Spleen and lung virome analysis of South American fur seals (Arctocephalus australis) collected on the southern Brazilian coast.

South American fur seals (Arctocephalus australis) are believed to reach the coast of Rio Grande do Sul (RS) through sea currents. They live in colonies and are frequently found resting on the beach. However, it is also common to find dead pinnipeds on beaches, sharing the environment with humans, domestic animals and other wild species on the coast and facilitating the transmission of pathogens. In the present study, a metagenomic approach was applied to evaluate the viral diversity in organs of fur seals found deceased along the coast of the state of RS, southern Brazil. The lungs and spleens of 29 animals were collected, macerated individually, pooled separately (one pool for lungs and another for spleens) and sequenced using the Illumina MiSeq platform. Sequences more closely related to members of the Anelloviridae and Circoviridae families were detected. Nine putative new species of anellovirus and one putative new genus, named Nitorquevirus, were described. Additionally, the circovirus sequences found in the lungs of A. australis have a common ancestor with PCV3, a proposed swine pathogen. Our study expanded the knowledge about viral communities in pinnipeds and could be useful for monitoring new viruses and potential viral sharing among wildlife, domestic animals, and humans.

Virome of crab-eating (Cerdocyon thous) and pampas foxes (Lycalopex gymnocercus) from southern Brazil and Uruguay.

Crab-eating (Cerdocyon thous) and Pampas foxes (Lycalopex gymnocercus) are wild canids distributed in South America. Domestic dogs (Canis lupus familiaris) and wild canids may share viral pathogens, including rabies virus (RABV), canine distemper virus (CDV), and canine parvovirus 2 (CPV-2). To characterize the virome of these wild canid species, the present work evaluated the spleen and mesenteric lymph node virome of 17 crab-eating and five Pampas foxes using high-throughput sequencing (HTS). Organ samples were pooled and sequenced using an Illumina MiSeq platform. Additional PCR analyses were performed to identify the frequencies and host origin for each virus detected by HTS. Sequences more closely related to the Paramyxoviridae, Parvoviridae and Anelloviridae families were detected, as well as circular Rep-encoding single-stranded (CRESS) DNA viruses. CDV was found only in crab-eating foxes, whereas CPV-2 was found in both canid species; both viruses were closely related to sequences reported in domestic dogs from southern Brazil. Moreover, the present work reported the detection of canine bocavirus (CBoV) strains that were genetically divergent from CBoV-1 and 2 lineages. Finally, we also characterized CRESS DNA viruses and anelloviruses with marked diversity. The results of this study contribute to the body of knowledge regarding wild canid viruses that can potentially be shared with domestic canids or other species.

Liver virome of healthy pigs reveals diverse small ssDNA viral genomes.

Brazil is a major exporter of pork meat worldwide. Swine liver is a common ingredient in food consumed by humans, thus emphasizing the importance of evaluating the presence of associated pathogens in swine liver. To obtain knowledge, this study aimed to provide insights into the viral communities of livers collected from slaughtered pigs from southern Brazil. The 46 livers were processed and submitted for high-throughput sequencing (HTS). The sequences were most closely related to Anelloviridae, Circoviridae and Parvoviridae families. The present work also describes the first Brazilian PCV1 and the first PPV6 and PPV7 from South America. Virus frequencies revelead 63% of samples positive for TTSuV1, 71% for TTSuVk2, 10.8% for PCV, 13% for PPV and 6% for PBov. This report addresses the diversity of the liver virome of healthy pigs and expands the number of viruses detected, further characterizing their genomes to assist future studies.

Highly divergent cattle hepacivirus N in Southern Brazil.

The genus Hepacivirus includes 14 species (Hepacivirus A-N). In this study, we determined a partial genome sequence of a highly divergent bovine hepacivirus (hepacivirus N, HNV) isolate from cattle in Southern Brazil. Previously described HNV isolates have shared 80-99.7% nucleotide sequence identity in the NS3 coding region. However, the sequence determined in this study had 72.6% to 73.8% nucleotide sequence identity to known HNV NS3 sequences. This high divergence could be seen in a phylogenetic tree, suggesting that it represents a new genotype of HNV. These data expand our knowledge concerning the genetic variability and evolution of hepaciviruses.

Samarium-153 for intravascular irradiation therapy with liquid-filled balloons to prevent restenosis: acute and long-term results in a hypercholesterolemic rabbit restenosis model.

BACKGROUND: It has been shown that irradiation with either beta and gamma sources inhibit neointimal formation. Samarium-153 ((153)Sm) is an isotope with 0.8 MeV, subdivided in three different beta energies and 103 keV of gamma energy. This compound has been tested and used in humans for palliation of pain from bone metastases. The aim of the present study was to evaluate the feasibility and efficacy of brachytherapy with (153)Sm-filled balloon to inhibit neointimal formation in rabbits after balloon overstretch injury. METHODS: Nineteen rabbits underwent balloon injury in their iliac arteries. In 12 animals (control), oversized balloons filled with saline solution were inflated up to 5 atm for a period of 5 min. In 7 rabbits, the same procedure was performed but using balloons filled with (153)Sm. In all cases, both iliac arteries were treated. The prescribed radiation dose was 15 Gy at 1 mm depth. After 30 days, the animals were sacrificed and their arterial segments were analyzed. Radiation exposure at the animal chest to the table and at a distance of 1 m from the table was measured. RESULTS: Histopathologic analysis showed a striking reduction in the amount of neointima in the irradiated arteries compared with control vessels (0.36+/-0.21 vs. 1.07+/-0.56 mm(2), P<.01). The dose delivered to the animal chest was 21.5 mR/h, whereas only 1.9 mR/h was measured at the table and virtually no radiation could be detected at a distance of 1 m from the table. CONCLUSIONS: Brachytherapy with (153)Sm was feasible with minimal personnel exposure radiation and effectively inhibited neointimal formation in this experimental model. These results warrant further experimental and clinical investigations.

Atrial natriuretic peptide in normotensive and hypertensive heart transplanted patients.

AIM OF THE STUDY: The present study is aimed to investigate the 24-hour pattern of Atrial Natriuretic Peptide (ANP) plasma concentration in normotensive (N) and hypertensive (H) heart transplanted patients (HTP) in order to detect the pathophysiological role of blood pressure regimen for ANP increase in HTP. METHODS: Eight NHTP and five HHTP have been investigated, all being hemodynamically compensated, without histological evidence of rejection and treated by Cyclosporine, Azathioprine and Prednisone. The control group was constituted by 10 clinically healthy subjects (CHS). ANP was assayed six times over the 24 h span. The contrasts between groups were statistically analyzed by means of Student's t-test for the 24 h mean values. RESULTS: The t-test found a statistically significant difference between the ANP 24 h mean levels either of CHS and HTP or NHTP and HHTP. The ANP 24 h mean plasma levels are increased of 190.4% in HHTP and of 44.3% in NHTP in comparison with CHS. DISCUSSION: The findings suggest that the further elevation of ANP in HHTP is a compensatory mechanism to antagonize high blood pressure. Moreover, being the ANP percent increase in HHTP three times more.

Twenty-four-hour pattern of atrial natriuretic peptide in heart transplantation: evidence for lack of circadian rhythm. Temporal inter-relationships with plasma renin activity, aldosterone and cortisol.

We have investigated the circadian rhythm of plasma atrial natriuretic peptide in 13 stable output heart transplanted patients, all without evidence of histological rejection and cardiac impairment, following antirejection therapy with Cyclosporine, Azathioprine and Prednisone. The 24-h pattern of plasma renin activity, plasma aldosterone and plasma cortisol has been studied as well. All the investigated variables were assayed six times over the 24-h span. The circadian time-qualified data were analyzed by ANOVA and Cosinor method. The 24-h mean levels of atrial natriuretic peptide, plasma renin activity and plasma aldosterone are significantly increased, while the concentrations of plasma cortisol are reduced in the heart transplanted recipients. ANOVA detected a significant within-day variability of all these humoral variables only in healthy subjects. A statistically significant circadian rhythm was validated by Cosinor procedure for all the investigated molecules in healthy subjects but not in heart transplanted patients. In our opinion, the increase of atrial natriuretic peptide is a counterregulatory mechanism aimed to compensate the cyclosporine-mediated activation of the renin-angiotensin-aldosterone system. The disappearance of the plasma renin activity, aldosterone and atrial natriuretic peptide circadian rhythm can be ascribed to the constant activation of the renin-angiotensin-aldosterone system. The hypocortisolism is due, in our opinion, both to glucocorticoid therapy and increase of plasma ANP concentration.

Lack of circadian rhythm of plasma concentrations of vasoactive intestinal peptide in patients with orthotopic heart transplants.

OBJECTIVE: To study the circadian pattern of plasma concentrations of vasoactive intestinal peptide (VIP) in patients with orthotopic heart transplants. Circulating VIP is known to have neural and immunological sources. PATIENTS AND METHODS: 13 patients with orthotopic heart transplants were studied 12-53 months (mean 31.8 months) after operation. All were haemodynamically compensated and had no histological evidence of rejection. They were being treated with cyclosporin, azathioprine, and prednisone. Ten healthy individuals were studied as controls. Circulating VIP was assayed six times within a 24h period. Time qualified data were analysed by ANOVA and the cosinor method. Student's t test for unpaired data and Bingham's test for cosinor-derived parameters were used for statistical comparisons. RESULTS: Plasma concentrations of VIP were lower in the patients with orthotopic heart transplants than in the controls (p < 0.001). ANOVA and the cosinor method respectively showed a statistically significant within-day variability and circadian rhythm in the controls but not in the patients with heart transplants. DISCUSSION: The low plasma concentrations of VIP in the patients with heart transplants could be the result of the lack of contribution by the cardiac VIPergic fibres, a reduction of VIP release by the pharmacologically suppressed immune system, the inhibitory effects of cyclosporin on neural function and humoral secretions, and the effects of negative feedback on VIP release of high concentrations of atrial natriuretic peptide. The lack of the circadian rhythm suggests a structural disorder, which should be further investigated.

Endothelial changes following repeated effect of vasoconstrictive substances in vitro.

Using perfusion method the reactivity of rabbit femoral and ear arteries was investigated following the administration of rising noradrenaline doses in four consecutive intervals. Progressive increase of vasoconstrictive activity of vessel segments was found. Results of histological examination showed a progressive de-endothelialization of perfused vessels. The loss of endothelium increased depending on time of action and on intensity of vasoconstrictive stimuli.

Effect of aging on circadian rhythm of atrial natriuretic peptide, plasma renin activity, and plasma aldosterone.

Atrial natriuretic peptide (ANP) shows a nychtohemeral fluctuation and an age-related trend. The aim of this study was to explore the circadian rhythm of ANP as a function of age. Circadian rhythms of plasma renin activity (PRA), aldosterone (PA), and cortisol (PC) were explored as well. Twenty clinically healthy subjects, 10 young (20-25 yrs) and 10 elderly (65-75 yrs), were investigated, while recumbent, after synchronization to light-dark regimen and meal timing. Blood samples for RIA tests were collected six times during the 24-hr span. The chronobiologic analysis in young subjects demonstrated a significant circadian rhythm for all the investigated variables with an acrophase-timing located at 16.48 for ANP, 4.44 for PRA, 5.32 for PA, and 7.12 for PC. In elderly subjects we documented an important increase of 24-hr mean plasma levels but not a statistically significant circadian rhythm for ANP, and a decrease in mean value of PRA which maintained, however, a significant periodic 24-hr oscillation in parallel with PA and PC. The results in young subjects reinforce the concept that ANP plays physiologically an inhibitory role on the phasic secretion of renin. The lack of the circadian rhythm for ANP along with the divergent changes in ANP and PRA 24-hr mean concentration of elderly subjects both suggest that ANP exerts with advancing age only a counterregulatory role on the tonic rather than the phasic release of renin.

[The circadian rhythm of plasma vasoactive intestinal peptide, atrial natriuretic peptide, renin activity, aldosterone and cortisol in heart transplantation. The effects of immunosuppressive therapy]. / Ritmo circadiano di peptide vasoattivo intestinale, peptide natriuretico atriale, attività reninica, aldosterone e cortisolo plasmatici nel trapianto di cuore. Effetti della terapia immunosopressiva.

Vasoactive intestinal peptide (VIP) is released both by neural endings and lymphocytes. Aim of our investigation was to study the effects of immunosuppressive therapy on VIP plasma concentrations. The research has been performed on 10 heart transplanted patients assuming cyclosporine (CYCL) and prednisone (PRED). The circulating T lymphocyte subsets, atrial natriuretic peptide (ANP), plasma renin activity (PRA), plasma aldosterone (PA) and plasma cortisol (PC) have been also assayed. Blood pressure (BP) and heart rate (HR) have been monitored over a 24-hour period to detect whether circulating VIP in heart transplanted patients is influenced by pharmacologically-induced interactions. Seriate samplings along the 24-hour span have been performed. Mean values of ANP, PRA and PA were increased, while VIP, PC and T lymphocyte subsets were decreased in heart transplanted patients as compared to clinically healthy subjects. ANOVA and Cosinor analysis showed, respectively, a statistically significant 24-hour variability and circadian rhythm for all the investigated variables only in normal subjects. BP and HR circadian rhythm in heart transplanted patients suggest that the adrenergic activity regulating the cardiovascular system is restored. This finding argues that the reduction in VIP plasma concentrations is likely due to the decreased lymphocyte production secondary to immunosuppressive therapy, or can also be ascribed to the inhibiting action of high circulating levels of ANP.

The circadian rhythm of atrial natriuretic peptide, vasoactive intestinal peptide, beta-endorphin and cortisol in healthy young and elderly subjects.

Atrial natriuretic peptide, vasoactive intestinal peptide, beta-endorphin and cortisol are humoral variables characterized by a 24-h periodicity. We evaluated the circadian rhythm of these peptides and hormones in healthy subjects who were young (between 20-25 years) or elderly (between 65-75 years). All were on controlled diets. Blood samples were collected six times during a 24-h period (at 06.00, 08.00, 12.00, 18.00, 20.00 and 24.00 h) beginning 8-h after start of recumbency. The time-related data were analysed by the Cosinor method in order to validate the circadian rhythm and to quantify rhythmometric parameters which included the midline estimate of rhythm (mesor). In contrast to the young subjects, Cosinor analysis failed to reveal a significant circadian rhythm in elderly subjects, for plasma cortisol. In elderly subjects oscillation (mesor) of atrial nutriuretic peptide was higher, while that of vasoactive intestinal peptide and beta-endorphins was lower. The results suggest changes in the physiological secretion of these three peptides in healthy elderly subjects.

Evaluation of endothelium loss by in vitro vessel perfusion using millipore filters.

Endothelial loss of isolated rabbit femoral artery and renal artery was evaluated during in vitro vessel perfusion. Desquamated endothelial cells were captured on millipore filters from the perfusion solution outflow of the vessel in 3 intervals lasting 5 minutes each. In the first 5 minutes of perfusion the endothelial loss was 1,289.2 +/- 166.5 cells: in the interval after a 30 minute perfusion 4,967.9 +/- 1,428.3 cells were caught on the filters, 3.9 times more than in the first interval. During and after the 2 minutes air bubble perfusion the endothelial catch was 5.5 times greater as compared to the second interval with the average of 27,473 +/- 6,209.6 cells. The present method of quantification of the endothelial cell loss in the in vitro vessel perfusion experiment makes it possible to obtain informations about the actual state of the endothelial lining and to contribute to more precise evaluation of the modulatory effect of the endothelium on vessel reactivity to pharmacological agents.

[Pulmonary surfactant. Properties, relation to the respiratory distress syndrome in neonates and possibilities of its prevention and therapy]. / Plúcny surfaktant. Vlastnosti, vzt'ah k RDS novorodencov a moznosti jeho prevencie a terapie.

The paper considers pulmonary surfactant with regard to the development of the lungs in the fetal period. Its biochemical properties are characterized and its value for respiratory function in neonates is assessed. Immaturity of lung tissue and of its function in preterm babies accounts for acute respiratory failure, the respiratory distress syndrome (RDS). Prevention of RDS in neonates took advantage of the knowledge obtained in experiments concerning the stimulatory effect of some mediators and hormones on the production and secretion of pulmonary surfactant. In clinical practice mostly glucocorticoids have been used, frequently in combination with beta 2 adrenergic mimetics, while aminophylline and thyroxine were used relatively rarely. The good effectiveness of ambroxol in RDS prevention has recently been demonstrated in clinical studies. The low occurrence of adverse effects potentiates the advantages of ambroxol administration. Both in RDS prevention and treatment the prospective lies with the administration of exogenous surfactant. At present effort is focusing on detecting means that would not only reduce the incidence and mortality for RDS but would also present a minimized risk for the developing organism. (Ref. 110.).

Vasoactive intestinal peptide fluctuates in human blood with a circadian rhythm.

The vasoactive intestinal peptide (VIP) may be radioimmunoassayed in systemic venous blood. The plasma concentrations of VIP were investigated in human blood according to a chronobiological design. The study documented a circadian rhythmicity in time-qualified concentrations of VIP. Accordingly, VIP may be ascribed to biological variables characterized by periodicity in their physiological attributes. The rhythmic physiology of VIP is, however, highly disturbed in its tonic and phasic properties during senescence.

Temporal interrelationships between circadian rhythms of vasoactive intestinal peptide and T lymphocyte subpopulations.

Recent investigations documented an immunosuppressive action of VIP. Previous studies demonstrated that T lymphocytes exhibit a circadian rhythm (CR). This investigation was, thus, performed with the aim of detecting the relationships intercurrent between the 24-h patterns of VIP and T lymphocyte subsets. The hypothesis was formulated that circulating VIP may play a role in controlling the CR of T cells. The research was carried out on 10 clinically healthy subjects, tested six times during the 24-h span by assaying circulating levels of VIP and total T (OKT3), T helper (OKT4), and T suppressor/cytotoxic (OKT8) lymphocytes. Time data series were analyzed by Cosinor method. All investigated variables were seen to be characterized by a statistically significant CR. While the acrophase of VIP CR was found to be located at 18.20, the crest of OKT3, OKT4, OKT8 CR was seen to occur at 03.04, 02.16 and 02.56, respectively. The phase shift was found to be significant, suggesting that VIP and T lymphocytes physiologically fluctuate with a phase angle during their nyctohemeral cyclicity. The finding can be regarded as an indirect evidence of a negative VIPergic chronoregulation of CR of T lymphocytes.

[Changes in the vascular endothelium in fetal rabbits during perfusion in vitro]. / Zmeny endotelu ciev plodov králika v podmienkach perfúzie in vitro.

Under perfusion conditions in vitro the morphologic state of preparations of fetal vessels of the ductus arteriosus and the pulmonary artery was studied by light and electron microscopy. At maintaining standard conditions of perfusion experiments, manipulation of vessels did not result in substantial changes of the endothelium. Storage of the vessel preparations at 4 degrees C for 24 hours did not induce marked morphologic changes, yet the vessels exhibited increased constriction reactivity to noradrenaline which resulted in severe damage of the endothelial lining. Perfusion of the ductus arteriosus by oxygen saturated solution led to vessel constriction and severe endothelial damage. The observed changes should be taken into account on evaluating responses of vascular segments to vasoactive substances.

Day-time changes for pituitary-thyroid hormones denote a rhythmic structure not detectable in elderly subjects.

The day-time concentrations of TSH, T3, T4, FT3 and FT4 were investigated in clinically healthy volunteers, 20 young individuals, aged from 20 to 27 years, and 12 elderly subjects, aged from 70 to 81 years. Nocturnal sampling was purposely avoided in order to eliminate the interindividual differences in sleep patterns. The day-time series were analysed by means of cosinor analysis to interpolate the best-fitting sinusoidal wave that covers the day-night period. The reconstruction of the circadian sinusoidal wave was validated for TSH, T3, T4, FT3 and FT4 in young subjects, and, with limitation, for T4 in elderly individuals. The findings suggest that the day time patterns of TSH, T3, FT4 and FT3 in elderly subjects are characterized by a dyschronism that is not detectable in young subjects. The investigation of the day-time concentrations is suggested as a cost-effective tool for exploring the circadian cycle of the pituitary-thyroid hormones in aged people.