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Expression of PC3, carboxypeptidase E and enkephalin in human monocyte-derived macrophages as a tool for genetic
J LaMendola1, S K Martin, D F Steiner
1Howard Hughes Medical Institute, University of Chicago, IL 60637, USA. jlamendo@midway.uchicago.edu
Abstract:
Circulating monocytes in human peripheral blood are readily available, easily obtained, and can be cultured in vitro. Once plated, the monocytes spontaneously differentiate into macrophages. Undifferentiated human monocytes do not express carboxypeptidase E (CPE), prohormone convertase 3 (PC3/PC1) or proenkephalin (ENK), suggesting that gene induction during differentiation results in the expression of these genes. RT-PCR of human monocyte-derived macrophage (HMDM) mRNA showed detectable levels of ENK mRNA at 48 h after plating, followed by PC3 and CPE mRNAs at 72 h. PC3 expression was confirmed by Western blotting in THP-1 cells. Similarities in expression of enzymes involved in the conversion of neuroendocrine precursors, such as proenkephalin, into functionally mature peptides underscores the close association HMDMs may share with the neuroendocrine system. Because of this, HMDMs may prove to be a valuable, non-surgical source of human tissue for clinical genetic diagnosis of some convertase disorders.
Insights
Human monocytes differentiate into macrophages, expressing key neuroendocrine genes like proenkephalin. This finding suggests human monocyte-derived macrophages are a valuable source for diagnosing convertase disorders.
Area of Science:
- Neuroendocrinology
- Cell Biology
Background:
- Circulating monocytes are accessible and differentiate into macrophages in vitro.
- Undifferentiated monocytes lack expression of carboxypeptidase E (CPE), prohormone convertase 3 (PC3), and proenkephalin (ENK).
Purpose of the Study:
- To investigate the gene expression of CPE, PC3, and ENK during monocyte differentiation into macrophages.
- To explore the potential of human monocyte-derived macrophages (HMDMs) as a diagnostic tool for neuroendocrine disorders.
Main Methods:
- Monocytes were isolated from human peripheral blood and cultured in vitro.
- Reverse transcription-polymerase chain reaction (RT-PCR) was used to detect mRNA levels of ENK, PC3, and CPE.
- Western blotting confirmed PC3 expression in THP-1 cells.
Main Results:
- ENK mRNA was detected at 48 hours post-plating.
- PC3 and CPE mRNAs were observed at 72 hours post-plating.
- PC3 expression was validated in THP-1 cells.
Conclusions:
- HMDMs express enzymes crucial for neuroendocrine precursor conversion, indicating a link to the neuroendocrine system.
- HMDMs represent a promising non-surgical source for clinical genetic diagnosis of convertase disorders.