Induction of cytokine messenger RNA transcripts in mouse macrophages by Listeria monocytogenes isolated from channel catfish

Li-Feng Zhang From Research Program, College of Veterinary Medicine and Mississippi Agricultural and Forestry Experiment Station (MAFES), Mississippi State University, PO Box 9825, Mississippi State, MS 39762-9825.

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 MD, MS
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Chinling Wang From Research Program, College of Veterinary Medicine and Mississippi Agricultural and Forestry Experiment Station (MAFES), Mississippi State University, PO Box 9825, Mississippi State, MS 39762-9825.

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 DVM, PhD

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Abstract

Objective

To determine whether differences exist in induction and quantity of tumor necrosis factor α (TNF-α), interleukin (IL)1ß, and IL-10 mRNA transcripts when mouse J774A.1 macrophages are infected with Listeria monocytogenes, including 2 isolates originating from channel catfish, the wild-type virulent (EGD) strain, and a nonhemolytic strain (ATCC 15313).

Samples

Listeria monocytogenes isolates from kidneys or fillets of channel catfish were used to stimulate cytokine production from mouse macrophages. The RNA from the infected macrophages was collected.

Procedure

Four hours after infection with L monocytogenes, total cellular RNA was extracted from the J774A.1 cells and reversed transcribed to cDNA, which was amplified, using specific primers for TNF- α, IL-1ß, or IL-10. The specific amplified DNA fragments were detected on polyacrylamide gels and quantified, using a reverse transcription polymerase chain reaction (PCR)-mediated ELISA.

Results

The wild-type hemolytic EGD strain and the 2 hemolytic catfish isolates of L monocytogenes induced higher amounts of TNF-α-, IL-1ß-, and IL-10- specific mRNA in J774A.1 cells than did the nonhemolytic strain.

Conclusions

Hemolysin-associated induction of TNF-α, IL-1ß, and IL-10 cytokines may be related to survival and replication of L monocytogenes in macrophages. It also suggests that the PCR-mediated ELISA procedure is a sensitive test to quantify cytokines from cell cultures. (Am J Vet Res 1998;59:717-721)

Abstract

Objective

To determine whether differences exist in induction and quantity of tumor necrosis factor α (TNF-α), interleukin (IL)1ß, and IL-10 mRNA transcripts when mouse J774A.1 macrophages are infected with Listeria monocytogenes, including 2 isolates originating from channel catfish, the wild-type virulent (EGD) strain, and a nonhemolytic strain (ATCC 15313).

Samples

Listeria monocytogenes isolates from kidneys or fillets of channel catfish were used to stimulate cytokine production from mouse macrophages. The RNA from the infected macrophages was collected.

Procedure

Four hours after infection with L monocytogenes, total cellular RNA was extracted from the J774A.1 cells and reversed transcribed to cDNA, which was amplified, using specific primers for TNF- α, IL-1ß, or IL-10. The specific amplified DNA fragments were detected on polyacrylamide gels and quantified, using a reverse transcription polymerase chain reaction (PCR)-mediated ELISA.

Results

The wild-type hemolytic EGD strain and the 2 hemolytic catfish isolates of L monocytogenes induced higher amounts of TNF-α-, IL-1ß-, and IL-10- specific mRNA in J774A.1 cells than did the nonhemolytic strain.

Conclusions

Hemolysin-associated induction of TNF-α, IL-1ß, and IL-10 cytokines may be related to survival and replication of L monocytogenes in macrophages. It also suggests that the PCR-mediated ELISA procedure is a sensitive test to quantify cytokines from cell cultures. (Am J Vet Res 1998;59:717-721)

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