1.Transcriptome and Proteome Profiling of Primary Human Gastric Interstitial Cells of Cajal Predicts Pacemaker Networks
Daphne FOONG ; Meena MIKHAEL ; Jerry ZHOU ; Ali ZARROUK ; Xiaodong LIU ; Jan SCHRÖDER ; Jose M POLO ; Vincent HO ; Michael D O’CONNOR
Journal of Neurogastroenterology and Motility 2023;29(2):238-249
Background/Aims:
Interstitial cells of Cajal (ICC) are specialized gastrointestinal (GI) pacemaker cells required for normal GI motility. Dysfunctions in ICC have been reported in patients with GI motility disorders, such as gastroparesis, who exhibit debilitating symptoms and greatly reduced quality of life. While the proteins, calcium-activated chloride channel anoctamin-1 (ANO1) and the receptor tyrosine kinase (KIT), are known to be expressed by human ICC, relatively little is known about the broad molecular circuitry underpinning human ICC functions. The present study therefore investigates the transcriptome and proteome of ANO1-expressing, KIT low /CD45- /CD11B- ICC obtained from primary human gastric tissue.
Methods:
Excess human gastric tissue resections were obtained from sleeve gastrectomy patients. ICC were purified using fluorescence-activated cell sorting (FACSorting). Then, ICC were characterized by using immunofluorescence, real-time polymerase chain reaction, RNAsequencing and mass spectrometry.
Results:
Compared to unsorted cells, real-time polymerase chain reaction showed the KIT low /CD45- /CD11B- ICC had: a 9-fold (P < 0.05) increase in ANO1 expression; unchanged KIT expression; and reduced expression for genes associated with hematopoietic cells (CD68, > 10-fold, P < 0.001) and smooth muscle cells (DES, > 4-fold, P < 0.05). RNA-sequencing and gene ontology analyses of the KIT low / CD45- /CD11B- cells revealed a transcriptional profile consistent with ICC function. Similarly, mass spectrometry analyses of the KIT low / CD45- /CD11B - cells presented a proteomic profile consistent with ICC activities. STRING-based protein interaction analyses using the RNA-sequencing and proteomic datasets predicted protein networks consistent with ICC-associated pacemaker activity and ion transport.
Conclusion
These new and complementary datasets provide a valuable molecular framework for further understanding how ICC pacemaker activity regulates smooth muscle contraction in both normal GI tissue and GI motility disorders.
2.Effects of Carpobrotus edulis Extract on Oxidative Stress and 158N Oligodendrocyte Death.
Amira ZARROUK ; Mohammed Ali SMACH ; Jawhar HAFSA ; Randa SGHAIER ; Hatem MAJDOUB ; Mohammed HAMMAMI ; Bassem CHARFEDDINE
Biomedical and Environmental Sciences 2019;32(4):291-299
OBJECTIVE:
Age-related diseases, including neurodegenerative diseases, are associated with oxidative stress and lipid peroxidation, and increase the levels of cholesterol auto-oxidation products such as 7β-hydroxycholesterol (7β-OHC). Thus, it is imperative to identify agents that can prevent 7β-OHC-induced side-effects.
METHODS:
We evaluated the potential protective effects of Carpobrotus edulis ethanol-water extract (EWe) on murine oligodendrocytes (158N) cultured in the absence or presence of 7β-OHC (20 μg/mL, 24 h). The cells were incubated with EWe (20-200 µg/mL) 2 h before 7β-OHC treatment. Mitochondrial activity and cell growth were evaluated with the MTT assay. Photometric methods were used to analyze antioxidant enzyme [catalase (CAT) and glutathione peroxidase (GPx)] activities and the generation of lipid and protein oxidation products [malondialdehyde (MDA), conjugated diene (CD), and carbonylated proteins (CPs)].
RESULTS:
Treatment with 7β-OHC induced cell death and oxidative stress (reflected by alteration in CAT and SOD activities). Overproduction of lipid peroxidation products (MDA and CDs) and CPs was also reported. The cytotoxic effects associated with 7β-OHC were attenuated by 160 μg/mL of EWe of C. edulis. Cell death induced by 7β-OHC treatment was ameliorated, GPx and CAT activities were restored to normal, and MDA, CD, and CP levels were reduced following C. edulis extract treatment.
CONCLUSION
These data demonstrate the protective activities of C. edulis EWe against 7β-OHC-induced disequilibrium in the redox status of 158N cells, indicative of the potential role of this plant extract in the prevention of neurodegenerative diseases.
Aizoaceae
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Animals
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Cell Line
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Drug Evaluation, Preclinical
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Hydroxycholesterols
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Mice
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Neurodegenerative Diseases
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prevention & control
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Neuroprotection
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Oligodendroglia
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drug effects
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metabolism
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Oxidative Stress
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drug effects
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Phytotherapy
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Plant Extracts
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pharmacology
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therapeutic use