Ignite Creation Date:
2024-05-05 @ 5:07 PM
Last Modification Date:
2024-10-26 @ 9:28 AM
Study NCT ID:
NCT00397280
Status:
RECRUITING
Last Update Posted:
2024-07-15
First Post:
2006-11-08
Brief Title:
Immune Cell Response to Stimuli
Sponsor:
National Institute of Environmental Health Sciences NIEHS
Organization:
National Institutes of Health Clinical Center CC
Study Overview
Official Title:
Innate Immunity Signal Transduction in Human Leukocytes
Status:
RECRUITING
Status Verified Date:
2024-06-13
Last Known Status:
None
Delayed Posting:
No
If Stopped, Why?:
Not Stopped
Has Expanded Access:
False
If Expanded Access, NCT#:
N/A
Has Expanded Access, NCT# Status:
N/A
Acronym:
None
Brief Summary:
This study will investigate the response of immune cells neutrophils monocytes to various signals in the test tube to determine how they sense the signals in the body and what substances they produce in response to them It will determine how the cells may under certain circumstances contribute to inflammation and will measure substances in the blood plasma the liquid non-cellular part of the blood that might stimulate white blood cells in order to understand how the blood responds to possible disease-related conditions
Healthy normal volunteers 18 years of age and older who weigh at least 110 pounds may be eligible for this study Participants give about 320 milliliters mL of blood about 1 13 cups or less at each donation They donate no more than once every 8 weeks and no more than six times a year On some occasions less than 320 mL of blood may be drawn The collected blood is separated into its components and specific cells are exposed to substances to examine their response
Healthy normal volunteers 18 years of age and older who weigh at least 110 pounds may be eligible for this study Participants give about 320 milliliters mL of blood about 1 13 cups or less at each donation They donate no more than once every 8 weeks and no more than six times a year On some occasions less than 320 mL of blood may be drawn The collected blood is separated into its components and specific cells are exposed to substances to examine their response
Detailed Description:
The objective of this study is several-fold We wish to determine the response of white blood cells neutrophils monocytes eosinophils lymphocytes to various signals in the test tube to test how they sense the signals in the body and what substances they produce We wish to determine how these cells may under certain circumstances contribute to inflammation We wish to measure the substances in the blood plasma that might contribute to stimulation of white blood cells to understand the response of the whole blood to potential disease-associated conditions
The objective is to define the signaling pathways activated by lipopolysaccharide LPS and other selected innate immunity stimuli and the downstream inflammatory functional consequences in human leukocytes in vitro Adult 18-65 yrs old nonpregnant healthy volunteers will have 320 ml of whole blood collected by venipuncture in a monitored setting no more frequently than once every 8 weeks No further interventions will be exercised upon the subjects The whole blood will be fractionated into neutrophil red blood cell mononuclear cell and plasma fractions using plasma-Percoll discontinuous centrifugation Leukocytes will be subjected in vitro to inflammatory stimuli eg LPS and selected signaling outcomes eg mitogen-activated protein kinase activation Rho GTPase activation protein-protein interactions and functional measures eg chemotaxis superoxide anion and cytokine production quantified in the absence and presence of relevant chemical inhibitors eg SB203580 a p38 MAPK inhibitor In each such experiment cells from the daily donor will be used as paired controls to the in vitro experimental intervention eg SB203580 inhibitor vs DMSO vehicle Three or more repetitions on different donors of each specific experimental outcome as necessary will be performed to establish statistical significance of findings
A specific focus of the studies planned will be to define the role of lipid raft membrane microdomains in transduction of the LPS signal in human leukocytes Lipid rafts are cholesterol-rich microdomains in the plasma membrane within which the LPS receptor Toll-like Receptor 4 TLR4 has been described to reside LPS signaling has been reported to be sensitive to raft cholesterol content presumably because the specific repertoire of proteins in rafts is sensitive to raft cholesterol content Rafts are thought to act as dynamic signaling platforms for co-segregation of proximal adaptor proteins kinases and other signaling proteins Of interest while LPS has been described to modulate the activity of proteins that determine raft cholesterol content egg Liver X Receptor ABCA1 virtually no work has been done to clarify 1 the mechanisms underlying LPS-induced intracellular cholesterol redistribution and more importantly 2 whether such intracellular redistribution of cholesterol is causal to the signaling events triggered by LPS or 3 whether innate immunity signaling is dependent upon inter-subject variations in raft cholesterol content
Furthermore we will investigate the role of the tumor suppressor gene p53 in the regulation of inflammation It is now widely accepted that inflammation and cancer development are interconnected Dr Menendez is one of the international leaders in the study of the tumor suppressor gene p53 His group has discovered that activation of p53 through exposure to carcinogenic stimuli leads to differential expression of genes that have a direct effect on the inflammatory response such as several toll-like-receptor genes Dr Menendez will use human leukocytes that will be isolated from whole blood He will expose these cells to stimuli that activate p53 such as doxorubicin a chemotherapy agent or radiation and examine the expression of toll-like-receptor genes as well as the response to LPS and other inflammatory agents in vitro
Finally we will investigate the role of zinc-finger proteins in gene expression in inflammatory cells ZFP36 is an RNA binding zinc finger protein that is involved in the turnover of mRNAs encoding several clinical important cytokines including TNFa Several missense promoter and 3-untranslated variants have been identified in humans in some cases these variants and their haplotypes have been associated with human inflammatory disease We will isolate peripheral blood macrophages and stimulate them with LPS and other cytokines then evaluate the behavior of cytokine mRNA and gene expression
The objective is to define the signaling pathways activated by lipopolysaccharide LPS and other selected innate immunity stimuli and the downstream inflammatory functional consequences in human leukocytes in vitro Adult 18-65 yrs old nonpregnant healthy volunteers will have 320 ml of whole blood collected by venipuncture in a monitored setting no more frequently than once every 8 weeks No further interventions will be exercised upon the subjects The whole blood will be fractionated into neutrophil red blood cell mononuclear cell and plasma fractions using plasma-Percoll discontinuous centrifugation Leukocytes will be subjected in vitro to inflammatory stimuli eg LPS and selected signaling outcomes eg mitogen-activated protein kinase activation Rho GTPase activation protein-protein interactions and functional measures eg chemotaxis superoxide anion and cytokine production quantified in the absence and presence of relevant chemical inhibitors eg SB203580 a p38 MAPK inhibitor In each such experiment cells from the daily donor will be used as paired controls to the in vitro experimental intervention eg SB203580 inhibitor vs DMSO vehicle Three or more repetitions on different donors of each specific experimental outcome as necessary will be performed to establish statistical significance of findings
A specific focus of the studies planned will be to define the role of lipid raft membrane microdomains in transduction of the LPS signal in human leukocytes Lipid rafts are cholesterol-rich microdomains in the plasma membrane within which the LPS receptor Toll-like Receptor 4 TLR4 has been described to reside LPS signaling has been reported to be sensitive to raft cholesterol content presumably because the specific repertoire of proteins in rafts is sensitive to raft cholesterol content Rafts are thought to act as dynamic signaling platforms for co-segregation of proximal adaptor proteins kinases and other signaling proteins Of interest while LPS has been described to modulate the activity of proteins that determine raft cholesterol content egg Liver X Receptor ABCA1 virtually no work has been done to clarify 1 the mechanisms underlying LPS-induced intracellular cholesterol redistribution and more importantly 2 whether such intracellular redistribution of cholesterol is causal to the signaling events triggered by LPS or 3 whether innate immunity signaling is dependent upon inter-subject variations in raft cholesterol content
Furthermore we will investigate the role of the tumor suppressor gene p53 in the regulation of inflammation It is now widely accepted that inflammation and cancer development are interconnected Dr Menendez is one of the international leaders in the study of the tumor suppressor gene p53 His group has discovered that activation of p53 through exposure to carcinogenic stimuli leads to differential expression of genes that have a direct effect on the inflammatory response such as several toll-like-receptor genes Dr Menendez will use human leukocytes that will be isolated from whole blood He will expose these cells to stimuli that activate p53 such as doxorubicin a chemotherapy agent or radiation and examine the expression of toll-like-receptor genes as well as the response to LPS and other inflammatory agents in vitro
Finally we will investigate the role of zinc-finger proteins in gene expression in inflammatory cells ZFP36 is an RNA binding zinc finger protein that is involved in the turnover of mRNAs encoding several clinical important cytokines including TNFa Several missense promoter and 3-untranslated variants have been identified in humans in some cases these variants and their haplotypes have been associated with human inflammatory disease We will isolate peripheral blood macrophages and stimulate them with LPS and other cytokines then evaluate the behavior of cytokine mRNA and gene expression
Study Oversight
Has Oversight DMC:
None
Is a FDA Regulated Drug?:
False
Is a FDA Regulated Device?:
False
Is an Unapproved Device?:
None
Is a PPSD?:
None
Is a US Export?:
None
Is an FDA AA801 Violation?:
None
Secondary IDs
| Secondary ID | Type | Domain | Link |
|---|---|---|---|
| 07-E-0023 | None | None | None |