Characterization of rat parotid and submandibular acinar cell apoptosis in primary culture

Kirsten Limesand, Katherine A. Barzen, Linda A. Sanders, Robert A. Sclafani, Mary V. Raynolds, Mary E. Reyland, Steven M. Anderson, David O. Quissell

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Apoptosis is a highly organized cellular process that is critical for maintaining glandular homeostasis. We have used primary rat salivary acinar cells from the parotid and submandibular glands to investigate the critical regulatory events involved in apoptosis. Caspase-3 activity, cleavage of caspase substrates, and deoxyribonucleic acid (DNA) fragmentation were assayed in cells treated with etoposide, a DNA-damaging agent, or brefeldin A (BFA), a Golgi toxin. Dose-response studies showed that the sensitivity of both cell types to etoposide and BFA was similar, with 150 μM etoposide or 1.5 μM BFA inducing maximal caspase activation. However, BFA induced a more robust activation of caspase and DNA fragmentation in both cell types. Similar results were observed when the caspase cleavage of poly(adenosine 5′-diphosphate ribose) polymerase and protein kinase C delta were analyzed by Western blot. Analysis of the kinetics of apoptosis showed that caspase-3 activation was maximal at 8 h of etoposide or BFA treatment in the parotid cells and at 8-18 h in the submandibular cells. A similar time course was observed when DNA fragmentation was assayed, although maximal DNA fragmentation in BFA-treated cells was two- to threefold higher than that observed in etoposide-treated cells. Despite slight kinetic differences, it would appear that the apoptotic cascade is very similar in both primary parotid and submandibular acinar cells. Although limited in their long-term stability in culture, the use of primary, nonimmortalized salivary acinar cultures will also permit the use of specific transgenic animals to further characterize the molecular events involved in the regulation of salivary gland acinar cell apoptosis.

Original languageEnglish (US)
Pages (from-to)170-177
Number of pages8
JournalIn Vitro Cellular and Developmental Biology - Animal
Volume39
Issue number3-4
StatePublished - Mar 2003
Externally publishedYes

Fingerprint

Brefeldin A
Acinar Cells
Etoposide
Rats
Apoptosis
Caspases
DNA
Chemical activation
Caspase 3
Cells
Protein Kinase C-delta
Kinetics
Ribose
Poly Adenosine Diphosphate Ribose
Diphosphates
Genetically Modified Animals
Submandibular Gland
Parotid Gland
Adenosine
Salivary Glands

Keywords

  • Apoptosis
  • Brefeldin A
  • Etoposide
  • Primary culture
  • Salivary gland

ASJC Scopus subject areas

  • Developmental Biology
  • Clinical Biochemistry
  • Cell Biology

Cite this

Limesand, K., Barzen, K. A., Sanders, L. A., Sclafani, R. A., Raynolds, M. V., Reyland, M. E., ... Quissell, D. O. (2003). Characterization of rat parotid and submandibular acinar cell apoptosis in primary culture. In Vitro Cellular and Developmental Biology - Animal, 39(3-4), 170-177.

Characterization of rat parotid and submandibular acinar cell apoptosis in primary culture. / Limesand, Kirsten; Barzen, Katherine A.; Sanders, Linda A.; Sclafani, Robert A.; Raynolds, Mary V.; Reyland, Mary E.; Anderson, Steven M.; Quissell, David O.

In: In Vitro Cellular and Developmental Biology - Animal, Vol. 39, No. 3-4, 03.2003, p. 170-177.

Research output: Contribution to journalArticle

Limesand, K, Barzen, KA, Sanders, LA, Sclafani, RA, Raynolds, MV, Reyland, ME, Anderson, SM & Quissell, DO 2003, 'Characterization of rat parotid and submandibular acinar cell apoptosis in primary culture', In Vitro Cellular and Developmental Biology - Animal, vol. 39, no. 3-4, pp. 170-177.
Limesand, Kirsten ; Barzen, Katherine A. ; Sanders, Linda A. ; Sclafani, Robert A. ; Raynolds, Mary V. ; Reyland, Mary E. ; Anderson, Steven M. ; Quissell, David O. / Characterization of rat parotid and submandibular acinar cell apoptosis in primary culture. In: In Vitro Cellular and Developmental Biology - Animal. 2003 ; Vol. 39, No. 3-4. pp. 170-177.
@article{8a000f78d2334d73a55b57990a50cdfe,
title = "Characterization of rat parotid and submandibular acinar cell apoptosis in primary culture",
abstract = "Apoptosis is a highly organized cellular process that is critical for maintaining glandular homeostasis. We have used primary rat salivary acinar cells from the parotid and submandibular glands to investigate the critical regulatory events involved in apoptosis. Caspase-3 activity, cleavage of caspase substrates, and deoxyribonucleic acid (DNA) fragmentation were assayed in cells treated with etoposide, a DNA-damaging agent, or brefeldin A (BFA), a Golgi toxin. Dose-response studies showed that the sensitivity of both cell types to etoposide and BFA was similar, with 150 μM etoposide or 1.5 μM BFA inducing maximal caspase activation. However, BFA induced a more robust activation of caspase and DNA fragmentation in both cell types. Similar results were observed when the caspase cleavage of poly(adenosine 5′-diphosphate ribose) polymerase and protein kinase C delta were analyzed by Western blot. Analysis of the kinetics of apoptosis showed that caspase-3 activation was maximal at 8 h of etoposide or BFA treatment in the parotid cells and at 8-18 h in the submandibular cells. A similar time course was observed when DNA fragmentation was assayed, although maximal DNA fragmentation in BFA-treated cells was two- to threefold higher than that observed in etoposide-treated cells. Despite slight kinetic differences, it would appear that the apoptotic cascade is very similar in both primary parotid and submandibular acinar cells. Although limited in their long-term stability in culture, the use of primary, nonimmortalized salivary acinar cultures will also permit the use of specific transgenic animals to further characterize the molecular events involved in the regulation of salivary gland acinar cell apoptosis.",
keywords = "Apoptosis, Brefeldin A, Etoposide, Primary culture, Salivary gland",
author = "Kirsten Limesand and Barzen, {Katherine A.} and Sanders, {Linda A.} and Sclafani, {Robert A.} and Raynolds, {Mary V.} and Reyland, {Mary E.} and Anderson, {Steven M.} and Quissell, {David O.}",
year = "2003",
month = "3",
language = "English (US)",
volume = "39",
pages = "170--177",
journal = "In Vitro Cellular and Developmental Biology - Animal",
issn = "1071-2690",
publisher = "Springer New York",
number = "3-4",

}

TY - JOUR

T1 - Characterization of rat parotid and submandibular acinar cell apoptosis in primary culture

AU - Limesand, Kirsten

AU - Barzen, Katherine A.

AU - Sanders, Linda A.

AU - Sclafani, Robert A.

AU - Raynolds, Mary V.

AU - Reyland, Mary E.

AU - Anderson, Steven M.

AU - Quissell, David O.

PY - 2003/3

Y1 - 2003/3

N2 - Apoptosis is a highly organized cellular process that is critical for maintaining glandular homeostasis. We have used primary rat salivary acinar cells from the parotid and submandibular glands to investigate the critical regulatory events involved in apoptosis. Caspase-3 activity, cleavage of caspase substrates, and deoxyribonucleic acid (DNA) fragmentation were assayed in cells treated with etoposide, a DNA-damaging agent, or brefeldin A (BFA), a Golgi toxin. Dose-response studies showed that the sensitivity of both cell types to etoposide and BFA was similar, with 150 μM etoposide or 1.5 μM BFA inducing maximal caspase activation. However, BFA induced a more robust activation of caspase and DNA fragmentation in both cell types. Similar results were observed when the caspase cleavage of poly(adenosine 5′-diphosphate ribose) polymerase and protein kinase C delta were analyzed by Western blot. Analysis of the kinetics of apoptosis showed that caspase-3 activation was maximal at 8 h of etoposide or BFA treatment in the parotid cells and at 8-18 h in the submandibular cells. A similar time course was observed when DNA fragmentation was assayed, although maximal DNA fragmentation in BFA-treated cells was two- to threefold higher than that observed in etoposide-treated cells. Despite slight kinetic differences, it would appear that the apoptotic cascade is very similar in both primary parotid and submandibular acinar cells. Although limited in their long-term stability in culture, the use of primary, nonimmortalized salivary acinar cultures will also permit the use of specific transgenic animals to further characterize the molecular events involved in the regulation of salivary gland acinar cell apoptosis.

AB - Apoptosis is a highly organized cellular process that is critical for maintaining glandular homeostasis. We have used primary rat salivary acinar cells from the parotid and submandibular glands to investigate the critical regulatory events involved in apoptosis. Caspase-3 activity, cleavage of caspase substrates, and deoxyribonucleic acid (DNA) fragmentation were assayed in cells treated with etoposide, a DNA-damaging agent, or brefeldin A (BFA), a Golgi toxin. Dose-response studies showed that the sensitivity of both cell types to etoposide and BFA was similar, with 150 μM etoposide or 1.5 μM BFA inducing maximal caspase activation. However, BFA induced a more robust activation of caspase and DNA fragmentation in both cell types. Similar results were observed when the caspase cleavage of poly(adenosine 5′-diphosphate ribose) polymerase and protein kinase C delta were analyzed by Western blot. Analysis of the kinetics of apoptosis showed that caspase-3 activation was maximal at 8 h of etoposide or BFA treatment in the parotid cells and at 8-18 h in the submandibular cells. A similar time course was observed when DNA fragmentation was assayed, although maximal DNA fragmentation in BFA-treated cells was two- to threefold higher than that observed in etoposide-treated cells. Despite slight kinetic differences, it would appear that the apoptotic cascade is very similar in both primary parotid and submandibular acinar cells. Although limited in their long-term stability in culture, the use of primary, nonimmortalized salivary acinar cultures will also permit the use of specific transgenic animals to further characterize the molecular events involved in the regulation of salivary gland acinar cell apoptosis.

KW - Apoptosis

KW - Brefeldin A

KW - Etoposide

KW - Primary culture

KW - Salivary gland

UR - http://www.scopus.com/inward/record.url?scp=0141566641&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0141566641&partnerID=8YFLogxK

M3 - Article

C2 - 14505429

AN - SCOPUS:0141566641

VL - 39

SP - 170

EP - 177

JO - In Vitro Cellular and Developmental Biology - Animal

JF - In Vitro Cellular and Developmental Biology - Animal

SN - 1071-2690

IS - 3-4

ER -