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Irradiation leads to apoptosis of Kupffer cells by a Hsp27-dependant pathway followed by release of TNF-alpha.

Tello K, Christiansen H, Gürleyen H, Dudas J, Rave-Fränk M, Hess CF, Ramadori G, Saile B - Radiat Environ Biophys (2008)

Bottom Line: The pathomechanisms underlying this phenomenon, however, remained to be elucidated.However, normalization of the TNF-alpha concentration with respect to cell number revealed that TNF-alpha concentration per attached cell remained constant during the observation period.We conclude that irradiation of Kupffer cells leads to apoptosis because of downregulation of Hsp27 and consecutive upregulation of bax expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine, Section of Gastroenterology and Endocrinology, Georg-August-University Göttingen, Robert-Koch-Strasse 40, 37075, Göttingen, Germany.

ABSTRACT
In a previous publication, we were able to show that irradiation of Kupffer cells, the liver resident macrophages, leads to an increased TNF-alpha concentration in the culture medium. The pathomechanisms underlying this phenomenon, however, remained to be elucidated. Here, we show that following irradiation of Kupffer cells, the apoptosis rate increased drastically within 48 h. At the same time, the total TNF-alpha concentration in cell lysates of Kupffer cells attached to the culture plate decreased. However, normalization of the TNF-alpha concentration with respect to cell number revealed that TNF-alpha concentration per attached cell remained constant during the observation period. Western blot analysis showed that heat shock protein 27 (Hsp27) is strongly downregulated and bax is upregulated in irradiated Kupffer cells as compared to sham-irradiated cells. Overexpression of Hsp27 in Kupffer cells was shown to prevent the effect of irradiation on bax expression, apoptosis and, at the same time, on increase of TNF-alpha concentration in the Kupffer cell medium. We conclude that irradiation of Kupffer cells leads to apoptosis because of downregulation of Hsp27 and consecutive upregulation of bax expression. Furthermore, we suggest that apoptosis of Kupffer cells leads to an increase of TNF-alpha concentration in the culture medium which may be due to cell death rather than active release or synthesis.

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a Survival of Kupffer cells which were transfected either with the control vector or the Hsp27 expression vector (Trypan blue exclusion after sham-irradiation and irradiation with at 8 Gy (c vector = empty control vector; Hsp27 Exp = Hsp27 expression vector). The values presented are mean values of seven independent Kupffer cell isolations. b Apoptosis of Kupffer cells. (c vector = empty control vector; Hsp27 Exp = Hsp27 expression vector) as measured by the AnnexinV/propidiumiodide method. Transfection was initiated 12 h prior to irradiation. Values presented are means ± SD of seven independent Kupffer cell isolations. Level of significance: **P < 0.01
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Fig4: a Survival of Kupffer cells which were transfected either with the control vector or the Hsp27 expression vector (Trypan blue exclusion after sham-irradiation and irradiation with at 8 Gy (c vector = empty control vector; Hsp27 Exp = Hsp27 expression vector). The values presented are mean values of seven independent Kupffer cell isolations. b Apoptosis of Kupffer cells. (c vector = empty control vector; Hsp27 Exp = Hsp27 expression vector) as measured by the AnnexinV/propidiumiodide method. Transfection was initiated 12 h prior to irradiation. Values presented are means ± SD of seven independent Kupffer cell isolations. Level of significance: **P < 0.01

Mentions: To gain evidence whether Hsp27 is capable to inhibit radiation-induced apoptosis in Kupffer cells, we overexpressed Hsp27 in Kupffer cells and irradiated the cells with 8 Gy. Transfection with a control vector did not alter cellular survival (Fig. 4a) or apoptosis rates of sham-irradiated cultures when compared to non-transfected cells (Fig. 4b). Transfection with the expression vector had very little effect on survival and apoptosis in sham-irradiated cultures (Fig. 4b). However, after irradiation overexpression of Hsp27 substantially increased cellular survival (Fig. 4a) and reduced radiation-induced increase of apoptosis to apoptosis levels observed in sham-irradiated cells (Fig. 4b).Fig. 4


Irradiation leads to apoptosis of Kupffer cells by a Hsp27-dependant pathway followed by release of TNF-alpha.

Tello K, Christiansen H, Gürleyen H, Dudas J, Rave-Fränk M, Hess CF, Ramadori G, Saile B - Radiat Environ Biophys (2008)

a Survival of Kupffer cells which were transfected either with the control vector or the Hsp27 expression vector (Trypan blue exclusion after sham-irradiation and irradiation with at 8 Gy (c vector = empty control vector; Hsp27 Exp = Hsp27 expression vector). The values presented are mean values of seven independent Kupffer cell isolations. b Apoptosis of Kupffer cells. (c vector = empty control vector; Hsp27 Exp = Hsp27 expression vector) as measured by the AnnexinV/propidiumiodide method. Transfection was initiated 12 h prior to irradiation. Values presented are means ± SD of seven independent Kupffer cell isolations. Level of significance: **P < 0.01
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Related In: Results  -  Collection

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Fig4: a Survival of Kupffer cells which were transfected either with the control vector or the Hsp27 expression vector (Trypan blue exclusion after sham-irradiation and irradiation with at 8 Gy (c vector = empty control vector; Hsp27 Exp = Hsp27 expression vector). The values presented are mean values of seven independent Kupffer cell isolations. b Apoptosis of Kupffer cells. (c vector = empty control vector; Hsp27 Exp = Hsp27 expression vector) as measured by the AnnexinV/propidiumiodide method. Transfection was initiated 12 h prior to irradiation. Values presented are means ± SD of seven independent Kupffer cell isolations. Level of significance: **P < 0.01
Mentions: To gain evidence whether Hsp27 is capable to inhibit radiation-induced apoptosis in Kupffer cells, we overexpressed Hsp27 in Kupffer cells and irradiated the cells with 8 Gy. Transfection with a control vector did not alter cellular survival (Fig. 4a) or apoptosis rates of sham-irradiated cultures when compared to non-transfected cells (Fig. 4b). Transfection with the expression vector had very little effect on survival and apoptosis in sham-irradiated cultures (Fig. 4b). However, after irradiation overexpression of Hsp27 substantially increased cellular survival (Fig. 4a) and reduced radiation-induced increase of apoptosis to apoptosis levels observed in sham-irradiated cells (Fig. 4b).Fig. 4

Bottom Line: The pathomechanisms underlying this phenomenon, however, remained to be elucidated.However, normalization of the TNF-alpha concentration with respect to cell number revealed that TNF-alpha concentration per attached cell remained constant during the observation period.We conclude that irradiation of Kupffer cells leads to apoptosis because of downregulation of Hsp27 and consecutive upregulation of bax expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine, Section of Gastroenterology and Endocrinology, Georg-August-University Göttingen, Robert-Koch-Strasse 40, 37075, Göttingen, Germany.

ABSTRACT
In a previous publication, we were able to show that irradiation of Kupffer cells, the liver resident macrophages, leads to an increased TNF-alpha concentration in the culture medium. The pathomechanisms underlying this phenomenon, however, remained to be elucidated. Here, we show that following irradiation of Kupffer cells, the apoptosis rate increased drastically within 48 h. At the same time, the total TNF-alpha concentration in cell lysates of Kupffer cells attached to the culture plate decreased. However, normalization of the TNF-alpha concentration with respect to cell number revealed that TNF-alpha concentration per attached cell remained constant during the observation period. Western blot analysis showed that heat shock protein 27 (Hsp27) is strongly downregulated and bax is upregulated in irradiated Kupffer cells as compared to sham-irradiated cells. Overexpression of Hsp27 in Kupffer cells was shown to prevent the effect of irradiation on bax expression, apoptosis and, at the same time, on increase of TNF-alpha concentration in the Kupffer cell medium. We conclude that irradiation of Kupffer cells leads to apoptosis because of downregulation of Hsp27 and consecutive upregulation of bax expression. Furthermore, we suggest that apoptosis of Kupffer cells leads to an increase of TNF-alpha concentration in the culture medium which may be due to cell death rather than active release or synthesis.

Show MeSH
Related in: MedlinePlus