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Hemoglobin induced changes in the transcriptome of normal subjects

ABSTRACT: Hemoglobin induced changes in the transcriptome of normal subjects

PROVIDER: PRJNA637391 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR11928393_1.fastq.gz	Fastqsanger.gz
	SRR11928393_2.fastq.gz	Fastqsanger.gz
	SRR11928394_1.fastq.gz	Fastqsanger.gz
	SRR11928394_2.fastq.gz	Fastqsanger.gz
	SRR11928395_1.fastq.gz	Fastqsanger.gz

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Project description:BACKGROUND & AIMS: Recent studies revealed that hemoglobin is expressed in some non-erythrocytes and it suppresses oxidative stress when overexpressed. Oxidative stress plays a critical role in the pathogenesis of non-alcoholic steatohepatitis (NASH). This study was to investigate whether hemoglobin is expressed in hepatocytes and how it is related to oxidative stress in NASH patients. METHODS: Microarray was performed to identify differentially expressed genes in NASH. Quantitative real time PCR (qRT-PCR) was used to examine gene expression levels. Western blotting and immunofluorescence staining were employed to examine hemoglobin proteins. Flow cytometry was used to measure intracellular oxidative stress. RESULTS: Analysis of microarray gene expression data has revealed a significant increase in the expression of hemoglobin alpha (HBA1) and beta (HBB) in liver biopspies from NASH patients. Increased hemoglobin expression in NASH was validated by qRT-PCR. However, the expression of erythrocyte specific marker genes such as SPTA, SPTB, GYPA, GATA1, and ALAS2 did not change, indicating that increased hemoglobin expression in NASH was not from erythropoiesis, but could result from increased expression in hepatocytes. Immunofluorescence staining demonstrated positive HBA1 and HBB expression in the hepatocytes of NASH livers. Hemoglobin expression was also observed in human hepatocellular carcinoma HepG2 cell line. Furthermore, treatment with hydrogen peroxide, a known oxidative stress inducer, induced a dose dependent increase in HBA1 expression in HepG2 cells. Intriguingly, forced hemoglobin expression suppressed oxidative stress. CONCLUSIONS: Oxidative stress upregulates hemoglobin expression in hepatocytes. Suppression of oxidative stress by hemoglobin could be a mechanism to protect hepatocytes from oxidative damage. These findings suggest that hemoglobin is an inducible antioxidant in hepatocytes in response to increased oxidative stress as found in NASH livers. Twelve biopsy diagnosed NASH patients were included in this study. For control groups, total RNA from 5 different subjects were purchased from ADMET. These subjects are free from liver disease.

Project description:Hypoxia, a naturally occurring phenomenon, results in tremendous losses in the growth of plants and in their productivity. On the physiological level oxygen deprivation results in distinct metabolic rearrangements, is accompanied by dehydration, and oxidative and nitrosative stresses. Six to seven core hypoxia-responsive genes, which are always induced under oxygen deprivation, have been shown to include hemoglobin 1 (Hb1) across a wide range of plant species. However, the function of this induced Hb1 is not completely understood .The importance of elevated Hb expression has been attributed mainly to its nitric oxide (NO) scavenging properties, and hence, to the elimination of this free-radical and signaling species. NO is known to accumulate under hypoxia and its level is in reciprocal relationship with the Hb expression levels. However, evidence has accumulated on Hbs as essential for normal plant development and they take part in the regulation of e.g. flowering, germination and lateral and adventitious root emergence. The physiological role of induced hemoglobin (Hb) expression and its role in NO scavenging were assessed by introducing bacterial (Vitreoscilla stercoraria) hemoglobin (VHb) to the cytoplasm and mitochondria of Arabidopsis thaliana. Heterologous expression was chosen to avoid interference with developmental functions of endogenous Hbs. Sixteen day old transformants were subjected to hypoxia or to NO donor treatment (DETA, 500 µM) for 2h and 24h and global changes in gene expression were assessed using a microarray approach. Both hypoxic and NO-treatment experiments were aimed also to reveal novel functions for Hb under abiotic stress conditions, exemplified by oxygen deprivation and NO/oxidative stresses and during normal development of Arabidopsis transformants on the background of normally expressed endogenous Hbs Hypoxia, a naturally occurring phenomenon, results in tremendous losses in the growth of plants and in their productivity. On the physiological level oxygen deprivation results in distinct metabolic rearrangements, is accompanied by dehydration, and oxidative and nitrosative stresses. Six to seven core hypoxia-responsive genes, which are always induced under oxygen deprivation, have been shown to include hemoglobin 1 (Hb1) across a wide range of plant species. However, the function of this induced Hb1 is not completely understood .The importance of elevated Hb expression has been attributed mainly to its nitric oxide (NO) scavenging properties, and hence, to the elimination of this free-radical and signaling species. NO is known to accumulate under hypoxia and its level is in reciprocal relationship with the Hb expression levels. However, evidence has accumulated on Hbs as essential for normal plant development and they take part in the regulation of e.g. flowering, germination and lateral and adventitious root emergence. The physiological role of induced hemoglobin (Hb) expression and its role in NO scavenging were assessed by introducing bacterial (Vitreoscilla stercoraria) hemoglobin (VHb) to the cytoplasm and mitochondria of Arabidopsis thaliana. Heterologous expression was chosen to avoid interference with developmental functions of endogenous Hbs. Sixteen day old transformants were subjected to hypoxia or to NO donor treatment (DETA, 500 µM) for 2h and 24h and global changes in gene expression were assessed using a microarray approach. Both hypoxic and NO-treatment experiments were aimed also to reveal novel functions for Hb under abiotic stress conditions, exemplified by oxygen deprivation and NO/oxidative stresses and during normal development of Arabidopsis transformants on the background of normally expressed endogenous Hbs

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Hemoglobin induced changes in the transcriptome of normal subjects

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