Postnatal Overfeeding during Lactation Induces Endothelial Dysfunction and Cardiac Insulin Resistance in Adult Rats.

Early overnutrition is associated with cardiometabolic alterations in adulthood, likely attributed to reduced insulin sensitivity due to its crucial role in the cardiovascular system. This study aimed to assess the long-term effects of early overnutrition on the development of cardiovascular insulin resistance. An experimental childhood obesity model was established using male Sprague Dawley rats. Rats were organized into litters of 12 pups/mother (L12-Controls) or 3 pups/mother (L3-Overfed) at birth. After weaning, animals from L12 and L3 were housed three per cage and provided ad libitum access to food for 6 months. L3 rats exhibited elevated body weight, along with increased visceral, subcutaneous, and perivascular fat accumulation. However, heart weight at sacrifice was reduced in L3 rats. Furthermore, L3 rats displayed elevated serum levels of glucose, leptin, adiponectin, total lipids, and triglycerides compared to control rats. In the myocardium, overfed rats showed decreased IL-10 mRNA levels and alterations in contractility and heart rate in response to insulin. Similarly, aortic tissue exhibited modified gene expression of TNFα, iNOS, and IL-6. Additionally, L3 aortas exhibited endothelial dysfunction in response to acetylcholine, although insulin-induced relaxation remained unchanged compared to controls. At the molecular level, L3 rats displayed reduced Akt phosphorylation in response to insulin, both in myocardial and aortic tissues, whereas MAPK phosphorylation was elevated solely in the myocardium. Overfeeding during lactation in rats induces endothelial dysfunction and cardiac insulin resistance in adulthood, potentially contributing to the cardiovascular alterations observed in this experimental model.

The CCN2 Polymorphism rs12526196 Is a Risk Factor for Ascending Thoracic Aortic Aneurysm.

Cellular communication network factor 2 (CCN2/CTGF) has been traditionally described as a downstream mediator of other profibrotic factors including transforming growth factor (TGF)-ß and angiotensin II. However, recent evidence from our group demonstrated the direct role of CCN2 in maintaining aortic wall homeostasis and acute and lethal aortic aneurysm development induced by angiotensin II in the absence of CCN2 in mice. In order to translate these findings to humans, we evaluated the potential association between three polymorphisms in the CCN2 gene and the presence of a thoracic aortic aneurysm (TAA). Patients with and without TAA retrospectively selected were genotyped for rs6918698, rs9402373 and rs12526196 polymorphisms related to the CCN2 gene. Multivariable logistic regression models were performed. In our population of 366 patients (69 with TAA), no associations were found between rs6918698 and rs9402373 and TAA. However, the presence of one C allele from rs12526196 was associated with TAA comparing with the TT genotype, independently of risk factors such as sex, age, hypertension, type of valvulopathy and the presence of a bicuspid aortic valve (OR = 3.17; 95% CI = 1.30-7.88; p = 0.011). In conclusion, we demonstrated an association between the C allele of rs12526196 in the CCN2 gene and the presence of TAA. This study extrapolates to humans the relevance of CCN2 in aortic aneurysm observed in mice and postulates, for the first time, a potential protective role to CCN2 in aortic aneurysm pathology. Our results encourage future research to explore new variants in the CCN2 gene that could be predisposed to TAA development.

Interplay between extracellular matrix components and cellular and molecular mechanisms in kidney fibrosis.

Chronic kidney disease (CKD) is characterized by pathological accumulation of extracellular matrix (ECM) proteins in renal structures. Tubulointerstitial fibrosis is observed in glomerular diseases as well as in the regeneration failure of acute kidney injury (AKI). Therefore, finding antifibrotic therapies comprises an intensive research field in Nephrology. Nowadays, ECM is not only considered as a cellular scaffold, but also exerts important cellular functions. In this review, we describe the cellular and molecular mechanisms involved in kidney fibrosis, paying particular attention to ECM components, profibrotic factors and cell-matrix interactions. In response to kidney damage, activation of glomerular and/or tubular cells may induce aberrant phenotypes characterized by overproduction of proinflammatory and profibrotic factors, and thus contribute to CKD progression. Among ECM components, matricellular proteins can regulate cell-ECM interactions, as well as cellular phenotype changes. Regarding kidney fibrosis, one of the most studied matricellular proteins is cellular communication network-2 (CCN2), also called connective tissue growth factor (CTGF), currently considered as a fibrotic marker and a potential therapeutic target. Integrins connect the ECM proteins to the actin cytoskeleton and several downstream signaling pathways that enable cells to respond to external stimuli in a coordinated manner and maintain optimal tissue stiffness. In kidney fibrosis, there is an increase in ECM deposition, lower ECM degradation and ECM proteins cross-linking, leading to an alteration in the tissue mechanical properties and their responses to injurious stimuli. A better understanding of these complex cellular and molecular events could help us to improve the antifibrotic therapies for CKD.

CCN2 Increases TGF-ß Receptor Type II Expression in Vascular Smooth Muscle Cells: Essential Role of CCN2 in the TGF-ß Pathway Regulation.

The cellular communication network factor 2 (CCN2/CTGF) has been traditionally described as a mediator of the fibrotic responses induced by other factors including the transforming growth factor ß (TGF-ß). However, several studies have defined a direct role of CCN2 acting as a growth factor inducing oxidative and proinflammatory responses. The presence of CCN2 and TGF-ß together in the cellular context has been described as a requisite to induce a persistent fibrotic response, but the precise mechanisms implicated in this relation are not described yet. Considering the main role of TGF-ß receptors (TßR) in the TGF-ß pathway activation, our aim was to investigate the effects of CCN2 in the regulation of TßRI and TßRII levels in vascular smooth muscle cells (VSMCs). While no differences were observed in TßRI levels, an increase in TßRII expression at both gene and protein level were found 48 h after stimulation with the C-terminal fragment of CCN2 (CCN2(IV)). Cell pretreatment with a TßRI inhibitor did not modify TßRII increment induced by CCN2(VI), demonstrating a TGF-ß-independent response. Secondly, CCN2(IV) rapidly activated the SMAD pathway in VSMCs, this being crucial in the upregulation of TßRII since the preincubation with an SMAD3 inhibitor prevented it. Similarly, pretreatment with the epidermal growth factor receptor (EGFR) inhibitor erlotinib abolished TßRII upregulation, indicating the participation of this receptor in the observed responses. Our findings suggest a direct role of CCN2 maintaining the TGF-ß pathway activation by increasing TßRII expression in an EGFR-SMAD dependent manner activation.

Interleukin-17A induces vascular remodeling of small arteries and blood pressure elevation.

An important link exists between hypertension and inflammation. Hypertensive patients present elevated circulating levels of proinflammatory cytokines, including interleukin-17A (IL-17A). This cytokine participates in host defense, autoimmune and chronic inflammatory pathologies, and cardiovascular diseases, mainly through the regulation of proinflammatory factors. Emerging evidence also suggests that IL-17A could play a role in regulating blood pressure and end-organ damage. Here, our preclinical studies in a murine model of systemic IL-17A administration showed that increased levels of circulating IL-17A raised blood pressure induced inward remodeling of small mesenteric arteries (SMAs) and arterial stiffness. In IL-17A-infused mice, treatment with hydralazine and hydrochlorothiazide diminished blood pressure elevation, without modifying mechanical and structural properties of SMA, suggesting a direct vascular effect of IL-17A. The mechanisms of IL-17A seem to involve an induction of vascular smooth muscle cell (VSMC) hypertrophy and phenotype changes, in the absence of extracellular matrix (ECM) proteins accumulation. Accordingly, treatment with an IL-17A neutralizing antibody diminished SMA remodeling in a model of angiotensin II (Ang II) infusion. Moreover, in vitro studies in VSMCs reported here, provide further evidence of the direct effects of IL-17A on cell growth responses. Our experimental data suggest that IL-17A is a key mediator of vascular remodeling of the small arteries, which might contribute, at least in part, to blood pressure elevation.

Molecular Regulation of Notch Signaling by Gremlin.

Gremlin is a member of the TGF-ß superfamily that can act as a BMP antagonist, and recently, has been described as a ligand of the vascular endothelial growth factor receptor 2 (VEGFR2). Gremlin shares properties with the Notch signaling pathway. Both participate in embryonic development and are reactivated in pathological conditions. Gremlin is emerging as a potential therapeutic target and biomarker of renal diseases. Here we review the role of the Gremlin-VEGFR2 axis in renal damage and downstream signaling mechanisms, such as Notch pathway.

Eligibility test for advanced chronic kidney disease: Revision and proposal.

Nowadays, chronic kidney disease (CKD) prevalence keeps increasing worldwide. The management of these patients usually requires renal replacement therapy (RRT). However, the complexity of patients' profiles comprises a great challenge to overcome. During the last decades, CKD units have been developed to offer multidisciplinary and coordinated attention to patients, helping in the decision-making of the RRT. Nevertheless, there is a huge variability in the performance and organization of care practice, implying an existing necessity to homogenize the RRT modality chosen. We propose a test composed of two parts: one to be completed by the medical staff (to evaluate contraindications for the different RRT techniques) and another by the patient or nursing staff (to consider patients' preferences). In this sense, it would be possible to have a common and useful tool to complement patient education in RRT, as well as sharing decision-making in the ACKD units taking into account patient preferences.

A new procedure to induce aortic aneurysms in mice.

Aortic aneurysms (AAs) are a major public health challenge, featured by a progressive impairs in aortic wall integrity that drives to aortic dilation and, in end stage, to its rupture. Despite important advances in the surgical treatment of aortic aneurysms, there is currently no pharmacological intervention that prevents their development, reduces their expansion, or avoids their rupture. In addition to classic risk factors such age or gender, several heritable connective tissue disorders have been associated with AA developing, highlighting the role of extracellular matrix (ECM) genes alterations in the developing of AA. In this sense, we have recently demonstrated that global deletion of the cellular communicating network factor 2 (CCN2), previously known as connective tissue growth factor (CTGF) due to its role in the extracellular matrix formation, predisposes to early and lethal AAs development after Angiotensin II (Ang II) infusion in mice. Here, we detail the protocol to induce and detect AAs generation in inducible global CCN2 knockout mice after Ang II infusion which allow the characterization of CCN role in AA development and may help to the development of pharmacological target for AA treatment.

[Survey to know the perception of sexual education in adolescents]. / Encuesta para conocer la percepción sobre la educación sexual en adolescentes.

OBJECTIVE: Sex education is essential for an accurate approach of sexuality in adolescents, being more effective when it is carried out at early ages. This study aimed to estimate the sexual health knowledge and the information in this regard perceived by adolescents, as well as where it came from. METHODS: A cross-sectional observational study was performed in students from two public high schools in Castilla-La Mancha who were surveyed about sexual knowledge. Comparisons were made by using the Student's t test or Mann Whitney U test, the Chi-square test (or Fisher's exact test) depending on the variables. RESULTS: 248 students were included (68.1% from Madridejos and 31.9% from Herencia) with a mean age of 14.67±1.1 years. The 47.2% were women. Only 4.8% of the participants affirm to talk about sexuality with their parents. The 56% declared had consumed pornographic content, with a mean first access age of 12.8±1 years. Significant statistically differences were found between the information received in schools and in the familiar surroundings, being alcohol and drug intake, sexually transmitted infections (STI), and contraceptive methods more frequently treated at the educational centers. A 25% of the surveyed students who reported having performed sexual relations stated not having used condom. Approximately, half of the participants did not perceive a high risk of getting STIs (HIV, herpes and chlamydia) in the case of having sex without condom. CONCLUSIONS: There is a lack of sexual knowledge in adolescents, perceiving a noteworthy heterogeneity in the offered information depending on it is received in high school or in familiar surroundings.

OBJECTIVE: La educación sexual es un pilar fundamental para un correcto enfoque de la sexualidad en los adolescentes, siendo más efectiva si se realiza en edades tempranas. El objetivo de este trabajo se fundamentó en estimar los conocimientos de salud sexual y la información percibida por los adolescentes, así como de dónde provenía. METHODS: Se realizó un estudio observacional transversal en estudiantes de dos centros educativos públicos de Castilla-La Mancha a los que se les realizó una encuesta sobre conocimientos de sexualidad. Se hicieron comparaciones mediante t de student o U de Mann Whitney, o test de Chi-cuadrado (o test exacto de Fisher), según variables. RESULTS: Se incluyeron 248 encuestados/as (68,1% de Madridejos y 31,9% de Herencia) con una edad media de 14,67±1,1 años. El 47,2% fueron mujeres. Solo un 4,8% de los/as encuestados/as afirmaron hablar sobre sexualidad con sus progenitores. Un 56% afirmaron haber consumido contenido pornográfico, siendo la edad media de la primera visualización de 12,8±1 años. Se encontraron diferencias estadísticamente significativas entre la información recibida en los centros educativos y el ámbito familiar, tratándose más frecuentemente el consumo de alcohol y drogas, las infecciones de transmisión sexual (ITS) y los métodos anticonceptivos en los centros. Un cuarto de los/as encuestados/as que afirmaron haber tenido relaciones sexuales no habían utilizado preservativo. Aproximadamente, la mitad de los/as participantes no percibieron un riesgo alto de contraer ITS (VIH, herpes y clamidia) en el caso de mantener relaciones sexuales sin preservativo. CONCLUSIONS: Existe una falta de educación sexual en los/as adolescentes, percibiendo gran heterogeneidad en los conocimientos ofrecidos en los centros educativos y el ámbito familiar.

Safe and Successful Treatment With Pcsk9 Inhibitors in Hypercholesterolemia and Renal Transplantation: A Case Report.

BACKGROUND: Patients with chronic kidney disease (CKD) are more prone to cardiovascular disease (CVD) risks, including CVD-derived sudden death. Additionally, patients with CKD also develop lipid metabolism abnormalities. Proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors (PCSK9i) are drugs capable of reducing CVD risk in patients with CKD, but their efficacy is scarcely assessed in transplant patients. CASE PRESENTATION: Here, we report a case of a 74-year-old man undergoing nephrology follow-up for a cadaver donor kidney transplant. The patient described an atorvastatin allergic reaction after an acute coronary syndrome. Because the patient had a very high risk for CVD, alirocumab was substituted for atorvastatin. The patient showed a well-tolerated and effective response and stable everolimus levels. CONCLUSION: PCSK9i may be considered a pharmacologic option for treating lipid metabolism disorder and reducing low-density lipoprotein cholesterol in transplant recipients.

[Cross-sectional study on sleep habits and new technologies use in high school students.] / Estudio transversal sobre hábitos de sueño y nuevas tecnologías en estudiantes de ciclos formativos.

OBJECTIVE: During lasts years, the use of information and communication technologies (ICTs) have boomed up, as well as their related- harmful behaviours. Parallel, time and quality of sleep has reduced along current society, which implies negatively in health in medium and long-terms. The present study aims to evaluate the association between lifestyle habits and quality of sleep of a subpopulation of young students. METHODS: An observational transversal study has been performed in Certificate of Medium and Higher Education from a High school of Alcázar de San Juan (Ciudad Real, Spain), who filled up a survey related to their lifestyle habits and the use of ICTs. Furthermore, the survey also included several variables related to quality of sleep by using Pittsburgh test. Bivariate comparisons using student test or Mann-Whitney U test, or Chi-square or exact test depending on the variable were performed. Afterwards, logistic regression was also done. RESULTS: The study sample were 286 students (43.4% women) were included being 22.2±7.3 years old on average. 99.7% of them owned mobile phone, using it 42 hours per week. Average total score of Pittsburgh test was 6.4±3.5, being higher in women (7.36±3.8) than in men (5.62±3.1). Moreover, 51.7% of surveyed students suffered from sleep disorders, being associated to several risk factors, such us using mobile phone while lying and without light (OR=2.04; CI95% [1.12-3.73]), using mobile phone in the middle of the night (OR=1.9; CI95% [1.06-3.42]) and drinking and smoking (OR=2.28; CI95% [1.14-4.55]). On the other hand, practising sports was defined as protector factor (OR=0.43; CI95% [0.26-0.72]). CONCLUSIONS: More than half of surveyed suffer from sleep disorders, mainly derived from the inadequate use of ICTs, showing differences between genders.

OBJETIVO: Durante los últimos años se ha incrementado el uso de nuevas tecnologías de la información y comunicación (TIC), así como los comportamientos nocivos en relación a su uso. Paralelamente, se ha descrito un descenso en el tiempo y calidad del sueño, lo cual repercute negativamente en la salud a medio y largo plazo. El objetivo del presente estudio fue evaluar la asociación de los diferentes hábitos con la calidad del sueño de una subpoblación de jóvenes estudiantes. METODOS: Se realizó un estudio observacional transversal de ciclos formativos en un instituto de educación secundaria de Alcázar de San Juan (Ciudad Real), basado en una encuesta sobre hábitos de vida y uso de nuevas tecnologías, además de diferentes variables relacionadas con el sueño, evaluadas mediante el índice de Pittsburgh. Se hicieron comparaciones bivariadas mediante t de student o U de Mann Whitney, o test de Chi-cuadrado (o test exacto) según variables. Posteriormente se realizó regresión logística. RESULTADOS: La muestra estuvo compuesta por 286 estudiantes con una edad media de 22,2±7,3 y siendo mujeres un 43,4%. El 99,7% tenía teléfono móvil, el cual utilizaban de media 42 horas a la semana. La puntuación media del test de Pittsburgh fue de 6,4±3,5, siendo mayor en las mujeres (7,36±3,8) que en los hombres (5,62±3,1). El 51,7% de los encuestados presentaban trastornos de sueño, siendo los factores asociados el uso del móvil acostado y sin luz (OR=2,04; IC95% [1,12-3,73]), el uso del móvil en mitad de la noche (OR=1,9; IC95% [1,06-3,42]), el consumo de tabaco y alcohol (OR=2,28; IC95% [1,14-4,55]) y hacer deporte como factor protector (OR=0,43 IC95% [0,26-0,72]). CONCLUSIONES: Más de la mitad de los encuestados presentan trastornos de sueño, fundamentalmente debido a un uso inadecuado de las TIC, existiendo diferencias claras entre hombres y mujeres.

Systematic Review and Meta-Analysis: Prevalence of Non-Alcoholic Fatty Liver Disease and Liver Fibrosis in Patients with Inflammatory Bowel Disease.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is a common concomitant condition in patients with inflammatory bowel disease (IBD). We aim to assess the magnitude of this association. METHODS: We searched MEDLINE, EMBASE and Scopus libraries for the period up to February 2023 to identify studies reporting cohorts of IBD patients in which NALFLD was evaluated. RESULTS: Eighty-nine studies were analyzed. The overall prevalence of NAFLD was 24.4% (95%CI, 19.3-29.8) in IBD, 20.2% (18.3-22.3) in Crohn's disease and 18.5% (16.4-20.8) for ulcerative colitis. Higher prevalence was found in male compared to female patients, in full papers compared to abstracts, and in cross-sectional studies compared to prospective and retrospective ones. The prevalence of NAFLD in IBD has increased in studies published from 2015 onwards: 23.2% (21.5-24.9) vs. 17.8% (13.2-22.9). Diagnostic methods for NAFLD determined prevalence figures, being highest in patients assessed by controlled attenuation parameter (38.8%; 33.1-44.7) compared to ultrasonography (28.5%; 23.1-34.2) or other methods. The overall prevalence of fibrosis was 16.7% (12.2-21.7) but varied greatly according to the measurement method. CONCLUSION: One-quarter of patients with IBD might present with NAFLD worldwide. This proportion was higher in recent studies and in those that used current diagnostic methods.

[Evolución de marcadores de laboratorio en pacientes con detección persistente de SARS-CoV-2.] / Laboratory markers evolution in patients with persistent detection of SARS-CoV-2.

OBJECTIVE: The study of the evolution of certain biomarkers in patients with persistent detection of SARS-CoV-2 could determine the profile of the pathology that these patients may suffer. The objective of this study was to describe the evolution of different laboratory markers in patients with persistent detection of SARS-CoV-2, and determining these parameters were into reference values. METHODS: Patients were divided into two groups: the control group (G0) included patients with a positive direct test for SARS-CoV-2 followed by 2 negative, while the problem group (G1) included patients with at least 3 consecutive positive tests. The time between consecutive samples was five to twenty days, and only patients with negative serology were included. Demographic data, comorbidities, symptoms, radiology and hospitalization were collected, as well as data from analytic and blood gases. The comparison between the study groups was realized using the t-student and U Mann-Whitney test for quantitative variables, and the χ2 test for qualitative variables. Results with p<0.05 were taken as significant. RESULTS: Ninety patients were included, thirty-eight in G0 and fifty-two in G1. D-dimer decreased 10.20 times more in G0 patients, and normal levels of this parameter at t1 were 1.46 times more frequent in these patients. The percentage of lymphocytes increased sixteen times more in G0, and the normal values in t1 were 10.40 times more common in these patients. C-reactive protein decreased significantly in both groups, and lactate increased more in G1 patients. CONCLUSIONS: The results of the study suggest that some biomarkers evolve differently in patients with persistent detection of SARS-CoV-2, which may have significant clinical impact. This information could help to determine the main organs or systems affected, allowing to anticipate socio-sanitary measures to prevent or compensate these alterations.

OBJETIVO: El estudio de la evolución de algunos biomarcadores en pacientes con detección persistente de SARS-CoV-2 permitiría determinar el perfil de las patologías que podrían padecer. El objetivo de este estudio fue describir la evolución de distintos marcadores de laboratorio en pacientes con detección persistente de SARS-CoV-2 y estudiar los cambios en la proporción de pacientes con valores considerados como normales. METODOS: Los pacientes se dividieron en dos grupos: el grupo control (G0) incluyó pacientes con una prueba de detección de infección activa positiva para SARS-CoV-2 seguida de dos negativas, mientras que el grupo problema (G1) incluyó pacientes con al menos tres pruebas positivas consecutivas. El tiempo entre muestras consecutivas fue de cinco a veinte días, y se incluyeron solamente pacientes con serología negativa. Se recogieron datos demográficos, comorbilidades, sintomatología, radiología y hospitalización, así como los datos de las analíticas y las gasometrías. La comparación entre los grupos de estudio se realizó mediante el test t-student y U Mann-Whitney para variables cuantitativas, y el test de χ2 para variables cualitativas. Se tomaron como significativos resultados con p<0,05. RESULTADOS: Se incluyeron noventa pacientes, treinta y ocho en G0 y cincuenta y dos en G1. El dímero D descendió 10,20 veces más en pacientes G0, y los niveles normales de este parámetro en t1 fueron 1,46 veces más frecuentes en estos pacientes. El porcentaje de linfocitos se elevó dieciséis veces más en G0, y los valores normales en t1 fueron 10,40 veces más habituales en estos pacientes. La proteína C reactiva descendió de manera importante en ambos grupos, y el lactato aumentó más en pacientes G1. CONCLUSIONES: Los resultados del estudio sugieren que algunos biomarcadores evolucionan de manera diferente en pacientes con detección persistente de SARS-CoV-2, lo que podría tener importantes repercusiones clínicas. Esta información podría ayudar a determinar los principales órganos o sistemas afectados, permitiendo anticipar medidas sociosanitarias para prevenir o compensar estas alteraciones.

CCN2 Binds to Tubular Epithelial Cells in the Kidney.

Cellular communication network-2 (CCN2), also called connective tissue growth factor (CTGF), is considered a fibrotic biomarker and has been suggested as a potential therapeutic target for kidney pathologies. CCN2 is a matricellular protein with four distinct structural modules that can exert a dual function as a matricellular protein and as a growth factor. Previous experiments using surface plasmon resonance and cultured renal cells have demonstrated that the C-terminal module of CCN2 (CCN2(IV)) interacts with the epidermal growth factor receptor (EGFR). Moreover, CCN2(IV) activates proinflammatory and profibrotic responses in the mouse kidney. The aim of this paper was to locate the in vivo cellular CCN2/EGFR binding sites in the kidney. To this aim, the C-terminal module CCN2(IV) was labeled with a fluorophore (Cy5), and two different administration routes were employed. Both intraperitoneal and direct intra-renal injection of Cy5-CCN2(IV) in mice demonstrated that CCN2(IV) preferentially binds to the tubular epithelial cells, while no signal was detected in glomeruli. Moreover, co-localization of Cy5-CCN2(IV) binding and activated EGFR was found in tubules. In cultured tubular epithelial cells, live-cell confocal microscopy experiments showed that EGFR gene silencing blocked Cy5-CCN2(IV) binding to tubuloepithelial cells. These data clearly show the existence of CCN2/EGFR binding sites in the kidney, mainly in tubular epithelial cells. In conclusion, these studies show that circulating CCN2(IV) can directly bind and activate tubular cells, supporting the role of CCN2 as a growth factor involved in kidney damage progression.

Oxidative Stress and Cellular Senescence Are Involved in the Aging Kidney.

Chronic kidney disease (CKD) can be considered as a clinical model for premature aging. However, non-invasive biomarkers to detect early kidney damage and the onset of a senescent phenotype are lacking. Most of the preclinical senescence studies in aging have been done in very old mice. Furthermore, the precise characterization and over-time development of age-related senescence in the kidney remain unclear. To address these limitations, the age-related activation of cellular senescence-associated mechanisms and their correlation with early structural changes in the kidney were investigated in 3- to 18-month-old C57BL6 mice. Inflammatory cell infiltration was observed by 12 months, whereas tubular damage and collagen accumulation occurred later. Early activation of cellular-senescence-associated mechanisms was found in 12-month-old mice, characterized by activation of the DNA-damage-response (DDR), mainly in tubular cells; activation of the antioxidant NRF2 pathway; and klotho downregulation. However, induction of tubular-cell-cycle-arrest (CCA) and overexpression of renal senescent-associated secretory phenotype (SASP) components was only found in 18-month-old mice. In aging mice, both inflammation and oxidative stress (marked by elevated lipid peroxidation and NRF2 inactivation) remained increased. These findings support the hypothesis that prolonged DDR and CCA, loss of nephroprotective factors (klotho), and dysfunctional redox regulatory mechanisms (NRF2/antioxidant defense) can be early drivers of age-related kidney-damage progression.

Epigenetic Modulation of Gremlin-1/NOTCH Pathway in Experimental Crescentic Immune-Mediated Glomerulonephritis.

Crescentic glomerulonephritis is a devastating autoimmune disease that without early and properly treatment may rapidly progress to end-stage renal disease and death. Current immunosuppressive treatment provides limited efficacy and an important burden of adverse events. Epigenetic drugs are a source of novel therapeutic tools. Among them, bromodomain and extraterminal domain (BET) inhibitors (iBETs) block the interaction between bromodomains and acetylated proteins, including histones and transcription factors. iBETs have demonstrated protective effects on malignancy, inflammatory disorders and experimental kidney disease. Recently, Gremlin-1 was proposed as a urinary biomarker of disease progression in human anti-neutrophil cytoplasmic antibody (ANCA)-associated crescentic glomerulonephritis. We have now evaluated whether iBETs could regulate Gremlin-1 in experimental anti-glomerular basement membrane nephritis induced by nephrotoxic serum (NTS) in mice, a model resembling human crescentic glomerulonephritis. In NTS-injected mice, the iBET JQ1 inhibited renal Gremlin-1 overexpression and diminished glomerular damage, restoring podocyte numbers. Chromatin immunoprecipitation assay demonstrated BRD4 enrichment of the Grem-1 gene promoter in injured kidneys, consistent with Gremlin-1 epigenetic regulation. Moreover, JQ1 blocked BRD4 binding and inhibited Grem-1 gene transcription. The beneficial effect of iBETs was also mediated by modulation of NOTCH pathway. JQ1 inhibited the gene expression of the NOTCH effectors Hes-1 and Hey-1 in NTS-injured kidneys. Our results further support the role for epigenetic drugs, such as iBETs, in the treatment of rapidly progressive crescentic glomerulonephritis.

CCN2 (Cellular Communication Network Factor 2) Deletion Alters Vascular Integrity and Function Predisposing to Aneurysm Formation.

BACKGROUND: CCN2 (cellular communication network factor 2) is a matricellular protein involved in cell communication and microenvironmental signaling responses. CCN2 is known to be overexpressed in several cardiovascular diseases, but its role is not completely understood. METHODS: Here, CCN2 involvement in aortic wall homeostasis and response to vascular injury was investigated in inducible <i>Ccn2</i>-deficient mice, with induction of vascular damage by infusion of Ang II (angiotensin II; 15 days), which is known to upregulate CCN2 expression in the aorta. RESULTS: Ang II infusion in CCN2-silenced mice lead to 60% mortality within 10 days due to rapid development and rupture of aortic aneurysms, as evidenced by magnetic resonance imaging, echography, and histological examination. <i>Ccn2</i> deletion decreased systolic blood pressure and caused aortic structural and functional changes, including elastin layer disruption, smooth muscle cell alterations, augmented distensibility, and increased metalloproteinase activity, which were aggravated by Ang II administration. Gene ontology analysis of RNA sequencing data identified aldosterone biosynthesis as one of the most enriched terms in CCN2-deficient aortas. Consistently, treatment with the mineralocorticoid receptor antagonist spironolactone before and during Ang II infusion reduced aneurysm formation and mortality, underscoring the importance of the aldosterone pathway in Ang II-induced aorta pathology. CONCLUSIONS: CCN2 is critically involved in the functional and structural homeostasis of the aorta and in maintenance of its integrity under Ang II-induced stress, at least, in part, by disruption of the aldosterone pathway. Thus, this study opens new avenues to future studies in disorders associated to vascular pathologies.

Kidney microRNA Expression Pattern in Type 2 Diabetic Nephropathy in BTBR Ob/Ob Mice.

Diabetic nephropathy (DN) is the main leading cause of chronic kidney disease worldwide. Although remarkable therapeutic advances have been made during the last few years, there still exists a high residual risk of disease progression to end-stage renal failure. To further understand the pathogenesis of tissue injury in this disease, by means of the Next-Generation Sequencing, we have studied the microRNA (miRNA) differential expression pattern in kidneys of Black and Tan Brachyury (BTBR) ob/ob (leptin deficiency mutation) mouse. This experimental model of type 2 diabetes and obesity recapitulates the key histopathological features described in advanced human DN and therefore can provide potential useful translational information. The miRNA-seq analysis, performed in the renal cortex of 22-week-old BTBR ob/ob mice, pointed out a set of 99 miRNAs significantly increased compared to non-diabetic, non-obese control mice of the same age, whereas no miRNAs were significantly decreased. Among them, miR-802, miR-34a, miR-132, miR-101a, and mir-379 were the most upregulated ones in diabetic kidneys. The in silico prediction of potential targets for the 99 miRNAs highlighted inflammatory and immune processes, as the most relevant pathways, emphasizing the importance of inflammation in the pathogenesis of kidney damage associated to diabetes. Other identified top canonical pathways were adipogenesis (related with ectopic fatty accumulation), necroptosis (an inflammatory and regulated form of cell death), and epithelial-to-mesenchymal transition, the latter supporting the importance of tubular cell phenotype changes in the pathogenesis of DN. These findings could facilitate a better understanding of this complex disease and potentially open new avenues for the design of novel therapeutic approaches to DN.

Acute Kidney Injury is Aggravated in Aged Mice by the Exacerbation of Proinflammatory Processes.

Acute kidney injury (AKI) is more frequent in elderly patients. Mechanisms contributing to AKI (tubular cell death, inflammatory cell infiltration, impaired mitochondrial function, and prolonged cell-cycle arrest) have been linked to cellular senescence, a process implicated in regeneration failure and progression to fibrosis. However, the molecular and pathological basis of the age-related increase in AKI incidence is not completely understood. To explore these mechanisms, experimental AKI was induced by folic acid (FA) administration in young (3-months-old) and old (1-year-old) mice, and kidneys were evaluated in the early phase of AKI, at 48 h. Tubular damage score, KIM-1 expression, the recruitment of infiltrating immune cells (mainly neutrophils and macrophages) and proinflammatory gene expression were higher in AKI kidneys of old than of young mice. Tubular cell death in FA-AKI involves several pathways, such as regulated necrosis and apoptosis. Ferroptosis and necroptosis cell-death pathways were upregulated in old AKI kidneys. In contrast, caspase-3 activation was only found in young but not in old mice. Moreover, the antiapoptotic factor BCL-xL was significantly overexpressed in old, injured kidneys, suggesting an age-related apoptosis suppression. AKI kidneys displayed evidence of cellular senescence, such as increased levels of cyclin dependent kinase inhibitors p16ink4a and p21cip1, and of the DNA damage response marker γH2AX. Furthermore, p21cip1 mRNA expression and nuclear staining for p21cip1 and γH2AX were higher in old than in young FA-AKI mice, as well as the expression of senescence-associated secretory phenotype (SASP) components (Il-6, Tgfb1, Ctgf, and Serpine1). Interestingly, some infiltrating immune cells were p21 or γH2AX positive, suggesting that molecular senescence in the immune cells ("immunosenescence") are involved in the increased severity of AKI in old mice. In contrast, expression of renal protective factors was dramatically downregulated in old AKI mice, including the antiaging factor Klotho and the mitochondrial biogenesis driver PGC-1α. In conclusion, aging resulted in more severe AKI after the exposure to toxic compounds. This increased toxicity may be related to magnification of proinflammatory-related pathways in older mice, including a switch to a proinflammatory cell death (necroptosis) instead of apoptosis, and overactivation of cellular senescence of resident renal cells and infiltrating inflammatory cells.

Role of Macrophages and Related Cytokines in Kidney Disease.

Inflammation is a key characteristic of kidney disease, but this immune response is two-faced. In the acute phase of kidney injury, there is an activation of the immune cells to fight against the insult, contributing to kidney repair and regeneration. However, in chronic kidney diseases (CKD), immune cells that infiltrate the kidney play a deleterious role, actively participating in disease progression, and contributing to nephron loss and fibrosis. Importantly, CKD is a chronic inflammatory disease. In early CKD stages, patients present sub-clinical inflammation, activation of immune circulating cells and therefore, anti-inflammatory strategies have been proposed as a common therapeutic target for renal diseases. Recent studies have highlighted the plasticity of immune cells and the complexity of their functions. Among immune cells, monocytes/macrophages play an important role in all steps of kidney injury. However, the phenotype characterization between human and mice immune cells showed different markers; therefore the extrapolation of experimental studies in mice could not reflect human renal diseases. Here we will review the current information about the characteristics of different macrophage phenotypes, mainly focused on macrophage-related cytokines, with special attention to the chemokine CCL18, and its murine functional homolog CCL8, and the macrophage marker CD163, and their role in kidney pathology.