Zn Protoporphyrin Research Articles

Quantitative determination of Zn protoporphyrin IX, heme and protoporphyrin IX in Parma ham by HPLC

We measured the contents of Zn protoporphyrin IX (ZPP), heme and protoporphyrin IX (PPIX) in Parma ham by simultaneous analysis using high-performance liquid chromatography (HPLC). Extraction with ethyl acetate–acetic acid (4:1) was suitable for the quantitative analysis of ZPP. The contents of heme, ZPP and PPIX in Parma ham were 15.0–29.9, 27.7–47.0 and 0.4–1.1 μg/g, respectively, and total content of porphyrin was 43.7–76.6 μg/g. The amount of ZPP in Parma ham was larger than that of heme, and ZPP accounted for 60–70% of all porphyrins.

Evaluation of serum transferrin receptor for iron deficiency in women of child-bearing age

The objective was to study the evaluation of serum transferrin receptor (sTfR) for Fe deficiency in women of child-bearing age. Primary screening was performed in 942 women ranging in child-bearing age. Serum ferritin (SF), Zn protoporphyrin (ZPP) and Hb were determined. Then the subjects were divided into four groups: normal, Fe store depletion (IDs), Fe-deficiency erythropoiesis and Fe-deficiency anaemia. sTfR was determined and sTfR/SF (sTfR/logSF and log(sTfR/SF)) was calculated. Changes of sTfR in women of different Fe status were observed. A receiver-operating characteristic (ROC) curve was used to evaluate whether sTfR had proper diagnostic efficacy for functional Fe deficiency. The levels of sTfR increased significantly along with the aggravation of Fe deficiency. Increase of STfR/SF along with the aggravation of Fe deficiency was more significant than that of sTfR. STfR had a significant negative correlation with SF and Hb, while it had a significant positive correlation with ZPP. The ROC curve showed that the diagnostic effective rate of sTfR for Fe deficiency could reach 83 %. At this point, the sensitivity was 79 % and the specificity was 63 %. Log(sTfR/SF) could be considered to have the highest effective ratio in detecting IDs, since it reached 99 %. STfR and sTfR/SF could both reflect body Fe-deficiency status specifically. They could be used as reliable indicators for evaluating Fe status and diagnosing Fe deficiency in women of child-bearing age.

Heme oxygenase-1 mediates the inhibitory actions of brazilin in RAW264.7 macrophages stimulated with lipopolysaccharide

Brazilin, the main constituent of Caesalpinia sappan L., is a natural red pigment that has been reported to possess anti-inflammatory properties. This study aimed to identify a novel anti-inflammatory mechanism of brazilin. We found that brazilin did not cause cytotoxicity below 300 μM, and activated heme oxygenase-1 (HO-1) protein synthesis in a concentration-dependent manner at 10–300 μM in RAW264.7 macrophages without affecting mRNA transcription of HO-1. Additionally, brazilin increased bilirubin production and HO-1 activity in RAW264.7 macrophages. In lipopolysaccharide (LPS)-stimulated macrophages, brazilin suppressed the release of nitric oxide (NO), prostaglandin E 2 (PGE 2), interleukin (IL)-1β and tumor necrosis factor-α (TNF-α), and reduced the expression of inducible nitric oxide synthase (iNOS). A specific inhibitor of HO-1, Zn(II) protoporphyrin IX, blocked the suppression of NO production, cytokines release and iNOS expression by brazilin. These results suggest that brazilin possesses anti-inflammatory actions in macrophages and works through a novel mechanism involving the action of HO-1.

Assessment and interpretation of micronutrient status during pregnancy

Accurate assessment of maternal micronutrient status is critical to the prevention of suboptimal micronutrient status and anaemia during pregnancy. Measurement of Fe, folate and vitamin B12 status is complicated by adaptive changes to maternal and placental physiology that markedly affect concentrations of circulating micronutrients and their functional biomarkers. Validation of new assessment methods by comparison with gold standards is often prevented by ethical considerations. Antenatal screening in the UK is predominantly concerned with the detection of anaemia, although estimation of maternal Fe stores by serum ferritin at the start of antenatal care may be a more effective preventive strategy. Functional assessment of maternal anaemia is highly problematic, so instead reference data are used for its definition. The effect of mild-to-moderate anaemia on pregnancy outcome is unclear because of the crude nature of its assessment and the influence of confounding factors. Fe-deficient erythropoiesis may be detected by assessment of erythrocyte Zn protoporphyrin and reticulocyte Hb, although such measures may be unavailable in many clinical laboratories. Serum soluble transferrin receptor is highly responsive to tissue Fe deficiency and is less affected by inflammation than most other indicators. Direct inter-assay comparison of serum and erythrocyte folate values is inadvisable since recovery rates differ greatly between methods. Serum total homocysteine is a useful functional biomarker of both folate and vitamin B12 status but during pregnancy is influenced by other factors that reduce its sensitivity. Isotope-dilution liquid chromatography-tandem MS and serum holo-transcobalamin provide new opportunities to gain detailed data of folate species and vitamin B12 fractions in large samples.

The Role of PDE5 Inhibitors in Heme Oxygenase–cGMP Relationship in Rat Cavernous Tissues

Heme oxygenase (HO) enzyme catalyzes oxidative degradation of heme to biliverdin and carbon monoxide (CO). CO shares many properties with nitric oxide (NO) including the activation of soluble guanyl cyclase. To assess cavernous tissue HO activity and cyclic guanosine monophosphate (cGMP) levels in response to oral phosphodiesterase type 5 (PDE5) inhibitors. Seven hundred twenty male Sprague-Dawley rats, divided into six groups, were investigated. Group 1, controls; group 2 received sildenafil citrate orally; group 3 received vardenafil hydrochloride; and group 4 received tadalafil. Group 5 was subdivided into three equal subgroups, received the same dose of each drug added to the HO inhibitor, Zn protoporphyrin. Group 6 was subdivided into three equal subgroups, received the same dose of each drug added to the NO inhibitor, L-nitroarginine methylester. Eight rats from each group/subgroup were sacrificed at 0.5, 1, 2, 3, 4, 6, 18, 24, and 36 hours, respectively. HO enzyme activity assay and cGMP tissue levels in dissected rat cavernous tissues. Both cavernous tissue HO enzyme activity and cGMP levels were increased significantly in sildenafil-, vardenafil-, and tadalafil-treated rats compared with the controls, with significant decreases after either HO or NO inhibition. Cavernous tissue HO enzyme activity and cGMP showed a positive significant correlation (r = 0.854, P < 0.001). The effects of PDE5 inhibitors in cavernous tissue are partly mediated through HO enzyme activity.

The responses of serum transferrin receptors to iron supplements in subjects with iron-deficiency erythropoiesis and iron-deficiency anaemia

We aimed to study the response of serum transferrin receptors (sTfR) to Fe supplementation in women of childbearing age with Fe-deficiency erythropoiesis (IDE) and Fe-deficiency anaemia (IDA). Primary screening was performed in 942 women ranging in age from 18 to 45 years. After Fe-related biochemical indices such as serum ferritin, Zn protoporphyrin and Hb were determined, the subjects were divided into four groups: normal, Fe store depletion, IDE and IDA. A total of 131 women were randomly selected from the normal, IDE and IDA groups. Subsequently, seventy-six women with IDE and IDA were given various doses of Fe (14 mg/d for IDE; 28 mg/d for IDA) with ferrous l-threonate capsules for twelve consecutive weeks. After receiving Fe supplements, the levels of Fe and sTfR were determined at weeks 0, 3, 6, 9 and 12. The levels of sTfR in women of childbearing age with IDE and IDA were significantly higher than those in the normal group. After receiving Fe supplements, the levels of sTfR were significantly decreased in women of childbearing age with IDE and IDA, while the levels of serum ferritin were significantly increased. In conclusion, STfR can be used as a reliable indicator for assessing the efficacy of Fe supplements.

Heme oxygenase-1 mediates the anti-inflammatory effect of propyl gallate in LPS-stimulated macrophages

This work aimed to elucidate the anti-inflammatory mechanism of the phenolic antioxidant propyl gallate (PG). PG-induced heme oxygenase-1 (HO-1) mRNA and protein of the RAW264.7 macrophages in time- and concentration-dependent manner. It suppressed induction of inducible nitric oxide synthase (iNOS) and subsequent production of nitric oxide (NO) through the downregulation of iNOS promoter activity in lipopolysaccharide (LPS)-activated macrophages. Western-blot data showed that induction of HO-1 expression was coincident with the anti-inflammatory action of PG. Inhibition of HO-1 activity by Zn(II) protoporphyrin IX (ZnPP), a specific inhibitor of HO-1, or knockdown of the HO-1 by small interfering RNA (siRNA) oligonucleotides significantly blocked suppression by PL on iNOS expression, NO production and NF-κB activation which were elevated in LPS-stimulated macrophages. Collectively, PG shows its anti-inflammatory activity via mediation of HO-1 in an in vitro inflammation model.

Zn protoporphyrin IX is formed not from heme but from protoporphyrin IX

We examined the effects of exogenous myoglobin, a bivalent chelator, and nitrite on Zn protoporphyrin IX (ZPP) formation by using model systems. ZPP was formed in a model solution without addition of exogenous myoglobin. After incubation, the amount of ZPP in a model solution was increased but that of heme was not decreased compared with the amounts before incubation. Protoporphyrin IX (PPIX) instead of ZPP also accumulated in a model solution with addition of EDTA, but the amount of heme was not reduced. These results suggested that ZPP was not formed by the Fe–Zn substitution in heme but was formed by the insertion of Zn into PPIX, which was formed independently. The fact that the effects of various factors in model systems with/without addition of a bivalent chelator were similar suggested that ZPP formation was strongly affected by PPIX formation. Inhibition of PPIX formation by nitrite might be the reason for the low levels of ZPP in cured meats.

Oral phosphodiesterase-5 inhibitors: effect of heme oxygenase inhibition on cGMP signalling in rat cavernous tissue

This work postulated that heme oxygenase (HO) is partly responsible for controlling phosphodiesterase-5 inhibitor actions by modulating cyclic guanosine monophosphate (cGMP) cavernous tissue levels. Five hundred and four male Sprague-Dawley rats, divided into five groups, were investigated. Group 1 (n=72) included controls, group 2 (n=72) received sildenafil citrate (Viagra) orally, group 3 (n=72) received vardenafil hydrochloride (Levitra), group 4 (n=72) received tadalafil (Cialis). Group 5 (n=216), subdivided into three subgroups (A, B and C, 72 each), received the same dose of each drug with the HO inhibitor, Zn protoporphyrin. Eight rats from each group/subgroup were killed at 0.5, 1, 2, 3, 4, 6, 18, 24 and 36 h when cGMP levels in the cavernous tissues were estimated. Cavernous tissue cGMP levels increased significantly in sildenafil, vardenafil and tadalafil-treated rats compared to the controls with significant decreases after HO inhibition. It is concluded that the effects of these PDE-5 inhibitors in rat cavernous tissue are partly mediated through HO activity via the cGMP signalling pathway.

Water-Soluble Polymer Conjugates of ZnPP for Photodynamic Tumor Therapy

Zn-protoporphyrin (ZnPP) is a promising candidate for cancer therapy. It is known to inhibit heme-oxygenase-1 (HO-1), resulting in suppressed biliverdin/bilirubin production accompanying lowered antioxidative capacity. As a consequence, a significant suppression of tumor growth in vivo was reported. Recent findings also showed that ZnPP efficiently generated reactive singlet oxygen under illumination of visible light. In the present report, we describe the photosensitizing capabilities of water-soluble polymer conjugates of ZnPP as novel compounds for photodynamic therapy against solid tumors. The polymer conjugation made ZnPP water-soluble, thus possible for injection for its aqueous solution. The cellular uptake and photobiological activity of ZnPP derivatives have been tested using a human T-cell leukemia cell line in vitro and demonstrated most potent phototoxic effects of SMA-ZnPP followed by PEG-ZnPP under aerobic conditions.

Heme oxygenase-1 inhibits cytokine production by activated mast cells

Heme oxygenase-1 (HO-1) is thought to contribute to host defense reactions against various stresses. In addition, recent reports have suggested that HO-1 modulates immunocyte activation and functions. HO-1 suppresses mast cell degranulation, but whether HO-1 suppresses cytokine synthesis as well is not yet known. We examined whether rat HO-1 cDNA transfected rat basophilic leukemia (RBL)-2H3 cells have altered cytokine production in response to stimulation with anti-ovalbumin (OA) serum/OA compared to Mock transfected RBL-2H3 cells. HO-1 inhibited anti-OA serum/OA-induced IL-3 and TNF-α production. Inhibition of HO-1 activity by Zn (II) protoporphyrin IX, a specific HO-1 inhibitor, prevented the suppression of TNF-α production. The cytokine inhibition by HO-1 was associated with selective suppression of the DNA-binding activity of AP-1 transcription factors. The suppression of mast cell cytokine production by HO-1 may be an important aspect of the processes that lead to resolution of allergic inflammation.

Mass spectrometric evidence for a zinc–porphyrin complex as the red pigment in dry-cured Iberian and Parma ham

Extracts containing red pigment complexes from the two types of dry-cured hams, Italian Parma and Spanish Iberian ham, were obtained using water and acetone as extraction solvents followed by a crude purification with C18 column filtration. The purified extracts were then analyzed spectroscopically by recording absorption and fluorescence spectra ( λ ex = 420 nm), which both indicate the presence of chemically identical red chromophores with properties similar to a complex of transition metals and protoporphyrin IX. Electrospray ionization mass spectrometry (ESI-MS) in the positive ion mode confirms the presence of identical chemical compounds. ESI-MS in the negative ion mode detects a cluster of seven isotopologue ions (that of m/ z 623.2 as the most intense) with a pattern matching that of a Zn protoporphyrin IX complex. Based on mass spectral data it is concluded that a Zn–porphyrin complex constitutes a major chromophore in dry-cured Iberian ham as well as in Parma ham.

Observation of the distribution of Zn protoporphyrin IX (ZPP) in Parma ham by using purple LED and image analysis

We investigated the distribution of Zn protoporphyrin IX (ZPP) in Parma ham by using purple LED light and image analysis in order to elucidate the mechanism of ZPP formation. Autofluorescence spectra of Parma ham revealed that ZPP was present in both lean meat and fat, while red emission other than that of ZPP was hardly detected. Although ZPP was found to be distributed widely in Parma ham, it was more abundant in intermuscular fat and subcutaneous fat than in lean meat. The intensity of red emission was weak in muscles that were exposed during the processing. ZPP in both lean meat and subcutaneous fat tended to be more abundant in the inner region than in the outer region. It was thought that ZPP is transferred from lean meat to fat tissue during the processing, resulting in the small amount of ZPP in the lean meat adjacent to subcutaneous fat. Our results led to a completely new hypothesis that ZPP is formed in lean meat and transferred to fat tissue.

Measurement of lipid oxidation and porphyrins in high oxygen modified atmosphere and vacuum-packed minced turkey and pork meat by fluorescence spectra and images

This paper illustrates that fluorescence spectroscopy and imaging can be used to measure the extent and distribution of lipid oxidation in meat. Minced turkey thighs and pork semimembranosus muscles were stored for 7 and 12 days at 4 °C in high oxygen (O 2) modified atmosphere packages and vacuum. Turkey meat packed in high O 2 atmosphere was oxidised already after 7 days of storage. The sensory rancid odour score was 4.7 (on a scale from 1 to 9) and the TBARS value was 1.86 mg MDA/kg. There was also an increase in fluorescence emission intensity in the 410–550 nm region, which arises from lipid oxidation products. The combination of unsaturated fatty acids and access to O 2 resulted in lipid oxidation gradients in the turkey meat samples, and these gradients were clearly visualised by fluorescence images. In comparison, pork meat was more stable against lipid oxidation, with TBARS values <0.2 mg MDA/kg and no development of fluorescent lipid oxidation products was detected. The fluorescence spectra measured in the present experiment suggest that turkey thighs and pork semimembranosus muscle in addition to protoporphyrin also have a natural content of Zn protoporphyrin. The porphyrin content was higher in pork meat than in turkey meat. It increased during storage time when the meat was packed in vacuum, and it decreased with O 2 availability. The distribution of porphyrins in the meat was visualised by fluorescence imaging.

Heme oxygenase-1 mediates the anti-inflammatory effect of mushroom Phellinus linteus in LPS-stimulated RAW264.7 macrophages

This work aimed to elucidate the anti-inflammatory mechanism of the n-BuOH subfraction (PL) prepared from fruiting bodies of Phellinus linteus. PL induced heme oxygenase-1 (HO-1) of the RAW264.7 macrophages in concentration- and time-dependent manner. It suppressed induction of inducible nitric oxide synthase (iNOS) and subsequent production of nitric oxide (NO) through down-regulation of iNOS promoter activity in lipopolysaccharide (LPS)-stimulated macrophages. Zn(II) protoporphyrin IX (ZnPP), a specific inhibitor of HO-1, partly blocked suppression by PL on iNOS promoter activity and NO production, which were elevated in LPS-stimulated macrophages. LPS was able to enhance NO production via reactive oxygen species (ROS) generation, c-Jun NH 2-terminal kinase (JNK) and c-Jun induction. ZnPP prevented PL from down-regulating ROS generation and JNK activation in LPS-stimulated macrophages. Taken together, PL shows its anti-inflammatory activity via mediation of HO-1 in an in vitro inflammation model.

Erythrocyte CuZn Superoxide Dismutase Activity Is Decreased in Iron-Deficiency Anemia

Iron and copper are essential microminerals that are intimately related. The present study was performed to determine the effect of iron-deficiency anemia (IDA) and treatment with iron on laboratory indicators of copper status. Hemoglobin, mean corpuscular volume erythrocyte Zn protoporphyrin, serum ferritin, serum copper, serum ceruloplasmin, and erythrocyte CuZn-superoxide dismutase (SOD) activity were studied in 12 adult women with IDA before and after iron treatment for 60-90 d (100 mg/d Fe, as ferric polymaltose) and in 27 women with normal iron status. Prior to treatment with iron, serum copper and ceruloplasmin were not different between the groups and treatment with iron did not affect these measures. IDA women, before and after treatment with iron, presented a 2.9- and 2-fold decrease in erythrocyte CuZn-SOD activity compared to women with normal iron status (p < 0.001). Treatment with iron increased erythrocyte CuZn-SOD activity of the IDA group; however, this change was not statistically significant. In conclusion, CuZn-SOD activity is decreased in IDA. Measurement of this enzyme activity is not useful for evaluating copper nutrition in iron-deficient subjects.

Pulmonary Expression of Inducible Heme-Oxygenase after Ischemia/Reperfusion of the Lower Extremities in Rats

Expression of inducible heme-oxygenase (HO-1) has been shown to be increased in various inflammatory disorders, which may confer a protective role. The aim of our study was to assess pulmonary expression of HO-1 after ischemia/reperfusion (I/R) of the lower limbs in rats. We compared three groups of rats (n = 5/group): one Sham group, and two I/R groups (aorta cross-clamped for 2 h followed by 2 h of reperfusion), one of which pre-treated with Zn-protoporphyrin (Zn-PP), a competitive inhibitor of HO (50 micromol/kg, i.p.). At the end of experiment, lungs were harvested for determination of HO activity and HO-1 expression by Western blot and immunohistochemistry. Lung injury was assessed by bronchoalveolar lavage, histological study, and determination of the lung Evans Blue dye content, an index of microvascular permeability. I/R of the lower limbs was responsible for acute lung injury (ALI), characterized by neutrophilic infiltration (87 +/- 20 x 10(3) neutrophils/mm(3), Sham group versus 191 +/- 38 x 10(3) neutrophils/mm(3), I/R group; P < 0.002) and an increase in lung Evans blue dye content: (74 +/- 6 microg/g, Sham group versus 122 +/- 48 microg/g, I/R group; P < 0.05). Pre-treatment with Zn-PP further increases the Evans Blue content (122 +/- 48 microg/g, I/R group versus 179 +/- 23 microg/g Zn-PP group P < 0.05) and the neutrophilic infiltration. Pulmonary heme-oxygenase activity, and HO-1 content were increased after I/R. (10.5 +/- 12 pmol bilirubin/mg protein/h, Sham group versus 101.2 +/- 66 pmol bilirubin/mg protein/h, I/R group; P < 0.02). Immunohistochemistry revealed that the expression of HO-1 was mainly localized to inflammatory cells. ALI following I/R of the lower limbs in rats is associated with an increase of pulmonary expression of HO-1, inhibition of this expression increase the severity of ALI.

Sequential induction of heme oxygenase-1 and manganese superoxide dismutase protects cultured astrocytes against nitric oxide

Nitric oxide (NO) is a widely recognized mediator of physiological and pathophysiological signal transmission. In an attempt to better understand the molecular actions of NO in astrocytes, stress protein expression in response to NO donor sodium nitroprusside was investigated. Heme oxygenase-1 (HO-1) has been identified as an inducer of manganese superoxide dismutase (MnSOD), playing a cytoprotective role under the condition of nitrosative stress. We present evidence that the sequential induction of HO-1 and MnSOD protects astrocytes from NO toxicity: (1) both HO-1 and MnSOD expression were induced by NO; (2) NO-mediated increase in MnSOD activity was partly abolished by HO-1 inhibitor Zn(II) protoporphyrin IX (ZnPP); (3) pretreatment of astrocytes with a nontoxic dose of NO protected the cells against the later treatment with a toxic dose of NO; (4) inhibition of HO-1 by ZnPP sensitized astrocytes to the nontoxic dose of NO resulting in a marked cytotoxicity; and (5) adenovirus-mediated overexpression of MnSOD protected astrocytes from the NO toxicity. The molecular action of NO in astrocytes appears to be dose-dependent. While a high dose of NO exerts cytotoxicity leading to the tissue damage in the central nervous system, a low dose of NO may act as an important signaling molecule in astrocytes with concurrent induction of cytoprotective proteins such as HO-1 and MnSOD.

Involvement of heme oxygenase as antioxidant defense in soybean nodules

Objective: We have previously demonstrated that the inducible form of heme oxygenase plays a critical role in protecting against oxidative stress in mammals. To gain further insight into the functions of this enzyme in plants, we have tested its activity and expression in soybean nodules subjected to cadmium (Cd) stress.Materials and methods: Four-weeks-old soybean nodulated plants were treated with different cadmium chloride concentrations (0, 50 and 200 μM) during 48 h. Oxidative stress parameters such as TBARS content, GSH levels and antioxidant enzyme activities were measured as well as heme oxygenase activity and expression. Besides, the effect of biliverdin and Zn-protophorphyrin IX were analized.Results: Treatment with 200 μM Cd during 48 h caused a 67% increase in TBARS content, whereas GSH decreased 44%, and total superoxide dismutase, gluthatione reductase and guaiacol peroxidase were also inhibited 54, 20 and 60%, respectively. A total of 200 μM Cd produced the overexpression of heme oxygenase-1, as well as a 10-fold enhancement of its activity. Co-administration of biliverdin (10 μM) completely prevented the effects caused by Cd. Treatment with Zn protoporphyrin IX, a strong inhibitor of heme oxygenase, expectedly decreased heme oxygenase-1 activity to half. When the inhibitor was given together with Cd, completely prevented the enzyme induction and oxidative stress parameters were significantly enhanced.Conclusion: Taking together, these results are indicating that heme oxygenase plays a protective role against oxidative cell damage in soybean nodules.

Effects of Nitric Oxide Synthase and Heme Oxygenase Inducers and Inhibitors on Molecular Signaling of Erectile Function

Changes in gene expression and the enzymatic activity of inducible nitric oxide synthase (iNOS), heme oxygenase isoenzymes 1 and 2 (HO-1 and HO-2) and cyclic GMP (cGMP) tissue levels in the corpora cavernosa (CC) were investigated experimentally in sixty Sprague-Dawley rats subjected to either HO or NOS inducers or inhibitors. They were divided into 5 groups (n = 12/group): Control group, L-arginine group which received L-arginine as NOS inducer, L-nitroarginine methyl ester (L-NAME) group which received L-NAME as NOS inhibitor, Hemin group which received hemin as HO inducer and Zn protoporphyrin (ZnPP) group which received ZnPP as HO inhibitor. The results of this study demonstrated that there was co-induction as well as co-repression of iNOS and HO-1 gene expression using either the inducers or inhibitors of both enzymes, whereas HO-2 gene expression was found to be upregulated by the use of L-arginine only in L-arginine rat group. HO-2 gene expression was not changed by the use of either HO inducer, suppressor or NOS suppressor. cGMP tissue levels in CC was significantly elevated in rats receiving HO inducer (hemin) as well as in those receiving NOS inducer (arginine) as compared with the control group, with a significant elevation in cGMP levels in hemin group as compared with L-arginine group. The use of hemin appears to be more efficient and dominates the use of arginine in inducing HO activity as well as cGMP tissue levels. These data indicate that HO and its product carbon monoxide (CO) are supervising nitric oxide as a signaling molecule in erectile function. On the other hand, There was a significant decrease in cGMP levels in HO inhibitor and in NOS inhibitor groups with a significant decrease in its levels in ZnPP group as compared with L-NAME group. HO enzymatic activity was significantly elevated in the hemin group, whereas it was significantly decreased in ZnPP and in arginine rat groups in comparison with the other groups. NOS activity was significantly elevated in arginine group and in ZnPP group and it was significantly decreased in L-NAME and in hemin rat groups in comparison with the other rat groups. Conclusion: The use of either NOS or HO inducers can equally enhance erectile function via upregulation of gene expression of the two signaling genes involved in erection as well as through upregulation of the tissue levels of cGMP, the mediator of vasodilatation. Our data indicate that HO/CO system is supervising and dominating nitric oxide as a signaling molecule in erectile function. Thus, induction of HO may have therapeutic implications for management of erectile dysfunction.