Digoxin-like Immunoreactive Factor Research Articles

Role of endogenous digitalis-like factors in the clinical manifestations of severe preeclampsia: a systematic review

Endogenous digitalis-like factor(s), originally proposed as a vasoconstrictor natriuretic hormone, was discovered in fetal and neonatal blood accidentally because it cross-reacts with antidigoxin antibodies (ADAs). Early studies using immunoassays with ADA identified the digoxin-like immuno-reactive factor(s) (EDLF) in maternal blood as well, and suggested it originated in the feto-placental unit. Mammalian digoxin-like factors have recently been identified as at least two classes of steroid compounds, plant derived ouabain (O), and several toad derived bufodienolides, most prominent being marinobufagenin (MBG). A synthetic pathway for MBG has been identified in mammalian placental tissue. Elevated maternal and fetal EDLF, O and MBG have been demonstrated in preeclampsia (PE), and inhibition of red cell membrane sodium, potassium ATPase (Na, K ATPase (NKA)) by EDLF is reversed by ADA fragments (ADA-FAB). Accordingly, maternal administration of a commercial ADA-antibody fragment (FAB) was tested in several anecdotal cases of PE, and two, small randomized, prospective, double-blind clinical trials. In the first randomized trial, ADA-FAB was administered post-partum, in the second antepartum. In the post-partum trial, ADA-FAB reduced use of antihypertensive drugs. In the second trial, there was no effect of ADA-FAB on blood pressure, but the fall in maternal creatinine clearance (CrCl) was prevented. In a secondary analysis using the pre-treatment maternal level of circulating Na, K ATPase (NKA) inhibitory activity (NKAI), ADA-FAB reduced the incidence of pulmonary edema and, unexpectedly, that of severe neonatal intraventricular hemorrhage (IVH). The fall in CrCl in patients given placebo was proportional to the circulating level of NKAI. The implications of these findings on the pathophysiology of the clinical manifestations PE are discussed, and a new model of the respective roles of placenta derived anti-angiogenic (AAG) factors (AAGFs) and EDLF is proposed.

In vivo effects of 17β-estradiol on cardiac Na+/K+-ATPase expression and activity in rat heart

In this study the in vivo effects of estradiol in regulating Na+/K+-ATPase function in rat heart was studied. Adult male Wistar rats were treated with estradiol (40μg/kg, i.p.) and after 24h the animals were sacrificed and the heart excised. Following estradiol administration, cardiac Na+/K+-ATPase activity, expression of the α1 subunit, and phosphorylation of the α1 subunit were significantly increased. These animals also had significantly decreased levels of digoxin-like immunoreactive factor(s). Na+ levels were also significantly reduced but to a level that was still within the normal physiological range, highlighting the ability of the Na+/K+-ATPase to balance the ionic composition following treatment with estradiol. Estradiol treated rats also showed increased phosphorylation of protein kinase B (Akt), and extracellular-signal-regulated kinase 1/2 (ERK1/2). We therefore suggest a role for Akt and/or ERK1/2 in estradiol-mediated regulation of cardiac Na+/K+-ATPase expression and activity in rat heart.

Digoxin and digoxin-like immunoreactive factors (DLIF) modulate the release of pro-inflammatory cytokines

Cardiac glycosides such as digoxin and their endogenous counterpart digoxin-like immunoreactive factor (DLIF) may possess anti-inflammatory properties. Pro-inflammatory cytokines from human peripheral blood mononuclear cells (PBMC) were measured by ELISA using specific antibodies. Immunocytochemistry was used to localize NF-K: B. Non-stimulated PBMC constitutively secreted minimum amounts of cytokines. LPS (1 mg/L) stimulation lead to steep increases in TNF-alpha, IL-6 and IL-8 concentrations with peak rises at 8 h. An 8 h delay was observed for IL-10. Increases in IL-10 were sustained for18 h period. Significant inhibition (P > 0.05) of TNF-alpha, IL-6 and IL-8 at non-toxic of digoxin concentration (< 100 nM) and DLIF (10 nM digoxin equivalent (de)) was observed whereas no such effect was seen for IL-10. Inhibition of the degradation of activated NF-K: B in the PBMC was observed with the indicated concentrations of digoxin, DLIF or Pyrrolidine dithiocarbamate (PDTC). Digoxin and DLIF inhibit the release of proinflammatory cytokines from PBMC via NF-K: B-dependent pathway suggesting an anti-inflammatory effect.

Mammalian Cardenolides in Cancer Prevention and Therapeutics

Digoxin-like immunoreactive factor (DLIF) and ouabain-like factor (OLF) are the mammalian counterparts to the plant-derived cardiotonic steroids digoxin and ouabain. Compelling evidence indicates that the cardiotonic steroids may have anticancer properties. Recent evidence indicates that low (nanomolar) concentrations of DLIF selectively induce cell death in transformed cells, while sparing normal cells, and is even more potent than the plant-derived compounds. The discovery that these endogenous molecules may play a role in the regulation of cancer cell proliferation provides a potentially new paradigm for the physiologic role of DLIF and OLF. In addition, the possible use of digoxin itself as a therapeutic agent in cancer has been explored, and evidence suggests that its conversion to dihydrodigoxin may be involved in regulating anticancer activity. The mechanism(s) for the pro-apoptotic property of these compounds is not known. In this brief review, we will discuss the proposed mechanism of action of digoxin, ouabain, DLIF, and OLF as anticancer compounds and discuss the effects that metabolic conversion to their dihydro-derivatives may have on this activity. From the perspective of therapeutic drug monitoring, these findings suggest some potential new challenges in the need to measure concentrations of digoxin and dihydrodigoxin as well as their endogenous counterparts DLIF and OLF in serum.

Digoxin-Like Immunoreactive Factors Induce Apoptosis in Human Acute T-Cell Lymphoblastic Leukemia

Plant-derived cardenolides reportedly possess anticancer properties in human leukemic cells via selective induction of apoptosis, cell cycle arrest, and differentiation. Selective induction of apoptosis with mammalian-derived digoxin-like immunoreactive factor (DLIF) could provide new strategies for anticancer drug development or the identification of biomarkers for cancer. We investigated whether DLIFs selectively induce apoptosis in human lymphoblastic leukemic cells. We compared the relative potencies of digoxin, ouabain, and DLIF on induction of programmed cell death in Jurkat cells (an acute T-leukemic cell line), K-562 (a myelogenous leukemia cell line), and nonpathologic human peripheral blood mononuclear cells (PBMCs). Apoptosis was measured by flow cytometry with the annexin V/propidium iodide method. Digoxin and ouabain induced apoptosis in Jurkat cells [digoxin 50% inhibitory concentration (IC(50)), 24 nmol/L; ouabain IC(50), 26 nmol/L]. Neither digoxin nor ouabain induced apoptosis in K-562 cells or PBMCs. DLIF was more potent (IC(50), 1.9 nmol/L) and >2-fold more effective than digoxin or ouabain at inducing maximum apoptosis in Jurkat cells. The IC(50) values in the apoptosis assays were >100-fold lower (DLIF) and 20-fold lower (digoxin and ouabain) than the IC(50) required for Na(+)- and K(+)-dependent ATPase (DLIF, 200 nmol/L; digoxin, 910 nmol/L; ouabain, 600 nmol/L). DLIF selectively induces apoptosis in a human acute T-cell lymphoblastic leukemia cell line but not in K-562 cells or PBMCs. These data suggest a new physiological role for these endogenous hormone-like factors.

Digoxin-Like Immunoreactive Factor in Human Cerebrospinal Fluid

Back to table of contents Previous article Next article LETTERFull AccessDigoxin-Like Immunoreactive Factor in Human Cerebrospinal FluidRif S. El-Mallakh M.D.James Miller Ph.D.Roland Valdes Jr., Ph.D.,Tamella Buss Cassis M.D.,Rena Li M.D., Ph.D.,Rif S. El-Mallakh M.D.Search for more papers by this authorJames Miller Ph.D.Search for more papers by this authorRoland Valdes Jr., Ph.D.Search for more papers by this author,Tamella Buss Cassis M.D.Search for more papers by this author,Rena Li M.D., Ph.D.Search for more papers by this author,Published Online:1 Jan 2007AboutSectionsPDF/EPUB ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinked InEmail SIR: Mammals produce a family of endogenous compounds that resemble the sodium pump-inhibiting cardiac glycosides, digoxin and ouabain. 1 These endogenous cardenolides have been proposed to be important in the pathophysiology of bipolar illness, where it is relatively deficient in the serum of manic subjects. 1 The measurement of these compounds in the CNS has not been previously documented. Refrigerated cerebrospinal fluid (CSF) waste was frozen at −70°C within 18 hours of collection. Digoxin-like immunoreactive factor (DLIF) was measured by radioimmunoassay as described by Grider et al., 1 using components provided by New England Nuclear (Billerica, Mass.). The correlation among four samples quantified twice was 0.88. Ten samples were studied from three women and seven men. Mean age among the women was 41.9 years (range=30 to 53), and among the men it was 61.6 years (range=32 to 94). Mean DLIF concentration was 91.7 pg digoxin equivalents/ml.Comment Human CSF contains a substance which cross reacts with antidigoxin antibodies. This supports previous reports of CSF factors that bear immunologic resemblance to cardiac glycosides 2 and sodium pump inhibitory activity of CSF. 3 Though the source of CSF DLIF is not known, similar compounds have been purified from mammalian hypothalamus. 4 Given that the hypothalamic-pituitary-adrenal axis may be dysregulated in bipolar illness, that DLIF production is responsive to dexamethasone and adreno-corticotropin hormone (ACTH), 5 and that DLIF may be dysregulated in bipolar subjects, 1 it would seem important to study CSF DLIF in bipolar illness. Department of Pathology and Laboratory Medicine, University of Louisville School of Medicine, Louisville, Ky.Advanced Dermatology and Dermaesthetics, Louisville, Ky.Sun Health Institute, Sun City, Ariz.

P.5.039 Acamprosate counteracts anxiety-like behaviorinduced by multiple ethanol withdrawals

In this study the in vivo effects of estradiol in regulating Na+/K+-ATPase function in rat heart was studied. Adult male Wistar rats were treated with estradiol (40 μg/kg, i.p.) and after 24 h the animals were sacrificed and the heart excised. Following estradiol administration, cardiac Na+/K+-ATPase activity, expression of the α1 subunit, and phosphorylation of the α1 subunit were significantly increased. These animals also had significantly decreased levels of digoxin-like immunoreactive factor(s). Na+ levels were also significantly reduced but to a level that was still within the normal physiological range, highlighting the ability of the Na+/K+-ATPase to balance the ionic composition following treatment with estradiol. Estradiol treated rats also showed increased phosphorylation of protein kinase B (Akt), and extracellular-signal-regulated kinase 1/2 (ERK1/2). We therefore suggest a role for Akt and/or ERK1/2 in estradiol-mediated regulation of cardiac Na+/K+-ATPase expression and activity in rat heart.

P.6.052 The effect of lamotrigine on depressiveand convulsive modifications induced by pentetrazole and reserpine in mice

In this study the in vivo effects of estradiol in regulating Na+/K+-ATPase function in rat heart was studied. Adult male Wistar rats were treated with estradiol (40 μg/kg, i.p.) and after 24 h the animals were sacrificed and the heart excised. Following estradiol administration, cardiac Na+/K+-ATPase activity, expression of the α1 subunit, and phosphorylation of the α1 subunit were significantly increased. These animals also had significantly decreased levels of digoxin-like immunoreactive factor(s). Na+ levels were also significantly reduced but to a level that was still within the normal physiological range, highlighting the ability of the Na+/K+-ATPase to balance the ionic composition following treatment with estradiol. Estradiol treated rats also showed increased phosphorylation of protein kinase B (Akt), and extracellular-signal-regulated kinase 1/2 (ERK1/2). We therefore suggest a role for Akt and/or ERK1/2 in estradiol-mediated regulation of cardiac Na+/K+-ATPase expression and activity in rat heart.

S.01.05 Are novel antipsychotics as effective as anticonvulsants or lithium on the long-term treatment of bipolar disorder?

In this study the in vivo effects of estradiol in regulating Na+/K+-ATPase function in rat heart was studied. Adult male Wistar rats were treated with estradiol (40 μg/kg, i.p.) and after 24 h the animals were sacrificed and the heart excised. Following estradiol administration, cardiac Na+/K+-ATPase activity, expression of the α1 subunit, and phosphorylation of the α1 subunit were significantly increased. These animals also had significantly decreased levels of digoxin-like immunoreactive factor(s). Na+ levels were also significantly reduced but to a level that was still within the normal physiological range, highlighting the ability of the Na+/K+-ATPase to balance the ionic composition following treatment with estradiol. Estradiol treated rats also showed increased phosphorylation of protein kinase B (Akt), and extracellular-signal-regulated kinase 1/2 (ERK1/2). We therefore suggest a role for Akt and/or ERK1/2 in estradiol-mediated regulation of cardiac Na+/K+-ATPase expression and activity in rat heart.

Endogenous digoxin-like immunoreactive factor (DLIF) serum concentrations are decreased in manic bipolar patients compared to normal controls

Background: A decrease in sodium pump activity in erythrocytes has been associated with manic episodes of bipolar illness relative to euthymic moods. Since red blood cells are long-lived and lack a nucleus, it is likely that a plasma factor is responsible for the observed decrease in sodium pump activity. Methods: Utilizing a radioimmunoassay, we examined the serum concentrations of the digoxin-like immunoreactive factor (DLIF) in ill and well bipolar patients and compared the values to those of normal controls. Results: DLIF was significantly decreased in manic individuals as compared to normal controls (143.6±S.E.M. 20.94 vs 296.6±12.76 pg digoxin equivalents/ml, respectively, F=4.77, P<0.05), but not compared to euthymic bipolar subjects 213.8±86.92, P=0.77). There were no significant differences in DLIF concentrations between manic and euthymic bipolar individuals ( P=0.8). Since relapse in bipolar patients appears to display a seasonal pattern, we also measured the plasma concentration of this factor over a 12-months period. Normal controls exhibited a seasonal pattern of change in serum DLIF concentrations with a nadir in the winter months. Plasma concentrations of DLIF in bipolar patients did not show a seasonal pattern and maintained low levels throughout the year. Limitations: Due to the nonspecificity of our antibody, we could measure only total DLIF. Furthermore, it is unclear what the role of circulating DLIF, if any, may be on brain function. Conclusion: DLIF may be involved in the pathophysiology of mania.

Analytic performance of two automated nonpretreatment digoxin immunoassays.

The analytic performance of two automated nonpretreatment digoxin methods, AxSYM Digoxin II and Vitros digoxin immunoassays, was assessed. Both assays had analytic sensitivities of less than 0.2 microg/L, were linear from digoxin concentrations of 0.5 to 4.0 microg/L, and showed acceptable precision, with a maximum total coefficient of variation (CV) of 8.9% and 6.4% for the AxSYM and Vitros, respectively. Comparison of the two methods using samples from patients receiving digoxin gave the following relationship: Vitros = 0.91 x AxSYM + 0.23 (r = 0.97, Sy,x = 0.12). Digoxinlike immunoreactive factor (DLIF) crossreactivity was examined in specimens from patients who had hepatic disease, renal insufficiency, had undergone cardiac surgery, and in neonatal cord blood samples. Minimal crossreactivity was observed for most samples and the average crossreactivity for each group of samples was comparable for the two methods. The recovery of digoxin added to samples from each group of DLIF was similar, except for that from cord blood samples, for which recovery was significantly lower with the AxSYM method. Titration of a digoxin-spiked serum pool with digoxin-immune Fab showed a similar decrease in the measured digoxin concentration for both methods. Overall, the analytic performance characteristics of these two methods were comparable.

Serum digoxin‐like immunoreactive factor in children and its relation to sodium metabolism

Serum digoxin‐like immunoreactive factor (DLIF), an endogenous substance that cross‐reacts with anti‐digoxin antibodies, was assessed (fluorescence polarization immunoassay) in children (n= 134) aged 5‐16 y, who had never been treated with cardiac glycosides. DLIF was found in 50% of serum samples at a mean concentration of 0.16 ± 0.06 ng/ml (range 0.03–0.35 ng/ml). Although the study population as a whole was apparently homogeneous with regard to serum sodium content, and none had clinical signs of sodium imbalance, children with DLIF showed significantly lower natraemia (p= 0:0002), higher urinary concentration (p= 0:001) and fractional excretion (p= 0:001) of sodium, and increased systolic blood pressure (p= 0:009) compared with children without DLIF. Inverse correlations were found between DLIF concentration and serum sodium (p &lt; 0:01), urine sodium content (p &lt; 0:001), 24‐h sodium excretion (p &lt; 0:001), systolic (p &lt; 0:001) and diastolic (p &lt; 0:01) blood pressure. These findings suggest that sodium handling is different in children with and without DLIF, since this material seems to be released preferably in subjects who show a trend towards negative sodium balance. Such an association suggests that DLIF may be a physiologically relevant material involved in sodium homeostasis.

Standards of laboratory practice: cardiac drug monitoring

In this Standard of Laboratory Practice we recommend guidelines for therapeutic monitoring of cardiac drugs. Cardiac drugs are primarily used for treatment of angina, arrhythmias, and congestive heart failure. Digoxin, used in congestive heart failure, is widely prescribed and therapeutically monitored. Monitoring and use of antiarrhythmics such as disopyramide and lidocaine have been steadily declining. Immunoassay techniques are currently the most popular methods for measuring cardiac drugs. Several reasons make measurement of cardiac drugs in serum important: their narrow therapeutic index, similarity in clinical complications and presentation of under- and overmedicated patients, need for dosage adjustments, and confirmation of patient compliance. Monitoring may also be necessary in other circumstances, such as assessment of acetylator phenotypes. We present recommendations for measuring digoxin, quinidine, procainamide (and N-acetylprocainamide), lidocaine, and flecainide. We discuss guidelines for measuring unbound digoxin in the presence of an antidote (Fab fragments), for characterizing the impact of digoxin-like immunoreactive factor (DLIF) and other cross-reactants on immunoassays, and for moni-toring the unbound (free fraction) of drugs that bind to alpha1-acid glycoprotein. We also discuss logistic, clinical, hospital, and laboratory practice guidelines needed for implementation of a successful therapeutic drug monitoring service for cardiac drugs.

Serum digoxin-like immunoreactive factor in children and its relation to sodium metabolism.

Serum digoxin-like immunoreactive factor (DLIF), an endogenous substance that cross-reacts with antidigoxin antibodies, was assessed (fluorescence polarization immunoassay) in children (n = 134) aged 5-16 y, who had never been treated with cardiac glycosides. DLIF was found in 50% of serum samples at a mean concentration of 0.16 +/- 0.06 ng/ml (range 0.03-0.35 ng/ml). Although the study population as a whole was apparently homogeneous with regard to serum sodium content, and none had clinical signs of sodium imbalance, children with DLIF showed significantly lower natraemia (p = 0.0002), higher urinary concentration (p = 0.001) and fractional excretion (p = 0.001) of sodium, and increased systolic blood pressure (p = 0.009) compared with children without DLIF. Inverse correlations were found between DLIF concentration and serum sodium (p < 0.01), urine sodium content (p < 0.001), 24-h sodium excretion (p < 0.001), systolic (p < 0.001) and diastolic (p < 0.01) blood pressure. These findings suggest that sodium handling is different in children with and without DLIF, since this material seems to be released preferably in subjects who show a trend towards negative sodium balance. Such an association suggests that DLIF may be a physiologically relevant material involved in sodium homeostasis.

Digoxinlike Immunoreactive Factor Isolated From Human Pleural Fluids is Structurally Similar to Digoxin

AbstractTo further define the chemical structure of human endogenous digoxinlike immunoreactive factors (DLIF) we used human pleural effusions as a source of the substance. Digoxinlike immunoreactive factor activity was detected by radioimmunoassay in the pleural fluid of each of four patients; average concentration was 0.35 ng/mL. The chemical profile of DLIF was determined by initial extraction and concentration of DLIF by ion exchange chromatography followed by reverse phase-high-pressure liquid chromatography (RP-HPLC) separation and purification. Using high-pressure liquid chromatography cochromatography of DLIF, together with several radioactively marked glycosides, we observed a single peak of DLIF activity that was chro- matographically identical to digoxin. The present study further supports the recent finding that DLIF is related structurally to the cardiac glycosides, and for the first time it has been proven that DLIF is present in pleural fluids

Comparability of a new turbidimetric digoxin test with other immunochemical tests and with HPLC—a multicenter evaluation

A new turbidimetric inhibition immunoassay for digoxin (Tina-quant [a] Digoxin, Boehringer Mannheim) was evaluated in seven laboratories. It can be performed without sample pretreatment with ready-to-use reagents on nondedicated analyzers in combination with routine clinical chemistry. The studies revealed a good analytical performance: lower limit of detection 0.12 microg/L (3 SD from mean of blank); linearity up to 7.5 microg/L; median between-run CVs 8.1% (0.6 microg/L), 2.8% (1.5 microg/L), 1.9% (3 microg/L); mean analytical recovery in control sera 98-102%; slopes from 0.97 to 1.09 and intercepts from -0.28 to 0.10 microg/L in comparison with four immunoassays; and a high resistance to common interferents. The test was more resistant to digoxin-like immunoreactive factor (DLIF) interference than other methods, showing cross-reactivity only in some intensive care patient samples. Among 192 patients in whom DLIF is expected (e.g., pregnant women, patients with renal failure, newborns), 90% of results were < or =0.26 microg/L digoxin. Cortisol showed no cross-reactivity and digoxigenin had a low reactivity. An interlaboratory survey revealed a good comparability of the Tina-quant [a] test with the median of all methods (slope 0.99, intercept -0.06 microg/L). An HPLC method for digoxin based on isocratic separation of samples on an RP-18 column followed by detection by an immunoassay yielded a reasonable comparability with the immunochemical tests with noncritical samples. Divergent results of immunoassays caused by DLIFs or different cross-reactivities with digoxin metabolites or derivatives can be explained by the use of this HPLC method.

Digoxin and Its Related Endogenous Factors

The digitalis drugs are plant-derived cardenolide compounds used medicinally for several hundred years. These drugs elicit inotropic and chronotropic effects on the heart, but they also affect many other tissues. The mechanism of action involves inhibition of the ion-transport activity of a membrane-associated protein called Na,K-ATPase (sodium pump). Present theory holds that the sodium pump is the principal molecular receptor for the digitalis drugs. Recent evidence indicates the presence of naturally occurring digitalis-like compounds in mammals. It is believed these compounds, collectively known as either digitalis-like (DLF) or ouabain-like (OLF) factors, may be endogenous hormones regulating the biological activity of the sodium pump and its isoforms. The presence of deglycosylated and other congeners of one specific DLF, the digoxin-like immunoreactive factor (DLIF), has very recently been described in humans. Digoxin as a drug is the most widely prescribed digitalis in the U.S., and its measurement in serum has established a model for present-day therapeutic drug monitoring (TDM). Historically, the accurate measurement of digoxin in blood has been difficult. This article focuses on the present understanding of the clinical use of digoxin, factors that affect the accuracy of measuring digoxin, the principle of measuring metabolically active species of digoxin, and the effects of DLIF and other interfering substances in digoxin immunoassay.

Atrial natriuretic factor and digoxin-like immunoreactive factor in diabetic patients: their interrelation and the influence of the autonomic nervous system.

In diabetic patients, several factors contribute to volume expansion and have to be counteracted by humoral and neuronal feedback control systems. We investigated N-terminal proatrial natriuretic factor (ANF1-98) and digoxin-like immunoreactive factor (DLIF), which are two counteracting hormones, and their interrelationship, with additional consideration given to autonomic nervous function in diabetic patients. ANF1-98 and DLIF were measured in 64 diabetic patients. Autonomic nervous function was assessed using nine autonomic nervous function tests. The patients were subdivided into two groups, one with four or more (group 1) and one with less than four abnormal results in autonomic function tests (group 2). Compared with group 2, group 1 demonstrated detectable DLIF levels less often (17.2 vs. 45.7, P = 0.0195) and increased levels of ANF1-98 (mean +/- SEM: 850.0 +/- 108.8 vs. 554.8 +/- 45.9 pmol/L, P = 0.0099). However, the groups did not differ in blood pressure, daily sodium, and daily potassium excretion. The number of abnormal autonomic function tests correlated significantly with ANF1-98 (P = 0.0002). In patients with detectable DLIF, DLIF correlated with ANF1-98 (P = 0.0080). These results demonstrate close interactions between the autonomic nervous system and the two natriuretic hormones. In patients with autonomic nervous dysfunction, higher levels of ANF may possibly compensate for the lack of the natriuretic DLIF to counteract hypertension and chronic volume expansion.

Atrial natriuretic factor and digoxin-like immunoreactive factor in diabetic patients: their interrelation and the influence of the autonomic nervous system

Atrial natriuretic factor and digoxin-like immunoreactive factor in diabetic patients: their interrelation and the influence of the autonomic nervous system

Evidence for presence of a reduced form of digoxin-like immunoreactive factor (dihydro-DLIF) in mammalian tissues

Digoxin-like immunoreactive factor (DLIF) from adrenal glands is an endogenous ligand structurally related to the plant-derived cardiac glycoside digoxin. Cardiac glycosides regulate the activity of the sodium pump and thus play key roles in disease processes involving regulation of ion transport. We now report the discovery of an endogenous dihydro-DLIF analogous to dihydrodigoxin. We used HPLC, ultraviolet spectrophotometry, and cross-reactivity with two antibodies, one specific for digoxin and one for dihydrodigoxin, to support the hypothesis that dihydro-DLIF contains a chemically reduced lactone ring. The spectral absorbance maximum for dihydro-DLIF is at 196 nm, identical to dihydrodigoxin. DLIF and dihydro-DLIF are 975- and 2588-fold less immunoreactive than digoxin and dihydrodigoxin for their respective antibodies. The molar ratio of dihydro-DLIF to DLIF is approximately 5.3 in bovine adrenocortical tissue and approximately 0.38 in human serum. Dihydrodigoxin (reduced lactone ring) added to microsomes isolated from bovine adrenal cortex produced a 4.5-fold increase in digoxin-like immunoreactivity (oxidized lactone ring) after 3 h of incubation. The biotransformation is likely mediated by a cytochrome P-450 NADPH-dependent process. Our findings demonstrate the presence of a dihydro-DLIF in mammals and suggest a metabolic route for synthesis of endogenous DLIF in mammalian tissue.