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- Title
- INTERLEUKIN-7 DIFFERENTIALLY REGULATES THE ACTIVATION, PROLIFERATION, AND HOMING OF T-CELLS: IMPLICATIONS FOR IMMUNOTHERAPY.
- Creator
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Kittipatarin, Christina, Khaled, Annette, University of Central Florida
- Abstract / Description
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Interleukin-7 (IL-7) is an essential lymphocyte growth factor required for the survival and proliferation of mature T-cells. As a therapeutic agent, IL-7 has the potential to restore T-cell numbers following immune depletion and to promote immunity against cancers. While the survival function of IL-7 is well established, less is known about how it supports T-cell expansion, a critical feature of the immune response. To study the biological effects of IL-7 on T-cell growth, we developed an in...
Show moreInterleukin-7 (IL-7) is an essential lymphocyte growth factor required for the survival and proliferation of mature T-cells. As a therapeutic agent, IL-7 has the potential to restore T-cell numbers following immune depletion and to promote immunity against cancers. While the survival function of IL-7 is well established, less is known about how it supports T-cell expansion, a critical feature of the immune response. To study the biological effects of IL-7 on T-cell growth, we developed an in vitro culture technique to expand T-cells ex vivo. A significant finding from our studies is that IL-7 did not induce the expansion of all T-cells, indicating that there are inherent differences in the response of individual T-cell subsets to IL-7. Culture with high doses of IL-7 (>150 ng/ml) preferentially expanded CD8 T-cells, but lead to the dramatic loss of CD4 T-cells which favored growth in lower dosages of IL-7 (<10 ng/ml). This effect was due to the regulation of LCK, a kinase predominantly associated with the CD4 co-receptor. We found that transgenic expression of the CD4 co-receptor onto CD8 T-cells promoted their growth in lower concentrations of IL-7. Conversely, inhibition of LCK activity in CD4 T-cells restored their responsiveness to high doses of IL-7 as indicated by the activation of the transcription factor STAT5, in a manner similar to CD8 T-cells. Interestingly, not all CD8 T-cells expanded in high doses of IL-7 and this effect was specific to CD8 T-cells that expressed an activated memory phenotype. We found that IL-7 promoted the proliferation of CD8 T-cells through Cdc25A, a phosphatase required for cell cycle progression. Expression of a constitutively active Cdc25A could maintain T-cell survival and proliferation in the absence of IL-7, demonstrating that Cdc25A is a crucial transducer of IL-7 growth signals. Inhibition of Cdc25A was sufficient to decrease proliferation and down-regulate the expression of activation/ memory markers on CD8 T-cells in the presence of IL-7. Upon further study, we identified a novel role for IL-7 through Cdc25A in the regulation of CD62L, an adhesion molecule required for lymph node entry. Culture with high doses of IL-7 down-regulated the expression of CD62L, suggesting that high doses of IL-7 could affect the ability of T-cells to enter or re-enter the lymph nodes. Collectively, our findings demonstrate that IL-7 administration at the supraphysiological doses currently used in the clinical trials could have a negative impact on the growth of CD4 T-cells and the homing of CD8 T-cells to the lymph nodes, effects which can impede the generation of an effective immune response.
Show less - Date Issued
- 2010
- Identifier
- CFE0003372, ucf:48449
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0003372
- Title
- An Exploratory Study of Physiologic Responses to a Passive Exercise Intervention in Mechanically-ventilated Critically Ill Adults.
- Creator
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Amidei, Christina, Sole, Mary, Byers, Jacqueline, Covelli, Maureen, Smith, Gerald, University of Central Florida
- Abstract / Description
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Muscle weakness is the most common and persistent problem after a critical illness. Early mobilization of the critically ill patient, beginning with passive exercise and progressing to ambulation, may mitigate muscle effects of the critical illness. However, mobilization may produce adverse effects, especially early in the illness when risk for physiologic deterioration is common. If safe, introducing a mobility intervention early in the illness may facilitate ventilator weaning, shorten...
Show moreMuscle weakness is the most common and persistent problem after a critical illness. Early mobilization of the critically ill patient, beginning with passive exercise and progressing to ambulation, may mitigate muscle effects of the critical illness. However, mobilization may produce adverse effects, especially early in the illness when risk for physiologic deterioration is common. If safe, introducing a mobility intervention early in the illness may facilitate ventilator weaning, shorten intensive care unit and hospitals stays, and improve functional status and quality of life for mechanically ventilated critically ill patients. The aim of this study was assess the cardiopulmonary and inflammatory responses to an early standardized passive exercise protocol (PEP) in mechanically ventilated critically ill patients. Using a quasi-experimental within-subjects repeated measures design, mechanically ventilated critically ill adults who were physiologically stable received a single standardized PEP within 72 hours of intubation. The PEP consisted of 20 minutes of bilateral passive leg movement delivered by continuous passive motion machines at a rate of 20 repetitions per minute, from 5-75 degrees, to simulate very slow walking. Physiologic parameters evaluated included heart rate (HR), mean blood pressure (MBP), oxygen saturation, and cytokine levels (IL-6 and IL-10), obtained before, during, and after the intervention. The Behavioral Pain Scale (BPS), administered before, during and after the intervention was used as a measure of participant comfort. The study sample was comprised of 18 (60%) males and 12 (40%) females, with a mean age of 56.5 years (SD 16.9 years), who were primarily Caucasian (N=18, 64%). Mean APACHE II scores for the sample were 23.8 (SD 6.2) with a mean predicted death rate of 48.8 (SD 19.8), indicating moderate mortality risk related to illness severity. Number of comorbidities ranged from 1-10 (X=4). All participants completed the intervention with no adverse events. Using repeated measures analysis of variance (rmANOVA), no significant differences were found in HR, MBP, or oxygen saturation at any of the four time points in comparison to baseline. BPS scores were significantly reduced (F(2.43, 70.42)=4.08, p=.02) at 5 and 10 minutes after the PEP was started, and were sustained at 20 minutes and for one hour after the PEP was completed. IL-6 was significantly reduced (F(1.60, 43.1)=4.351, p=.03) at the end of the intervention but not at the end of the final rest period. IL-10 values were not significantly different at any of the three time points, but IL-6 to IL-10 ratios did decrease significantly (F(1.61, 43.38)=3.42, p=.05) at the end of the PEP and again after a 60 minute rest period. Passive leg exercise was well tolerated by study participants. HR, MBP, and oxygen saturation were maintained within order set-specified ranges during and for one hour after activity, and patient comfort improved during and after the intervention. A downward trend in HR was noted in participants, which is contrary to usual HR response during exercise, and may represent clinical improvement in this population related to reduction in pain. Reduction of mean IL-6 values at the end of the PEP, but not after the rest period, suggests that the PEP was responsible for the initial IL-6 improvement. Improvement of IL-6 to IL-10 ratios from the end of the PEP to the end of the final rest period suggests that IL-10, although non-significant, may have had some effect, indicating that IL-10 increases may occur later than the time period of study.Passive exercise can be used as an approach to facilitating mobilization in mechanically ventilated critically ill adults until they are ready to participate in more active exercise. It could be that more frequent and aggressive exercise, such as passive cycling at faster rates, four times daily, will be tolerated in this population. While the understanding of clinical significance of cytokine profiles in critically ill patients is still evolving, cytokine levels may be useful in explaining benefits of mobilization in this population. Further study is required to replicate the impact of passive exercise on pain, and it may represent a novel approach to pain management in critically ill patients.
Show less - Date Issued
- 2012
- Identifier
- CFE0004350, ucf:49424
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0004350
- Title
- Bone Morphogenetic Protein-7 (BMP-7) Polarizes Monocytes into M2 Macrophages.
- Creator
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Rocher, Crystal, Singla, Dinender, Siddiqi, Shadab, Sugaya, Kiminobu, University of Central Florida
- Abstract / Description
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Atherosclerosis is an inflammatory disease in which an accumulation of fatty acids and cholesterol occurs to form a plaque in small and large arteries. Monocyte polarization to classic M1 macrophages or alternative M2 macrophages is an important area of research that can determine the severity of disease progression. BMP-7 is a key growth factor responsible for directing differentiation of mesenchymal stem cells into brown fat cells, suggesting a role of BMP-7 in cellular plasticity; however,...
Show moreAtherosclerosis is an inflammatory disease in which an accumulation of fatty acids and cholesterol occurs to form a plaque in small and large arteries. Monocyte polarization to classic M1 macrophages or alternative M2 macrophages is an important area of research that can determine the severity of disease progression. BMP-7 is a key growth factor responsible for directing differentiation of mesenchymal stem cells into brown fat cells, suggesting a role of BMP-7 in cellular plasticity; however, its role in monocyte polarization is yet to be revealed. In the current study, we hypothesize that monocyte treatment with BMP-7 will significantly result in increased polarization of monocytes into M2 macrophages and increased expression of anti-inflammatory cytokines. To that effect, we have established a stress induced cell culture system with monocytes (THP-1 cells) and apoptotic conditioned medium (ACM), simulating injury, to understand the effects of BMP-7 on M2 macrophage polarization from monocytes. Our data demonstrates that the BMP type 2 receptor (BMPR2) is found on monocytes and its activation is significantly (p(<)0.05) increased in both monocytes and M2 macrophages following treatment with BMP-7. Furthermore, a significant (p(<)0.05) increase of M2 macrophages in the BMP-7 treated group was shown following immunostaining with CD206 and arginase-1, two M2 macrophage markers, whereas a significant (p(<)0.05) decrease of iNOS expression, an M1 macrophage marker, was shown. Moreover, treatment with BMP-7 resulted in significantly (p(<)0.05) increased expression of IL-10 and IL-1ra, two anti-inflammatory cytokines, but significantly (p(<)0.05) decreased levels of the pro-inflammatory cytokines, MCP-1, IL-6 and TNF-?. We also hypothesize that polarization of monocytes to M2 macrophages occurs through activation of SMAD1/5/8 and PI3K-Akt-mTOR pathways. Upon BMP-7 binding to its receptor, BMPR2, activation of SMAD1/5/8 occurs which then activates the p85 subunit of PI3K resulting in downstream activation of Akt and mTOR. Our data shows that following treatment with BMP-7, expression of p-SMAD1/5/8, p-PI3K, p-Akt and p-mTOR is significantly (p(<)0.05) increased compared to controls whereas p-PTEN, an inhibitor of the PI3K pathway, is significantly (p(<)0.05) decreased in the BMP-7 treated group compared to controls. In conclusion, our data reveals that BMP-7 polarizes monocytes into M2 macrophages and it achieves this through activation of the PI3K-Akt-mTOR pathway, which will have significant applications for atherosclerosis treatment.
Show less - Date Issued
- 2013
- Identifier
- CFE0004922, ucf:49617
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0004922
- Title
- THE PRESENCE OF PAIN RELATED CYTOKINES AND CHEMOKINES IN SCHWANNOMAS AND THEIR POTENTIAL ASSOCIATION WITH CHRONIC PAIN IN SCHWANNOMATOSIS.
- Creator
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Nagamoto, Jackson D, Fernandez-Valle, Cristina, University of Central Florida
- Abstract / Description
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Schwannomatosis (SWN) is a genetic disorder that predisposes affected individuals to develop multiple Schwannomas anywhere in the peripheral nervous system. This can be due to a mutation in the LZTR1 or SMARCB1 genes on chromosome 22. SWN has the defining clinical symptom of chronic pain and a lack of vestibular schwannomas, which sets it apart from other, related disorders such as Neurofibromatosis Type II (NF2). Currently, it is unknown what causes the chronic pain of SWN patients but it is...
Show moreSchwannomatosis (SWN) is a genetic disorder that predisposes affected individuals to develop multiple Schwannomas anywhere in the peripheral nervous system. This can be due to a mutation in the LZTR1 or SMARCB1 genes on chromosome 22. SWN has the defining clinical symptom of chronic pain and a lack of vestibular schwannomas, which sets it apart from other, related disorders such as Neurofibromatosis Type II (NF2). Currently, it is unknown what causes the chronic pain of SWN patients but it is hypothesized that cytokines may have promote the neuropathic pain experienced by patients. This study investigates the presence of the chemokine CCL2 and the cytokine IL6 in human SWN schwannomas and non-SWN schwannomas to determine if there is a difference in the presence of these cytokines between the two tumor types. It was demonstrated that all of the SWN schwannomas expressed both CCL2 and IL6 whereas the non-SWN schwannomas expressed only one or the other protein if either. These results indicate that the presence of these cytokines within the SWN schwannomas is different from non-SWN schwannomas and could be a potential contributing factor in the occurrence of neuropathic pain experienced by SWN which is part of the differential diagnosis for NF2 and SWN.
Show less - Date Issued
- 2019
- Identifier
- CFH2000525, ucf:45627
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFH2000525
- Title
- Signals Delivered By Interleukin-7 Regulate The Activities Of Bim And JunD In T Lymphocytes.
- Creator
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Ruppert, Shannon, Khaled, Annette, Self, William, Zervos, Antonis, Teter, Kenneth, University of Central Florida
- Abstract / Description
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Interleukin-7 (IL-7) is an essential cytokine for lymphocyte growth that has the potential for promoting proliferation and survival. While the survival and proliferative functions of IL-7 are well established, the identities of IL-7 signaling components in pathways other than JAK/STAT, that accomplish these tasks remain poorly defined. To this end, we used IL-7 dependent T-cells to examine those components necessary for cell growth and survival. Our studies revealed two novel signal...
Show moreInterleukin-7 (IL-7) is an essential cytokine for lymphocyte growth that has the potential for promoting proliferation and survival. While the survival and proliferative functions of IL-7 are well established, the identities of IL-7 signaling components in pathways other than JAK/STAT, that accomplish these tasks remain poorly defined. To this end, we used IL-7 dependent T-cells to examine those components necessary for cell growth and survival. Our studies revealed two novel signal transducers of the IL-7 growth signal: BimL and JunD. IL-7 promoted the activity of JNK (Jun N-terminal Kinase), and that JNK, in turn, drove the expression of JunD, a component of the Activating Protein 1 (AP-1) transcription factors. Inhibition of JNK/JunD blocked glucose uptake and HXKII gene expression, indicating that this pathway was responsible for promoting HXKII expression. After a bioinformatics survey to reveal possible JunD-regulated genes activated early in the IL-7 signaling cascade, our search revealed that JunD could control the expression of proteins involved in signal transduction, cell survival and metabolism, including Pim-1. Pim-1, an IL-7 induced protein, was inhibited upon JNK or JunD inhibition. Our hypothesis that JunD positively regulated proliferation was confirmed when the proliferation of primary CD8+ T-cells cultured with IL-7 was impaired upon treatment with JunD siRNA. These results show that the IL-7 signal is more complex than the JAK/STAT pathway, activating JNK and JunD to induce rapid growth through the expression of metabolic factors like HXKII and Pim-1. When metabolic activities are inhibited, cells undergo autophagy, or cell scavenging, to provide essential nutrients. Pro-apoptotic Bim was evaluated for its involvement in autophagy. Bim is a BH3-only member of the Bcl-2 family that contributes to T-cell death. Partial rescue of T-cells occurs when Bim and the interleukin-7 receptor are deleted, implicating Bim in IL-7-deprived T-cell apoptosis. Alternative splicing results in three different isoforms: BimEL, BimL, and BimS. To study the effect of Bim deficiency and define the function of the major isoforms, Bim-containing and Bim-deficient T-cells, dependent on IL-7 for growth, were used. Loss of Bim in IL-7-deprived T-cells delayed apoptosis, but blocked the degradative phase of autophagy. The conversion of LC3-I to LC3-II was observed in Bim-deficient T-cells, but p62, which is degraded in autolysosomes, accumulated. To explain this, BimL, was found to support acidification of lysosomes associated with autophagic vesicles. Key findings showed that inhibition of lysosomal acidification accelerated death upon IL-7 withdrawal only in Bim-containing T-cells, indicating that in these cells autophagy was protective. IL-7 dependent T-cells lacking Bim were insensitive to inhibition of autophagy or lysosomal acidification. BimL co-immunoprecipitated with dynein and Lamp1-containing vesicles, indicating BimL could be an adaptor for dynein to facilitate loading of lysosomes. In Bim deficient T-cells, lysosome-tracking probes revealed vesicles of less acidic pH. Over-expression of BimL restored acidic vesicles in Bim deficient T-cells, while other isoforms, BimEL and BimS, associated with intrinsic cell death. These results reveal a novel role for BimL in lysosomal positioning that may be required for the formation of functional autolysosomes during autophagy.
Show less - Date Issued
- 2012
- Identifier
- CFE0004435, ucf:49331
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0004435
