Identification of Predictive Markers of Response to Immunomodulators in the Treatment of COVID-19 Patients.

BACKGROUND: Proper identification of patients at risk of developing serious disease in the context of SARS-CoV-2 infection, as well as the initiation of early treatment, is one of the fundamental elements for successful management of COVID-19. The main objective of this study was to evaluate the usefulness of serum biomarkers (neutrophils, lymphocytes, C-reactive protein, lactate dehydrogenase, D-dimer, ferritin, and interleukin-6) to predict the early response to immunosuppressant therapy in COVID-19 patients. METHODS: This is a case-control study nested in a retrospective cohort, which included hospitalized patients with interstitial pneumonia and with elevation of some proinflammatory parameters. Each of the individuals who died during the 28-day follow-up was defined as a case. For each case, 4 controls were selected, matched by age, gender, and comorbidities. RESULTS: The initial cohort included 856 patients. The incidence of therapeutic failure in the cohort was 14%, thus we identified a total of 120 cases. After the application of a Cox regression model, high serum concentrations of LDH (> 451 IU/L), ferritin (> 1,014 ng/mL) and D-Dimer (> 1,300 ng/mL) were identified as predictors of poor response to treatment. Highly-specific cut-off points could not be established for any of these biomarkers. CONCLUSIONS: Some inflammatory biomarkers, such as LDH, ferritin, and D-dimer, may be helpful in identifying patients for whom an early immunomodulatory therapeutic intervention should be considered in the treatment of COVID-19 patients with pneumonia.

Bronchoalveolar Lavage Fluid Cytology in Culture-Documented Invasive Pulmonary Aspergillosis in Patients with Hematologic Diseases: Analysis of 67 Episodes.

There is a paucity of studies on the yield of Gomori-methenamine-silver (GMS) staining in bronchoalveolar lavage (BAL) fluid cytology and its comparison with fluorescent dye staining for the diagnosis of invasive pulmonary aspergillosis (IPA) in patients with hematologic malignancies. To that end, we analyzed the yield of direct fungal visualization in BAL fluid cytology with GMS staining, in a series of culture-positive IPA cases in 67 patients with hematologic malignancies, and we compared the results with those of direct examination with calcofluor white staining and BAL fluid galactomannan assays, when available. GMS staining in BAL fluid cytology was positive in 42% of the 67 cases and revealed coinfections in 7 cases. In contrast, only 2/67 (3.6%) BAL fluid samples were positive in direct smears stained with the fluorescent dye calcofluor white. Positive GMS staining results were significantly more frequent in IPA cases with cavitary lesions and IPA cases caused by >1 Aspergillus species, but the proportions of positive cytology results among Aspergillus species were not different.

Are incidence and epidemiology of anaerobic bacteremia really changing?

Incidence, prognosis and need of performing blood cultures for anaerobic bacteria are under debate, mainly due to the belief that the presence of anaerobes in blood can be easily suspected on clinical basis. We aimed to assess these three points in a retrospective analysis of a 10-year experience in our tertiary hospital. All episodes of significant anaerobic bacteremia diagnosed from 2003 to 2012 were included. Risk factors for mortality and clinical predictability of anaerobic bacteremia were evaluated in 113 randomly selected episodes. Overall incidence of anaerobic bacteremia was 1.2 episodes/1000 admissions, with no significant changes during the 10-year study period. B. fragilis group (38.1 %) and Clostridium spp. (13.7 %) were the most frequent isolated microorganisms. As for the clinical study, 43.4 % of the patients had a comorbidity classified as ultimately fatal or rapidly fatal according to the McCabe and Jackson scale. Clinical manifestations suggestive of anaerobic involvement were present in only 55 % of the patients. Twenty-eight patients (24.8 %) died during the hospitalization. Independent predictive factors of mortality were a high Charlson's comorbidity index and presentation with septic shock, whereas, an adequate source control of the infection was associated with a better outcome. In our centre, incidence of anaerobic bacteremia remained stable during the last decade. The routine use of anaerobic BCs seems to be adequate, since in about half of the cases anaerobes could not be suspected on clinical bases. Moreover, prompt source control of infection is essential in order to reduce mortality of patients with anaerobic bacteremia.

The search for endocarditis in patients with candidemia: a systematic recommendation for echocardiography? A prospective cohort.

Most current guidelines do not recommend systematic screening with echocardiography in patients with candidemia, as Candida infective endocarditis (CIE) is considered an uncommon disease. During the study period, we recommended echocardiography systematically to all candidemic patients that did not have contraindications and accepted to participate in the study. We intended to assess the incidence of unrecognized CIE in adult patients with candidemia. Our institution is a tertiary teaching hospital in which we follow all patients with candidemia. From January 2007 to October 2012, echocardiography was systematically recommended to suitable candidates. We recorded 263 cases of candidemia in adult patients. Echocardiography was not performed in 76 of these patients for the following reasons: patients had died when blood cultures became positive (17), patients were critically or terminally ill (38), or the patient or physician refused the procedure (21). The remaining 187 patients constitute the basis of this report. CIE was diagnosed in 11 cases (4.2 % of the whole candidemic population and 5.9 % of the population with echocardiographic study). The results of transthoracic echocardiography (TTE) suggested infective endocarditis (IE) in 5/172 patients (2.9 %), and the result of transesophageal echocardiography (TEE) was positive in 10/87 (11.5 %). Among 11 confirmed cases of CIE, the disease was clinically unsuspected in three patients. At least 4.2 % of all candidemic patients have CIE. CIE is frequently clinically unsuspected and echocardiography is required to demonstrate a high proportion of cases.

Micafungin at physiological serum concentrations shows antifungal activity against Candida albicans and Candida parapsilosis biofilms.

We assessed the in vitro activity of micafungin against preformed Candida biofilms by measuring the concentration of drug causing the most fungal damage and inhibition of regrowth. We studied 37 biofilm-producing Candida spp. strains from blood cultures. We showed that micafungin was active against planktonic and sessile forms of Candida albicans strains and moderately active against Candida parapsilosis sessile cells. Concentrations of micafungin above 2 µg/ml were sufficiently high to inactivate regrowth of Candida sessile cells.

The CD4:CD8 ratio is associated with markers of age-associated disease in virally suppressed HIV-infected patients with immunological recovery.

OBJECTIVES: Inversion of the CD4:CD8 ratio (< 1) has been identified as a hallmark of inmmunosenescence and an independent predictor of mortality in the general population. We aimed to assess the association between the CD4:CD8 ratio and markers of age-associated disease in treated HIV-infected patients with good immunovirological response. METHODS: A cross-sectional analysis was conducted in 132 HIV-infected adults on antiretroviral therapy (ART), with plasma HIV RNA < 50 HIV-1 RNA copies/mL for at least 1 year, CD4 count > 350 cells/µL and age < 65 years. We analysed the associations between the CD4:CD8 ratio and subclinical atherosclerosis [assessed using carotid intima-media thickness (IMT)], arterial stiffness [assessed using the augmentation index (AIx)], the estimated glomerular filtration rate (eGFR), muscle wasting and sarcopenia [assessed using appendicular lean mass/height(2) (ALM) measured by dual-energy X-ray absorptiometry (DEXA)]. RESULTS: CD4:CD8 ratio inversion was associated with higher IMT, lower eGFR and lower ALM (all values P < 0.05), but not with AIx. In multivariate analyses adjusted for age, sex, hypertriglyceridaemia, tobacco use and cumulative ART exposure, inversion of the CD4:CD8 ratio was independently associated with higher IMT [odds ratio (OR) 2.9; 95% confidence interval (CI) 1.2-7.1], arterial stiffness (OR 4.8; 95% CI 1.0-23.5) and lower eGFR (OR 5.2; 95% CI 1.0-64.4), but not sarcopenia (OR 0.7; 95% CI 0.2-2.7). These associations persisted when models were applied to subjects with nadir CD4 counts > 200 cells/µL and those with CD4 counts > 500 cells/µL. CONCLUSIONS: The CD4:CD8 ratio in treated HIV-infected subjects with good immunovirological response is independently associated with markers of age-associated disease. Hence, it might be a clinically useful predictor of non-AIDS-defining conditions.

Is catheter-related candidemia a polyclonal infection?

Diagnosis of catheter-related candidemia (CRC) requires the simultaneous isolation of Candida spp. from both blood and catheter samples. We previously observed that in most CRC cases, the genotype of the yeast found in catheter samples is also recovered from blood. However, it is not clear whether CRC is a polyclonal infection. We prospectively studied 20 patients with CRC caused by Candida albicans, C. parapsilosis, or C. glabrata to analyze whether their infections were polyclonal. As many as 10 colonies per sample (n = 475) isolated from blood (n = 220) and catheter (n = 255) specimens were studied using species-specific microsatellite markers. Genotyping always revealed matches between the Candida spp. from blood and catheter samples. However, 15% of patients had a polyclonal pattern of infection or catheter colonization that was species specific. An additional genotype was found exclusively in the catheters of two patients infected with C. albicans, whereas an additional genotype was noted in the blood culture of a patient infected with C. parapsilosis. Considering only the presence of different genotypes in blood samples, 5% of patients had polyclonal infections. We conclude that most cases of CRC are caused by a single genotype.

Clinical experience in the treatment of COVID-19 with monoclonal antibodies in solid organ transplant recipients.

Solid organ transplant (SOT) recipients are at high risk for complications from coronavirus disease 2019 (COVID-19). SOT recipients mount lower immunological responses to vaccines than general population and are at high risk for breakthrough COVID-19 infections. Passive immunotherapy in the form of anti-Spike monoclonal antibodies (MoAbs) may be an alternative for the prophylaxis and treatment of COVID-19 in these patients. SARS-CoV-2 has evolved by accumulating resistance mutations that have escaped the neutralizing action of most MoAbs. However, MoAbs directed at more conserved epitopes and that maintain effector functions could maintain efficacy in the treatment of these patients. According to published data, SOT recipients with low anti-spike antibody responses to vaccination could benefit from the use of MoAbs in pre-exposure prophylaxis, in the treatment of COVID-19 mild to moderate and severe COVID-19 with less than 15 days of symptom duration and low oxygen requirements. Combination therapy could be more effective than monotherapy for the treatment of mild-to-moderate SARS-CoV-2 infection.

Smoking cessation in HIV patients: rate of success and associated factors.

BACKGROUND: Smoking is the modifiable cardiovascular (CV) risk factor that contributes most to causing premature CV disease. Prevalence of smoking in patients with HIV infection is double that of the general population. OBJECTIVES: To determine the rate of patients succeeding in quitting smoking after 12 months, factors associated with this success, and the characteristics of tobacco consumption and nicotine dependence. METHODS: Longitudinal descriptive study. Three hundred and sixty-eight HIV-infected patients were interviewed. Smokers in Prochaska's stage of action began a programme to quit smoking. We registered the variables related to tobacco consumption and the level of success of cessation. RESULTS: 63.9% of the patients were active smokers and 14% of them began the cessation programme. Average motivation for cessation was 7.8 +/- 1.4 (Richmond) and nicotine dependence rate 5.5 +/- 3.0 (Fagerström). After 1 year, 25% had quit smoking. Those patients who stopped smoking presented a higher motivation level (8.8 +/- 1.3 vs. 7.5 +/- 1.5, P=0.048). Cessation significantly reduced their CV risk at 12 months [2.5 [interquartile range (IQR) 2.0-5.2] vs. 1.7 [IQR 1.0-3.5], P=0.026]. CONCLUSIONS: The prevalence of smokers in our population of HIV-infected patients was 63.9%. Only 14% began a smoking cessation programme. Twelve months after a programme to quit smoking, cessation rate was 25%; this was influenced mostly by the level of motivation of the patient.

Fosfomycin in antimicrobial stewardship programs.

Due to the increase in antimicrobial resistance, strategies such as antimicrobial stewardship programs (ASP) have been developed to improve the clinical results, decrease the adverse effects and the development of resistances and ensure cost-effective therapies. Fosfomycin has a unique mechanism of action against Gram-positive and Gram-negative bacteria. Cross-resistance is uncommon; however, fosfomycin should be used in combination in severe infections to avoid selecting resistant mutations. Fosfomycin's oral formulation facilitates sequential treatment, has low toxicity and high tissue penetration, even in the central nervous system and bone. Fosfomycin is active against resistant Gram-positive bacteria such as methicillin-resistant Staphylococcus aureus (MRSA), vancomycin- resistant enterococci and penicillin-resistant Streptococcus pneumoniae, as well as against resistant Gram-negative bacteria such as extended-spectrum beta-lactamase-producing and carbapenemase-producing enterobacteria. Fosfomycin is therefore useful for cases of persistent bacteremia, skin and soft tissue infections, as a glycopeptide-sparing and carbapenem-sparing drug for healthcare-associated infections and for polymicrobial infections. Published studies have demonstrated the synergy between fosfomycin and beta-lactams, daptomycin and glycopeptides against MSSA and MRSA; with linezolid in biofilm-associated infections and with aminoglycosides and colistin against Gram-negative bacteria, providing a nephroprotective effect.

Biofilm production is not associated with poor clinical outcome in 485 patients with Staphylococcus aureus bacteraemia.

OBJECTIVES: Staphylococcus aureus biofilm may constitute a major cause of virulence. Our main objective was to analyse whether there was an association between biofilm production and poor outcome in patients with S. aureus bacteraemia. METHODS: We studied 485 S. aureus strains isolated from the blood of patients with bacteraemia from 2012 to 2015. We assessed in vitro biomass production using crystal violet assay and metabolic activity using tetrazolium salt assay. Strains were classified in tertile ranks as follows: low biomass producers, moderate biomass producers, high biomass producers, low metabolic activity, moderate metabolic activity and high metabolic activity. We excluded from analysis strains with moderate crystal violet and tetrazolium salt values. We defined poor outcome as fulfillment of one or more of the following conditions: 30-day attributable mortality, infective endocarditis, persistent bacteraemia and recurrent bacteraemia. RESULTS: Outcome was poor in 199 (41.0%) of 485 S. aureus bacteraemia episodes. The distribution of poor outcome with respect to biomass production and metabolic activity was as follows: low biomass producers, 36.6% vs. high biomass producers, 43.2% (p 0.26); and low metabolic activity, 43.5% vs. high metabolic activity, 36.2% (p 0.91). The presence of methicillin-resistant S. aureus was the only characteristic that was more likely to be present in the high metabolic activity group (17.4% vs. 39.3%, p < 0.001). CONCLUSIONS: Biofilm production, as determined by any of the methods used in the present study, is not associated with poor outcome in patients with S. aureus bacteraemia.

Age and the renin-aldosterone system.

Age effect on plasma renin activity (PRA) and PRA classification was studied in young and older normotensive volunteers. Ambulatory PRA was lower in the older age group than in the younger with both on an unrestricted diet and a low-sodium diet. Renal function, aldosterone excretion, and plasma renin substrate were comparable in both groups. Age had a substantial effect on PRA classification. When the young normotensives were controls, 32% (6/19) older normotensives had abnormally low PRA, or "low renin normotension." Similarly, 18% (2/11) of young patients with essential hypertension but 80% (12/15) of older hypertensives had low PRA. When the older volunteers were controls, however, the incidence of low renin hypertension (LRH) decreased to 53% in the older patients. The use of predominantly young controls for defining normal limits of PRA may result in an overestimate of the incidence of LRH and may contribute to the heterogeneity of LRH.

Plasma C3d levels and ischemic heart disease in type II diabetes.

OBJECTIVE--To test the hypothesis that the complement system may be activated in patients with type II diabetes and CAD. RESEARCH DESIGN AND METHODS--The plasma C3d concentration was measured in 106 type II diabetic patients and 25 nondiabetic control subjects. The patient group was subdivided according to AER, and the groups were adjusted for age, sex, and known duration of diabetes. For the assignment to a given subgroup, normoalbuminuria was defined as AER < 15 microns/min, microalbuminuria as AER 16-250 micrograms/min, and macroalbuminuria as AER > 250 micrograms/min. The presence or absence of coronary disease was assessed through clinical examination, ECG, and coronary angiography. An RIA system was used for measurement of urinary albumin levels, and the plasma C3d concentrations were measured by ELISA. RESULTS--Within each of the AER-defined subgroups, the plasma C3d levels were significantly higher in patients with IHD than in those without. Thus, in the normoalbuminuric group, plasma C3d levels were 16.3 AU/ml (95% CI 13.9-19) in patients with IHD vs. 11.6 AU/ml (95% CI 10.5-12.7) in those without (P < 0.001). The corresponding data for the microalbuminuric and macroalbuminuric groups were 21.8 (95% CI 18.1-26.3) vs. 13.6 (95% CI 12.3-15.1) and 31.6 (95% CI 24.9-40) vs. 17.5 (13.6-22.6) AU/ml (P < 0.01), respectively. Patients with IHD also had significantly higher plasma C3d levels than normal control subjects, regardless of AER subgroup. A multiple logistic regression analysis demonstrated an association between the plasma C3d concentration and IHD and AER. CONCLUSIONS--Activation of the complement system may play a role in the development of macrovascular disease in type II diabetes.

Antifungal stewardship in a tertiary-care institution: a bedside intervention.

Antifungal stewardship (AFS) programmes are needed in tertiary-care hospitals. Our aim is to describe a bedside non-restrictive AFS programme, and to evaluate its economic impact. During the first year of the AFS a bundle of non-interventional measures were implemented. During the second year an infectious diseases specialist visited 453 patients receiving candins, liposomal amphotericin B, voriconazole or posaconazole. Monthly costs were studied with an interrupted time series (ITS) analysis. The main prescribing departments were haematology (35%), medical departments (23%), and intensive care units (20%). Reasons to start antifungal therapy were: targeted therapy (36%), prophylaxis (32%), empirical therapy (20%) and pre-emptive therapy (12%). At the initial visit, diagnostic advice was provided in 40% of cases. The most common therapeutic recommendations were to de-escalate the antifungal drug (17%) or to suspend it (7%). Annual total antifungal expenditure was reduced from US$3.8 million to US$2.9 million over the first 2 years, generating net savings of US$407,663 and US$824,458 per year after considering the cost of additional staff required. The ITS analyses showed a significant economic impact after the first 12 months of the intervention (p 0.042 at month 13), which was enhanced in the following 24 months (p 0.006 at month 35). The number of defined daily doses decreased from 66.4 to 54.8 per 1000 patient-days. Incidence of candidaemia was reduced from 1.49 to 1.14 (p 0.08) and related mortality was reduced from 28% to 16% (p 0.1). A collaborative and non-compulsory AFS program based on bedside intervention is an efficacious and cost-effective approach that optimizes the use of AF drugs.

Atrial natriuretic peptide in rats with experimental cirrhosis of the liver without ascites.

Plasma levels of atrial natriuretic peptide (ANP) have been measured in eight sodium-retaining cirrhotic nonascitic rats and eight control animals before and after extracellular volume expansion by isotonic saline infusion (30 ml/kg BW, 20 min). In addition, disappearance of [125I]ANP was studied in six control and six cirrhotic rats. The effect of infusing synthetic rat ANP-(1-28) (1 microgram) on mean arterial pressure and renal function has been also studied. In basal conditions, cirrhotic rats showed higher ANP plasma levels than control animals (71.1 +/- 16.6 vs. 43.9 +/- 5.1 pg/ml; P less than 0.05). After extracellular volume expansion, ANP increased in both control and cirrhotic rats; the increase in cirrhotic was higher than that in control rats (88 +/- 27% vs. 33 +/- 8%; P less than 0.05). Disappearance of iodinated ANP from plasma was identical in control and cirrhotic rats. ANP infusion induced a larger decrease in mean arterial pressure in control (21 +/- 5%) than in cirrhotic rats (9 +/- 2.5%). ANP induced comparable increases in glomerular filtration rate and renal plasma flow in both groups of animals. However, diuretic and natriuretic effects were markedly impaired in cirrhotic animals. Thus, urinary flow increased by 91 +/- 18 microliters/min in control animals, but only by 37 +/- 7 microliters/min in cirrhotic animals. Fractional sodium excretion increased to 1.7 +/- 0.44% in controls and to 0.54 +/- 0.12% in cirrhotic rats (P less than 0.05). It is concluded that the defect in renal handling of sodium in cirrhotic rats is not due to a lack of ANP synthesis or release. In addition, these animals show an impaired renal response to ANP.

Severe hypertension in the Spanish population. Association with specific HLA antigens.

There is now increasing evidence for immunological changes in essential hypertension. Immunological response is determined in part by genes linked to the HLA system. It has been reported a positive association between HLA B15 and the risk for cerebral events in essential hypertensive (EH) patients. We studied the distribution of HLA antigens in 128 EH (age range, 13-85 years) and 1000 normotensive controls. EH were classified in accordance with the World Health Organization (WHO) criteria: in WHO Stages I and II, there were 100 patients; in WHO Stage III, there were 28 patients. HLA-A and B antigens of peripheral blood lymphocytes were studied according to the microlymphocytotoxicity test. The results were compared by chi-square analysis, and the p value was multiplied by the number of antigens studied at each locus, to avoid overestimation of an association. Frequency of HLA-BW 22 was higher in EH compared with controls (5.4% vs 1.2%, p less than 0.01). Frequency of HLA-B12 in EH with WHO Stage III hypertension (64.2%) was significantly increased compared either with EH in WHO Stage I or II (29%, p less than 0.01) or the control group (26.9% p less than 0.001). The incidence of HLA-B15 antigen in the whole hypertensive group was 3.1%, lower than in normotensive controls (6.4%, p less than 0.8). None of the patients with WHO Stage III hypertension had the HLA-B15 antigen. In conclusion, the results seemed to indicated that the Spanish population had an association between HLA-B12 and severe hypertension.

Low plasma renin activity in normotensive patients with diabetes mellitus: relationship to neuropathy.

To determine the effect of diabetes mellitus on the renin-aldosterone system, independent of age, nephropathy, or hypertension, 16 normotensive diabetics with long-term disease (mean duration, 15 years) and no (14) or minimal (2) proteinuria, were compared to nine age-matched, normotensive controls. Plasma renin activity (PRA) measured supine and after 4 hours of quiet ambulation, both on an ad libitum diet and on Day 4 of a 10 mEq low sodium diet, was always lower in the diabetics (31%-56% of control values). After the combined stimulus of sodium depletion and ambulation, PRA was 2.2 +/- 0.4 in the diabetics compared to 3.4 +/- 0.2 ng/ml/hr in controls (p less than 0.025). On the low sodium diet, PRA and the postural response of PRA correlated directly with the degree of autonomic dysfunction as quantitated by the velocity of esophageal peristalsis (r = 0.60, p less than 0.05; r = 0.75, p less than 0.005 respectively), suggesting that autonomic neuropathy was an important factor contributing to low PRA in these patients. No other parameters correlated with PRA. Plasma renin substrate (PRS) tended to be lower in diabetics (1053 +/- 95 vs 1358 +/- 132 ng AI/ml; p less than 0.07) but not sufficiently so to account for the substantial difference in PRA. Furthermore, PRS did not correlate with PRA. Fasting blood sugar, while higher in diabetics (209 vs 96 mg/dl), and creatinine clearance, which was lower (112 +/- 13 vs 78 +/- 4 ml/min; p less than 0.01), also did not correlate with PRA. Other factors, including serum creatinine, serum potassium, urinary aldosterone, blood pressure, and body weight, and the responses of these parameters to sodium depletion, were similar in diabetics and controls. These data implicate visceral neuropathy as a major factor in the hyporeninemia of these diabetics.

Cholinergic mediation of growth hormone secretion elicited by arginine, clonidine, and physical exercise in man.

The role of acetylcholine in human GH secretion was studied with atropine, which selectively blocks cholinergic muscarinic receptors and crosses the blood-brain barrier. Paired tests were performed in 22 normal subjects divided into 4 groups. The stimuli employed were arginine (30 g/30 min, iv), clonidine (300 micrograms, orally), physical exercise for 20 min, and saline. In the second test, atropine (1 mg, im) was administered before GH stimulation. Arginine elicited a GH secretory peak of 16.6 +/- 5 ng/ml (mean +/- SEM), which was completely blocked when atropine was administered with arginine (0.9 +/- 0.1 ng/ml). Atropine did not, however, modify the PRL secretory response; peak levels after arginine and atropine plus arginine were 16.3 +/- 3.1 and 16.8 +/- 2.5 ng/ml, respectively. Clonidine elicited a GH secretory peak (11.8 +/- 2.7 ng/ml) which also was blocked by pretreatment with atropine (1.2 +/- 0.2 ng/ml). Neither clonidine nor clonidine plus atropine altered PRL secretion. GH levels also were sharply increased after physical exercise, with a peak level of 19.4 +/- 4.9 ng/ml. Atropine completely blocked exercise-induced GH secretion (2 +/- 0.9 ng/ml). Atropine alone did not modify GH or PRL values compared with saline administration. The potency of the atropine-induced suppression of GH secretion by three different stimuli, each with presumably different mechanisms of action, suggests that acetylcholine plays an important role in the regulation of GH secretion.

Free fatty acids block growth hormone (GH) releasing hormone-stimulated GH secretion in man directly at the pituitary.

Increases in plasma FFA levels inhibit GH responses to a variety of pharmacological and physiological stimuli. To gain further insight into the mechanism by which FFA exert their effect, we studied the plasma GH responses to GHRH-(1-44) (1 microgram/kg, iv) in normal subjects in whom plasma FFA levels were raised by a lipid-heparin infusion (250 mL 10% Intralipid plus 2500 U heparin). Paired tests were performed in 10 normal subjects, with and without lipid-heparin pretreatment. Lipid-heparin infusion from -30 to 120 min increased mean FFA levels from 0.41 +/- 0.03 (+/- SEM) to 3.12 +/- 0.40 mmol/L at 120 min. The mean plasma GH levels after GHRH administration were lower at all times; however, the values were significantly different (P less than 0.05) only at the later times (45, 60, and 90 min). When considered individually, an all or none pattern was observed; 5 subjects had no plasma GH response to GHRH, and 5 had no reduction. To investigate the time relationships between the FFA peak and subsequent GH blockade, a different protocol of paired tests was performed with GHRH with or without a different lipid-heparin infusion protocol. Lipid-heparin was infused from -90 to 0 min, with an additional heparin pulse at -15 min, to obtain a higher and earlier (0 min) FFA increase. FFA increased from 1.06 +/- 0.19 to 11.61 +/- 0.83 mmol/L at zero time. The GHRH-induced GH secretory peak (15.8 +/- 3.5 ng/ml) at 15 min was completely blocked (0.9 +/- 0.2 ng/ml), and the mean plasma GH levels were also lower at 30, 45, and 60 min. To determine whether the FFA-induced blockade of GH secretion was exerted in the pituitary, a series of in vitro studies was conducted using monolayer cultures of rat anterior pituitary glands, with GHRH concentrations of both 10(-10) and 10(-8) M and 10(-5) M forskolin to stimulate GH release. Both caprylic and oleic acid inhibited basal GH release and GHRH- or forskolin-induced GH release. PRL release was not altered, nor were toxic actions noted on the cells. In conclusion, FFA are able to block GH secretion directly at the pituitary level.

Atropine blockade of growth hormone (GH)-releasing hormone-induced GH secretion in man is not exerted at pituitary level.

The role of acetylcholine (Ach) in the regulation of human GH secretion was assessed using atropine, which selectively blocks cholinergic muscarinic receptors. Paired tests were performed in seven normal subjects using GH-releasing hormone (GHRH) 1-44 (1 microgram/kg iv), with and without atropine pretreatment (1 mg im). The GHRH 1-44-induced GH secretory peak [20.7 +/- 4.5 (SEM) ng/ml] was completely blocked by atropine administration (2.3 +/- 0.6 ng/ml) (P less than 0.01). To determine whether this atropine blockade was at the pituitary level, a series of in vitro studies were conducted using monolayer cultures of cells from bovine anterior pituitary glands. GHRH 1-44 (10(-8) M) stimulated bovine GH release (11.1 +/- 1.5 micrograms/ml) as compared to control values (5.1 +/- 0.4 microgram/ml) (P less than 0.01). This response was not altered by 10(-6) M atropine (14.9 +/- 0.9 microgram/ml). Similar results were obtained with GHRH, 10(-9) M, with or without atropine, 10(-7) M. Addition of 10(-6) M Ach to the incubation medium significantly increased bovine GH release (12.7 +/- 1.2 microgram/ml) and the effect of 10(-6) M Ach and 10(-8) M GHRH was additive (20.9 +/- 2.1 micrograms/ml) (P less than 0.01). Similar results were obtained with Ach, 10(-5) M, and GHRH, 10(-9) M. Atropine or eserine alone did not alter basal GH secretion, and atropine blocked Ach-stimulating activity. In conclusion, atropine blockade of GHRH-induced GH secretion appears to be exerted at a site other than pituitary.