27 results on '"Rolf O. Ehrhardt"'
Search Results
2. Cell therapy post-production technologies: a select review of key innovations in the field
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Rolf O. Ehrhardt, Brian Schryver, and Maria L. Thompson
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Engineering ,Engineering management ,Management science ,business.industry ,Field (Bourdieu) ,Key (cryptography) ,business ,Post-production - Published
- 2017
3. Standardized Cryopreservation of Stem Cells
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Maria L. Thompson, Eric J. Kunkel, and Rolf O. Ehrhardt
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0301 basic medicine ,Andrology ,03 medical and health sciences ,030104 developmental biology ,0302 clinical medicine ,030204 cardiovascular system & hematology ,Biology ,Stem cell ,Cryopreservation - Published
- 2017
4. Process automation helps standardize and de-risk thawing of cellular therapy products
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Maria L. Thompson, Brian Schryver, and Rolf O. Ehrhardt
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Cell therapy ,Cancer Research ,Transplantation ,Oncology ,Risk analysis (engineering) ,Computer science ,Immunology ,Immunology and Allergy ,Cell Biology ,Process automation system ,Genetics (clinical) - Published
- 2017
5. Technological developments for small-scale downstream processing of cell therapies
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Maria L. Thompson, Lee Buckler, Eric J. Kunkel, and Rolf O. Ehrhardt
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0301 basic medicine ,Cancer Research ,Computer science ,Immunology ,Cytological Techniques ,Cell- and Tissue-Based Therapy ,Specimen Handling ,03 medical and health sciences ,0302 clinical medicine ,Bioreactors ,Downstream (manufacturing) ,Immunology and Allergy ,Humans ,Genetics (clinical) ,Cryopreservation ,Transplantation ,Downstream processing ,Scale (chemistry) ,Cell Biology ,030104 developmental biology ,Oncology ,Risk analysis (engineering) ,Batch Cell Culture Techniques ,030220 oncology & carcinogenesis ,Microtechnology - Abstract
Despite considerable regulatory and clinical hurdles, the development and use of cell-based therapies are gaining momentum. As more of these therapies move toward commercial approval and larger-scale distribution, associated manufacturing and processing technologies are being advanced. Modern technologies directed at downstream processing seek to distribute such therapies from the manufacturing site to the patient more efficiently and reliably. Novel small-scale downstream solutions boost the transformation of cell therapies from abstraction to reality.
- Published
- 2015
6. Blockade of Experimental Atopic Dermatitis via Topical NF-κB Decoy Oligonucleotide
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Kim Mai, Tony Muchamuel, Susan Hudak, Christopher G. De Vry, Debra Mutnick, Jie Zhang, Christi Parham, Leslie M. McEvoy, Brian Schryver, Maya Dajee, Timothy Colby, Srinivasa Prasad, Tina Le, László G. Kömüves, Aung Oo, Donald Ruhrmund, Jennifer Alleman-Sposeto, Radha Shyamsundar, and Rolf O. Ehrhardt
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Male ,Allergy ,Administration, Topical ,Apoptosis ,Mice, Inbred Strains ,Inflammation ,Dermatology ,Biochemistry ,Permeability ,Dermatitis, Atopic ,Ointments ,Mice ,chemistry.chemical_compound ,Immune system ,Antigen ,medicine ,Animals ,Edema ,Ear, External ,Molecular Biology ,Skin ,integumentary system ,business.industry ,NF-κB ,Atopic dermatitis ,Cell Biology ,medicine.disease ,Betamethasone valerate ,Oligodeoxyribonucleotides ,chemistry ,Immunology ,Atrophy ,medicine.symptom ,business ,Decoy ,Cell Division ,Immunosuppressive Agents - Abstract
Atopic dermatitis (AD) is a common chronic skin inflammatory disease. Long-term use of topical corticosteroids in skin inflammation poses risks of systemic and local side effects. The NF-kappaB transcription factor family plays a central role in the progression and maintenance of AD. This study explores the possibility of using topical NF-kappaB Decoy as a novel therapeutic alternative for targeting Th1/Th2-driven skin inflammation in experimental AD. A high-affinity, topical NF-kappaB Decoy developed for human efficacy demonstrates: (i) efficient NF-kappaB Decoy penetration in pig skin, (ii) NF-kappaB Decoy nuclear localization in keratinocytes and key immune cells, and (iii) potent "steroid-like" efficacy in a chronic dust-mite antigen skin inflammation treatment model. NF-kappaB Decoy exerts its anti-inflammatory action through the effective inhibition of essential regulators of inflammation and by induction of apoptosis of key immune cells. Unlike betamethasone valerate (BMV), long-term NF-kappaB Decoy treatment does not induce skin atrophy. Moreover, topical NF-kappaB Decoy, in contrast to BMV, restores compromised stratum corneum integrity and barrier function. Steroid withdrawal causes rapid rebound of inflammation, while the NF-kappaB Decoy therapeutic benefit was maintained for weeks. Thus, topical NF-kappaB Decoy provides a novel mechanism of reducing chronic skin inflammation with improved skin homeostasis and minimal side effects.
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- 2006
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7. Cryopreservation and Thawing of Mammalian Cells
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Eric J. Kunkel, Maria L. Thompson, and Rolf O. Ehrhardt
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Cryoprotectant ,Cell culture ,Botany ,Viability assay ,Sample integrity ,Biology ,Stem cell ,Metabolic activity ,Cryopreservation ,Cell biology - Abstract
Cryopreservation is the process by which intact living cells are preserved at cryogenic temperatures in liquid nitrogen. Freezing cells allow them to be stored, often for years, in a state where their normal metabolic activity is suspended in order to protect them from damage due to chemical reactivity and time. The cells can then be thawed and resuscitated as needed. Cells are cryopreserved in order to guard against genetic drift in continuous cell lines, or against transformation or differentiation in noncontinuous cell lines, such as stem cells and primary cells. In the case of primary cells that are isolated directly from the tissue of interest, they have a finite ability to divide; therefore, cryopreservation is necessary to preserve their unique characteristics for future experiments. Optimal cryopreservation of cells relies on proper freezing and thawing methods. To be protected from structural damage during the freeze-thaw process, mammalian cells are frozen in the presence of cryoprotectant. Post-thaw viability assays are then conducted to measure the success of the cryopreservation techniques by calculating the percentage of frozen cells that are alive and able to recover normal function once thawed. Key Concepts: Cryopreservation is used to enable long-term storage, preserve current genetic state, prevent transformation or differentiation during subculture and provide a back-up stock in case of infection/contamination during culture. A controlled rate of freezing and rapid thawing is necessary for optimal cryopreservation and recovery. Care must be taken to minimize transient warming events during transfer and storage, as it impacts viability and recovery. Active thawing results in higher cell viability and recovery than passive thawing. Timing is critical to all stages of the cryopreservation process. All materials should be ready before beginning the procedure, and steps should be taken to ensure each sample is handled with minimal delay. Method standardisation and quality control are necessary to maintain sample integrity and reproducibility. Keywords: cryopreservation; controlled-rate freezing, cryoprotectant; viability; freezing; cell thawing
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- 2014
8. The ThawSTARTM Automated Cell Thawing Platform: De-Risking Thawing From Research to the Clinic
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Maria L. Thompson, Rolf O. Ehrhardt, Brian Schryver, and Eric J. Kunkel
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Andrology ,Cancer Research ,Transplantation ,Oncology ,business.industry ,Immunology ,Immunology and Allergy ,Medicine ,Cell Biology ,business ,Genetics (clinical) - Published
- 2016
9. Standardized cryopreservation of pluripotent stem cells
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Rick I. Cohen, Maria L. Thompson, Brian Schryver, and Rolf O. Ehrhardt
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Cryopreservation ,Pluripotent Stem Cells ,Cell type ,business.industry ,Cell ,Cold storage ,Cell Biology ,General Medicine ,Biology ,Reference Standards ,Embryonic stem cell ,Cell biology ,Biotechnology ,Cell Line ,medicine.anatomical_structure ,Freezing ,medicine ,Humans ,Stem cell line ,Viability assay ,Induced pluripotent stem cell ,business ,Developmental Biology - Abstract
The successful exploitation of human cells for research, translational, therapeutic, and commercial purposes requires that effective and simple cryopreservation methods be applied for storage in local and master cell banks. Of all the cell types utilized in modern research, human embryonic stem cells and their more recent relatives, induced pluripotent stem cells, are two of the most sensitive to cryopreservation. It is frequently observed that the lack of quality control and proper processing techniques yield poor recovery of pluripotent stem cells. The procedures in this unit have been optimized for handling some of the most recalcitrant stem cell lines, and provide a method for controlled-rate freezing, using minimal equipment that affords levels of cell viability comparable to expensive controlled-rate freezers. The protocol also eliminates the requirement for isopropanol, avoiding the hazards, on-going cost, and inconsistencies associated with its use and disposal. It provides a clinically relevant, inexpensive, reliable, and user-friendly method that successfully prepares cells for long-term cold storage and ensures maximum levels of cell viability post thaw.
- Published
- 2014
10. Tissue Specificity of E- and P-Selectin Ligands in Th1-Mediated Chronic Inflammation
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Alvina Chu, Kenneth Hong, Ellen L. Berg, and Rolf O. Ehrhardt
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Immunology ,Immunology and Allergy - Abstract
The demonstrated role of E- and P-selectin ligands in the recruitment of Th1 cells raises the question of tissue specificity determination by pathogenic T cells. We took advantage of the fact that chronic Th1-mediated inflammation in the scid/scid CD4+CD45RBhigh T cell transfer model can occur at multiple tissue sites, resembling inflammatory bowel disease in the colon and psoriasis in the skin. We show that the majority of infiltrating effector T cells from psoriatic skin expresses high levels of functional P-selectin ligand (87 ± 3%), detected by P-selectin-Ig (PIg), while a significantly smaller subset of T cells from colitic lesions expresses this ligand (24 ± 2%). Similarly, E-selectin ligand is preferentially expressed on CD4+ T cells infiltrating the skin (24 ± 2%), but only on very few CD4+ T cells infiltrating the colon (CIT; 1.3 ± 0.8%). In contrast, CD4+ T cells infiltrating the skin express α4β7 at a significantly lower level than CIT (mean fluorescence intensity, 28 vs 61, respectively), although, interestingly, αEβ7 was expressed at high levels on both populations. Analysis of the disease-inducing potential of PIg+ and PIg− CD4+ CIT cells revealed that both populations not only express similar levels of the gut-homing molecule α4β7 (mean fluorescence intensity, 50 vs 56, respectively), but do not differ in their capacity to express IFN-γ. Furthermore, CIT depleted of cells expressing functional P-selectin ligand were able to induce colitis upon transfer, suggesting that induction of colitis in this model may be independent of E- and P-selectin. These results indicate that adhesion molecule expression and the homing pattern of inflammatory T cells are regulated by the local environment independently of their inflammatory capacity.
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- 1999
11. IL-12, Independently of IFN-γ, Plays a Crucial Role in the Pathogenesis of a Murine Psoriasis-Like Skin Disorder
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Kenneth Hong, Alvina Chu, Björn R. Lúdvíksson, Ellen L. Berg, and Rolf O. Ehrhardt
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Immunology ,Immunology and Allergy - Abstract
The onset of acute psoriasis and the exacerbation of chronic psoriasis are often associated with a history of bacterial infection. We demonstrate that while only few scid/scid mice develop disease when CD4+CD45Rbhigh T cells are transferred alone, coadministration of LPS plus IL-12 or staphylococcal enterotoxin B into scid/scid mice 1 day after CD4+CD45Rbhigh T cell transfer greatly enhances disease penetrance and severity. Most importantly, the skin lesions induced by this method exhibit many of the histologic hallmarks observed in human psoriasis. Skin infiltrating CD4+ T cells were predominantly memory/effector cells (CD45Rblow) and exhibited a highly polarized Th1 phenotype. To test whether the development of pathogenic T cells was dependent on their production of IFN-γ, we transferred IFN-γ−/− CD4+CD45Rbhigh T cells into scid/scid or into T, B and NK cell-deficient scid/beige mice. Surprisingly, the incidence of psoriasis was similar to scid/scid animals that received IFN-γ+/+ T cells, although acanthosis of the skin was attenuated. In contrast, the development of psoriasis was abolished if anti-IL-12 mAb was administered on day 7 and 35 after T cell transfer. Skin-derived IFN-γ−/− inflammatory cells, but not cells from anti-IL-12-treated animals, secreted substantial amounts of TNF-α, suggesting that the inflammatory effect of IFN-γ−/− T cells may be partly exerted by TNF-α and that the therapeutic effect of anti-IL-12 may depend on its ability to down-regulate both TNF-α and IFN-γ. Overall, these results suggest that IL-12, independently of IFN-γ, is able to induce pathogenic, inflammatory T cells that are able to induce psoriasiform lesions in mice.
- Published
- 1999
12. Administration of mAb Against αEβ7 Prevents and Ameliorates Immunization-Induced Colitis in IL-2−/− Mice
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Björn R. Lúdvíksson, Warren Strober, Ryuta Nishikomori, Syed K. Hasan, and Rolf O. Ehrhardt
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Immunology ,Immunology and Allergy - Abstract
We previously demonstrated that 2,4,6-trinitrophenol (TNP)-OVA immunization leads to a transmural colitis in the IL-2−/− mouse that is caused by IL-12-driven CD4+ Th1 T cells and resembles human Crohn’s disease. The integrin αEβ7 is highly expressed on colonic intraepithelial lymphocytes and has been suggested to function as a homing or retention molecule for intraepithelial lymphocytes. To evaluate the role of αEβ7 in colitis, we administered a mAb against αEβ7 to IL-2−/− mice that were immunized at the same time with TNP-OVA in CFA. To our surprise, this treatment resulted in a significantly reduced colitis severity score, 0–2 vs 3–4, that was associated with a significant reduction in CD4+ lamina propria lymphocyte subpopulation (p < 0.01). In contrast, the total number of splenic CD4+ T cells of treated animals was significantly elevated compared with that of untreated animals (3.2 ± 0.6 × 107 vs 1.2 ± 0.2 × 107; p < 0.05). Similarly, functional studies revealed that IFN-γ production by lamina propria lymphocytes isolated from IL-2−/− TNP-OVA-immunized mice treated with anti-αEβ7 was significantly lower than in untreated IL-2−/− TNP-OVA-immunized mice. In contrast, IFN-γ production by splenic cells isolated from treated IL-2−/− TNP-OVA-immunized mice was significantly higher than in untreated mice. Finally, TNP-OVA-immunized IL-2−/− mice that were treated after the colitis had been established also showed a significant decrease in mucosal inflammation after αEβ7 mAb administration. Thus, the above findings demonstrate that the onset and maintenance of inflammatory bowel disease depends on the colonic localization of lamina propria CD4+ lymphocytes expressing αEβ7.
- Published
- 1999
13. Active Wegener’s Granulomatosis Is Associated with HLA-DR+ CD4+ T Cells Exhibiting an Unbalanced Th1-Type T Cell Cytokine Pattern: Reversal with IL-10
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Björn R. Lúdvíksson, Michael C. Sneller, Kevin S. Chua, Cheryl Talar-Williams, Carol A. Langford, Rolf O. Ehrhardt, Anthony S. Fauci, and Warren Strober
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Immunology ,Immunology and Allergy - Abstract
Wegener’s granulomatosis (WG) is a granulomatous vasculitis that affects the upper respiratory tract, lung, and kidney. Since T cells make up a significant proportion of cells infiltrating granulomatous lesions in WG, we investigated the proliferative response and cytokine profile of T cells from these patients. PBMCs were isolated from 12 patients with active WG, 7 patients with inactive disease, and 12 healthy normal donors. PBMCs from clinically active WG patients exhibited increased proliferation following stimulation with either PMA/ionomycin or anti-CD2 and anti-CD28, when compared with normal donors. In addition, these PBMCs exhibited increased secretion of IFN-γ, but not of IL-4, IL-5, or IL-10. Furthermore, TNF-α production from PBMCs and CD4+ T cells isolated from patients with WG was elevated, when compared with healthy donors. In further studies, we investigated the ability of WG patients’ monocytes to produce IL-12 and showed that both inactive and active patients produced increased amounts of IL-12. Finally, the in vitro IFN-γ production by WG PBMC is inhibited in a dose-dependent manner by exogenous IL-10. These data suggest that T cells from WG patients overproduce IFN-γ and TNF-α, probably due to dysregulated IL-12 secretion, and that IL-10 may therefore have therapeutic implications for this disease.
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- 1998
14. Cryopreservation and thawing of cells
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Wayne M. Yokoyama, Rolf O. Ehrhardt, and Maria L. Thompson
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Cryopreservation ,Hybridomas ,business.industry ,Cell Survival ,Cells ,Immunology ,Rate control ,Biology ,Freezing methods ,Biotechnology ,Cell Line ,Cryoprotective Agents ,Cryoprotective Agent ,Animals ,Humans ,Dimethyl Sulfoxide ,Viability assay ,Primary cell ,Stem cell ,business ,Process engineering ,Cell Proliferation - Abstract
Successful cryopreservation of cells requires not only that the cells be handled in a proper fashion for harvesting with equipment in place to ensure consistency, reproducibility, and sterility, but also that a correct choice and amount of cryoprotective agent is added. In general, a controlled freezing rate of 1°C/min is necessary to retain optimal viability of the recovered cells. There are many variations of cell freezing methods in use, including costly electronically regulated control rate freezers, unstandardized, passive isopropyl alcohol freezing containers, and crude rudimentary devices constructed from Styrofoam boxes or paper insulation. However, for the freezing and recovery of cell lines, primary cells, and stem cell cultures, the protocol described in this unit is simple, reproducible, and successful. Not only does it eliminate the need for isopropanol, as well as the costs and hazards associated with its use and disposal, but it provides a uniform method with improved cell viability and recovery.
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- 2012
15. The Role of sIgA + B Cells in Oral Immunity
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Warren Strober and Rolf O. Ehrhardt
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Immunoglobulin A ,Cellular differentiation ,Lymphocyte Cooperation ,Administration, Oral ,Lymphocyte Activation ,General Biochemistry, Genetics and Molecular Biology ,History and Philosophy of Science ,T-Lymphocyte Subsets ,Immunity ,Animals ,Humans ,Medicine ,B-Lymphocytes ,biology ,business.industry ,General Neuroscience ,Cell Differentiation ,Immunoglobulin Isotypes ,Immunoglobulin M ,Immunoglobulin A, Secretory ,Immunology ,biology.protein ,Lymphocyte activation ,Cytokines ,business - Published
- 1995
16. The ThawSTARTM automated cell thawing system: standardizing the last step in the cryopreservation work flow
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Eric J. Kunkel, Rolf O. Ehrhardt, Brian Schryver, and Maria L. Thompson
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Cancer Research ,Transplantation ,Oncology ,Computer science ,business.industry ,Immunology ,Immunology and Allergy ,Work flow ,Cell Biology ,Process engineering ,business ,Genetics (clinical) ,Cryopreservation - Published
- 2015
17. BioT™ cryopod carrier: standardized cryogenic temperature handling of cellular therapies
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Scott Comiso, John Fink, Maria L. Thompson, Rolf O. Ehrhardt, and Eric J. Kunkel
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Cancer Research ,Transplantation ,Biot number ,Computer science ,Immunology ,Airlock ,Critical zone ,Process (computing) ,Sampling (statistics) ,Classification scheme ,Cell Biology ,Reliability engineering ,Oncology ,Immunology and Allergy ,Work site ,Cryogenic temperature ,Genetics (clinical) - Abstract
personnel and process flows, and resulting air flow patterns. Within the BSC critical zone, sites should be in close proximity to operations such as centrally located settle plates and particle sensors not more than 1 foot from the work site. Surface viable sites should be in close proximity to processing activity, at locations contacted by operator gloves, areas where materials transfer from lower classification, and other facility surfaces (e.g. floors, walls) to assess disinfectant efficacy. Suggested sampling frequencies are shown in Tables 1 and 2. EM Levels: Cell therapy areas typically will have ISO 5 BSCs placed within an ISO 7 room with ISO 8 support such as a gowning airlock. This classification scheme is outlined in USP Cellular and Tissue-Based Products. Suggested EM levels for cell therapy aseptic processing are shown in Table 3. Conclusion: Considerations are offered to help cell therapy manufacturers to set-up and an appropriate risk-based EM sampling program that meets criteria of being meaningful, manageable, and defendable.
- Published
- 2015
18. Non-viral delivery of nuclear factor-kappaB decoy ameliorates murine inflammatory bowel disease and restores tissue homeostasis
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Christopher G. De Vry, Christi Parham, Sarvesh Adda, Leslie M. McEvoy, Kim Mai, Srinivasa Prasad, Nicole Kahoud, Tony Muchamuel, Jennifer Hoffman, Radhika Garlapati, Jie Zhang, Brian Schryver, Rolf O. Ehrhardt, Radha Shyamsundar, László G. Kömüves, Tina Le, Carlos Lorenzana, and Maya Dajee
- Subjects
Budesonide ,Male ,Colon ,Inflammatory bowel disease ,Oxazolone ,chemistry.chemical_compound ,Mice ,Medicine ,Animals ,Homeostasis ,Colitis ,Interleukin 6 ,Tissue homeostasis ,Mice, Inbred BALB C ,Microscopy, Confocal ,biology ,business.industry ,Dextran Sulfate ,Inflammatory Bowel Disease ,Gastroenterology ,Gene Transfer Techniques ,NF-kappa B ,T helper cell ,Genetic Therapy ,medicine.disease ,Inflammatory Bowel Diseases ,Ulcerative colitis ,Mice, Inbred C57BL ,Disease Models, Animal ,medicine.anatomical_structure ,chemistry ,Oligodeoxyribonucleotides ,Trinitrobenzenesulfonic Acid ,Immunology ,Cancer research ,biology.protein ,Female ,Inflammation Mediators ,business ,medicine.drug - Abstract
Background: Nuclear factor-κB (NF-κB) is a key transcriptional regulator of inflammatory bowel disease (IBD). Aim: To investigate the therapeutic potential of a locally administered “non-viral” nuclear factor-κB decoy (NFκBD) in multiple experimental models of IBD. Methods: A fully phosphorothioated decoy oligonucleotide with improved stability that specifically binds NF-κB and blocks inflammatory mediators regulated by this transcription factor without the help of viral envelope-assisted delivery was developed. The therapeutic effects of NFκBD were studied in the trinitrobenzene sulphonic acid, oxazolone and dextran sodium sulphate induced colitis models. Results: Intracolonic administration of NFκBD results in the delivery of NFκBD to inflammatory cells and a reduction of NF-κB heterodimers. In the T helper cell 1-driven trinitrobenzene sulphonic acid-induced colitis model, mice receiving NFκBD treatment exhibit a dose-dependent reduction in disease severity and a more rapid recovery to normal body weight, similar to a clinically relevant dose of budesonide. Clinical efficacy was corroborated by considerable reductions in colitis pathology and tissue levels of several pro-inflammatory markers, including tumour necrosis factor α, interleukin 6, interleukin 1β and monocyte chemotactic protein 1. NFκBD also mitigates disease activity in the T helper cell 2-like oxazolone colitis and epithelial injury-related acute dextran sodium sulphate colitis models. Interestingly, restoration of tissue homeostasis is observed in NFκBD-treated animals with the rapid re-emergence of functional goblet cells and a return to normal patterns of cell proliferation in the mucosal epithelium and smooth muscle cell layers. Conclusions: These data support the potential use of “naked” NFκBD as a cross-functional therapeutic in IBD, and show for the first time that it can facilitate the restoration of colon homeostasis and function.
- Published
- 2006
19. Role of IFN-gamma in Th1 differentiation: IFN-gamma regulates IL-18R alpha expression by preventing the negative effects of IL-4 and by inducing/maintaining IL-12 receptor beta 2 expression
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Ronald B. Smeltz, June Chen, Rolf O. Ehrhardt, and Ethan M. Shevach
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Mice, Inbred A ,Immunology ,Mice, Transgenic ,Biology ,Neutralization ,Interferon-gamma ,Mice ,Adjuvants, Immunologic ,Immunology and Allergy ,Animals ,Receptor ,Interleukin 4 ,Cells, Cultured ,Mice, Inbred BALB C ,Receptors, Interleukin-18 ,Interleukin-18 ,Receptors, Interleukin-12 ,Cell Differentiation ,Receptors, Interleukin ,Th1 Cells ,Interleukin-12 ,Cell biology ,DNA-Binding Proteins ,Mice, Inbred C57BL ,Interleukin 12 ,Female ,Interleukin-4 ,Signal transduction ,Interleukin-18 Receptor alpha Subunit - Abstract
Two key events occur during the differentiation of IFN-gamma-secreting Th1 cells: up-regulation of IL-12Rbeta2 and IL-12-driven up-regulation of IL-18Ralpha. We previously demonstrated that IL-12-driven up-regulation of IL-18Ralpha expression is severely impaired in IFN-gamma(-/-) mice. However, it was unclear from these studies how IFN-gamma influenced IL-18Ralpha since IFN-gamma alone had no direct effect on IL-18Ralpha expression. In the absence of IL-4, IL-12-dependent up-regulation of IL-18Ralpha/IL-12Rbeta2 was independent of IFN-gamma. However, in the presence of IL-4, IFN-gamma functions to limit the negative effects of IL-4 on both IL-18Ralpha and IL-12Rbeta2. Neutralization of IL-4 restored IL-12-driven up-regulation of IL-18Ralpha/IL-12Rbeta2 in an IFN-gamma-independent fashion. In the absence of both IL-12 and IL-4, IFN-gamma up-regulates IL-12beta2 expression and primes IFN-gamma-producing Th1 cells. When T cells were primed in the presence of IL-4, no correlation was found between the levels of expression of the IL-18Ralpha or the IL-12Rbeta2 and the capacity of these cells to produce IFN-gamma, suggesting that IL-4 may also negatively affect IL-12-mediated signal transduction and thus Th1 differentiation. These data clarify the role of IFN-gamma in regulation of IL-18Ralpha/IL-12Rbeta2 during both IL-12-dependent and IL-12-independent Th1 differentiation.
- Published
- 2002
20. Validation of a novel portable freezing device in the optimal freezing of peripheral blood mononuclear cells for potential cell therapy use
- Author
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Qizhi Tang, Maria L. Thompson, Rolf O. Ehrhardt, and Brian Schryver
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Cancer Research ,Transplantation ,business.industry ,Immunology ,Cell Biology ,Peripheral blood mononuclear cell ,Cryopreservation ,Cell therapy ,Clinical study ,Oncology ,Immunology and Allergy ,Medicine ,Drug product ,business ,Genetics (clinical) ,Biomedical engineering - Abstract
In order to ensure a standardized and consistent Drug Product freezing and to minimize batch-to-batch differences in cell recovery and viability postthaw, TxCell scientists tested the CoolCell container, a passive freezing device, as an alternative to the classical controlled rate freezer. Results showed that by using a CoolCell freezing device, Ag-Tregs can be successfully cryopreserved and recovered in a standardized way acceptable to the processing and manufacturing of cell therapies. TxCell now intends to use the CoolCell device in its phase IIb clinical study with its lead AgTreg cell product candidate, Ovasave . Use of the CoolCell passive freezing device for cell therapy manufacturing of Ag-Treg represents a new standardized method of cell therapy product cryopreservation. The ability to develop and store functional Treg in a cost-effective and reproducible manner is an important milestone in the ultimate use of these cells in the clinic.
- Published
- 2014
21. Persistence of pathogenic CD4+ Th1-like cells in vivo in the absence of IL-12 but in the presence of autoantigen
- Author
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Kenneth Hong, Rolf O. Ehrhardt, and Ellen L. Berg
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CD4-Positive T-Lymphocytes ,Adoptive cell transfer ,Cell Survival ,Immunology ,Inflammation ,Mice, SCID ,Biology ,Autoantigens ,Lymphocyte Depletion ,Mice ,In vivo ,Recurrence ,T-Lymphocyte Subsets ,medicine ,Immunology and Allergy ,Animals ,Psoriasis ,Lymph node ,Immunization Schedule ,Autoimmune disease ,Mice, Inbred BALB C ,Anti-Inflammatory Agents, Non-Steroidal ,Antibodies, Monoclonal ,Th1 Cells ,medicine.disease ,Adoptive Transfer ,Interleukin-12 ,Disease Models, Animal ,medicine.anatomical_structure ,Chronic Disease ,biology.protein ,Interleukin 12 ,Female ,Lymph ,Lymph Nodes ,medicine.symptom ,Antibody ,Injections, Intraperitoneal - Abstract
Despite recent successful treatment of murine autoimmune disease with anti-IL-12 mAb, it has not yet been addressed whether anti-IL-12 mAb can also be effective in late stages of disease and whether it can provide lasting protection against recurrence, especially during continued presence of autoantigen. We used a newly developed psoriasis model in scid/scid mice, which allows easy tracking of pathogenic T cells, to show that when anti-IL-12 mAb is given for 2 wk (1 mg/wk) in the late stage of severe disease, inflammation is greatly reduced, as measured by ear thickness and histology (scores, 1.1 ± 0.1 vs 2.0 ± 0.4). Moreover, prolonged treatment (4 wk) of chronic psoriatic mice with high doses of mAb (1 mg/wk; prolonged active anti-inflammatory treatment (PAAIT)) results in the almost complete resolution of lesions (scores, 0.3 ± 0.1 vs 2.7 ± 0.2). Surprisingly, however, despite these significant treatment results, the psoriasis-like lesions return soon after the anti-IL-12 mAb treatment is discontinued. This rapid relapse of disease may be attributed to large populations of activated CD4+ T cells present in the lymph nodes of PAAIT animals still expressing an effector/memory phenotype (CD45RBlow, L-selectinlow). Upon stimulation in vitro such PAAIT lymph node cells secrete high amounts of IFN-γ (129 ng/ml); when transferred into naive scid/scid animals they are able to rapidly induce disease without costimulation. Our data indicates an alternative IL-12-independent pathway for pathogenic Th-1-like cells in vivo during the chronic phase of disease that allows these cells to persist and maintain their pathogenicity in the draining lymph tissue of the autoimmune site.
- Published
- 2001
22. T helper type 2 cell differentiation occurs in the presence of interleukin 12 receptor beta2 chain expression and signaling
- Author
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Rolf O. Ehrhardt, Ryuta Nishikomori, and Warren Strober
- Subjects
CD4-Positive T-Lymphocytes ,Immunology ,Mice, Transgenic ,Lymphocyte Activation ,interleukin 4 ,Interleukin 21 ,Interferon-gamma ,Mice ,Th2 Cells ,reversibility ,cytokine ,Immunology and Allergy ,Cytotoxic T cell ,Animals ,IL-2 receptor ,Interleukin 4 ,Interleukin 3 ,CD40 ,biology ,Receptors, Interleukin-12 ,Cell Differentiation ,Receptors, Interleukin ,STAT4 Transcription Factor ,Th1 Cells ,Molecular biology ,DNA-Binding Proteins ,Interleukin 15 ,signal transducer and activator of transcription 4 ,biology.protein ,Interleukin 12 ,Trans-Activators ,Original Article ,Interleukin-4 ,T helper type 1 ,Signal Transduction - Abstract
The differentiation of CD4(+) T cells into T helper type 1 (Th1) cells is driven by interleukin (IL)-12 through the IL-12 receptor beta2 (IL-12Rbeta2) chain, whereas differentiation into Th2 cells is driven by IL-4, which downregulates IL-12Rbeta2 chain. We reexamined such differentiation using IL-12Rbeta2 chain transgenic mice. We found that CD4(+) T cells from such mice were able to differentiate into Th2 cells when primed with IL-4 or IL-4 plus IL-12. In the latter case, the presence of IL-4 suppressed interferon (IFN)-gamma production 10-100-fold compared with cells cultured in IL-12 alone. Finally, in studies of the ability of IL-12 to convert Th2 cells bearing a competent IL-12R to the Th1 cells, we showed that: (a) T cells bearing the IL-12Rbeta2 chain transgene and primed under Th2 conditions could not be converted to Th1 cells by repeated restimulation under Th1 conditions; and (b) established Th2 clones transfected with the IL-12Rbeta2 chain construct continued to produce IL-4 when cultured with IL-12. These studies show that IL-4-driven Th2 differentiation can occur in the presence of persistent IL-12 signaling and that IL-4 inhibits IFN-gamma production under these circumstances. They also show that established Th2 cells cannot be converted to Th1 cells via IL-12 signaling.
- Published
- 2000
23. When immunization leads to autoimmunity: chronic inflammation as a result of thymic and mucosal dysregulation in IL-2 knock-out mice
- Author
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Rolf O. Ehrhardt and Bjorn R. Ludviksson
- Subjects
Lymphocyte ,Immunology ,Inflammation ,Autoimmunity ,Thymus Gland ,medicine.disease_cause ,Mice ,Immune system ,Immunity ,medicine ,Immunology and Allergy ,Animals ,Immunity, Mucosal ,Mice, Knockout ,biology ,Chemistry ,medicine.anatomical_structure ,Viral replication ,Polyclonal antibodies ,biology.protein ,Interleukin 12 ,Interleukin-2 ,Immunization ,medicine.symptom - Abstract
In 1965 Wheelock identified IFN--/ as a new soluble substance produced by activated T cells that inhibited viral replication in fibroblasts [ I ] . This seminal observation led to the explosive research on lymphocyte mediated activators during various aspects of the cellular immune response [2-71, and was followed in 1976 by the discovery of IL-2 by Morgan et al. [S]. IL-2 was discovered as a secreted product of T cells activated by a polyclonal mitogen, phytohemagglutinin (PHA), that stimulated the proliferation of
- Published
- 2000
24. Differential activation requirements of isotype-switched B cells
- Author
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Nils Lycke, Belinda Gray, Rolf O. Ehrhardt, John K. Inman, Warren Strober, and G R Harriman
- Subjects
T cell ,Immunology ,Naive B cell ,Receptors, Antigen, B-Cell ,Biology ,Lymphocyte Activation ,Cell Line ,Mice ,Peyer's Patches ,medicine ,Immunology and Allergy ,Animals ,Intestinal Mucosa ,Antigen-presenting cell ,B cell ,B-Lymphocytes ,Mice, Inbred BALB C ,CD40 ,Lymphokine ,Germinal center ,Immunoglobulin D ,Germinal Center ,Molecular biology ,Immunoglobulin Class Switching ,Immunoglobulin A ,B-1 cell ,medicine.anatomical_structure ,Immunoglobulin M ,biology.protein ,Female ,Spleen - Abstract
In the present studies, we compared the activation requirements of sIgM+/sIgD+ B cells with those of isotype-switched sIgM-/sIgA+ B cells. We found that whereas sIgM+ B cells respond to T cell-independent (TI) and T cell-dependent (TD) Ag with no significant bias toward one stimulus, sIgA+ B cells were deficient in their ability to respond to antigen receptor cross-linking but responded remarkably well to TD stimuli. Thus, dextran-conjugated anti-IgA antibody (anti-IgA-dextran), anti-kappa-dextran, or various immobilized anti-IgA antibodies (Ab) induced only low-level IgA B cell proliferation and no IgA secretion in the presence of various lymphokines; in marked contrast, sIgA+ B cells responded to cognate and noncognate T cell stimulation as well as to stimulation by CD40 ligand-bearing fibroblasts by secreting large amounts of IgA (up to 240 000 ng/ml per 10(5) cells). This pattern of sIgA+ B cell responsiveness was noted with both germinal center peanut agglutininhi (PNAhi) and non-germinal center PNAlo B cells. In confirmation of these results, whole Peyer's patch or lamina propria cell populations containing less than 15% sIgA+ B cells stimulated with a noncognate T cell stimulus or T cell membranes secreted mainly IgA (68%-94% of the total Ig secreted) and relatively little IgM. The strict T cell dependence of IgA B cell activation and differentiation provides important insights into immune responses of mucosal tissues and must be considered in the development of vaccines, particularly those designed to stimulate mucosal tissues containing large numbers of isotype-switched B cells.
- Published
- 1996
25. Regulation of IgA B Cell Development
- Author
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Rolf O. Ehrhardt and Warren Strober
- Subjects
medicine.anatomical_structure ,Chemistry ,medicine ,B cell ,Cell biology - Published
- 1994
26. Chronic intestinal inflammation: an unexpected outcome in cytokine or T cell receptor mutant mice
- Author
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Warren Strober and Rolf O. Ehrhardt
- Subjects
Mechanosensation ,Ratón ,medicine.medical_treatment ,T-cell receptor ,Mutant ,Receptors, Antigen, T-Cell ,T lymphocyte ,Biology ,Inflammatory Bowel Diseases ,Models, Biological ,General Biochemistry, Genetics and Molecular Biology ,Mice, Mutant Strains ,Pathogenesis ,Disease Models, Animal ,Mice ,Immune system ,Cytokine ,Immunology ,medicine ,Animals ,Cytokines ,Receptor ,Neuroscience - Published
- 1993
27. Induction and Regulation of Colitis in IL-2-/- Mice
- Author
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Belinda Gray, Bjorn R. Ludviksson, Warren Strober, and Rolf O. Ehrhardt
- Subjects
business.industry ,Immunology ,Emergency Medicine ,Medicine ,Colitis ,Critical Care and Intensive Care Medicine ,business ,medicine.disease - Published
- 1997
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