In order to investigate the relationship between the expression of heme oxygenase-2 (HO-2) mRNA and the pathogenesis of Hirschsprung's disease (HD), total ribonucleic acid (RNA) was extracted in the aganglionic and ganglionic segments of colon respectively from 15 cases of HD. The single-stranded cDNA of HO-2 was synthesized and further amplified by reverse transcription-polymerase chain reaction (RT-PCR). The expression of HO-2 mRNA was normal in ganglionic segments, but absent in aganglionic segments. It is concluded that the absence of HO-2 mRNA expression may be an important mechanism responsible for HD.
Colon/enzymology ; Heme Oxygenase (Decyclizing)/biosynthesis ; Heme Oxygenase (Decyclizing)/*genetics ; Hirschsprung Disease/*enzymology ; Hirschsprung Disease/etiology ; Hirschsprung Disease/genetics ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics

To investigate the expression of the HO-1 gene in PC12 cells in hypoxic environment and gain further insight to the role of HO-1 in cerebral ischemia, PC12 cells were exposed to hypoxia environment (95% N2, 5% CO2) for 0.5 h, 1 h, 4 h, 8 h, 12 h, 24 h respectively. The level of HO-1 mRNA was examined by reverse transcriptase polymerase chain reaction (RT-PCR); the volume of COHb in the media were measured spectrophotometrically and the cGMP concentration of PC12 cell extracts was determined by radioimmunoassay. We found that after exposure to hypoxia for 1 h, 4 h, 8 h, 12 h, 24 h, HO-1 mRNA increased by 3%, 4%, 17%, 31% 36% as compared with that in control group respectively (P < 0.01 or P < 0.05); the COHb increased by 12%, 29%, 59%, 88%, 94% as compared with that in control group respectively (P < 0.01 or P < 0.05), and the cGMP concentration were 2.2, 3.4, 5.2, 8.1, 10.9-fold as that of the control group (P < 0.01). We are led to conclude that hypoxia induced the expression of HO-1 gene, the production of endogenous CO, and the concentration of cGMP was elevated as well.
Carbon Monoxide/metabolism ; Cell Hypoxia ; Cyclic GMP/metabolism ; Heme Oxygenase (Decyclizing)/biosynthesis ; Heme Oxygenase (Decyclizing)/*genetics ; Heme Oxygenase-1 ; PC12 Cells ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics ; Up-Regulation

To investigate the expression of the HO-1 gene in PC12 cells in hypoxic environment and gain further insight to the role of HO-1 in cerebral ischemia, PC12 cells were exposed to hypoxia environment (95% N2, 5% CO2) for 0.5 h, 1 h, 4 h, 8 h, 12 h, 24 h respectively. The level of HO-1 mRNA was examined by reverse transcriptase polymerase chain reaction (RT-PCR); the volume of COHb in the media were measured spectrophotometrically and the cGMP concentration of PC12 cell extracts was determined by radioimmunoassay. We found that after exposure to hypoxia for 1 h, 4 h, 8 h, 12 h, 24 h, HO-1 mRNA increased by 3%, 4%, 17%, 31% 36% as compared with that in control group respectively (P < 0.01 or P < 0.05); the COHb increased by 12%, 29%, 59%, 88%, 94% as compared with that in control group respectively (P < 0.01 or P < 0.05), and the cGMP concentration were 2.2, 3.4, 5.2, 8.1, 10.9-fold as that of the control group (P < 0.01). We are led to conclude that hypoxia induced the expression of HO-1 gene, the production of endogenous CO, and the concentration of cGMP was elevated as well.
Animals ; Carbon Monoxide ; metabolism ; Cell Hypoxia ; Cyclic GMP ; metabolism ; Heme Oxygenase (Decyclizing) ; biosynthesis ; genetics ; Heme Oxygenase-1 ; PC12 Cells ; RNA, Messenger ; biosynthesis ; genetics ; Rats ; Up-Regulation

OBJECTIVESTo observe the function of heme oxygenase (HO) in the lung damage in hepatic cirrhosis rats.

METHODSLiver cirrhosis model rats were made by CCl4. Lung samples taken from normal and cirrhotic rats were examined for HO-1 and HO-2 protein and expression distribution with immunohistochemical staining and western blot.

RESULTSLiver cirrhosis model rats were successfully constructed. There was a notable increase of HO-1 staining (0.062+/-0.021 vs 0.185+/-0.044, t=11.24, P<0.01) and protein expression (0 vs 5294.92+/-46.02, t=11.45, P<0.01) in both vascular and bronchial smooth muscle cells and endothelium in cirrhotic rats, however, no statistical difference of HO-2 between cirrhotic and normal rats was observed.

CONCLUSIONThe HO-CO pathway is probably involved in the pathogenesis of lung damage in hepatic cirrhosis rats.

Animals ; Carbon Tetrachloride Poisoning ; Heme Oxygenase (Decyclizing) ; analysis ; biosynthesis ; Heme Oxygenase-1 ; Liver Cirrhosis, Experimental ; enzymology ; Lung ; enzymology ; Male ; Nitric Oxide ; metabolism ; Rats ; Rats, Sprague-Dawley

To confirm the existence of heme oxygenase (HO)- carbon monoxide (CO)- cyclic guanosine monophosphate (cGMP) pathway in the cultured human trabecular meshwork cells (HT-MCs) in vitro, and to evaluate the inductive role of hemin on this pathway, HTMCs of the third to fourth generation were cultured in vitro. Reverse transcripase-polymerase chain reaction (RT-PCR) was employed for detection of HO-1 and HO-2 mRNA. Immunohistochemical staining was used to detect HO-1 and HO-2 proteins. Hemin was added into the culture solution. The HO-1 mRNA levels were quantified by RT-PCR. The relative amount of carbon monoxide released into the media was measured with the quantifying carbon monoxide hemoglobin (HbCO) by spectrophotometry. Radioimmunoassay was used to determine changes of cGMP in HTMCs. The results showed that cultured cells had the specific characteristics of HTMCs. Both HO-1 and HO-2 genes were expressed in HTMCs, as well as HO-1 and HO-2 proteins in HTMCs. Hemin induced HO-1 mRNA, HbCO and cGMP in a dose-dependent manner. In conclusion, HO-CO-cGMP pathway exists in the cultured HTMCs and can be induced by hemin. Pharmacological stimulation of HO-CO-cGMP pathway may constitute a novel therapeutic approach to rescuing glaucoma.
Carbon Monoxide/*metabolism ; Cells, Cultured ; Cyclic GMP/*biosynthesis ; Cyclic GMP/genetics ; Heme Oxygenase (Decyclizing)/*biosynthesis ; Heme Oxygenase (Decyclizing)/genetics ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics ; Signal Transduction ; Trabecular Meshwork/cytology ; Trabecular Meshwork/*metabolism

To confirm the existence of heme oxygenase (HO)- carbon monoxide (CO)- cyclic guanosine monophosphate (cGMP) pathway in the cultured human trabecular meshwork cells (HT-MCs) in vitro, and to evaluate the inductive role of hemin on this pathway, HTMCs of the third to fourth generation were cultured in vitro. Reverse transcripase-polymerase chain reaction (RT-PCR) was employed for detection of HO-1 and HO-2 mRNA. Immunohistochemical staining was used to detect HO-1 and HO-2 proteins. Hemin was added into the culture solution. The HO-1 mRNA levels were quantified by RT-PCR. The relative amount of carbon monoxide released into the media was measured with the quantifying carbon monoxide hemoglobin (HbCO) by spectrophotometry. Radioimmunoassay was used to determine changes of cGMP in HTMCs. The results showed that cultured cells had the specific characteristics of HTMCs. Both HO-1 and HO-2 genes were expressed in HTMCs, as well as HO-1 and HO-2 proteins in HTMCs. Hemin induced HO-1 mRNA, HbCO and cGMP in a dose-dependent manner. In conclusion, HO-CO-cGMP pathway exists in the cultured HTMCs and can be induced by hemin. Pharmacological stimulation of HO-CO-cGMP pathway may constitute a novel therapeutic approach to rescuing glaucoma.
Carbon Monoxide ; metabolism ; Cells, Cultured ; Cyclic GMP ; biosynthesis ; genetics ; Heme Oxygenase (Decyclizing) ; biosynthesis ; genetics ; Humans ; RNA, Messenger ; biosynthesis ; genetics ; Signal Transduction ; Trabecular Meshwork ; cytology ; metabolism

In order to investigate the role of heme oxygenase-1 (HO-1) in the molecular mechanism of experimental allergic encephalomyelitis (EAE), which was induced by guinea pig spinal cord homogenate + complete freund adjuvant on Wistar rats, we observed the gene of HO-1 and its protein expression with reverse transcriptase polymerase chain reaction(RT-PCR) and immunohistochemistry 1, 7, 14, and 21 d after EAE induction in rats. The relationship between HO-1 and the symptoms of EAE was also observed. The results showed that the levels of HO-1 mRNA and its protein expression were very low in the brains of the control group, whereas they were enhanced gradually with pathological course in the brain and onsets of symptoms, signs of EAE. On day 7, the level of HO-1 mRNA reached the peak, but the expression level of HO-1 protein in the brains reached the peak on day 14. The immunoreactive cells of HO-1 were mainly located at the choroid plexuses and subfornical organ (SFO), as well as in regions around the "sleeve-like" lesion foci, all of which were coincident with the locations of lesions of EAE. The levels of HO-1 mRNA and its protein expression were lowered gradually on day 21, which were in parallel with the severities of symptoms and signs of EAE. After a specific inhibitor of HO-1, Snpp-9, was applied, both of the symptoms and pathological lesions of EAE in the rat brains were mitigated markedly. Therefore, these results may suggest that the dynamic changes of HO-1 mRNA and its protein expression are in parallel with the changes of symptoms and pathological lesions of EAE in the brain. In conclusion, the levels of HO-1 mRNA and its protein expression in brains may play an important role in the pathogenesis of EAE, and application of inhibitors of HO-1 may be one of the potential therapeutic ways for the prevention and treatment of EAE.
Animals ; Brain ; enzymology ; metabolism ; Encephalomyelitis, Autoimmune, Experimental ; enzymology ; genetics ; physiopathology ; Female ; Heme Oxygenase (Decyclizing) ; biosynthesis ; genetics ; Heme Oxygenase-1 ; RNA, Messenger ; biosynthesis ; genetics ; Rats ; Rats, Wistar ; Subfornical Organ ; metabolism ; pathology

We investigated the expression of heme oxygenase-1 (HO-1) gene and production of endogenous carbon monoxide (CO) in the rat lung tissue at different time points of chronic hypoxic pulmonary hypertension and the effect of hemin on the expression of HO-1 gene and pulmonary hypertension. A rat model of hypoxic pulmonary hypertension was recreated by exposure to intermittent normobaric hypoxic environment (10% O2). Reverse transcriptase polymerase chain reaction (RT-PCR) was performed to determine the level of HO-1 mRNA in the rat lung tissue and double wave length spectrophotometry was used to evaluate the quantity of COHb in arterial blood. Cardiac catheterization was employed to measure the right ventricular systolic pressure (RVSP) and HE staining was performed in dissected lung tissue to observe the pathological changes of the intra-acinar pulmonary arteries (IAPA). It was found that (1) There was a low level of HO-1 mRNA in normal rat lung tissue, but the level of HO-1 mRNA increased by 2-4 times in the lung tissue of hypoxic rats (P<0.01). The quantity of COHb was 2-3 times those of control group (P<0.01 or P<0.05). These were accompanied by the increased of RVSP and the thickened IAPA; (2) Hemin could keep the HO-1 mRNA and COHb in the hypoxic rat lung tissue at a high level, and partially suppressed the increase of rat RVSP, thereby ameliorating the pathological changes of IAPA. In conclusion, the upregulation of the expression of HO-1 gene and production of CO in the rat lung of hypoxic pulmonary hypertension plays a role of inhibition in the development of hypoxic pulmonary hypertension. Hemin has a therapeutic effect on hypoxic pulmonary hypertension.
Animals ; Carbon Monoxide ; metabolism ; physiology ; Heme Oxygenase (Decyclizing) ; biosynthesis ; genetics ; Heme Oxygenase-1 ; Hypertension, Pulmonary ; etiology ; metabolism ; Hypoxia ; complications ; Lung ; metabolism ; Myocytes, Smooth Muscle ; drug effects ; pathology ; Pulmonary Artery ; metabolism ; pathology ; RNA, Messenger ; biosynthesis ; genetics ; Rats

Heme oxygenase (HO) is a rate-limiting enzyme for endogenous carbon monoxide (CO) production. Recently it has been suggested that endogenous CO plays an important role in regulating smooth muscle tone. The development of bladder outlet obstruction in men with benign prostates hyperplasia was shown to be related to the prostates smooth muscle tone, but it was not clear whether endogenous HO/CO system mediates prostates smooth muscle activity. To investigate the influence of sex hormone on the expression of heme oxygenase (HO) gene in rat ventral prostate, we created the model of castrated male rats to test the mRNA levels of HO-1 and HO-2 by RT-PCR, and used immunohistochemical staining procedures with image analytic technical system to confirm the effects of exogenous androgen and estrogen on the expression of HO-1 and HO-2 protein in rat ventral prostate. The results showed that two isoforms of HO were present in rat ventral prostate. The epithelial cells of acini and fibromuscular stroma of the rat prostate displayed HO-1 immunoreactivity, whereas HO-2 immunostaining was only examined to be in the acinar cells. Both at protein and transcript levels, HO-1 in castrated group was markedly decreased compared with the normal control group (p<0.01). In groups of exogenous administration of androgen and estrogen HO-1 was much higher than that in the control groups (p<0.01). However, estrogen increased HO-1 protein level in prostate stroma, while the levels of HO-2 did not give any evidence of change among all groups (p>0.05). These findings suggest that expression of HO-1 gene is induced by sex hormones, in contrast, there is no change in HO-2 expression. We speculate that CO-HO system is possibly involved in the pathologic processes of prostates abnormal proliferation induced by sex hormones and that CO derived from HO-1 may play an important role in the regulation of smooth muscle activity in rat prostate.
Animals ; Carbon Monoxide ; metabolism ; Heme Oxygenase (Decyclizing) ; biosynthesis ; genetics ; Heme Oxygenase-1 ; Male ; Orchiectomy ; Prostate ; enzymology ; metabolism ; RNA, Messenger ; biosynthesis ; genetics ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Testosterone Propionate ; pharmacology

We investigated the expression of heme oxygenase-1 (HO-1) gene and production of endogenous carbon monoxide (CO) in the rat lung tissue at different time points of chronic hypoxic pulmonary hypertension and the effect of hemin on the expression of HO-1 gene and pulmonary hypertension. A rat model of hypoxic pulmonary hypertension was recreated by exposure to intermittent normobaric hypoxic environment (10% O2). Reverse transcriptase polymerase chain reaction (RT-PCR) was performed to determine the level of HO-1 mRNA in the rat lung tissue and double wave length spectrophotometry was used to evaluate the quantity of COHb in arterial blood. Cardiac catheterization was employed to measure the right ventricular systolic pressure (RVSP) and HE staining was performed in dissected lung tissue to observe the pathological changes of the intra-acinar pulmonary arteries (IAPA). It was found that (1) There was a low level of HO-1 mRNA in normal rat lung tissue, but the level of HO-1 mRNA increased by 2-4 times in the lung tissue of hypoxic rats (P<0.01). The quantity of COHb was 2-3 times those of control group (P<0.01 or P<0.05). These were accompanied by the increased of RVSP and the thickened IAPA; (2) Hemin could keep the HO-1 mRNA and COHb in the hypoxic rat lung tissue at a high level, and partially suppressed the increase of rat RVSP, thereby ameliorating the pathological changes of IAPA. In conclusion, the upregulation of the expression of HO-1 gene and production of CO in the rat lung of hypoxic pulmonary hypertension plays a role of inhibition in the development of hypoxic pulmonary hypertension. Hemin has a therapeutic effect on hypoxic pulmonary hypertension.
Anoxia/complications ; Carbon Monoxide/*metabolism ; Carbon Monoxide/physiology ; Heme Oxygenase (Decyclizing)/*biosynthesis ; Heme Oxygenase (Decyclizing)/genetics ; Heme Oxygenase-1 ; Hypertension, Pulmonary/etiology ; Hypertension, Pulmonary/*metabolism ; Lung/metabolism ; Myocytes, Smooth Muscle/drug effects ; Myocytes, Smooth Muscle/pathology ; Pulmonary Artery/metabolism ; Pulmonary Artery/*pathology ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics