Evangelic Diaconic Hospital

Leipzig, Germany

Evangelic Diaconic Hospital

Leipzig, Germany

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Fricke S.,Fraunhofer Institute for Cell Therapy and Immunology | Fricke S.,University of Leipzig | Hilger N.,Fraunhofer Institute for Cell Therapy and Immunology | Fricke C.,Ambulante Dienste | And 9 more authors.
Cellular and Molecular Life Sciences | Year: 2014

This is the first report showing that an epitope-specific ex vivo modulation of an allogeneic hematopoietic stem cell graft by the anti-human CD4 antibody MAX.16H5 IgG1 simultaneously facilitates the anti-tumor capacity of the graft (Graft-versus-leukemia effect, GvL) and the long-term suppression of the deleterious side effect Graft-versus-host-disease (GvHD). To distinguish and consolidate GvL from GvHD, the anti-human CD4 antibody MAX16.H5 IgG1 was tested in murine GvHD and tumor models. The survival rate was significantly increased in recipients receiving a MAX.16H5 IgG1 short-term (2 h) pre-incubated graft even when tumor cells were co-transplanted or when recipient mice were treated by MAX.16H5 IgG1 before transplantation. After engraftment, regulatory T-cells are generated only supporting the GvL effect. It was also possible to transfer the immune tolerance from GvHD-free recipient chimeras into third party recipient mice without the need of reapplication of MAX.16H5 IgG1 anti-human CD4 antibodies. These findings are also benefical for patients with leukemia when no matched related or unrelated donor is available and provides a safer allogeneic HSCT, which is more effective against leukemia. It also facilitates allogeneic (stem) cell transplantations for other indications (e.g., autoimmune-disorders). © 2013 Springer Basel.


Fricke S.,Fraunhofer Institute for Cell Therapy and Immunology | Fricke S.,University of Leipzig | Fricke C.,Solothurner Spitaler AG | Oelkrug C.,Fraunhofer Institute for Cell Therapy and Immunology | And 11 more authors.
Cellular and Molecular Life Sciences | Year: 2010

Non-adherent bone marrow-derived cells (NA-BMCs) are a mixed cell population that can give rise to multiple mesenchymal phenotypes and that facilitates hematopoietic recovery. We characterized NA-BMCs by flow cytometry, fibroblast colony-forming units (CFU-f), real-time PCR, and in in vivo experiments. In comparison to adherent cells, NA-BMCs expressed high levels of CD11b+ and CD90+ within the CD45+ cell fraction. CFU-f were significantly declining over the cultivation period, but NA-BMCs were still able to form CFU-f after 5 days. Gene expression analysis of allogeneic NA-BMCs compared to bone marrow (BM) indicates that NA-BMCs contain stromal, mesenchymal, endothelial cells and monocytes, but less osteoid, lymphoid, and erythroid cells, and hematopoietic stem cells. Histopathological data and analysis of weight showed an excellent recovery and organ repair of lethally irradiated mice after NA-BMC transplantation with a normal composition of the BM. © Springer Basel AG 2010.


Fricke S.,Fraunhofer Institute for Cell Therapy and Immunology | Fricke S.,University of Leipzig | Fricke C.,Medizinische Dienste SRO | Oelkrug C.,Fraunhofer Institute for Cell Therapy and Immunology | And 6 more authors.
Mycoses | Year: 2012

An early diagnosis of an invasive fungal infection is essential for the initiation of a specific antifungal therapy and to avoid unnecessary discontinuation of a baseline therapy for haematological or oncological diseases. A real-time PCR assay for the detection and strain identification of Aspergillus species from culture strains was evaluated. DNA preparation was evaluated in contaminated culture media, urine and serum. A LightCycler PCR to differentiate various Aspergillus species was established. A real-time PCR assay for the detection of Aspergillus species was improved and was able to detect and differentiate medically important Aspergillus spp. The sensitivity of the test was <10 plasmid equivalents/assay. The real-time PCR assay is a useful tool for the rapid identification of Aspergillus species and might be useful as an early diagnostic tool to detect an invasive fungal infection. A real-time PCR protocol was improved by generating plasmid standards, additional generation of melting curves for species identification and the correlation between the melting temperature and the nucleotide exchanges within the used 18S rRNA gene region. © 2011 Blackwell Verlag GmbH.


Fricke S.,Fraunhofer Institute for Cell Therapy and Immunology | Fricke S.,University of Leipzig | Rothe K.,Fraunhofer Institute for Cell Therapy and Immunology | Hilger N.,Fraunhofer Institute for Cell Therapy and Immunology | And 9 more authors.
Cytometry Part A | Year: 2012

Regulatory CD4 +CD25 +FoxP3 + T cells (T regs) suppress immunological reactions. However, the effect of adding T regs to hematopoietic stem cell grafts on recovery and graft versus host disease (GvHD) is unknown. T regs from splenocytes of C57Bl/6 and Balb/c wild-type mice were isolated by MACS separation and analyzed by flow cytometry. Using a murine syngeneic transplantation model that clearly distinguishes between donor and host hematopoiesis, we showed that co-transplantation of bone marrow cells (BMCs) with high levels of T regs leads to a 100% survival of the mice and accelerates the hematopoietic recovery significantly (full donor chimerism). In allogeneic transplantation, bone marrow and T regs co-transplantation were compared to allogeneic bone marrow transplantation with or without the addition of splenocytes. Survival, leukocyte recovery, chimerism at days -2, 19, 33, and 61 for murine CD4, human CD4, HLA-DR3, murine CD3, murine CD8, murine Balb/c-H2K d, murine C57Bl/6-H2K b, and GvHD appearance were analyzed. Allogeneic bone marrow transplantation requires the addition of splenocytes to reach engraftment. Mice receiving grafts with bone marrow, splenocytes and high levels of allogeneic T regs died within 28 days (hematopoietic failure). Here, we show also detailed flow cytometric data reagarding analysis of chimerism after transplantation in unique murine hematopoietic stem cell transplantation models. Our findings showed that the syngeneic co-transplantation of CD4 +, CD25 +, FoxP3 + T-cells and BMCs induced a stimulating effect on reconstitution of hematopoiesis after irradiation. However, in the allogeneic setting the co-transplantation of T regs aggravates the engraftment of transplanted cells. © 2012 International Society for Advancement of Cytometry.


Oelkrug C.,Fraunhofer Institute for Cell Therapy and Immunology | Hilger N.,Fraunhofer Institute for Cell Therapy and Immunology | Schonfelder U.,Evangelic Diaconic Hospital | Boltze J.,Fraunhofer Institute for Cell Therapy and Immunology | And 10 more authors.
International Journal of Radiation Biology | Year: 2014

Purpose: The time- and dose-dependent reconstitution of hematopoiesis after radiation exposure is strongly related to the stem cell population and can be used to predict hematological parameters. These parameters allow further insight into the hematopoietic system and might lead to the development of novel stem cell transplantation models. Materials and methods: CD4-/- C57Bl/6 mice, transgenic for human CD4 and HLA-DR3, were irradiated in a single (3, 6, 8 and 12 Gy) and fractionated (6 × 1 Gy, 6 × 1.5 Gy, 6 × 2 Gy; twice daily) dose regimen. Blood was analyzed weekly for red blood cells (RBC), hemoglobin concentration (Hb), hematocrit (HCT) and white blood cells (WBC). Organ and tissue damage after irradiation were examined by histopathology. Results: The recovery curves for RBC, Hb, HCT and WBC showed the same velocity (< 1 week) for all radiation doses (3-12 Gy) starting at different, dose-dependent times. The only dose-dependent parameter was defined by the beginning of the recovery process (dose-dependent shift) and higher doses were related to a later recovery of the hematopoietic system. The RBC, Hb and HCT recovery was followed by a saturation curve reaching a final concentration independent of the radiation dose. Histological analysis of the bone marrow in the single dose cohort showed a dose-dependent reduction of the cellularity in the bone marrow cavities. The fractioned radiation dose cohort resulted in a regeneration of all bone marrow cavities. Conclusion: Specific functions were developed to describe the reconstitution of hematological parameters after total body irradiation. © 2014 Informa UK, Ltd.


Fricke S.,Fraunhofer Institute for Cell Therapy and Immunology | Fricke S.,University of Leipzig | Fricke C.,Ambulante Dienste LUPS | Schimmelpfennig C.,Fraunhofer Institute for Cell Therapy and Immunology | And 8 more authors.
Journal of Applied Microbiology | Year: 2010

Aims: We established a real-time PCR assay for the detection and strain identification of Candida species and demonstrated the ability to differentiate between Candida albicans the most common species, and also Candida parapsilosis, Candida glabrata, Candida tropicalis and Candida dubliniensis by LightCycler PCR and melting curve analysis. Methods and Results: The DNA isolation from cultures and serum was established using the QIAmp Tissue Kit. The sensitivity of the assay was ≥ 2 genome equivalentsassay. It was possible to differentiate all investigated Candida species by melting curve analysis, and no cross-reaction to human DNA or Aspergillus species could be observed. Conclusions: The established real-time PCR assay is a useful tool for the rapid identification of Candida species and a base technology for more complex PCR assays. Significance and Impact of the Study: We carried out initial steps in validation of a PCR assay for the detection and differentiation of medically relevant Candida species. The PCR was improved by generating PCR standards, additional generation of melting curves for species identification and the possibility to investigate different specimens simultaneously. © 2010 The Society for Applied Microbiology.

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