| 1. |
- Hoogduijn, Martin J, et al.
(författare)
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Effects of freeze-thawing and intravenous infusion on mesenchymal stromal cell gene expression.
- 2016
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Ingår i: Stem Cells and Development. - : Mary Ann Liebert Inc. - 1557-8534 .- 1547-3287.
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Tidskriftsartikel (refereegranskat)abstract
- Mesenchymal stromal cells (MSC) are increasingly used as an investigative therapeutic product for immune disorders and degenerative disease. Typically, MSC are isolated from human tissue, expanded in culture and cryopreserved until usage. The safety and efficacy of MSC therapy will depend on the phenotypical and functional characteristics of MSC. The freeze-thawing procedure may change these characteristics. Furthermore, the cells encounter a microenvironment after administration that may impact their properties. It has been demonstrated that the majority of MSC localize to the lungs after intravenous infusion, making this the site to study the effects of the in vivo milieu on administered MSC. In the present study we investigated the effect of freeze-thawing and the mouse lung microenvironment on human adipose tissue-derived MSC. There were effects of freeze-thawing on the whole genome expression profile of MSC, although the effects did not exceed inter-donor differences. There were no major changes in the expression of hemostatic regulators on transcriptional level, but significantly increased expression of procoagulant tissue factor on the surface of thawed adipose MSC, correlating with increased procoagulant activity of thawed cells. Exposure for 2h to the lung microenvironment had a major effect on MSC gene expression and affected several immunological pathways. This indicates that MSC undergo functional changes shortly after infusion and this may influence the efficacy of MSC to modulate inflammatory responses. The results of this study demonstrate that MSC rapidly alter in response to the local milieu and that disease specific conditions may shape MSC after administration.
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| 2. |
- Ignatowicz, Lech, et al.
(författare)
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Mycobacterium tuberculosis infection interferes with HIV vaccination in mice.
- 2012
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Ingår i: PloS one. - : Public Library of Science (PLoS). - 1932-6203. ; 7:7, s. e41205-
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Tidskriftsartikel (refereegranskat)abstract
- Tuberculosis (TB) has emerged as the most prominent bacterial disease found in human immunodeficiency virus (HIV)-positive individuals worldwide. Due to high prevalence of asymptomatic Mycobacterium tuberculosis (Mtb) infections, the future HIV vaccine in areas highly endemic for TB will often be administrated to individuals with an ongoing Mtb infection. The impact of concurrent Mtb infection on the immunogenicity of a HIV vaccine candidate, MultiHIV DNA/protein, was investigated in mice. We found that, depending on the vaccination route, mice infected with Mtb before the administration of the HIV vaccine showed impairment in both the magnitude and the quality of antibody and T cell responses to the vaccine components p24Gag and gp160Env. Mice infected with Mtb prior to intranasal HIV vaccination exhibited reduced p24Gag-specific serum IgG and IgA, and suppressed gp160Env-specific serum IgG as compared to respective titers in uninfected HIV-vaccinated controls. Importantly, in Mtb-infected mice that were HIV-vaccinated by the intramuscular route the virus neutralizing activity in serum was significantly decreased, relative to uninfected counterparts. In addition mice concurrently infected with Mtb had fewer p24Gag-specific IFN-γ-expressing T cells and multifunctional T cells in their spleens. These results suggest that Mtb infection might interfere with the outcome of prospective HIV vaccination in humans.
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| 3. |
- Ignatowicz, Lech
(författare)
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Studies on the host immune response during pulmonary TB and during M.tuberculosis / HIV co-infection
- 2012
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Population studies have shown that HIV-infected individuals co-infected with Mycobacterium tuberculosis (MTB) are at up to 10-fold higher risk of developing active tuberculosis (TB) than their seronegative counterparts. Co-infected individuals also progress faster to AIDS than patients infected only with the virus. Therefore, understanding the relation between these two diseases is of a crucial importance. The overall aim of this thesis was to investigate the interaction between HIV and MTB infection from the perspective of host immunity. Using in vitro and in vivo models, we investigated innate and adaptive immune responses to better understand the mechanisms behind the increased susceptibility to MTB and HIV during co-infection. In the first study we examined regulation of the host immune response during pulmonary TB mediated by CD103+ dendritic cells (αE-DC) and CD4+ Foxp3+ regulatory T (Treg) cells. We found that in resistant mice, the number of lung αE-DCs increased dramatically during MTB infection. In contrast, susceptible DBA/2 mice which failed to recruit αE-DCs even during chronic infection. αE-DCs remained essentially TNF negative but contained a high number of TGFβ-producing cells in the lungs of infected mice. Further, we show that Treg cells in resistant and susceptible mice induced interferon IFNγ during pulmonary TB. We also showed that the Treg cell population was diminished in the lungs, but not in the draining pulmonary lymph nodes, of susceptible mice. The reduced number of lung Treg cells in susceptible mice coincided with an increased bacterial load and exacerbated lung pathology compared to resistant strains. These results demonstrate that immune-regulatory mechanisms may play an important role in protective immune responses during pulmonary TB. In study II we analyzed the impact of MTB-infected macrophages on DC functionality, including HIV transinfecting ability. An in vitro system was used in which human monocyte-derived DCs were exposed to soluble factors released by MTB- or BCG-infected macrophages. Soluble factors secreted from infected macrophages resulted in the production of proinflammatory cytokines and partial upregulation of maturation markers on DCs. Interestingly, the HIV transinfecting ability of DCs was enhanced In summary, this study shows that DCs respond to soluble factors released by mycobacteria-infected macrophages. These findings highlight the important role of bystander effects mediated by MTB-infected macrophages, and suggest it contributes to DC-mediated HIV spread and amplification in co-infected individuals. In study III we assessed the impact of MTB infection on the immunogenicity of a HIV vaccine. We found that, depending on the vaccination route, mice infected with MTB before the administration of the HIV vaccine showed impairment in both the magnitude and the quality of antibody and T cell responses directed towards the vaccine. Mice infected with MTB prior to HIV vaccination exhibited reduced IgG and IgA antibody levels and neutralizing activity of serum against the virus In addition mice infected with MTB had significantly lower HIV-specific multifunctional T cells. These results suggest that chronic MTB infection might interfere with the outcome of prospective HIV vaccination in humans. In study IV we developed a new murine model of MTB/HIV co-infection using conventional, immunocompetent mouse strains. We utilized the chimeric EcoNDK virus together with a low dose MTB aerosol infection. To date, we have observed that the viral burden increased in the spleen and in the lungs of animals infected with MTB prior to virus inoculation. We have also shown that co-infection did not affect control of bacterial growth. During the early stage of co-infection, EcoNDK induced a significantly higher frequency of MTB antigen-specific CD8+ T-cells in the spleen. Also, MTB-specific CD8+ T-cells in both MTB-infected and MTB/EcoNDK co-infected animals were enriched for Tim3- and PD1 expressing cells. In conclusion, this small animal model may provide a useful tool to increase our understanding of how MTB and HIV influence the host immune response during co-infection in vivo.
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| 4. |
- Mazurek, Jolanta, et al.
(författare)
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Mycobacteria-infected bystander macrophages trigger maturation of dendritic cells and enhance their ability to mediate HIV transinfection
- 2012
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Ingår i: European Journal of Immunology. - : Wiley. - 1521-4141 .- 0014-2980. ; 42:5, s. 1192-1202
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Tidskriftsartikel (refereegranskat)abstract
- Synergistic interplay between Mycobacterium tuberculosis (Mtb) and HIV in coinfected ind-ividuals leads to the acceleration of both tuberculosis and HIVdisease. Mtb, as well as HIV, may modulate the function of many immune cells, including DCs. To dissect the bystander impact of Mfs infected with Mtb on DC functionality, we here investigated changes in DC phenotype, cytokine profiles, and HIV-1 transinfecting ability. An in vitro system was used in which human monocyte-derived DCs were exposed to soluble factors released by Mfs infected with mycobacteria, including virulent clinical Mtb isolates and nonvirulent BCG. Soluble factors secreted from Mtb-infected Mfs, and to a lesser extent BCG-infected Mfs, resulted in the production of proinflammatory cytokines and partial upregulation of DC maturation markers. Interestingly, the HIV-1 transinfecting ability of DCs was enhanced upon exposure to soluble factors released by Mtb-infected Mfs. In summary, our study shows that DCs exposed to soluble factors released by mycobacteria-infected Mfs undergo maturation and display an augmented ability to transmit HIV-1 in trans. These findings highlight the important role of bystander effects during the course of MtbHIV coinfection and suggest that Mtb-infected Mfs may contribute to an environment that supports DC-mediated spread and amplification of HIV in coinfected individuals.
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| 5. |
- Moll, Guido, et al.
(författare)
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Cryopreserved or fresh mesenchymal stromal cells : Only a matter of taste or key to unleash the full clinical potential of MSC therapy?
- 2016
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Ingår i: Advances in Experimental Medicine and Biology. - Cham : Springer International Publishing. - 2214-8019 .- 0065-2598. ; 951, s. 77-98
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Bokkapitel (refereegranskat)abstract
- Mesenchymal stromal cells (MSCs) harbor great therapeutic potential for numerous diseases. From early clinical trials, success and failure analysis, bench-to-bedside and back-to-bench approaches, there has been a great gain in knowledge, still leaving a number of questions to be answered regarding optimal manufacturing and quality of MSCs for clinical application. For treatment of many acute indications, cryobanking may remain a prerequisite, but great uncertainty exists considering the therapeutic value of freshly thawed (thawed) and continuously cultured (fresh) MSCs. The field has seen an explosion of new literature lately, outlining the relevance of the topic. MSCs appear to have compromised immunomodulatory activity directly after thawing for clinical application. This may provide a possible explanation for failure of early clinical trials. It is not clear if and how quickly MSCs recover their full therapeutic activity, and if the “cryo stun effect” is relevant for clinical success. Here, we will share our latest insights into the relevance of these observations for clinical practice that will be discussed in the context of the published literature. We argue that the differences of fresh and thawed MSCs are limited but significant. A key issue in evaluating potency differences is the time point of analysis after thawing. To date, prospective double-blinded randomized clinical studies to evaluate potency of both products are lacking, although recent progress was made with preclinical assessment. We suggest refocusing therapeutic MSC development on potency and safety assays with close resemblance of the clinical reality.
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| 6. |
- Moll, Guido, et al.
(författare)
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Different Procoagulant Activity of Therapeutic Mesenchymal Stromal Cells Derived from Bone Marrow and Placental Decidua
- 2015
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Ingår i: Stem Cells and Development. - : Mary Ann Liebert Inc. - 1547-3287 .- 1557-8534. ; 28:S2, s. 50-51
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Tidskriftsartikel (refereegranskat)abstract
- While therapeutic mesenchymal stromal/stem cells (MSCs) have usually been obtained from bone marrow, perinatal tissues have emerged as promising new sources of cells for stromal cell therapy. In this study, we present a first safety follow-up on our clinical experience with placenta-derived decidual stromal cells (DSCs), used as supportive immunomodulatory and regenerative therapy for patients with severe complications after allogeneic hematopoietic stem cell transplantation (HSCT). We found that DSCs are smaller, almost half the volume of MSCs, which may favor microvascular passage. DSCs also show different hemocompatibility, with increased triggering of the clotting cascade after exposure to human blood and plasma in vitro. After infusion of DSCs in HSCT patients, we observed a weak activation of the fibrinolytic system, but the other blood activation markers remained stable, excluding major adverse events. Expression profiling identified differential levels of key factors implicated in regulation of hemostasis, such as a lack of prostacyclin synthase and increased tissue factor expression in DSCs, suggesting that these cells have intrinsic blood-activating properties. The stronger triggering of the clotting cascade by DSCs could be antagonized by optimizing the cell graft reconstitution before infusion, for example, by use of low-dose heparin anticoagulant in the cell infusion buffer. We conclude that DSCs are smaller and have stronger hemostatic properties than MSCs, thus triggering stronger activation of the clotting system, which can be antagonized by optimizing the cell graft preparation before infusion. Our results highlight the importance of hemocompatibility safety testing for every novel cell therapy product before clinical use, when applied using systemic delivery.
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| 7. |
- Moll, Guido, et al.
(författare)
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Do Cryopreserved Mesenchymal Stromal Cells Display Impaired Immunomodulatory and Therapeutic Properties?
- 2014
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Ingår i: Stem Cells. - : Oxford University Press (OUP). - 1066-5099 .- 1549-4918. ; 32:9, s. 2430-2442
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Tidskriftsartikel (refereegranskat)abstract
- We have recently reported that therapeutic mesenchymal stromal cells (MSCs) have low engraftment and trigger the instant blood mediated inflammatory reaction (IBMIR) after systemic delivery to patients, resulting in compromised cell function. In order to optimize the product, we compared the immunomodulatory, blood regulatory, and therapeutic properties of freeze-thawed and freshly harvested cells. We found that freeze-thawed MSCs, as opposed to cells harvested from continuous cultures, have impaired immunomodulatory and blood regulatory properties. Freeze-thawed MSCs demonstrated reduced responsiveness to proinflammatory stimuli, an impaired production of anti-inflammatory mediators, increased triggering of the IBMIR, and a strong activation of the complement cascade compared to fresh cells. This resulted in twice the efficiency in lysis of thawed MSCs after 1 hour of serum exposure. We found a 50% and 80% reduction in viable cells with freshly detached as opposed to thawed in vitro cells, indicating a small benefit for fresh cells. In evaluation of clinical response, we report a trend that fresh cells, and cells of low passage, demonstrate improved clinical outcome. Patients treated with freshly harvested cells in low passage had a 100% response rate, twice the response rate of 50% observed in a comparable group of patients treated with freeze-thawed cells at higher passage. We conclude that cryobanked MSCs have reduced immunomodulatory and blood regulatory properties directly after thawing, resulting in faster complement-mediated elimination after blood exposure. These changes seem to be paired by differences in therapeutic efficacy in treatment of immune ailments after hematopoietic stem cell transplantation.
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| 8. |
- Strömdahl, Ann Charlotte, et al.
(författare)
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Peptide-coated polyurethane material reduces wound infection and inflammation
- 2021
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Ingår i: Acta Biomaterialia. - : Elsevier BV. - 1742-7061. ; 128, s. 314-331
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Tidskriftsartikel (refereegranskat)abstract
- There is an urgent need for treatments that not only reduce bacterial infection that occurs during wounding but that also target the accompanying excessive inflammatory response. TCP-25, a thrombin-derived antibacterial peptide, scavenges toll-like receptor agonists such as endotoxins and lipoteichoic acid and prevents toll-like receptor-4 dimerization to reduce infection-related inflammation in vivo. Using a combination of biophysical, cellular, and microbiological assays followed by experimental studies in mouse and pig models, we show that TCP-25, when delivered from a polyurethane (PU) material, exerts anti-infective and anti-inflammatory effects in vitro and in vivo. Specifically, TCP-25 killed the common wound pathogens, Pseudomonas aeruginosa and Staphylococcus aureus, in both in vitro and in vivo assays. Furthermore, after its release from the PU material, the peptide retained its capacity to induce its helical conformation upon endotoxin interaction, yielding reduced activation of NF-κB in THP-1 reporter cells, and diminished accumulation of inflammatory cells and subsequent release of IL-6 and TNF-α in subcutaneous implant models in vivo. Moreover, in a porcine partial thickness wound infection model, TCP-25 treated infection with S. aureus, and reduced the concomitant inflammatory response. Taken together, these findings demonstrate a combined antibacterial and anti-inflammatory effect of TCP-25 delivered from PU in vitro, and in mouse and porcine in vivo models of localized infection-inflammation. Statement of significance: Local wound infections may result in systemic complications and can be difficult to treat due to increasing antimicrobial resistance. Surgical site infections and biomaterial-related infections present a major challenge for hospitals. In recent years, various antimicrobial coatings have been developed for infection prevention and current concepts focus on various matrices with added anti-infective components, including various antibiotics and antiseptics. We have developed a dual action wound dressing concept where the host defense peptide TCP-25, when delivered from a PU material, targets both bacterial infection and the accompanying inflammation. TCP-25 PU showed efficacy in in vitro and experimental wound models in mouse and minipigs.
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