Primary hematopoietic progenitors from fanconi anemia complementation type c deficient mice are hypersensitive to oxidative injury

Laura Haneline, Kelly Hiatt, Troy A. Gobbett, D. Clapp

Research output: Contribution to journalArticle

Abstract

Fanconi anemia (FA) is the most common genetic cause of bone marrow (BM) failure. The most frequently encountered DNA damage in eukaryotic cells results from reactive oxygen species (ROS) generated during normal oxidative metabolism. It remains controversial Whether FA cells are predisposed to oxidative injury. However, since FA cells have increased cytokine mediated apoptosis, chromosomal aberrations, and an increased incidence of malignancy, all of which occur in response to oxidative damage, we evaluated whether primary hematopoietic progenitors are predisposed to oxidative injury using a murine model containing a disruption of the murine homologue (Fac) of Fanconi Anemia group C. Fac -/- and Fac +/+ BM cells were incubated with increasing concentrations of hydrogen peroxide (H2O2), a widely used reagent to induce oxidative damage, and plated in a semisolid medium. Clonogenic growth of Fac -/- progenitors was significantly lower than Fac +/+ progenitors over multiple concentrations of H2O2- Because H2Ü2 is such a strong oxidizing agent and is potentially cytotoxic, elevated oxygen tension was evaluated as a second oxidative agent. We exposed liquid cultures of spleen and BM cells to 50% oxygen for 414 hours before establishing methylceüulose cultures. After a 4 hour incubation , Fac +/+ clonogenic growth was exactly the same as the 21% O2 condition, but Fac -/- clonogenic growth was reduced to 60% of colony formation in room air. A further reduction in clonogenic growth was observed when Fac -I- cells were incubated for 14 hours in 50% O2- We hypothesized that Fac -/- hematopoietic cells may have increased ROS. To test this hypothesis, we purified Scal+ cells and evaluated baseline ROS and ROS after 16 hours in liquid culture. Preliminary data suggest that Scal+ cells from Fac -/- mice have elevated baseline levels of ROS as compared to Fac +/+ cells, and that ROS are preferentially increased in Fac -/- cells following culture in cytokines. In summary, the reduction in clonogenic growth in response to two independent oxidative stimuli in primary Fac -/progenitors together with an increase in ROS, implicate oxidative injury either directly or indirectly in the pathogenesis of BM failure in FA.

Original languageEnglish
Pages (from-to)747
Number of pages1
JournalExperimental Hematology
Volume26
Issue number8
StatePublished - 1998

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Fanconi Anemia
Reactive Oxygen Species
Wounds and Injuries
Growth
Bone Marrow Cells
Bone Marrow
Cytokines
Oxygen
Eukaryotic Cells
Oxidants
Chromosome Aberrations
Hydrogen Peroxide
DNA Damage
Spleen
Cell Culture Techniques
Air
Apoptosis
Incidence

ASJC Scopus subject areas

  • Cancer Research
  • Cell Biology
  • Genetics
  • Hematology
  • Oncology
  • Transplantation

Cite this

Primary hematopoietic progenitors from fanconi anemia complementation type c deficient mice are hypersensitive to oxidative injury. / Haneline, Laura; Hiatt, Kelly; Gobbett, Troy A.; Clapp, D.

In: Experimental Hematology, Vol. 26, No. 8, 1998, p. 747.

Research output: Contribution to journalArticle

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abstract = "Fanconi anemia (FA) is the most common genetic cause of bone marrow (BM) failure. The most frequently encountered DNA damage in eukaryotic cells results from reactive oxygen species (ROS) generated during normal oxidative metabolism. It remains controversial Whether FA cells are predisposed to oxidative injury. However, since FA cells have increased cytokine mediated apoptosis, chromosomal aberrations, and an increased incidence of malignancy, all of which occur in response to oxidative damage, we evaluated whether primary hematopoietic progenitors are predisposed to oxidative injury using a murine model containing a disruption of the murine homologue (Fac) of Fanconi Anemia group C. Fac -/- and Fac +/+ BM cells were incubated with increasing concentrations of hydrogen peroxide (H2O2), a widely used reagent to induce oxidative damage, and plated in a semisolid medium. Clonogenic growth of Fac -/- progenitors was significantly lower than Fac +/+ progenitors over multiple concentrations of H2O2- Because H2{\"U}2 is such a strong oxidizing agent and is potentially cytotoxic, elevated oxygen tension was evaluated as a second oxidative agent. We exposed liquid cultures of spleen and BM cells to 50{\%} oxygen for 414 hours before establishing methylce{\"u}ulose cultures. After a 4 hour incubation , Fac +/+ clonogenic growth was exactly the same as the 21{\%} O2 condition, but Fac -/- clonogenic growth was reduced to 60{\%} of colony formation in room air. A further reduction in clonogenic growth was observed when Fac -I- cells were incubated for 14 hours in 50{\%} O2- We hypothesized that Fac -/- hematopoietic cells may have increased ROS. To test this hypothesis, we purified Scal+ cells and evaluated baseline ROS and ROS after 16 hours in liquid culture. Preliminary data suggest that Scal+ cells from Fac -/- mice have elevated baseline levels of ROS as compared to Fac +/+ cells, and that ROS are preferentially increased in Fac -/- cells following culture in cytokines. In summary, the reduction in clonogenic growth in response to two independent oxidative stimuli in primary Fac -/progenitors together with an increase in ROS, implicate oxidative injury either directly or indirectly in the pathogenesis of BM failure in FA.",
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N2 - Fanconi anemia (FA) is the most common genetic cause of bone marrow (BM) failure. The most frequently encountered DNA damage in eukaryotic cells results from reactive oxygen species (ROS) generated during normal oxidative metabolism. It remains controversial Whether FA cells are predisposed to oxidative injury. However, since FA cells have increased cytokine mediated apoptosis, chromosomal aberrations, and an increased incidence of malignancy, all of which occur in response to oxidative damage, we evaluated whether primary hematopoietic progenitors are predisposed to oxidative injury using a murine model containing a disruption of the murine homologue (Fac) of Fanconi Anemia group C. Fac -/- and Fac +/+ BM cells were incubated with increasing concentrations of hydrogen peroxide (H2O2), a widely used reagent to induce oxidative damage, and plated in a semisolid medium. Clonogenic growth of Fac -/- progenitors was significantly lower than Fac +/+ progenitors over multiple concentrations of H2O2- Because H2Ü2 is such a strong oxidizing agent and is potentially cytotoxic, elevated oxygen tension was evaluated as a second oxidative agent. We exposed liquid cultures of spleen and BM cells to 50% oxygen for 414 hours before establishing methylceüulose cultures. After a 4 hour incubation , Fac +/+ clonogenic growth was exactly the same as the 21% O2 condition, but Fac -/- clonogenic growth was reduced to 60% of colony formation in room air. A further reduction in clonogenic growth was observed when Fac -I- cells were incubated for 14 hours in 50% O2- We hypothesized that Fac -/- hematopoietic cells may have increased ROS. To test this hypothesis, we purified Scal+ cells and evaluated baseline ROS and ROS after 16 hours in liquid culture. Preliminary data suggest that Scal+ cells from Fac -/- mice have elevated baseline levels of ROS as compared to Fac +/+ cells, and that ROS are preferentially increased in Fac -/- cells following culture in cytokines. In summary, the reduction in clonogenic growth in response to two independent oxidative stimuli in primary Fac -/progenitors together with an increase in ROS, implicate oxidative injury either directly or indirectly in the pathogenesis of BM failure in FA.

AB - Fanconi anemia (FA) is the most common genetic cause of bone marrow (BM) failure. The most frequently encountered DNA damage in eukaryotic cells results from reactive oxygen species (ROS) generated during normal oxidative metabolism. It remains controversial Whether FA cells are predisposed to oxidative injury. However, since FA cells have increased cytokine mediated apoptosis, chromosomal aberrations, and an increased incidence of malignancy, all of which occur in response to oxidative damage, we evaluated whether primary hematopoietic progenitors are predisposed to oxidative injury using a murine model containing a disruption of the murine homologue (Fac) of Fanconi Anemia group C. Fac -/- and Fac +/+ BM cells were incubated with increasing concentrations of hydrogen peroxide (H2O2), a widely used reagent to induce oxidative damage, and plated in a semisolid medium. Clonogenic growth of Fac -/- progenitors was significantly lower than Fac +/+ progenitors over multiple concentrations of H2O2- Because H2Ü2 is such a strong oxidizing agent and is potentially cytotoxic, elevated oxygen tension was evaluated as a second oxidative agent. We exposed liquid cultures of spleen and BM cells to 50% oxygen for 414 hours before establishing methylceüulose cultures. After a 4 hour incubation , Fac +/+ clonogenic growth was exactly the same as the 21% O2 condition, but Fac -/- clonogenic growth was reduced to 60% of colony formation in room air. A further reduction in clonogenic growth was observed when Fac -I- cells were incubated for 14 hours in 50% O2- We hypothesized that Fac -/- hematopoietic cells may have increased ROS. To test this hypothesis, we purified Scal+ cells and evaluated baseline ROS and ROS after 16 hours in liquid culture. Preliminary data suggest that Scal+ cells from Fac -/- mice have elevated baseline levels of ROS as compared to Fac +/+ cells, and that ROS are preferentially increased in Fac -/- cells following culture in cytokines. In summary, the reduction in clonogenic growth in response to two independent oxidative stimuli in primary Fac -/progenitors together with an increase in ROS, implicate oxidative injury either directly or indirectly in the pathogenesis of BM failure in FA.

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