Quantitative comparison of click beetle and firefly luciferases for in vivo bioluminescence imaging

Miloud, Tewfik; Henrich, Carmen; Hämmerling, Günter J.

doi:10.1117/1.2800386

Cited by 43 publications

(41 citation statements)

References 7 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…For generation of Foxp3.LuciDTR BAC transgenic mice, a construct composed of the cDNAs for enhanced GFP (eGFP), the human DTR, and CBGr99 luciferase (27) was generated. Coding sequences were separated by a self-cleaving 2A peptide sequence from porcine teschovirus-1 for stoichiometric production of the three individual transgenic proteins (28).…”

Section: Mice and Treg Depletionmentioning

confidence: 99%

See 1 more Smart Citation

Dendritic Cells Support Homeostatic Expansion of Foxp3+ Regulatory T Cells in Foxp3.LuciDTR Mice

Suffner

Hochweller

Kühnle

et al. 2010

The Journal of Immunology

123

132

View full text Add to dashboard Cite

Foxp3+CD4+ regulatory T cells (Tregs) are crucial in maintaining self-tolerance and limiting immune responses to pathogens. Shifting the sensitive balance between Tregs and effector T cells requires extensive knowledge of the homeostatic properties of the different T cell populations. For the investigation of Treg homeostatic expansion, we introduce in this study novel BAC transgenic mice, designated Foxp3.LuciDTR, coexpressing enhanced GFP, luciferase for bioluminescence imaging of Tregs, and the diphtheria toxin receptor (DTR) for specific ablation of Tregs. Of several founder lines, Foxp3.LuciDTR-4 mice displayed ∼95% Treg depletion following injection of DT, resulting in activation of conventional CD4+ T cells, probably due to lack of control by Tregs. In contrast, Foxp3.LuciDTR-3 mice displayed only ∼70% Treg depletion without concomitant activation of CD4+ T cells and represented, therefore, a suitable model to study Treg homeostasis in an environment where other T cell populations were not altered. After depletion, the Treg compartment recovered to its original size in ∼2 wk. This recovery was mediated in a thymus-independent fashion by homeostatic proliferation of the surviving, nondepleted Tregs. The proliferating Tregs acquired an activated phenotype and maintained their suppressive capacity. Studies involving DT-mediated depletion of dendritic cells in CD11c.DOG mice showed that dendritic cells were required for optimal Treg homeostasis. In addition, IL-2 was identified as an essential factor for homeostatic recovery of the Treg compartment. These results show that Treg homeostasis is specifically regulated by the size of the Treg compartment and is independent of proliferation of conventional T cells.

show abstract

Section: Mice and Treg Depletionmentioning

confidence: 99%

“…injection of 4.5 mg D-luciferin (SynChem, Elk Grove Village, IL) using the IVIS 100 imaging system (Xenogen, Hopkinton, MA) as described (27). Analysis was performed with LivingImage software (v2.50, Xenogen).…”

Section: Bioluminescence Imagingmentioning

confidence: 99%

Dendritic Cells Support Homeostatic Expansion of Foxp3+ Regulatory T Cells in Foxp3.LuciDTR Mice

Suffner

Hochweller

Kühnle

et al. 2010

The Journal of Immunology

123

132

View full text Add to dashboard Cite

show abstract

“…In vivo bioluminescence imaging was performed as described previously (42). Mice were injected i.p with D-Luciferin (150 mg/kg of body weight); 5 min later mice were imaged using an IVIS 100 charge-coupled device imaging system (Xenogen) for 1 min.…”

Section: Bioluminescence Imagingmentioning

confidence: 99%

Organ-Specific Cellular Requirements for In Vivo Dendritic Cell Generation

Miloud

Fiegler

Suffner

et al. 2012

The Journal of Immunology

Self Cite

View full text Add to dashboard Cite

Bone marrow-derived dendritic cell (DC) precursors seed peripheral organs, where they encounter diverse cellular environments during their final differentiation into DCs. Flt3 ligand (Flt3-L) is critical for instructing DC generation throughout different organs. However, it remains unknown which cells produce Flt3-L and, importantly, which cellular source drives DC development in such a variety of organs. Using a novel BAC transgenic Flt3-L reporter mouse strain coexpressing enhanced GFP and luciferase, we show ubiquitous Flt3-L expression in organs and cell types. These results were further confirmed at the protein level. Although Flt3-L was produced by immune and nonimmune cells, the source required for development of the DC compartment clearly differed among organs. In lymphoid organs such as the spleen and bone marrow, Flt3-L production by hemopoietic cells was critical for generation of normal DC numbers. This was unexpected for the spleen because both immune and nonimmune cells equally contributed to the Flt3-L content in that organ. Thus, localized production rather than the total tissue content of Flt3-L in spleen dictated normal splenic DC development. No differences were observed in the number of DC precursors, suggesting that the immune source of Flt3-L promoted pre-cDC differentiation in spleen. In contrast, DC generation in the lung, kidney, and pancreas was mostly driven by nonhematopoietic cells producing Flt3-L, with little contribution by immune cells. These findings demonstrate a high degree of flexibility in Flt3-L–dependent DC generation to adapt this process to organ-specific cellular environments encountered by DC precursors during their final differentiation.

show abstract

“…Tumor-bearing mice were shaved and imaged 5 min after i.p. injection of 4.5 mg D-luciferin (SynChem, Elk Grove Village, IL, USA) using the IVIS 100 imaging system (Xenogen, Hopkinton, MA, USA) as previously described [31,47]. Analysis was performed with LivingImage software (v2.50, Xenogen).…”

Section: Blimentioning

confidence: 99%

Efficient Treg depletion induces T‐cell infiltration and rejection of large tumors

et al. 2010

View full text Add to dashboard Cite

There are a number of factors that hamper immunotherapy of cancer. For example, tumors exhibit an aberrant vasculature that appears to form a barrier against T-cell infiltration. Another major obstacle is created by Treg. So far, conventional depletion of Treg with anti-CD25 antibodies, which eliminate only 70% of Treg, has failed to significantly reduce the growth of established tumors. Using Foxp3.LuciDTR-4 mice, we show here that 90-95% Treg depletion resulted in complete regression of large established tumors, whereas 70% depletion was ineffective. The extensive Treg depletion induced a number of processes that are critical for tumor rejection, including activation of tumorspecific CD81 T cells and enhanced infiltration of these cells into the tumor. The precise mechanism of enhanced infiltration is not known, but normalization of the tumor vasculature is assumed to assist infiltration. Indeed, we observed that 90% Treg depletion caused normalization of the tumor vasculature as indicated by a reduction in leakiness and the number of dilated vessels. These results suggest that for clinical immunotherapy of cancer, it would be desirable to have reagents that allow high-level depletion of Treg, which, in conjunction with treatment modalities such as vaccination, may concomitantly increase T-cell activation and infiltration.Key words: Immunotherapy . Treg . Tumor immunology Supporting Information available online IntroductionGrowing evidence from various malignant entities including melanoma, colorectal, ovarian, cervical, hepatocellular, gastric and urothelial carcinoma indicates that an increased number of tumor infiltrating lymphocytes correlates with favorable disease course and longer survival [1][2][3][4][5][6][7][8][9]. A major obstacle for infiltration appears to be the tumor endothelial barrier. Neoangiogenesis of blood vessels is necessary for tumor survival, but the resulting tumor vasculature usually displays an abnormal architecture characterized by dilated and fragile vessels resulting in leakiness, hypoxia, acidosis and high interstitial pressure [10,11]. Interestingly, normalization of the tumor vasculature by experimental manipulation leads to increased infiltration and tumor destruction [12]. Thus, normalization of the tumor vasculature emerges as a novel and promising approach for immunotherapy. Several studies have attempted to interfere with the growth of mouse transplantation tumors by PC-61 monotherapy. Usually an effect on tumor growth has been observed only in prophylactic settings when Treg were depleted before or no later than 2 days after tumor inoculation, whereas PC-61-mediated Treg depletion in animals carrying established tumors had generally no or only little effect on tumor growth [25][26][27][28]. Significant therapeutic success was only observed when complex combinatorial approaches were applied such as total body irradiation combined with adoptive T-cell transfer and vaccination [29], and additional treatment with anti-PDL1 antibody [30], but usually mice were treated when tu...

show abstract

Quantitative comparison of click beetle and firefly luciferases for in vivo bioluminescence imaging

Cited by 43 publications

References 7 publications

Dendritic Cells Support Homeostatic Expansion of Foxp3+ Regulatory T Cells in Foxp3.LuciDTR Mice

Dendritic Cells Support Homeostatic Expansion of Foxp3+ Regulatory T Cells in Foxp3.LuciDTR Mice

Organ-Specific Cellular Requirements for In Vivo Dendritic Cell Generation

Efficient Treg depletion induces T‐cell infiltration and rejection of large tumors

Contact Info

Product

Resources

About