Serpin-A3 is Associated with Persistent Albuminuria in Adolescents with Secondary Podocytopathy, in a Region with High Prevalence of Chronic Kidney Disease of Unknown Origin

Abstract Background: The state of Aguascalientes, Mexico, has been recognized as a chronic kidney disease hotspot. Screening studies have revealed a high prevalence of persistent albuminuria (pA), histologically characterized by glomerulomegaly, and incomplete podocyte fusion, probably associated with oligonephrony. To date, urinary biomarkers have not been explored in this population. Objective: The aim of the study was to identify the presence of potential biomarkers of early renal injury in patients with pA (pACR) and that correspond with the characteristic nephropathy profile that prevails in this entity. Methods: This is a cross-sectional, analytical, and comparative study. Four groups were recruited: adolescents aged 10-17 years with pACR, isolated albuminuria (iACR), no albuminuria (negative control), and adults with biopsy-confirmed glomerulopathy (positive control). Urinary excretion of SerpinA3, heat-shock protein-72 (HSP-72), podocalyxin (PCX), and nephrin was evaluated in urine samples. SerpinA3 and HSP-72 were analyzed by Western blot, and PCX and nephrin were quantified by enzyme-linked immunosorbent assay. Results: The mean GFR in the pACR group was 113.4 mL/min/1.73m2 and differed significantly only from that of the positive control group (65.1 mL/min/1.73m2). The mean albuminuria value in the pACR group was 48.9 mg/g. SerpinA3 concentration differed between groups (0.08 vs. 0.25 ng/mL, p < 0.001): it was significantly higher in the pACR group compared to the negative controls (p = 0.037). Conclusion: SerpinA3 was significantly associated with pA and could become a biomarker of early kidney injury. Further investigations are required to determine whether SerpinA3 precedes the development of albuminuria and its pathogenic role.

Ethical considerations in animal research: the principle of 3r's

In the last century, progress in the knowledge of human diseases, their diagnosis and treatment have grown exponentially, due in large part to the introduction and use of laboratory animals. Along with this important progress, the need to provide training and guidance to the scientific community in all aspects related to the proper use of experimental animals has been indispensable. Animal research committees play a primary role in evaluating experimental research protocols, from their feasibility to the rational use of animals, but above all in seeking animal welfare. The Institutional Committee for the Care and Use of Animals (IACUC) has endeavored to share several relevant aspects in conducting research with laboratory animals. Here, we present and discuss the topics that we consider of utmost importance to take in the account during the design of any experimental research protocol, so we invite researchers, technicians, and undergraduate and graduate students to dive into the fascinating subject of proper animal care and use for experimentation. The main intention of these contributions is to sensitize users of laboratory animals for the proper and rational use of them in experimental research, as well as to disseminate the permitted and unpermitted procedures in laboratory animals. In the first part, the significance of experimental research, the main functions of IACUC, and the principle of the three R's (replacement, reduction, and refinement) are addressed.

Una nueva percepción del papel de la aldosterona en la fisiopatología de la nefrotoxicidad crónica inducida por la ciclosporina / A new perception of aldosterone role in the pathophysiology of cyclosporin induced chronic nephrotoxicity

La ciclosporina A (CsA), un inhibidor de la calcineurina, ha mejorado la supervivencia de injertos del transplante de órganos sólidos y su uso para manejar las enfermedades autoinmunitarias es cada vez más común. Si bien las tasas de supervivencia de pacientes y de injertos han aumentado, el uso clínico de la CsA es limitado ya que frecuentemente tiene efectos nefrotóxicos que se pueden presentar en dos formas distintas y bien caracterizadas: nefrotoxicidad aguda y crónica. La forma aguda se caracteriza por vasoconstricción renal inducida por un desequilibrio en la liberación de sustancias vasoactivas, lo que conduce a una disfunción renal. Esta forma de nefrotoxicidad es reversible. La toxicidad crónica, por su parte, se caracteriza por vasoconstricción y daño estructural que incluye patologías de las arteriolas y fibrosis tubulointersticial, las cuales, por lo general, no son reversibles. Los mecanismos de estos efectos perjudiciales no se conocen con exactitud aunque, en los últimos años, se han hecho grandes avances. En este artículo revisamos la literatura actual sobre la patogenia y las estrategias de tratamiento que se han utilizado para mejorar el daño renal causado por la nefrotoxicidad crónica por CsA. Recientemente se ha sugerido que la aldosterona juega un papel central en la patogenia de la nefrotoxicidad causada por CsA y que la espironolactona puede ser útil para prevenirla. Se discuten estos resultados y el uso del bloqueo de los receptores de mineralocorticoides.

Cyclosporine A (CsA) is a calcineurin inhibitor widely use to prevent organ transplant rejection and for treating autoimmune diseases. Since CsA introduction, organ transplant and patient survival significantly increased and this outcome has not been modified by the new immunosuppressive agents. The long use of CsA, however has been limited because its nephrotoxic side effect. Two forms of renal toxicity have been described, the acute and the chronic toxicity

Acciones de endotelina 1 y angiotensina II en embarazos complicados con preeclampsia / Endothelin 1 and angiotensin II in preeeclampsia

Introduction. It is generally thought that development of hypertension in preeclampsia (PE) is due to generalized endothelial dysfunction and/or results from an imbalance in the production and/or action of vasoactive factors, resulting in higher citosolic Ca2+ concentration which in turn leads to vasoconstriction and decreased blood pressure perfusion in organs, including the fetoplacental unit. Among vasoactive factors involved in blood pressure regulation, endothelin 1 (ET-1) and angiotensin II (Ang II) regulate citosolic Ca2+ concentrations and therefore are considered in this review. PE is associated with higher circulating and placental ET-1 levels, observation that explains, at least in part, vasoconstriction and oxidative stress. Higher and lower Ang II sensitivity seen in PE and normal pregnancy, respectively, could not be explained by changes in renin-angiotensin system components, including Ang II receptors (ATI). During normal pregnancy, ATI receptors are found as monomers and are inactivated by reactive oxygen species (ROS) leading to lower Ang II sensitivity. In contrast, PE is associated with increased ATl/bradicinin receptors (B2) heterodimers which are resistant to inactivation by ROS, maintaining increased ATI-receptor stimulated signaling in PE. In adittion, AT-1 agonistic antibodies (AT1-AA) obtained from PE women increases intracellular Ca2+, NADPH oxidase components and ROS, effects not observed with normal pregnancy AT1-AA. Conclusion. High ET-1 levels, the presence of AT1/B2 receptor heterodimers and increased AT1-AA are involved, at least in part, in the hypertensive and oxidative stress states in PE.

Introducción. Se reconoce que el desarrollo de la hipertensión en la preeclampsia (PE) resulta del daño endotelial generalizado y/o de la falta de equilibrio en la producción y/o acción de agentes vasoactivos, lo que conlleva al incremento en la concentración citosólica de Ca2+ que resulta en vasoconstricción y disminución de la perfusión sanguínea en los órganos, incluyendo la unidad fetoplacentaria. Dentro de los factores vaso-activos que regulan la presión arterial, en la presente revisión se consideró a la endotelina 1 (ET-1) y a la angiotensina II (Ang II), factores que regulan la concentración citosólica de Ca2+. En comparación con el embarazo normal, la PE se asocia con mayor concentración en suero y placenta de ET-1, lo que explica en parte la vasoconstricción y el estado de estrés oxidativo. La respuesta exagerada en la PE y el estado de refractariedad en el embarazo normal a la Ang II no pueden explicarse por componentes del sistema renina-angiotensina, incluyendo a los receptores de Ang II (ATI). Durante el embarazo normal los receptores AT-1 se encuentran en forma de monómeros y son inactivados por las especies reactivas de oxígeno (ROS), lo que se asocia con menor respuesta a Ang II. En cambio, la respuesta exagerada a la Ang II durante la PE puede deberse a la heterodimerizacion de los receptores ATI con los de bradicinina (B2), estado que les confiere resistencia a la inactivación por las especies reactivas de oxígeno (ROS), lo que explica el incremento en la concentración del Ca2+ intracelular. Además, los anticuerpos agonistas del receptor ATI (AT1-AA) de mujeres PE aumenta la concentración de Ca2+ intracelular, de la NADPH oxidasa y de ROS, efectos que no se presentan al utilizar AT1-AA de embarazadas normotensas. Conclusión. Las altas concentraciones de ET-1, la presencia de receptores ATI en forma de heterodimeros ATI/ B2 y el aumento en los AT1-AA explican en parte, el estado de hipertensión y de estrés oxidativo de la PE.

Acción novedosa de la aldosterona en la nefrotoxicidad por ciclosporina / Novel action of aldosterone in CsA nephrotoxicity

Calcineurin inhibitors are helpful ímmunosuppressíve agents in clinical practice. Thanks to their lower cost respect with new ímmunosuppressíve therapy, calcineurin inhibitors in our country continue being the most used treatment in solid organ transplant recipients or patients with autoimmune disease. In the 80's decade cyclosporine A (CsA) was introduced as the first calcineurin inhibitor transforming the immunosuppression therapy. Up to date, many articles evaluating beneficial and adverse effects of CsA have been published. In this review, basic aspects and actions of CsA are analyzed together with studies from our laboratory that pointed out the pathophysiological role of aldosterone as a mediator of functional and structural changes that are observed in CsA nephrotoxicity. Based in our findings, we proposed that in CsA nephrotoxicity, the aldosterone mediates renal vasoconstriction and enhances TGFJ3 expression promoting the development of nefrotoxicity. Finally, results from our laboratory and others allow us to suggest that aldosterone receptors blockade with spironolactone or eplerone could be a pharmacological therapy to reduce or prevent acute and chronic CsA nephrotoxicity in transplant recipients.

Los inhibidores de calcineurina son los agentes inmunosupresores más potentes con los que se cuenta en la práctica clínica, y gracias a su bajo costo respecto a las nuevas terapias inmunosupresoras, en nuestro país continúan siendo los agentes terapéuticos más utilizados para el manejo de pacientes con enfermedades autoinmunes o que reciben trasplantes. En la década de los 80's se introdujo la ciclosporina A (CsA) como primer inhibidor de calcineurina, lo cual revolucionó la terapia inmunosupresora. Desde entonces se han publicado muy variados artículos donde se han evaluado los efectos benéficos y deletéreos de estos inhibidores; específicamente nos enfocaremos a revisar las acciones de CsA y, en particular, los resultados de nuestro laboratorio que muestran el papel fisiopatológico que juega la aldosterona como mediador de los cambios funcionales y estructurales que se observan en la nefrotoxicidad por ciclosporina. Específicamente su participación en promover la vasoconstricción renal asociada a CsA y en el desarrollo de fibrosis al inducir la expresión de TGFβ. Por lo tanto, nuestros resultados y los de otros autores nos permiten proponer el bloqueo de los receptores de aldosterona con espironolactona o eplerone como un tratamiento farmacológico útil para reducir la incidencia de nefrotoxicidad aguda y crónica, inducida por CsA en pacientes con enfermedades autoinmunes o que reciben trasplante de órganos.

Semi-quantitative PCR: A tool to study low abundance messages in the kidney

Some specific functions are often localized to unique cellular types or structures in organs such as kidney, brain, blood, and endocrine glands. As a result, it is not ucommon that gene producta, although heavily expressed in some cell type within these organs, ultimately appear as low abundance products when total RNA is probed, resulting in decreased power of the conventional Northern blot analysis. To study gene expression in these circumstances, more sensitive techniques like RNAse protection assay and quantitative or semi-quantitative PCR strategies have been developed. In the present study, we provede a detailed descriptio of the semiquantitative PCR strategy in our laboratory. Using specific primers to amplify fragments from the neuronal isoform of the nitric oxide synthase and the thiazide-sensitive Na+:Cl- contransporter (low abundance messages in the kidney), we show that the semi-quantitative PCR strategy is a valuable tool when low abundance messages are to be studied