Efferent projections of Nps-expressing neurons in the parabrachial region.

In the brain, connectivity determines function. Neurons in the parabrachial nucleus (PB) relay diverse information to widespread brain regions, but the connections and functions of PB neurons that express Nps (neuropeptide S) remain mysterious. Here, we use Cre-dependent anterograde tracing and whole-brain analysis to map their output connections. While many other PB neurons project ascending axons through the central tegmental tract, NPS axons reach the forebrain via distinct periventricular and ventral pathways. Along the periventricular pathway, NPS axons target the tectal longitudinal column and periaqueductal gray then continue rostrally to target the paraventricular nucleus of the thalamus. Along the ventral pathway, NPS axons blanket much of the hypothalamus but avoid the ventromedial and mammillary nuclei. They also project prominently to the ventral bed nucleus of the stria terminalis, A13 cell group, and magnocellular subparafasciular nucleus. In the hindbrain, NPS axons have fewer descending projections, targeting primarily the superior salivatory nucleus, nucleus of the lateral lemniscus, and periolivary region. Combined with what is known about NPS and its receptor, the output pattern of Nps-expressing neurons in the PB region predicts a role in threat response and circadian behavior.

Sodium appetite and thirst do not require angiotensinogen production in astrocytes or hepatocytes.

In addition to its renal and cardiovascular functions, angiotensin signalling is thought to be responsible for the increases in salt and water intake caused by hypovolaemia. However, it remains unclear whether these behaviours require angiotensin production in the brain or liver. Here, we use in situ hybridization to identify tissue-specific expression of the genes required for producing angiotensin peptides, and then use conditional genetic deletion of the angiotensinogen gene (Agt) to test whether production in the brain or liver is necessary for sodium appetite and thirst. In the mouse brain, we identified expression of Agt (the precursor for all angiotensin peptides) in a large subset of astrocytes. We also identified Ren1 and Ace (encoding enzymes required to produce angiotensin II) expression in the choroid plexus, and Ren1 expression in neurons within the nucleus ambiguus compact formation. In the liver, we confirmed that Agt is widely expressed in hepatocytes. We next tested whether thirst and sodium appetite require angiotensinogen production in astrocytes or hepatocytes. Despite virtually eliminating expression in the brain, deleting astrocytic Agt did not reduce thirst or sodium appetite. Despite markedly reducing angiotensinogen in the blood, eliminating Agt from hepatocytes did not reduce thirst or sodium appetite, and in fact, these mice consumed the largest amounts of salt and water after sodium deprivation. Deleting Agt from both astrocytes and hepatocytes also did not prevent thirst or sodium appetite. Our findings suggest that angiotensin signalling is not required for sodium appetite or thirst and highlight the need to identify alternative signalling mechanisms. KEY POINTS: Angiotensin signalling is thought to be responsible for the increased thirst and sodium appetite caused by hypovolaemia, producing elevated water and sodium intake. Specific cells in separate brain regions express the three genes needed to produce angiotensin peptides, but brain-specific deletion of the angiotensinogen gene (Agt), which encodes the lone precursor for all angiotensin peptides, did not reduce thirst or sodium appetite. Double-deletion of Agt from brain and liver also did not reduce thirst or sodium appetite. Liver-specific deletion of Agt reduced circulating angiotensinogen levels without reducing thirst or sodium appetite. Instead, these angiotensin-deficient mice exhibited an enhanced sodium appetite. Because the physiological mechanisms controlling thirst and sodium appetite continued functioning without angiotensin production in the brain and liver, understanding these mechanisms requires a renewed search for the hypovolaemic signals necessary for activating each behaviour.

Neuropeptide S (NPS) neurons: Parabrachial identity and novel distributions.

Neuropeptide S (NPS) increases wakefulness. A small number of neurons in the brainstem express Nps. These neurons are located in or near the parabrachial nucleus (PB), but we know very little about their ontogeny, connectivity, and function. To identify Nps-expressing neurons within the molecular framework of the PB region, we used in situ hybridization, immunofluorescence, and Cre-reporter labeling in mice. The primary concentration of Nps-expressing neurons borders the lateral lemniscus at far-rostral levels of the lateral PB. Caudal to this main cluster, Nps-expressing neurons scatter through the PB and form a secondary concentration medial to the locus coeruleus (LC). Most Nps-expressing neurons in the PB region are Atoh1-derived, Foxp2-expressing, and mutually exclusive with neurons expressing Calca or Lmx1b. Among Foxp2-expressing PB neurons, those expressing Nps are distinct from intermingled subsets expressing Cck or Pdyn. Examining Nps Cre-reporter expression throughout the brain identified novel populations of neurons in the nucleus incertus, anterior hypothalamus, and lateral habenula. This information will help focus experimental questions about the connectivity and function of NPS neurons.

Transconjunctival and Subciliary Approach in the Treatment of Orbital Fractures: A Study on Oculoplastic Complication.

(1) Background: The aim of the present study was to compare lower eyelid post-operative complications, such as ectropion, entropion, and scleral show of orbital floor fractures, associated to the subciliary vs transconjunctival approaches. (2) Materials and Methods: A retrospective comparative study of patients who underwent surgery for orbital fractures by means of a transconjunctival or a subciliary approach at the Clinic of Dentistry and Maxillofacial Surgery of the University of Verona from January 2013 through September 2018 was designed. Data related to the trauma and to surgical procedures were retrieved, as well as a series of anthropometric parameters extrapolated from standardized photographs. Statistical analysis was performed on the outcomes. (3) Results: 33 patients underwent surgery by means of a transconjunctival approach and 36 patients by means of a subciliary approach. Ectropion was observed to a greater extent in the subciliary group, however the difference resulted to be not statistically significant. Patients in which osteosynthesis devices were used presented with a greater incidence of scleral show with respect to the remaining patients. No statistically significant difference was observed for any of the parameters taken into account. (4) Conclusions: Since the two approaches does not seem to be associated with remarkable differences in terms of outcomes, the choice of technique should be tailored to the patient's features and the surgeon's experience.

Pre-locus coeruleus neurons in rat and mouse.

Previously, we identified a population of neurons in the hindbrain tegmentum, bordering the locus coeruleus (LC). We named this population the pre-locus coeruleus (pre-LC) because in rats its neurons lie immediately rostral to the LC. In mice, however, pre-LC and LC neurons intermingle, making them difficult to distinguish. Here, we use molecular markers and anterograde tracing to clarify the location and distribution of pre-LC neurons in mice, relative to rats. First, we colocalized the transcription factor FoxP2 with the activity marker Fos to identify pre-LC neurons in sodium-deprived rats and show their distribution relative to surrounding catecholaminergic and cholinergic neurons. Next, we used sodium depletion and chemogenetic activation of the aldosterone-sensitive HSD2 neurons in the nucleus of the solitary tract (NTS) to identify the homologous population of pre-LC neurons in mice, along with a related population in the central lateral parabrachial nucleus. Using Cre-reporter mice for Pdyn, we confirmed that most of these sodium-depletion-activated neurons are dynorphinergic. Finally, after confirming that these neurons receive excitatory input from the NTS and paraventricular hypothalamic nucleus, plus convergent input from the inhibitory AgRP neurons in the arcuate hypothalamic nucleus, we identify a major, direct input projection from the medial prefrontal cortex. This new information on the location, distribution, and input to pre-LC neurons provides a neuroanatomical foundation for cell-type-specific investigation of their properties and functions in mice. Pre-LC neurons likely integrate homeostatic information from the brainstem and hypothalamus with limbic, contextual information from the cerebral cortex to influence ingestive behavior.

The Secret Garden of Neuronal circRNAs.

High-throughput transcriptomic profiling approaches have revealed that circular RNAs (circRNAs) are important transcriptional gene products, identified across a broad range of organisms throughout the eukaryotic tree of life. In the nervous system, they are particularly abundant, developmentally regulated, region-specific, and enriched in genes for neuronal proteins and synaptic factors. These features suggested that circRNAs are key components of an important layer of neuronal gene expression regulation, with known and anticipated functions. Here, we review major recognized aspects of circRNA biogenesis, metabolism and biological activities, examining potential new functions in the context of the nervous system.

Central afferents to the nucleus of the solitary tract in rats and mice.

The nucleus of the solitary tract (NTS) regulates life-sustaining functions ranging from appetite and digestion to heart rate and breathing. It is also the brain's primary sensory nucleus for visceral sensations relevant to symptoms in medical and psychiatric disorders. To better understand which neurons may exert top-down control over the NTS, here we provide a brain-wide map of all neurons that project axons directly to the caudal, viscerosensory NTS, focusing on a medial subregion with aldosterone-sensitive HSD2 neurons. Injecting an axonal tracer (cholera toxin b) into the NTS produces a similar pattern of retrograde labeling in rats and mice. The paraventricular hypothalamic nucleus (PVH), lateral hypothalamic area, and central nucleus of the amygdala (CeA) contain the densest concentrations of NTS-projecting neurons. PVH afferents are glutamatergic (express Slc17a6/Vglut2) and are distinct from neuroendocrine PVH neurons. CeA afferents are GABAergic (express Slc32a1/Vgat) and are distributed largely in the medial CeA subdivision. Other retrogradely labeled neurons are located in a variety of brain regions, including the cerebral cortex (insular and infralimbic areas), bed nucleus of the stria terminalis, periaqueductal gray, Barrington's nucleus, Kölliker-Fuse nucleus, hindbrain reticular formation, and rostral NTS. Similar patterns of retrograde labeling result from tracer injections into different NTS subdivisions, with dual retrograde tracing revealing that many afferent neurons project axon collaterals to both the lateral and medial NTS subdivisions. This information provides a roadmap for studying descending axonal projections that may influence visceromotor systems and visceral "mind-body" symptoms.

Renovascular hypertension elevates pulmonary ventilation in rats by carotid body-dependent mechanisms.

The two kidney-one clip (2K1C) renovascular hypertension depends on the renin-angiotensin system and sympathetic overactivity. The maintenance of 2K1C hypertension also depends on inputs from the carotid bodies (CB), which when activated stimulate the respiratory activity. In the present study, we investigated the importance of CB afferent activity for the ventilatory responses in 2K1C hypertensive rats and for phrenic and hypoglossal activities in in situ preparations of normotensive rats treated with angiotensin II. Silver clips were implanted around the left renal artery of male Holtzman rats (150 g) to induce renovascular hypertension. Six weeks after clipping, hypertensive 2K1C rats showed, in conscious state, elevated resting tidal volume and minute ventilation compared with the normotensive group. 2K1C rats also presented arterial alkalosis, urinary acidification, and amplified hypoxic ventilatory response. Carotid body removal (CBR), 2 wk before the experiments (4th week after clipping), significantly reduced arterial pressure and pulmonary ventilation in 2K1C rats but not in normotensive rats. Intra-arterial administration of angiotensin II in the in situ preparation of normotensive rats increased phrenic and hypoglossal activities, responses that were also reduced after CBR. Results show that renovascular hypertensive rats exhibit increased resting ventilation that depends on CB inputs. Similarly, angiotensin II increases phrenic and hypoglossal activities in in situ preparations of normotensive rats, responses that also depend on CB inputs. Results suggest that mechanisms that depend on CB inputs in renovascular hypertensive rats or during angiotensin II administration in normotensive animals increase respiratory drive.

Differential Expression of Hippocampal Circular RNAs in the BTBR Mouse Model for Autism Spectrum Disorder.

Autism spectrum disorder (ASD) is a heterogeneous neurodevelopmental condition with unknown etiology. Recent experimental evidences suggest the contribution of non-coding RNAs (ncRNAs) in the pathophysiology of ASD. In this work, we aimed to investigate the expression profile of the ncRNA class of circular RNAs (circRNAs) in the hippocampus of the BTBR T + tf/J (BTBR) mouse model and age-matched C57BL/6J (B6) mice. Alongside, we analyzed BTBR hippocampal gene expression profile to evaluate possible correlations between the differential abundance of circular and linear gene products. From RNA sequencing data, we identified circRNAs highly modulated in BTBR mice. Thirteen circRNAs and their corresponding linear isoforms were validated by RT-qPCR analysis. The BTBR-regulated circCdh9 was better characterized in terms of molecular structure and expression, highlighting altered levels not only in the hippocampus, but also in the cerebellum, prefrontal cortex, and amygdala. Finally, gene expression analysis of the BTBR hippocampus pinpointed altered biological and molecular pathways relevant for the ASD phenotype. By comparison of circRNA and gene expression profiles, we identified 6 genes significantly regulated at either circRNA or mRNA gene products, suggesting low overall correlation between circRNA and host gene expression. In conclusion, our results indicate a consistent deregulation of circRNA expression in the hippocampus of BTBR mice. ASD-related circRNAs should be considered in functional studies to identify their contribution to the etiology of the disorder. In addition, as abundant and highly stable molecules, circRNAs represent interesting potential biomarkers for autism.

Importance of the commissural nucleus of the solitary tract in renovascular hypertension.

The rodent renovascular hypertension model has been used to investigate the mechanisms promoting hypertension. The importance of the carotid body for renovascular hypertension has been demonstrated. As the commissural NTS (cNTS) is the first synaptic site in the central nervous system that receives information from carotid body chemoreceptors, we evaluated the contribution of cNTS to renovascular hypertension in the present study. Normotensive male Holtzman rats were implanted with a silver clip around the left renal artery to induce two-kidney, one-clip (2K1C) hypertension. Six weeks later, isoguvacine (a GABAA agonist) or losartan (an AT1 antagonist) was injected into the cNTS, and the effects were compared with carotid body removal. Immunohistochemistry for Iba-1 and GFAP to label microglia and astrocytes, respectively, and RT-PCR for components of the renin-angiotensin system and cytokines in the NTS were also performed 6 weeks after renal surgery. The inhibition of cNTS with isoguvacine or the blockade of AT1 receptors with losartan in the cNTS decreased the blood pressure and heart rate of 2K1C rats even more than carotid body removal did. The mRNA expression of NOX2, TNF-α and IL-6, microglia, and astrocytes also increased in the cNTS of 2K1C rats compared to that of normotensive rats. These results indicate that tonically active neurons within the cNTS are essential for the maintenance of hypertension in 2K1C rats. In addition to signals from the carotid body, the present results suggest that angiotensin II directly activates the cNTS and may also induce microgliosis and astrogliosis within the NTS, which, in turn, cause oxidative stress and neuroinflammation.

Aldosterone-sensitive HSD2 neurons in mice.

Sodium deficiency elevates aldosterone, which in addition to epithelial tissues acts on the brain to promote dysphoric symptoms and salt intake. Aldosterone boosts the activity of neurons that express 11-beta-hydroxysteroid dehydrogenase type 2 (HSD2), a hallmark of aldosterone-sensitive cells. To better characterize these neurons, we combine immunolabeling and in situ hybridization with fate mapping and Cre-conditional axon tracing in mice. Many cells throughout the brain have a developmental history of Hsd11b2 expression, but in the adult brain one small brainstem region with a leaky blood-brain barrier contains HSD2 neurons. These neurons express Hsd11b2, Nr3c2 (mineralocorticoid receptor), Agtr1a (angiotensin receptor), Slc17a6 (vesicular glutamate transporter 2), Phox2b, and Nxph4; many also express Cartpt or Lmx1b. No HSD2 neurons express cholinergic, monoaminergic, or several other neuropeptidergic markers. Their axons project to the parabrachial complex (PB), where they intermingle with AgRP-immunoreactive axons to form dense terminal fields overlapping FoxP2 neurons in the central lateral subnucleus (PBcL) and pre-locus coeruleus (pLC). Their axons also extend to the forebrain, intermingling with AgRP- and CGRP-immunoreactive axons to form dense terminals surrounding GABAergic neurons in the ventrolateral bed nucleus of the stria terminalis (BSTvL). Sparse axons target the periaqueductal gray, ventral tegmental area, lateral hypothalamic area, paraventricular hypothalamic nucleus, and central nucleus of the amygdala. Dual retrograde tracing revealed that largely separate HSD2 neurons project to pLC/PB or BSTvL. This projection pattern raises the possibility that a subset of HSD2 neurons promotes the dysphoric, anorexic, and anhedonic symptoms of hyperaldosteronism via AgRP-inhibited relay neurons in PB.

Novel regulator of vasopressin secretion: phoenixin.

The newly described hypothalamic peptide, phoenixin, is produced in the hypothalamus and adenohypophysis, where it acts to control reproductive hormone secretion. Both phoenixin and its receptor GPR173 are expressed in the hypothalamic supraoptic (SON) and paraventricular (PVN) nuclei, suggesting additional, nonreproductive effects of the peptide to control vasopressin (AVP) or oxytocin (OT) secretion. Hypothalamo-neurohypophysial explants released AVP but not OT in response to phoenixin. Intracerebroventricular administration of phoenixin into conscious, unrestrained male and female rats significantly increased circulating AVP, but not OT, levels in plasma, and it increased immediate early gene expression in the supraoptic nuclei of male rats. Bath application of phoenixin in hypothalamic slice preparations resulted in depolarization of PVN neurons, indicating a direct, neural action of phoenixin in the hypothalamus. Our results suggest that the newly described, hypothalamic peptide phoenixin, in addition to its effects on hypothalamic and pituitary mechanisms controlling reproduction, may contribute to the physiological mechanisms regulating fluid and electrolyte homeostasis.

Lateral parabrachial nucleus and opioid mechanisms of the central nucleus of the amygdala in the control of sodium intake.

Facilitatory and inhibitory mechanisms in the central nucleus of the amygdala (CeA) and the lateral parabrachial nucleus (LPBN), respectively, are important for the control of sodium and water intake. Here we investigated the importance of the opioid mechanisms in the CeA for water and 0.3M NaCl intake in euhydrated or hyperosmotic rats treated with injections of muscimol (GABAA agonist) or moxonidine (α2 adrenergic/imidazoline agonist) into the LPBN, respectively. Male Holtzman rats (n=4-8/group) with stainless steel cannulas implanted bilaterally in the CeA and in the LPBN were used. The ingestion of 0.3M NaCl and water by euhydrated rats treated with muscimol (0.5nmol/0.2µl) into the LPBN (29.4±2.7 and 15.0±2.4ml/4h, respectively) was abolished by the previous injections of naloxone (opioid antagonist, 40µg/0.2µl) into the CeA (0.7±0.3 and 0.3±0.1ml/4h, respectively). The ingestion of 0.3M NaCl by rats treated with intragastric 2M NaCl (2ml/rat) combined with moxonidine (0.5nmol/0.2µl) into the LPBN (17.0±3.8ml/2h) was also strongly reduced by the previous injections of naloxone into the CeA (3.2±2.5ml/2h). Sucrose intake was not affected by naloxone injections into the CeA, which minimized the possibility of non-specific inhibition of ingestive behaviors with this treatment. The present results suggest that opioid mechanisms in the CeA are essential for hypertonic NaCl intake when the LPBN inhibitory mechanisms are deactivated or attenuated with injections of muscimol or moxonidine in this area.

Crioglobulina con propiedad crioaglutinina en un paciente con linfoma esplénico de zona marginal / Cryoglobulin with cold agglutinin properties in a patient with splenic marginal zone lymphoma / Crioglobulina com propriedade crioaglutinina em um paciente com linfoma esplênico de zona marginal

El objetivo del trabajo fue describir un caso de linfoma esplénico de zona marginal (LEZM) con un componente monoclonal que posee propiedades de crioglobulina y crioaglutinina, un hallazgo de muy baja frecuencia. Una paciente con LEZM padeció una anemia hemolítica autoinmune 5 años después del diagnóstico, con hematocrito de 0,15 L/L, hemoglobina 49 g/L, lactato deshidrogenasa 16,82 μkat/L, prueba de Coombs directa positiva con anti- IgG/C3d, bilirrubina total 90,6 μmol/L e indirecta de 58,1 μmol/L. No presentó evidencia clínica ni serológica de infección por VIH, hepatitis B ni C. El proteinograma sérico presentó un pico monoclonal de 14 g/L, con crioglobulinemia positiva a las 24 h, y un criocrito de 30%. La crioglobulina purificada fue de tipo I con un componente monoclonal IgM-lambda, coincidente al observado en suero. El suero, el eluato a 37 °C y la crioglobulina purificada de la paciente presentaron actividad de crioaglutinina con especificidad anti-I, fenómeno producido por la misma inmunoglobulina. El hallazgo de una crioglobulina con propiedades de crioaglutinina en pacientes con LEZM no ha sido descrito previamente en la bibliografía.

The aim of this paper is to describe a case of splenic marginal zone lymphoma (SMZL) with monoclonal component having properties of cryoglobulin and cold agglutinin, a finding of very low frequency. A patient with SMZL suffered autoimmune hemolytic anemia five years after diagnosis, with hematocrit 0.15 L/L, hemoglobin 49 g/L, lactate dehydrogenase 16.82 μkat/L, direct Coombs test with anti-IgG/C3d positive, total bilirubin 90.6 μmol/L and indirect 58.1 μmol/L. She presented no clinical or serological evidence of HIV, hepatitis B or C infection. Serum protein electrophoresis showed a monoclonal peak of 14 g/L, with positive cryoglobulinemia at 24 hours, and 30% cryocrit. Purified cryoglobulin was type I with a monoclonal IgM-lambda component coincident with that observed in serum. The patient serum, eluate at 37 °C and purified cryoglobulin showed cold agglutinin activity with anti-I specificity, phenomenon produced by the same immunoglobulin. The finding of a cryoglobulin with cold agglutinin properties in patients with SMZL has not been previously described in the literature.

O objetivo do trabalho foi descrever um caso de linfoma esplênico de zona marginal (LEZM) com um componente monoclonal com propriedades do crioglobulina e crioaglutinina, um achado de muito baixa frequência. Um doente com LEZM sofreu uma anemia hemolítica autoimune cinco anos após o diagnóstico, com hematócrito de 0,15 L/L, hemoglobina 49 g/L, lactato desidrogenase de 16,82 μkat/L, teste de Coombs direto positivo com anti-IgG/C3d, bilirrubina total 90,6 μmol/L e indireta 58,1 μmol/L. Não apresentou evidência clínica ou sorológica de infecção por HIV, hepatite B ou C. O proteinograma sérico mostrou um pico monoclonal de 14 g/L, com crioglobulinemia positiva 24 horas, e um criocrito de 30%. Crioglobulina purificada foi tipo I com o componente monoclonal IgM-lambda, coincidente com a observada no soro. O soro, o eluato a 37 ° C e a crioglobulina purificada do paciente mostraram atividade de crioaglutinina com especificidade anti-I, fenômeno produzido pela mesma imunoglobulina. O achado de uma crioglobulina com propriedades de crioaglutinina em pacientes com LEZM não foi previamente descrito na literatura.

Importance of the central nucleus of the amygdala on sodium intake caused by deactivation of lateral parabrachial nucleus.

The lateral parabrachial nucleus (LPBN) and the central nucleus of the amygdala (CeA) are important central areas for the control of sodium appetite. In the present study, we investigated the importance of the facilitatory mechanisms of the CeA on NaCl and water intake produced by the deactivation of LPBN inhibitory mechanisms. Male Holtzman rats (n=7-14) with stainless steel cannulas implanted bilaterally in the CeA and LPBN were used. Bilateral injections of moxonidine (α2-adrenoceptor/imidazoline agonist, 0.5 nmol/0.2 µl) into the LPBN increased furosemide+captopril-induced 0.3M NaCl (29.7 ± 7.2, vs. vehicle: 4.4 ± 1.6 ml/2h) and water intake (26.4 ± 6.7, vs. vehicle: 8.2 ± 1.6 ml/2h). The GABAA agonist muscimol (0.25 nmol/0.2 µl) injected bilaterally into the CeA abolished the effects of moxonidine into the LPBN on 0.3M NaCl (2.8 ± 1.6 ml/2h) and water intake (3.3 ± 2.3 ml/2h). Euhydrated rats treated with muscimol (0.5 nmol/0.2 µl) into the LPBN also ingested 0.3M NaCl (19.1 ± 6.4 ml/4h) and water (8.8 ± 3.2 ml/4h). Muscimol (0.5 nmol/0.2 µl) into the CeA also abolished 0.3M NaCl (0.1 ± 0.04 ml/4h) and water intake (0.1 ± 0.02 ml/4h) in euhydrated treated with muscimol into the LPBN. The present results show that neuronal deactivation of the CeA abolishes NaCl intake produced by the blockade of LPBN inhibitory mechanisms, suggesting an interaction between facilitatory mechanisms of the CeA and inhibitory mechanisms of the LPBN in the control of NaCl intake.

Microproteínas urinarias en diabéticos tipo 2 normoalbuminúricos / Urinary microproteins in normoalbuminuric type 2 diabetic patients / Microproteínas urinárias em diabéticos tipo 2 normoalbuminúricos

En pacientes diabéticos tipo 2 normoalbuminúricos se observa la presencia de Microproteínas Urinarias (MU) en el rango 68-25 kDa. El objetivo del trabajo fue identificar en distintos estadios de la nefropatía diabética si dicho rango corresponde a un marcador de daño tubular. Se estudiaron 119 orinas espontáneas de pacientes diabéticos tipo 2; se les midió la relación albúmina/creatinina urinaria y la creatinina sérica. Se dividieron en 5 grupos: 71 normoalbuminúricos, 28 microalbuminúricos, 12 macroalbuminúricos, 2 urémicos en pre-diálisis y 6 en hemodiálisis. Las MU se detectaron en geles de poliacrilamida en 2 dimensiones para uso clínico y se analizaron con el programa Image J 1.30v. La identificación de las MU se realizó por “immunoblotting” o por espectrometría de masa MALDI-TOF-TOF. El 66% de los normoalbuminúricos presentaron las siguientes MU: orosomucoide, fragmento de 35 kDa de la cadena pesada H4 del inter alfa I inhibidor de tripsina y Beta Trace, las cuales no reflejaron daño tubular debido a que la concentración de las mismas no se incrementó en los pacientes en hemodiálisis, en comparación con los normoalbuminúricos. Dichas proteínas están vinculadas al endotelio vascular y podrían constituir un marcador urinario vascular-tubular renal de utilidad clínica en patologías sistémicas con riesgo cardiovascular y funcionalidad renal conservada.

Urinary excretion of microproteins (MU) was detected in normoalbuminuric type 2 diabetic patients, in the range of 68-25 kDa using SDS-PAGE with silver staining. The purpose of this study was to identify MU in diabetic patients in different grades of diabetic nephropathy, in order to clarify the diagnostic relevance as a marker of renal tubular damage. In the spontaneous urine of 119 type 2 diabetic patients, urinary albumin, urinary creatinine and serum were determined. Five groups were formed: 71 normoalbuminuric, 28 microalbuminuric, 12 macroalbuminuric, 2 in pre-dialysis and 6 in hemodialysis. The MU were separated by two-dimensional polyacrylamid gel electrophoresis for clinical use (2D UC) and were identified by immunoblotting or MALDI-TOF-TOF mass spectrometry and analyzed using Image J version 1.30v. Of the normoalbuminurics patients studied, 66% excreted the following MU: orosomucoid, 35 kDa fragment of inter-alpha-trypsin inhibitor heavy chain H4 (ITIH4) and Beta Trace; but their concentrations did not reflect tubular damage because they exhibited a progressive downregulation. These proteins are involved in vascular endothelium, and they could be a marker of renal tubular-microvascular disease that would be useful in systemic diseases with cardiovascular risk and with preserved renal function.

Em pacientes diabéticos tipo 2 “normoalbuminúricos” se observa a presença de microproteínas urinárias (MU) em um intervalo compreendido entre 68 e 25 kDa. O objetivo do trabalho é identificar em distintos estágios da nefropatia diabética se tal intervalo corresponde a um marcador de dano tubular. Foram estudadas 119 amostras de urina espontânea de pacientes diabéticos tipo 2; foi medida a reação albumina/creatinina urinária e a creatinina sérica. Dividiram-se em 5 grupos: 71 normoalbuminúricos, 28 microalbuminúricos, 12 macroalbuminúricos, 2 urêmicos pré-dialise e 6 em hemodiálise. Foram detectadas as MU em géis de poliacrilamida em duas dimensões para o uso clínico e analisadas com o programa Image J versão 1.30v. A identificação das MU foi realizada por “immunoblotting” ou por espectrometria de massa MALDI-TOF-TOF. 66% de normoalbuminúricos apresentaram as seguintes MU: orosomucoide, Fragmento de 35 kDa da cadeia pesada H4 do interalfa inibidor da tripsina e Beta Trace, as quais não mostraram dano tubular devido a que a concentração das mesmas não está aumentada nos pacientes em hemodiálise, em comparação com os normoalbuminúricos. Estas proteínas estão envolvidas com o endotélio vascular e poderiam constituir um marcador urinário vascular-tubular renal de utilidade clínica em patologias sistêmicas com risco cardiovascular e funcionalidade renal conservada.

Receptor soluble de transferrina y eritropoyetina en la anemia de los procesos crónicos con y sin deficiencia de hierro / Soluble transferrin receptor and erythropoietin in chronic disease anemia with or without iron deficiency

La regulación del receptor de transferrina (RTF) está relacionada con los depósitos de hierro (Fe) intracelular y guarda una relación constante con el receptor soluble presente en plasma. Se ha demostrado que en las anemias por deficiencia de Fe (AF) cuando se produce la disminución de los depósitos de Fe, aumenta la expresión del receptor. En las anemias de los procesos crónicos (APC) establecer el verdadero estado del Fe es complejo, debido a la influencia que tienen los procesos inflamatorios ó infecciosos en el equilibrio del Fe orgánico. Se estudiaron 30 sujetos sanos normales (grupo control) y 42 pacientes anémicos ( hemoglobina menor de 120 g/L) que presentaban APC con y sin deficiencia de hierro, a fin de establecer el valor diagnóstico del receptor soluble de transferrina (RTFs). Se correlacionó la eritropoyetina (EPO) como factor estimulador de la eritropoyesis , con los descensos de hemoglobina que se producen en ambos grupos. Los resultados fueron analizados aplicando el test estadístico ANOVA, no encontrándose diferencia significativa en los valores de RTFs entre los grupos de APC con y sin deficiencia de Fe. La relación log EPO versus hemoglobina (Hb) en ambos grupos mostró una correlación inversa estadísticamente significativa. Se concluye que de acuerdo a los resultados obtenidos, los valores de RTFs en estos pacientes se encuentran dentro de los rangos normales y no se relacionan con el estado del hierro orgánico. Por consiguiente, su empleo como parámetro diferencial para establecer deficiencia de hierro en las APC, no tendría aplicación diagnóstica.

Elevada concentracion de inmunoglobulinas séricas en distintos estadios clínicos de infección por HIV-1 / Concentration elevated of serum immunoglobulins in different clinical stages of infection for HIV-1

La concentración de IgG, IgA e IgM fue determinada por inmunodifusión radial el suero de 127 pacientes, 77 infectados por HIV-1 (Grupo A), agrupados en distintos estadios clínicos, y 50 no infectados por HIV-1 (Grupo B). Se encontraron diferencias estadísticamente significativas entre los niveles séricos de IgG e IgM (Grupo A), y los del Grupo B. En el contraste, discriminando por estadios clínicos, la concentración de IgG Estadio III y IV (Grupo A), fue mayor que IgG Grupo B, al igual que para IgM Estadio IV fue mayor que IgM Grupo B. Las medias de los niveles de inmunoglobulinas entre estadios clínicos, arrojaron diferencias significativas para las distintas inmunoglobulinas, y, especialmente, para IgA Estadio IV, mayor que IgA Estadio III. Resultó notable el aumento de IgM en el estadio IV, el de mayor compromiso clínico e inmunológico; esto podría indicar una respuesta paradógica remanente a nuevos antígenos. Las concentraciones elevadas de inmunoglobulinas séricas estarían dadas por una respuesta inmune inusual, que no es exclusiva de los linfocitos-B, y que ocurre desde los primeros estadios de infección por HIV-1.