Electrostatically stabilized hybrids of carbon and maghemite nanoparticles: electrochemical study and application.

Binary hybrids have been investigated for the past few decades due to the emerging properties of nanoparticle composites. Electrostatically stabilized core-shell nanostructures composed of surface active magnetic nanoparticles (SAMNs) and differently charged carbon nanomaterials display specific electrochemical properties. In this work, a set of binary hybrids that include a new class of magnetic nanoparticles is presented and the electrochemical features of the hybrids are reported. Gallic acid derived carbon dots (GA-CDs), PEG derived graphene dots (PEG-GDs), and quaternized carbon dots (Q-CDs) characterized by different charged groups were used for the preparation of different complexes with SAMNs. Thus, a set of six binary nanomaterials was obtained, and characterized by electrochemical impedance spectroscopy, cyclic voltammetry and chronoamperometry, demonstrating significant differences in the charge transfer resistance, capacitive current, electrochemical performance, and reversibility with respect to the isolated subunits. Among them, the combination of Q-CDs with an excess of SAMNs led to a Q-CD@SAMN hybrid, which displayed peculiar electrocatalytic properties attributable to the influence of the strong electrostatic interactions exerted by Q-CDs on the SAMN surface. Notwithstanding their small fraction (around 1% w/w), Q-CDs oriented the electrocatalysis of SAMNs toward the selective electro-oxidation of polyphenols at low applied potentials (+0.1 V vs. SCE). Finally, the Q-CD@SAMN hybrid was used for the development of a coulometric sensor for polyphenols, composed of a simple carbon paste electrode in a small volume electrochemical flow cell (1 µL), and used for the complete direct electro-oxidation of polyphenols from plant extracts.

Long-term chikungunya infection clinical manifestations after an outbreak in Italy: a prognostic cohort study.

OBJECTIVES: Following a Chikungunya (CHIKV) outbreak in Italy, a cohort study was conducted to describe the infection long-term clinical course and outcome. METHODS: Persons identified through active and passive surveillance as confirmed or possible CHIKV cases during the outbreak were enrolled and interviewed by trained public health nurses, between 4-5 and 12-13 months following the acute stage. Patients reporting persistent clinical symptoms were evaluated by rheumatologists. Serum samples were obtained and anti-CHIKV specific IgG and IgM immune responses detected. Only confirmed cases who completed the follow-up were analysed. RESULTS: Out of 250 patients, 66.5% still reported myalgia, asthenia or arthralgia (most frequent sign) after 12 months. Functional ability, measured by the ROAD index, was more impaired for lower extremities (3.75; Inter Quartile Range - IQR 4.4), and the activities of daily living (average 4.2; IQR 5). Variables independently associated with the presence of joint pain at 12-13 months were increasing age, and history of rheumatologic diseases). Elderly, females, and persons with history of rheumatologic diseases had higher anti-CHIKV IgG titres at 12-13 months. CONCLUSIONS: This study confirms, in an unselected population, that the long-lasting burden of CHIKV infection is significant.

Scleroderma fibroblasts constitutively express the long pentraxin PTX3.

OBJECTIVE: PTX3 is a secreted molecule which consists of a C-terminal domain similar to classical pentraxins (e.g. C-reactive protein) and of an unrelated N-terminal domain. Unlike the classical pentraxins, PTX3 is expressed in response to IL-1beta and TNF-alpha but not to IL-6. The present study was designed to investigate the expression of PTX3 in normal and scleroderma fibroblasts. METHODS: Normal and SSc fibroblasts were cultured in the presence and absence of inflammatory cytokines. PTX3 mRNA expression in fibroblasts was evaluated by Northern analysis. PTX3 protein levels in fibroblast culture medium were estimated by ELISA. RESULTS: Normal fibroblasts were induced to express high levels of P7X3 mRNA by IL-1beta and TNF-alpha but not by other cytokines or growth factors. Scleroderma fibroblasts, unlike normal fibroblasts, constitutively expressed high levels of PTX3 in the absence of deliberate stimulation. The constitutive expression of PTX3 in SSc fibroblasts was not modified by anti-TNF-alpha antibodies or IL-1 receptor antagonist. In contrast, IFN-gamma and TGF-beta inhibited the constitutive but not the stimulated expression of PTX3 in SSc fibroblasts. CONCLUSIONS: PTX3 is a main feature of activated scleroderma fibroblasts.

Oxidative stress in scleroderma: maintenance of scleroderma fibroblast phenotype by the constitutive up-regulation of reactive oxygen species generation through the NADPH oxidase complex pathway.

OBJECTIVE: To explore the role of reactive oxygen species (ROS) in the in vitro activation of skin fibroblasts from patients with systemic sclerosis (SSc). METHODS: Fibroblasts were obtained from involved skin of patients with limited or diffuse SSc. Oxidative activity imaging in living cells was carried out using confocal microscopy. Levels of O2- and H2O2 released from fibroblasts were estimated by the superoxide dismutase (SOD)-inhibitable cytochrome c reduction and homovanilic acid assays, respectively. To verify NADPH oxidase activation, the light membrane of fibroblasts was immunoblotted with an anti-p47phox-specific antibody. Fibroblasts were stimulated with various cytokines and growth factors to determine whether any of these factors modulate ROS generation. Cell proliferation was estimated by 3H-thymidine incorporation. Northern blot analysis was used to study alpha1 and alpha2 type I collagen gene expression. RESULTS: Unstimulated skin fibroblasts from SSc patients released more O2- and H2O2 in vitro through the NADPH oxidase complex pathway than did normal fibroblasts, since incubation of SSc fibroblasts with diphenylene iodonium, a flavoprotein inhibitor, suppressed the generation of ROS. This suppression was not seen with rotenone, a mitochondrial oxidase inhibitor, or allopurinol, a xanthine oxidase inhibitor. Furthermore, the cytosolic component of NADPH oxidase, p47phox, was translocated to the plasma membrane of resting SSc fibroblasts. A transient increase in ROS production was induced in normal but not in SSc fibroblasts by interleukin-1beta (IL-1beta), platelet-derived growth factor type BB (PDGF-BB), transforming growth factor beta1 (TGFbeta1), and H2O2. Treatment of normal and SSc fibroblasts with tumor necrosis factor a (TNFalpha), IL-2, IL-4, IL-6, IL-10, interferon-alpha (IFNalpha), IFNgamma, granulocyte-macrophage colony-stimulating factor (GM-CSP), G-CSF, or connective tissue growth factor (CTGF) had no effect on ROS generation. Constitutive ROS production by SSc fibroblasts was not inhibited when these cells were treated with catalase, SOD, IL-1 receptor antagonist, or antibodies blocking the effect of TGFbeta1, PDGF-BB, and other agonists (IL-4, IL-6, TNFalpha, CTGF). In contrast, treatment of SSc fibroblasts with the membrane-permeant antioxidant N-acetyl-L-cysteine inhibited ROS production, and this was accompanied by decreased proliferation of these cells and down-regulation of alpha1(I) and alpha2(I) collagen messenger RNA. CONCLUSION: The constitutive intracellular production of ROS by SSc fibroblasts derives from the activation of an NADPH oxidase-like system and is essential to fibroblast proliferation and expression of type I collagen genes in SSc cells. Our results also exclude O2-, H2O2, IL-1beta, TGFbeta1, PDGF-BB, IL-4, IL-6, TNFalpha, or CTGF as mediators of a positive, autocrine feedback mechanism of ROS generation.

Intravenous N-acetylcysteine for treatment of Raynaud's phenomenon secondary to systemic sclerosis: a pilot study.

OBJECTIVE: To assess the efficacy and tolerability of N-acetylcysteine (NAC) in patients with Raynaud's phenomenon (RP) secondary to systemic sclerosis (scleroderma; SSc). METHODS: Twenty-two patients with RP secondary to SSc were enrolled in a multicenter, open clinical trial lasting 11 weeks and conducted in winter. Primary outcome measures were frequency and severity of RP attacks, and number of digital ulcers. Secondary outcome measure was improvement in digital cold challenge test assessed by photoelectric plethysmography. Patients received a continuous 5 day intravenous infusion of NAC starting with a 2 h loading dose of 150 mg/kg subsequently adjusted to 15 mg/kg/h. RESULTS: All 22 patients completed the 5 day infusion and 20 of them the posttreatment followup. Both frequency and severity of RP attacks decreased significantly compared to pretreatment values. Active ulcers were significantly less numerous at all followup visits (25.18% of baseline count on Day 33 from the beginning of infusion). In the cold challenge test mean recovery time fell by 69.56%, 67.70%, 71.42%, and 71.05% on Days 12, 19. 33, and 61 from the beginning of treatment. Side effects were minor, easily controlled, and reversible. CONCLUSION: N-acetylcysteine appears to be safe for the treatment of RP secondary to SSc. These preliminary data warrant further controlled studies.

Expression and production of the long pentraxin PTX3 in rheumatoid arthritis (RA).

PTX3 is a secreted molecule which consists of a C-terminal domain similar to classical pentraxins (e.g. C-reactive protein (CRP)) and of an unrelated N-terminal domain. Unlike the classical pentraxins, the long pentraxin PTX3 is expressed in response to IL-1beta and tumour necrosis factor-alpha (TNF-alpha), but not to IL-6, in various cell types. The present study was designed to investigate the expression of PTX3 in RA. Dissociated RA and osteoarthritis (OA) type B synoviocytes were cultured in the presence and in the absence of inflammatory cytokines. PTX3 mRNA expression in synoviocytes was evaluated by Northern analysis. PTX3 protein levels in synovial cell cultures and synovial fluid were estimated by ELISA, and PTX3 distribution in synovial tissues by immunohistochemical techniques. OA synoviocytes were induced to express high levels of PTX3 mRNA by TNF-alpha, but not by other cytokines including IL-1beta and IL-6. RA synoviocytes, unlike OA synoviocytes, constitutively expressed high levels of PTX3 in the absence of deliberate stimulation. The constitutive expression of PTX3 in RA synoviocytes was not modified by anti-TNF-alpha antibodies, IL-1 receptor antagonist or a combination of the two agents. In contrast, interferon-gamma and transforming growth factor-beta inhibited PTX3 constitutive expression in RA synoviocytes. The joint fluid from RA patients contained higher levels of immunoreactive PTX3 than controls and the synovial tissue contained endothelial cells and synoviocytes positive for PTX3 by immunohistochemistry. In conclusion, PTX3 may play a role in inflammatory circuits of RA, and its relevance as a marker of disease activity deserves further study.

Monocytes of patients wiht systemic sclerosis (scleroderma spontaneously release in vitro increased amounts of superoxide anion.

It has been suggested that toxic oxygen free radicals can be involved in the pathogenesis of systemic sclerosis (scleroderma) (SSc). Because the cells that contribute to the generation of free radicals are not known, our aim was (i) to evaluate the ability of unmanipulated and phorbol 12-myristate 13-acetate-stimulated monocytes and polymorphonucleate neutrophils of SSc patients to generate superoxide anion (O2*-); and (ii) to investigate whether the O2*- produced by these cells involved the activation of nicotinamide-adenine dinucleotide diphosphate oxidase biochemical pathway. Employing the superoxide dismutase-inhibitable reduction of cytochrome c to evaluate the generation of O2*-, unmanipulated monocytes of SSc patients generated more O2*- than primary Raynaud's phenomenon patients and normal control monocytes (p = 0.0001), and the release was higher in patients with diffuse cutaneous involvement and 5 y or less disease duration (p = 0.02). The involvement of nicotinamide-adenine dinucleotide diphosphate oxidase in the enhanced 02*- production was demonstrated by the finding that the cytosolic components of the enzyme, p47phox and p67phox, were both translocated to the plasma membrane of enriched but otherwise unmanipulated monocytes of SSc patients. The involvement of mitochondrial oxidases was excluded by the lack of inhibition of O2*- production when monocytes were incubated in the presence of rotenone, a mitochondrial oxidase inhibitor. Upon stimulation with phorbol 12-myristate 13-acetate, monocytes of SSc patients produced more O2*- than controls. In SSc patients untreated polymorphonucleate neutrophils generated significantly less O2*- than monocytes (p = 0.0001) and only slightly more than polymorphonucleate neutrophils of primary Raynaud's phenomenon patients and normal controls (p = 0.03). In conclusion, we demonstrate that in patients with scleroderma, unmanipulated and phorbol 12-myristate 13-acetate-stimulated monocytes release in vitro increased amounts of superoxide anion through the activation of nicotinamide-adenine dinucleotide diphosphate oxidase and, thus, contribute to the oxidative stress found in this disease.

Reactive oxygen intermediates cause rapid release of the interleukin-1 decoy receptor from human myelomonocytic cells.

Free radicals play an important role in inflammation. We found that reactive oxygen intermediates (ROI) inhibit interleukin-1beta (IL-1beta) binding on human myelomonocytes. Production of superoxide anion (O2-) by Xanthine (X) and Xanthine-Oxidase (XO) or NADPH caused a reduction (48% +/- 15% in 25 experiments) in the IL-1beta binding of polymorphonuclear cells (PMN) and monocytes that was inhibited by superoxide dismutase (SOD). Hydrogen peroxide (H2O2) was only active on monocytes and this effect was prevented by catalase. O2(-)-induced loss of IL-1beta binding on PMN reached half maximum at 5 minutes and peaked after 30 minutes. The reduction of IL-1beta binding was due to reduction of IL-1beta receptors (R) on PMN surface without any change in affinity. ROI-induced reduction of surface IL-1R was not caused by receptor internalization, but rather by the release of a soluble form (45 kD) of the type II decoy R. The action of ROI on IL-1 binding was selective because major histocompatibility complex class I, CD18 and CD16 were unaffected. The O2(-)-induced release of IL-1 decoy R was not affected by protein synthesis inhibitors, but was partially blocked by protease inhibitors. Release of the IL-1 type II decoy R might represent one mechanism by which ROI antagonize and limit the proinflammatory effects of IL-1.

Humoral immune response and natural killer activity in patients with mixed cryoglobulinemia.

OBJECTIVE: Based on serological and molecular evidence of hepatitis C virus (HCV) infection in a significant proportion of patients with mixed cryoglobulinemia (MC), a direct association between HCV and MC has been suggested. The goal of the present study was to investigate the role played by HCV and by the immune response to the virus in the pathogenesis of mixed cryoglobulinemia. METHODS: A competitive reverse transcription polymerase chain reaction was employed to evaluate the concentrations of specific HCV RNA sequences in different clinical specimens (plasma, sera, cryoprecipitates, bone marrow and peripheral blood cells). Using recombinant and synthetic peptides covering the HCV core, envelope 1 (E1) and nonstructural regions 4 (NS4) and 5 (NS5), the humoral immune response in a group of MC patients was assessed with an enzyme-linked immunosorbent assay. Natural killer (NK) cell activity was estimated using a 4 hr 51 Cr release assay. RESULTS: Quantitation of the RNA molecules in the biological samples confirmed an increased virion concentration in cryoprecipitates from 13/15 patients with mixed cryoglobulinemia. Analysis of the humoral immune response against the synthetic peptides suggested a distinct response to HCV antigens in MC patients when compared to patients with HCV infection but without serological evidence of cryoglobulinemia. Unstimulated NK cell functioning was below the normal range in all patients tested. However, peripheral blood mononuclear cells showed no enhancement of NK activity by the interferon inducer polyinosinic acid:polycytidilic acid. Enhancement by interferon-alpha was normal, suggesting an impairment in interferon production. CONCLUSION: The quantitative data are in line with the hypothesis of a direct or indirect role of HCV in mixed cryoglobulinemia. The abnormal immune response could be involved in the onset and persistence of HCV infection, and possibly in the appearance of cryoglobulinemia.

Antibodies against terminal galactosyl alpha(1-3) galactose epitopes in patients with idiopathic myelofibrosis.

Sera from patients with myelofibrosis were analysed for circulating antibodies against an antigenic determinant characterized by two molecules of galactose in alpha 1-3 linkage (anti-Gal antibodies). 50% of patients were found to have values above the 90th percentile of the values of control sera chosen as a cut-off. The median level of the antibodies was significantly higher than the value detected in normal controls, but no difference could be found between patients with idiopathic myelofibrosis and those with myelofibrosis associated to a chronic myeloproliferative disorder. Anti-Gal antibodies were found to correlate with disease activity and with platelet count whereas no correlation was detected with other haematological parameters. Furthermore, for evaluation of disease activity, determination of serum anti-Gal antibodies was a sensitive and specific parameter. We conclude that humoral immunity against Gal alpha 1-3Gal may provide a sensitive tool to detect disease activity in patients with idiopathic myelofibrosis and may be important for understanding its pathogenesis.

Immunohistochemical localization of intracellular and extracellular associated TGF beta in the skin of patients with systemic sclerosis (scleroderma) and primary Raynaud's phenomenon.

We have investigated the distribution of TGF beta using antibodies specific for its intracellular and extracellular forms in full-thickness biopsies of patients with SSc, primary Raynaud's phenomenon (PRP), systemic lupus erythematosus (SLE), and from normal subjects. Nine of 11 SSc biopsies demonstrated intracellular TGF beta in endothelial cells while only 6 exhibited extracellular TGF beta. Endothelial cells in skin biopsies of all PRP patients displayed both intracellular and extracellular TGF beta. All other control biopsies were negative. In patients with PRP, some positively staining fibroblasts were found scattered throughout the dermis. Lastly, extracellular TGF beta was localized in the papillary dermis of PRP and SSc biopsies and in all the dermal layers of SLE patients. No significant staining of TGF beta was observed in the endothelial cells, fibroblasts, or in the extracellular matrix of the majority of biopsies from normal subjects. These data suggest that TGF beta may be one of the cytokines involved in the early stages of pathogenesis of SSc, and that endothelial cells in SSc and PRP may be a source and/or a target of TGF beta.

Influence of naloxone infusion on prolactin and growth hormone response to growth hormone-releasing hormone in anorexia nervosa.

Anorexia nervosa (AN) is frequently associated with anomalies of growth hormone (GH) and prolactin (PRL) secretion. We studied the GH and PRL responses to GHRH1-44 (50 micrograms IV) and the effect of a naloxone infusion (1.6 mg/hr), started 1 hr before GHRH administration, on this response in 12 female patients with AN, aged 15-30 yr, and in seven normal women, aged 19-27 yr, during the follicular phase as controls. In AN, GHRH induced an increase in GH levels similar to that observed in normal subjects. A significant inhibition of the GH response to GHRH was observed during naloxone infusion, similar to the inhibition in normal female subjects during the follicular phase. PRL levels showed a significant increment after GHRH alone and a slight, nonsignificant, PRL increment after GHRH during naloxone infusion in AN patients. In contrast a slight PRL decrease was observed after GHRH, both before and during naloxone infusion, in the normal subjects. Our study demonstrates that endogenous opioids play a role in influencing PRL secretion in patients with AN different from their role in normal subjects.

[The role of autoimmune mechanisms in the hyperthyroidism-viral hepatitis syndrome: a possible noncausal association. A review of the literature and the authors' own cases]. / Ruolo dei meccanismi autoimmuni nella sindrome ipertiroidismo-epatite virale: una possibile non casuale associazione. Revisione della letteratura e casistica personale.

Several patients with both Basedow-Graves' hyperthyroidism and viral hepatitis were observed and it was hypothesised that this association could be explained by the individual genetic pattern in which an immunological fragility conditioned a predisposition to autoimmune diseases. This paper reports on the cases observed, the patients' histocompatibility antigen profile and the experimental data found in the literature on autoimmune involvement in the two diseases, whose association may not be coincidental.

[Role of endogenous opioids in the modulation of hypophyseal hormone secretion]. / Ruolo degli oppioidi endogeni nella modulazione della secrezione ormonale ipofisaria.

The aim of the present study is to review current knowledge of the endogenous opiates in order to identify both their basic features and their receptors' properties, including the biological effects of their stimulation, and finally their endocrine actions. Since the identification of methionine-enkephalin and leucin-enkephalin, many opioid peptides with higher molecular weight have been characterized, and their origin from specific precursor has been recognized: proopiomelanocortin, preproenkephalin A, preproenkephalin B. In particular we have analyzed the pharmacological properties and the biological effects of beta-endorphin and met -and leu-enkephalins, the most diffuse of opioids. The use of naloxone has permitted the study of endogenous opioid tone and its effects on the release of the pituitary hormones. The present study reports a summary of data in the literature and personal observation indicating that the peripheral sexual steroids are the most important modulators of naloxone effects on endogenous opioid tone. Finally, the paper reports preliminary observations indicating that during naloxone infusion, females with hypothalamic amenorrhoea show a hormonal profile which differs from normal subjects.

Naloxone inhibition of postprandial growth hormone releasing hormone-induced growth hormone release in obesity.

The effects of opiate receptor antagonist naloxone on growth hormone (GH) release after growth hormone-releasing hormone (GHRH) administration were investigated, before or after feeding, at 13.00 h, in 20 obese women and in 10 normal women. When GHRH was administered to obese women before a meal at lunch time, the mean peak plasma GH levels were very low, while plasma GH responses significantly increased after feeding. Naloxone, infused at a rate of 1.6 mg/h starting 1 h before GHRH administration (50 micrograms i.v. as a bolus), was capable of inhibiting GH release induced by administration of GHRH after feeding. On the contrary, naloxone did not induce significant variations on the fasting GHRH-induced GH release. In normal women, naloxone did not significantly modify the GH response to GHRH, both before and after lunch. The inhibitory effect of naloxone indicates that in obese women there is an increased opioid activity, which could represent an abnormal response of the gastrointestinal tract to food ingestion.

Corticotropin-releasing hormone inhibition of gonadotropin secretion during the menstrual cycle.

To determine whether corticotropin-releasing hormone (CRH) exerts an inhibitory action on gonadotropin secretion in normal fertile women, the effects of CRH on luteinizing hormone (LH), follicle-stimulating hormone (FSH), and cortisol secretion were studied during the menstrual cycle. CRH had no effect on LH release during the midfollicular phase of the cycle. By contrast, IV injection of 100 micrograms CRH elicited significant decreases in LH concentrations during late follicular (-50%) and midluteal (-52%) phases of the cycle. LH concentrations decreased during the four-hours following injection of CRH and returned to those observed during the control period five hours after injection. Similarly, CRH elicited a significant decrease in FSH secretion during the midluteal phase of the cycle. CRH injection induced an increase in cortisol release during all phases of the cycle. These data demonstrate that exogenous CRH administration results in inhibition of gonadotropin secretion in late follicular and midluteal phases of the cycle. These results suggest that elevated endogenous CRH levels resulting in increased cortisol secretion could contribute to decreased gonadotropin secretion and, thus, disruption of reproductive function during stressful conditions in women.

[Growth hormone-releasing hormone. The physiopathologic aspects and its diagnostic-therapeutic use]. / Growth hormone releasing hormone. Aspetti fisiopatologici ed impiego diagnostico-terapeutico.

The object of the present study is to review all that in the last years has been discovered about growth hormone-releasing hormone (GHRH), in order to point out both its physiopathological characteristics and its possible diagnostic and therapeutic use. In the first section are summarily reviewed the different studies that culminated in 1982 with the identification of three GRF: GRF(1-37)-OH, GRF(1-40)-OH and GRF(1-44)-NH2, the last of which, by immunohistochemical methods, resulted to be similar to the hypothalamic hGHRH. Then we describe the anatomic distribution of GHRH in man, and its mechanism of action at both receptor and postreceptor levels. On the other hand, the control of the GHRH secretion by peptidergic hypothalamic neurons occurs through four principal monoaminergic systems such as dopaminergic, noradrenergic, adrenergic and serotoninergic ones, and also by cholinergic fibers and by endogenous opiates, all acting to cause the release, into the hypothalamo-hypophyseal portal circulation, of GHRH. In the second section is attracted attention on the GHRH as a diagnostic agent in the two diseases that represent the main alterations of the GH secretion: acromegaly and short stature. According to the different studies considered, it may be concluded that GHRH testing has limited diagnostic usefulness in the clinical evaluation of acromegaly, but allows to discriminate acromegalic patients with ectopic production of GHRH from those with pituitary tumors. For what concerns short stature, the results of observation realized both in adult subjects and in children, all with GH deficiency, by exogenous administration of GHRH, have pointed out that the majority of the GH deficiency patients have hypothalamic disregulation, and not a pure pituitary deficiency as it has been supposed before GHRH discovery. In the third section is attracted attention on the GHRH as a therapeutic agent. Its possible use in the therapy of the children with GH deficiency is of considerable interest, above all in relation to the hypothalamic pathogenesis of their short stature.

Periovulatory plasma prolactin response to synthetic growth hormone-releasing hormone in normal women.

Following the demonstration of a positive prolactin (PRL) response to growth hormone-releasing hormone (GHRH) in acromegalic and anorexic women, we have injected GHRH (50 micrograms intravenously as a bolus) in normal women during various phases of their menstrual cycle in order to establish whether a positive response was present also in normal subjects. Synthetic GHRH 1-44 elicited a significant increase in circulating PRL levels in eight women studied during the periovulatory phase of the menstrual cycle. In contrast, no significant changes in circulating PRL levels after GHRH administration were found in nine women during the midfollicular phase or in five women during the midluteal phase. A temporal correlation between the midcycle gonadotropin peak and the positive response to GHRH has been observed. Synthetic GHRH elicited the expected increase in GH levels during all phases of the cycle studied. Our data demonstrate that GHRH is capable of stimulating a PRL response in normal subjects and raise the possibility that PRL secretion is regulated by several hormones of hypothalamic origin.