, the aim of this study was to verify whether Kupffer cells are involved in the induction of liver apoptosis after i.v. injection of Pb(NO3)2 likely by secretion mechanisms. RESULTS: The in vivo hepatic apoptosis, induced by Pb(NO3)2 was prevented by a pre-treatment with gadolinium chloride (GdCl3), a Kupffer cells toxicant, that suppresses Kupffer cell activity and reduces to a half the apoptotic rate. In addition, in vivo Pb(NO3)2 administration deprives hepatocytes of reduced glutathione, whereas the loss of this important oxidation-preventing agent is considerably mitigated or abolished by pre-treatment with GdCl3. However, incubation of isolated hepatocytes and Kupffer cells and HepG2 cells with Pb(NO3)2 for 24 hours induced necrotic but not apoptotic cells. Apoptosis of hepatocytes and HepG2 cells was observed only after the addition of conditioned medium obtained from Kupffer cells cultured for 24 hours with Pb(NO3)2, thus indicating the secretion of soluble mediators of apoptosis by Kupffer cells. Apoptosis in the HepG2 cells was observed upon 24-hours incubation of HepG2 cells with 1 mM buthionine sulfoximine, a glutathione depleting agent, thus showing that there is an oxidative apoptogenic pathway in HepG2 cells. CONCLUSION: Pb(NO3)2 has, at most, a direct necrotic (but not apoptogenic) effect on hepatocytes and HepG2 cells, giving a clue about the regulatory role of Kupffer cells in the induction of liver apoptosis after a single Pb(NO3)2 injection without pre-treatment with GdCl3, probably via secreting soluble factors that trigger oxidative stress in target cells.

No MeSH data available.


Related in: MedlinePlus