Limits...
A focused Real Time PCR strategy to determine GILZ expression in mouse tissues.

Cari L, Ricci E, Gentili M, Petrillo MG, Ayroldi E, Ronchetti S, Nocentini G, Riccardi C - Results Immunol (2015)

Bottom Line: Glucocorticoid-Induced Leucine Zipper (GILZ) is a glucocorticoid-inducible gene that mediates glucocorticoid anti-inflammatory effects.An inappropriate primer pair amplified also GILZ pseudogene and TSC22d1iso3, thus producing misleading results.This quantitative evaluation may be used to better characterize the role of GILZ and L-GILZ in mice and may be translated to humans.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Section of Pharmacology, University of Perugia Medical School, P.le L. Severi 1, 06132 Perugia, Italy.

ABSTRACT
Glucocorticoid-Induced Leucine Zipper (GILZ) is a glucocorticoid-inducible gene that mediates glucocorticoid anti-inflammatory effects. GILZ and the isoform L-GILZ are expressed in a variety of cell types, especially of hematopoietic origin, including macrophages, lymphocytes and epithelial cells, and strongly upregulated upon glucocorticoid treatment. A quantitative analysis of GILZ expression in mouse tissues is technically difficult to perform because of the presence of a pseudogene and the high homology of GILZ gene with other genes of TSC22 family. We here propose specific primer pairs to be used in Real Time PCR to avoid unwanted amplification of GILZ pseudogene and TSC-22 family member d1iso3. These primer pairs were used to determine GILZ and L-GILZ expression, in either untreated or in vivo and in vitro dexamethasone-treated tissues. Results indicate that GILZ and L-GILZ are upregulated by glucocorticoids, being GILZ more sensitive to glucocorticoid induction than L-GILZ, but they are differently expressed in all examined tissues, confirming a different role in specific cells. An inappropriate primer pair amplified also GILZ pseudogene and TSC22d1iso3, thus producing misleading results. This quantitative evaluation may be used to better characterize the role of GILZ and L-GILZ in mice and may be translated to humans.

No MeSH data available.


GILZ and L-GILZ expression investigated by Real Time PCR in in vitro DEX-treated lymphoid tissues. Cells from thymus, spleen, bone marrow and lymph nodes were treated with two doses of DEX, for 3 h. The fold values in each group were calculated versus the same untreated cells, set equal to 1, on a base 2 log scale. Results are mean of triplicates±SD of two experiments.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4664734&req=5

f0020: GILZ and L-GILZ expression investigated by Real Time PCR in in vitro DEX-treated lymphoid tissues. Cells from thymus, spleen, bone marrow and lymph nodes were treated with two doses of DEX, for 3 h. The fold values in each group were calculated versus the same untreated cells, set equal to 1, on a base 2 log scale. Results are mean of triplicates±SD of two experiments.

Mentions: Lymphoid tissues from untreated mice were further characterized in vitro by 3 h DEX treatment. Two doses of DEX were used and results shown in Fig. 4 demonstrate that there is always upregulation of GILZ and L-GILZ after DEX exposure. L-GILZ and GILZ upregulation is dose-dependent only in thymus and bone marrow, while is dose-independent in spleen and lymph nodes, suggesting that a low DEX dose is enough to induce both genes.


A focused Real Time PCR strategy to determine GILZ expression in mouse tissues.

Cari L, Ricci E, Gentili M, Petrillo MG, Ayroldi E, Ronchetti S, Nocentini G, Riccardi C - Results Immunol (2015)

GILZ and L-GILZ expression investigated by Real Time PCR in in vitro DEX-treated lymphoid tissues. Cells from thymus, spleen, bone marrow and lymph nodes were treated with two doses of DEX, for 3 h. The fold values in each group were calculated versus the same untreated cells, set equal to 1, on a base 2 log scale. Results are mean of triplicates±SD of two experiments.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4664734&req=5

f0020: GILZ and L-GILZ expression investigated by Real Time PCR in in vitro DEX-treated lymphoid tissues. Cells from thymus, spleen, bone marrow and lymph nodes were treated with two doses of DEX, for 3 h. The fold values in each group were calculated versus the same untreated cells, set equal to 1, on a base 2 log scale. Results are mean of triplicates±SD of two experiments.
Mentions: Lymphoid tissues from untreated mice were further characterized in vitro by 3 h DEX treatment. Two doses of DEX were used and results shown in Fig. 4 demonstrate that there is always upregulation of GILZ and L-GILZ after DEX exposure. L-GILZ and GILZ upregulation is dose-dependent only in thymus and bone marrow, while is dose-independent in spleen and lymph nodes, suggesting that a low DEX dose is enough to induce both genes.

Bottom Line: Glucocorticoid-Induced Leucine Zipper (GILZ) is a glucocorticoid-inducible gene that mediates glucocorticoid anti-inflammatory effects.An inappropriate primer pair amplified also GILZ pseudogene and TSC22d1iso3, thus producing misleading results.This quantitative evaluation may be used to better characterize the role of GILZ and L-GILZ in mice and may be translated to humans.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Section of Pharmacology, University of Perugia Medical School, P.le L. Severi 1, 06132 Perugia, Italy.

ABSTRACT
Glucocorticoid-Induced Leucine Zipper (GILZ) is a glucocorticoid-inducible gene that mediates glucocorticoid anti-inflammatory effects. GILZ and the isoform L-GILZ are expressed in a variety of cell types, especially of hematopoietic origin, including macrophages, lymphocytes and epithelial cells, and strongly upregulated upon glucocorticoid treatment. A quantitative analysis of GILZ expression in mouse tissues is technically difficult to perform because of the presence of a pseudogene and the high homology of GILZ gene with other genes of TSC22 family. We here propose specific primer pairs to be used in Real Time PCR to avoid unwanted amplification of GILZ pseudogene and TSC-22 family member d1iso3. These primer pairs were used to determine GILZ and L-GILZ expression, in either untreated or in vivo and in vitro dexamethasone-treated tissues. Results indicate that GILZ and L-GILZ are upregulated by glucocorticoids, being GILZ more sensitive to glucocorticoid induction than L-GILZ, but they are differently expressed in all examined tissues, confirming a different role in specific cells. An inappropriate primer pair amplified also GILZ pseudogene and TSC22d1iso3, thus producing misleading results. This quantitative evaluation may be used to better characterize the role of GILZ and L-GILZ in mice and may be translated to humans.

No MeSH data available.