Our 7-year experience supporting the Ross autograft with the novel technique of Personalized External Aortic Root Support.

Objective: The Ross operation is a widely accepted option for aortic valve replacement in children, and evidence shows its excellent results in terms of hemodynamics and durability. However, indications are still limited due to the fact that it is a technically demanding procedure, only performed by specialized surgeons. On top of that, and despite numerous techniques being applied, autograft dilatation remains a key disadvantage, which can lead to graft failure. In recent years, the ExoVasc Personalized External Aortic Root Support (PEARS) has proven to be a safe and effective option to prevent aortic root dilatation in various aortopathies and is a technique that lends itself to support the pulmonary autograft in the Ross operation. Methods: During the past 7 years, we have used the ExoVasc PEARS graft, manufactured from the patients' pulmonary artery measurements from computed tomography scan data, to support the pulmonary autograft in the Ross operation. This graft (manufactured by Exstent Ltd, UK) is implanted at the same time as the autograft. We have reviewed all the patients who underwent this surgery, including demographic data, aorta measurements, operative data, and follow-up assessment consisting of periodic echocardiograms and magnetic resonance imaging scans. Results: Fifty patients were included in the study. Mean age at the time of the operation was 29.84 years, the youngest patient was 9 years-old. Nineteen patients (38%) had previous sternotomies; 11 of them having had a previous aortic valve replacement. Seventy-two percent of patients had initially a bicuspid aortic valve. Mean diameter of the ascending aorta was 3.83 cm. Forty-four percent of patients required a concomitant reduction aortoplasty due to mismatch sizes between the ascending aorta and the autograft. Mean bypass and crossclamp times were 200.66 and 151.14 minutes, respectively. Median length of stay was 6 days. Mean follow-up was 16.88 months. Two patients required subsequent aortic valve replacement (1 had rheumatic valve disease and the other had iatrogenic damage in his autograft valve leaflet). Ascending aorta dimensions remain stable when compared with immediate postoperative studies. There were no deaths. Conclusions: The ExoVasc PEARS graft has proven to be an excellent support in the Ross operation to prevent the autograft failure related to autograft dilatation that can offer several advantages compared with other existing techniques. With this type of support, we believe the Ross indications can be expanded to multiple clinical scenarios, given the good long-term results this operation offers in terms of durability, life expectancy, and hemodynamics.

The evolutionary conserved miR-137/325 tandem mediates obesity-induced hypogonadism and metabolic comorbidities by repressing hypothalamic kisspeptin.

BACKGROUND: Obesity-induced hypogonadism (OIH) is a prevalent, but often neglected condition in men, which aggravates the metabolic complications of overweight. While hypothalamic suppression of Kiss1-encoded kisspeptin has been suggested to contribute to OIH, the molecular mechanisms for such repression in obesity, and the therapeutic implications thereof, remain unknown. METHODS: A combination of bioinformatic, expression and functional analyses was implemented, assessing the role of the evolutionary-conserved miRNAs, miR-137 and miR-325, in mediating obesity-induced suppression of hypothalamic kisspeptin, as putative mechanism of central hypogonadism and metabolic comorbidities. The implications of such miR-137/325-kisspeptin interplay for therapeutic intervention in obesity were also explored using preclinical OIH models. RESULTS: MiR-137/325 repressed human KISS1 3'-UTR in-vitro and inhibited hypothalamic kisspeptin content in male rats, while miR-137/325 expression was up-regulated, and Kiss1/kisspeptin decreased, in the medio-basal hypothalamus of obese rats. Selective over-expression of miR-137 in Kiss1 neurons reduced Kiss1/ kisspeptin and partially replicated reproductive and metabolic alterations of OIH in lean mice. Conversely, interference of the repressive actions of miR-137/325 selectively on Kiss1 3'-UTR in vivo, using target-site blockers (TSB), enhanced kisspeptin content and reversed central hypogonadism in obese rats, together with improvement of glucose intolerance, insulin resistance and cardiovascular and inflammatory markers, despite persistent exposure to obesogenic diet. Reversal of OIH by TSB miR-137/325 was more effective than chronic kisspeptin or testosterone treatments in obese rats. CONCLUSIONS: Our data disclose that the miR-137/325-Kisspeptin repressive interaction is a major player in the pathogenesis of obesity-induced hypogonadism and a putative druggable target for improved management of this condition and its metabolic comorbidities in men suffering obesity. SIGNIFICANCE STATEMENT: Up to half of the men suffering obesity display also central hypogonadism, an often neglected complication of overweight that can aggravate the clinical course of obesity and its complications. The mechanisms for such obesity-induced hypogonadism remain poorly defined. We show here that the evolutionary conserved miR137/miR325 tandem centrally mediates obesity-induced hypogonadism via repression of the reproductive-stimulatory signal, kisspeptin; this may represent an amenable druggable target for improved management of hypogonadism and other metabolic complications of obesity.

Resolvin D2 prevents vascular remodeling, hypercontractility and endothelial dysfunction in obese hypertensive mice through modulation of vascular and proinflammatory factors.

During resolution of inflammation, specialized proresolving mediators (SPMs), including resolvins, are produced to restore tissue homeostasis. We hypothesized that there might be a dysregulation of SPMs pathways in pathological vascular remodeling and that resolvin D2 (RvD2) might prevent vascular remodeling and contractile and endothelial dysfunction in a model of obesity and hypertension. In aortic samples of patients with or without abdominal aortic aneurysms (AAA), we evaluated gene expression of enzymes involved in SPMs synthesis (ALOXs), SPMs receptors and pro-inflammatory genes. In an experimental model of aortic dilation induced by high fat diet (HFD, 60%, eighteen weeks) and angiotensin II (AngII) infusion (four weeks), we studied the effect of RvD2 administration in aorta and small mesenteric arteries structure and function and markers of inflammation. In human macrophages we evaluated the effects of AngII and RvD2 in macrophages function and SPMs profile. In patients, we found positive correlations between AAA and obesity, and between AAA and expression of ALOX15, RvD2 receptor GPR18, and pro-inflammatory genes. There was an inverse correlation between the expression of aortic ALOX15 and AAA growth rate. In the mice model, RvD2 partially prevented the HFD plus AngII-induced obesity and adipose tissue inflammation, hypertension, aortic and mesenteric arteries remodeling, hypercontratility and endothelial dysfunction, and the expression of vascular proinflammatory markers and cell apoptosis. In human macrophages, RvD2 prevented AngII-induced impaired efferocytosis and switched SPMs profile. RvD2 might represent a novel protective strategy in preventing vascular damage associated to hypertension and obesity likely through effects in vascular and immune cells.

Targeting cancer stem cell OXPHOS with tailored ruthenium complexes as a new anti-cancer strategy.

BACKGROUND: Previous studies by our group have shown that oxidative phosphorylation (OXPHOS) is the main pathway by which pancreatic cancer stem cells (CSCs) meet their energetic requirements; therefore, OXPHOS represents an Achille's heel of these highly tumorigenic cells. Unfortunately, therapies that target OXPHOS in CSCs are lacking. METHODS: The safety and anti-CSC activity of a ruthenium complex featuring bipyridine and terpyridine ligands and one coordination labile position (Ru1) were evaluated across primary pancreatic cancer cultures and in vivo, using 8 patient-derived xenografts (PDXs). RNAseq analysis followed by mitochondria-specific molecular assays were used to determine the mechanism of action. RESULTS: We show that Ru1 is capable of inhibiting CSC OXPHOS function in vitro, and more importantly, it presents excellent anti-cancer activity, with low toxicity, across a large panel of human pancreatic PDXs, as well as in colorectal cancer and osteosarcoma PDXs. Mechanistic studies suggest that this activity stems from Ru1 binding to the D-loop region of the mitochondrial DNA of CSCs, inhibiting OXPHOS complex-associated transcription, leading to reduced mitochondrial oxygen consumption, membrane potential, and ATP production, all of which are necessary for CSCs, which heavily depend on mitochondrial respiration. CONCLUSIONS: Overall, the coordination complex Ru1 represents not only an exciting new anti-cancer agent, but also a molecular tool to dissect the role of OXPHOS in CSCs. Results indicating that the compound is safe, non-toxic and highly effective in vivo are extremely exciting, and have allowed us to uncover unprecedented mechanistic possibilities to fight different cancer types based on targeting CSC OXPHOS.

Trabectedin modulates macrophage polarization in the tumor-microenvironment. Role of KV1.3 and KV1.5 channels.

Immune cells have an important role in the tumor-microenvironment. Macrophages may tune the immune response toward inflammatory or tolerance pathways. Tumor-associated macrophages (TAM) have a string of immunosuppressive functions and they are considered a therapeutic target in cancer. This study aimed to analyze the effects of trabectedin, an antitumor agent, on the tumor-microenvironment through the characterization of the electrophysiological and molecular phenotype of macrophages. Experiments were performed using the whole-cell configuration of the patch-clamp technique in resident peritoneal mouse macrophages. Trabectedin does not directly interact with KV1.5 and KV1.3 channels, but their treatment (16 h) with sub-cytotoxic concentrations of trabectedin increased their KV current due to an upregulation of KV1.3 channels. In vitro generated TAM (TAMiv) exhibited an M2-like phenotype. TAMiv generated a small KV current and express high levels of M2 markers. K+ current from TAMs isolated from tumors generated in mice is a mixture of KV and KCa, and in TAM isolated from tumors generated in trabectedin-treated mice, the current is mostly driven by KCa. We conclude that the antitumor capacity of trabectedin is not only due to its effects on tumor cells, but also to the modulation of the tumor microenvironment, due, at least in part, to the modulation of the expression of different macrophage ion channels.

How Frequently Benign Uterine Myomas Appear Suspicious for Sarcoma as Assessed by Transvaginal Ultrasound?

BACKGROUND: Uterine myomas may resemble uterine sarcomas in some cases. However, the rate of benign myomas appearing as sarcomas at an ultrasound examination is not known. The objective of this study is to determine the percentage of benign myomas that appear suspicious for uterine sarcoma on ultrasound examination. This is a prospective observational multicenter study (June 2019-December 2021) comprising a consecutive series of patients with histologically proven uterine myoma after hysterectomy or myomectomy who underwent transvaginal and/or transabdominal ultrasound prior to surgery. All ultrasound examinations were performed by expert examiners. MUSA criteria were used to describe the lesions (1). Suspicion of sarcoma was established when three or more sonographic features, described by Ludovisi et al. as "frequently seen in uterine sarcoma", were present (2). These features are no visible myometrium, irregular cystic areas, non-uniform echogenicity, irregular contour, "cooked" appearance, and a Doppler color score of 3-4. In addition, the examiners had to classify the lesion as suspicious based on her/his impression, independent of the number of features present. Eight hundred and ten women were included. The median maximum diameter of the myomas was 58.7 mm (range: 10.0-263.0 mm). Three hundred and forty-nine (43.1%) of the patients had more than one myoma. Using the criterion of >3 suspicious features, 40 (4.9%) of the myomas had suspicious appearance. By subjective impression, the examiners considered 40 (4.9%) cases suspicious. The cases were not exactly the same. We conclude that approximately 5% of benign uterine myomas may exhibit sonographic suspicion of sarcoma. Although it is a small percentage, it is not negligible.

Concomitant lymphoplasmacytic lymphoma, multiple myeloma, and amyloidosis: A diagnostic and therapeutic challenge.

We report a case based on simultaneous occurrence of Waldenström macroglobulinemia, myeloma and amyloidosis as a collision neoplasm. The strangeness and severity of the case presented a diagnostic and therapeutic challenge, which required individualised treatment and close follow-up to achieved stringent complete response.

KV 1.3 channels are novel determinants of macrophage-dependent endothelial dysfunction in angiotensin II-induced hypertension in mice.

BACKGROUND AND PURPOSE: KV 1.3 channels are expressed in vascular smooth muscle cells (VSMCs), where they contribute to proliferation rather than contraction and participate in vascular remodelling. KV 1.3 channels are also expressed in macrophages, where they assemble with KV 1.5 channels (KV 1.3/KV 1.5), whose activation generates a KV current. In macrophages, the KV 1.3/KV 1.5 ratio is increased by classical activation (M1). Whether these channels are involved in angiotensin II (AngII)-induced vascular remodelling, and whether they can modulate the macrophage phenotype in hypertension, remains unknown. We characterized the role of KV 1.3 channels in vascular damage in hypertension. EXPERIMENTAL APPROACH: We used AngII-infused mice treated with two selective KV 1.3 channel inhibitors (HsTX[R14A] and [EWSS]ShK). Vascular function and structure were measured using wire and pressure myography, respectively. VSMC and macrophage electrophysiology were studied using the patch-clamp technique; gene expression was analysed using RT-PCR. KEY RESULTS: AngII increased KV 1.3 channel expression in mice aorta and peritoneal macrophages which was abolished by HsTX[R14A] treatment. KV 1.3 inhibition did not prevent hypertension, vascular remodelling, or stiffness but corrected AngII-induced macrophage infiltration and endothelial dysfunction in the small mesenteric arteries and/or aorta, via a mechanism independent of electrophysiological changes in VSMCs. AngII modified the electrophysiological properties of peritoneal macrophages, indicating an M1-like activated state, with enhanced expression of proinflammatory cytokines that induced endothelial dysfunction. These effects were prevented by KV 1.3 blockade. CONCLUSIONS AND IMPLICATIONS: We unravelled a new role for KV 1.3 channels in the macrophage-dependent endothelial dysfunction induced by AngII in mice which might be due to modulation of macrophage phenotype.

Wilms Tumor 1b Expression Defines a Pro-regenerative Macrophage Subtype and Is Required for Organ Regeneration in the Zebrafish.

Organ regeneration is preceded by the recruitment of innate immune cells, which play an active role during repair and regrowth. Here, we studied macrophage subtypes during organ regeneration in the zebrafish, an animal model with a high regenerative capacity. We identified a macrophage subpopulation expressing Wilms tumor 1b (wt1b), which accumulates within regenerating tissues. This wt1b+ macrophage population exhibited an overall pro-regenerative gene expression profile and different migratory behavior compared to the remainder of the macrophages. Functional studies showed that wt1b regulates macrophage migration and retention at the injury area. Furthermore, wt1b-null mutant zebrafish presented signs of impaired macrophage differentiation, delayed fin growth upon caudal fin amputation, and reduced cardiomyocyte proliferation following cardiac injury that correlated with altered macrophage recruitment to the regenerating areas. We describe a pro-regenerative macrophage subtype in the zebrafish and a role for wt1b in organ regeneration.

Regulator of calcineurin 1 modulates vascular contractility and stiffness through the upregulation of COX-2-derived prostanoids.

Cyclooxygenase-2 (COX-2) derived-prostanoids participate in the altered vascular function and mechanical properties in cardiovascular diseases. We investigated whether regulator of calcineurin 1 (Rcan1) participates in vascular contractility and stiffness through the regulation of COX-2. For this, wild type (Rcan1+/+) and Rcan1-deficient (Rcan1-/-) mice untreated or treated with the COX-2 inhibitor rofecoxib were used. Vascular function and structure were analysed by myography. COX-2 and phospo-p65 expression were studied by western blotting and immunohistochemistry and TXA2 production by ELISA. We found that Rcan1 deficiency increases COX-2 and IL-6 expression and NF-κB activation in arteries and vascular smooth muscle cells (VSMC). Adenoviral-mediated re-expression of Rcan1.4 in Rcan1-/- VSMC normalized COX-2 expression. Phenylephrine-induced vasoconstrictor responses were greater in aorta from Rcan1-/- compared to Rcan1+/+ mice. This increased response were diminished by etoricoxib, furegrelate, SQ 29548, cyclosporine A and parthenolide, inhibitors of COX-2, TXA2 synthase, TP receptors, calcineurin and NF-κB, respectively. Endothelial removal and NOS inhibition increased phenylephrine responses only in Rcan1+/+ mice. TXA2 levels were greater in Rcan1-/- mice. In small mesenteric arteries, vascular function and structure were similar in both groups of mice; however, vessels from Rcan1-/- mice displayed an increase in vascular stiffness that was diminished by rofecoxib. In conclusion, our results suggest that Rcan1 might act as endogenous negative modulator of COX-2 expression and activity by inhibiting calcineurin and NF-kB pathways to maintain normal contractility and vascular stiffness in aorta and small mesenteric arteries, respectively. Our results uncover a new role for Rcan1 in vascular contractility and mechanical properties.

Mortality risk prediction in infective endocarditis surgery: reliability analysis of specific scores.

OBJECTIVES: We assessed the prognostic utility of risk scores in surgery for infective endocarditis (IE) to evaluate their reliability in mortality risk prediction. METHODS: An observational retrospective study was developed to include all patients who underwent surgery for active IE from 2002 to 2016. Classical and endocarditis-specific risk scores were calculated. RESULTS: A total of 180 patients were included in the study. The 30-day mortality rate was 26.82% [95% confidence interval (CI) 20.26-33.20%]. Classical risk scores were confirmed to have a suboptimal prognostic ability. Therefore, 4 IE-specific risk scores were calculated. Discrimination was evaluated using the area under the receiver operating characteristic curve. It was 0.76 (95% CI 0.68-0.82) for the Society of Thoracic Surgeons-IE (STS-IE) score; 0.68 (95% CI 0.58-0.76) for the De Feo-Cotrufo score; 0.73 (95% CI 0.66-0.79) for the PALSUSE score and 0.65 (95% CI 0.57-0.72) for the Costa score. The STS-IE score had higher discrimination when compared with the De Feo-Cotrufo score (P = 0.055) and the Costa score (P = 0.024); however, there was no significant difference when we compared the STS-IE score with the PALSUSE score (P = 0.58). Calibration was assessed using the Hosmer-Lemeshow test; an adequate calibration was confirmed in all 4 scores. CONCLUSIONS: Specific risk scores had better prognostic performance than classical risk scores. The STS-IE score had the highest discrimination and was adequately calibrated. The PALSUSE score also showed optimal discrimination and calibration. The De Feo-Cotrufo score had a lower discrimination in our sample; however, the De Feo-Cotrufo score is recommended in the current guidelines. The Costa score had the lowest discrimination.

Caracterização da evolução clínica dos recém-nascidos com gastrosquise em uma unidade de terapia intensiva neonatal de referência da américa latina / Characteristics of the clinical development of a newborn with gastroschisis in an intensive care unit in latin america

INTRODUCTION: Congenital malformations are major diseases observed at birth. They are the second most common cause of death in the neonatal population, the first one being prematurity. OBJECTIVE: To characterise the clinical outcome of newborns with gastroschisis (GS) in a neonatal intensive care unit. METHODS: A retrospective observational clinical study in 50 infants with GS using the association of intestinal abnormalities, impossibility of primary closure of the abdominal defect and reoperation necessity as classification criteria for the disease. The significance level was p < 0.05. RESULTS: The hospitalisation to primary surgery occurred with a median age of 2 hours. Fourteen percent of children were subjected to a primary silo interposition and 24% had associated intestinal malformation. Nineteen newborns (NB) required more than one surgery. The median length of stay was 33 days, higher in patients with complex GS (56 days). All NB recovered from urine output 48 hours after surgery and 40% had hyponatraemia and oligoanuria in this period. There was no difference between the natraemia and fasting time (p = 0.79). Weight gain was similar in both groups with total parenteral nutrition and became significantly higher in patients with simple GS after enteral feeding (p = 0.0046). These NB evolved 2.4 times less cholestasis. Late-onset sepsis occurred in 58% of patients and was related to the infection of the central venous catheter in 37.9% of cases. Mortality was higher in infants infected with complex GS and the overall mortality rate was 14%. CONCLUSION: Clinical characterisation of newborns with gastroschisis depends on the complexity and the knowledge and conduct of morbidities to reduce mortality.

INTRODUÇÃO: As malformações congênitas fazem parte das principais doenças observadas ao nascimento. Entre as causas de óbito no período neonatal as malformações foram a segunda causa, sendo ainda a primeira, a prematuridade. OBJETIVOS: Caracterizar a evolução clínica dos recém-nascidos (RN) com gastrosquise (GTQ) em uma unidade de terapia intensiva neonatal e descrever as morbidades renal, nutricional e infecciosa relacionados ao manejo clínico pós-natal na unidade de terapia intensiva neonatal MÉTODO: Foi realizado estudo observacional retrospectivo em 50 RN com GTQ, utilizando a associação de anormalidades intestinais, impossibilidade de fechamento primário do defeito abdominal e necesidade de reoperação como critérios de classificação para a doença. O nível de significância foi p < 0,05. RESULTADOS: A admissão hospitalar para cirurgia primária ocorreu com mediana de idade de 2 horas. O total de 14% das crianças foram submetidas a uma interposição de silo primária e 24% apresentaram malformação intestinal associada. Dezenove RN necessitaram mais de uma intervenção cirúrgica. A mediana do tempo de estadia foi de 33 dias, sendo maior nos pacientes com GTQ complexa (56 dias). Todos os RN recuperaram o débito urinário a partir de 48 horas do pós-operatório e 40% apresentaram hiponatremia e oligoanúria nesse período. Não houve diferença entre a natremia e o tempo de jejum (p = 0,79). O ganho ponderal foi similar em ambos os grupos com nutrição parenteral total e tornou-se significativamente maior nos pacientes com GTQ simples após a alimentação enteral (p = 0,0046). Esses RN evoluíram 2,4 vezes com menos colestase. Sepse tardia ocorreu em 58% dos pacientes e foi relacionada à infecção do CVC em 37,9% dos casos. A mortalidade foi maior nos RN infectados com GTQ complexa e a taxa global de mortalidade foi de 14%. CONCLUSÃO: A caracterização clínica dos RN com GTG depende da complexidade e do conhecimento e condução das morbidades para diminuir a mortalidade.

Otx2 is a target of N-myc and acts as a suppressor of sensory development in the mammalian cochlea.

Transcriptional regulatory networks are essential during the formation and differentiation of organs. The transcription factor N-myc is required for proper morphogenesis of the cochlea and to control correct patterning of the organ of Corti. We show here that the Otx2 gene, a mammalian ortholog of the Drosophila orthodenticle homeobox gene, is a crucial target of N-myc during inner ear development. Otx2 expression is lost in N-myc mouse mutants, and N-myc misexpression in the chick inner ear leads to ectopic expression of Otx2. Furthermore, Otx2 enhancer activity is increased by N-myc misexpression, indicating that N-myc may directly regulate Otx2. Inactivation of Otx2 in the mouse inner ear leads to ectopic expression of prosensory markers in non-sensory regions of the cochlear duct. Upon further differentiation, these domains give rise to an ectopic organ of Corti, together with the re-specification of non-sensory areas into sensory epithelia, and the loss of Reissner's membrane. Therefore, the Otx2-positive domain of the cochlear duct shows a striking competence to develop into a mirror-image copy of the organ of Corti. Taken together, these data show that Otx2 acts downstream of N-myc and is essential for patterning and spatial restriction of the sensory domain of the mammalian cochlea.

Resistance to pathogens in terpene down-regulated orange fruits inversely correlates with the accumulation of D-limonene in peel oil glands.

Volatile organic compounds (VOCs) are secondary metabolites acting as a language for the communication of plants with the environment. In orange fruits, the monoterpene D-limonene accumulates at very high levels in oil glands from the peel. Drastic down-regulation of D-limonene synthase gene expression in the peel of transgenic oranges harboring a D-limonene synthase transgene in antisense (AS) configuration altered the monoterpene profile in oil glands, mainly resulting in reduced accumulation of D-limonene. This led to fruit resistance against Penicillium digitatum (Pd), Xanthomonas citri subsp. citri (Xcc) and other specialized pathogens. Here, we analyze resistance to pathogens in independent AS and empty vector (EV) lines, which have low, medium or high D-limonene concentrations and show that the level of resistance is inversely related to the accumulation of D-limonene in orange peels, thus explaining the need of high D-limonene accumulation in mature oranges in nature for the efficient attraction of specialized microorganism frugivores.

The C-terminal module IV of connective tissue growth factor, through EGFR/Nox1 signaling, activates the NF-κB pathway and proinflammatory factors in vascular smooth muscle cells.

AIMS: Connective tissue growth factor (CTGF/CCN2) is a developmental gene upregulated in pathological conditions, including cardiovascular diseases, whose product is a matricellular protein that can be degraded to biologically active fragments. Among them, the C-terminal module IV [CCN2(IV)] regulates many cellular functions, but there are no data about redox process. Therefore, we investigated whether CCN2(IV) through redox signaling regulates vascular responses. RESULTS: CCN2(IV) increased superoxide anion (O2(•-)) production in murine aorta (ex vivo and in vivo) and in cultured vascular smooth muscle cells (VSMCs). In isolated murine aorta, CCN2(IV), via O2(•-), increased phenylephrine-induced vascular contraction. CCN2(IV) in vivo regulated several redox-related processes in mice aorta, including increased nonphagocytic NAD(P)H oxidases (Nox)1 activity, protein nitrosylation, endothelial dysfunction, and activation of the nuclear factor-κB (NF-κB) pathway and its related proinflammatory factors. The role of Nox1 in CCN2(IV)-mediated vascular responses in vivo was investigated by gene silencing. The administration of a Nox1 morpholino diminished aortic O2(•-) production, endothelial dysfunction, NF-κB activation, and overexpression of proinflammatory genes in CCN2(IV)-injected mice. The link CCN2(IV)/Nox1/NF-κB/inflammation was confirmed in cultured VSMCs. Epidermal growth factor receptor (EGFR) is a known CCN2 receptor. In VSMCs, CCN2(IV) activates EGFR signaling. Moreover, EGFR kinase inhibition blocked vascular responses in CCN2(IV)-injected mice. INNOVATION AND CONCLUSION: CCN2(IV) is a novel prooxidant factor that in VSMCs induces O2(•-) production via EGFR/Nox1 activation. Our in vivo data demonstrate that CCN2(IV) through EGFR/Nox1 signaling pathway induces endothelial dysfunction and activation of the NF-κB inflammatory pathway. Therefore, CCN2(IV) could be considered a potential therapeutic target for redox-related cardiovascular diseases.

TLR-2 and TLR-4 expression in monocytes of newborns with late-onset sepsis, / Expressão do TLR-2 e do TLR-4 em monócitos de recém-nascidos a termo com sepse tardia

Objective: To analyze toll-like receptor (TLR)-2 and TLR-4 expression in monocytes of newborns with late-onset sepsis. Methods: This prospective study included 27 full-term newborns aged 8 to 29 days, with clinical and laboratory diagnosis of late-onset sepsis. Ten newborns (37%) had positive cultures. Cytokines were measured by cytometric bead array in peripheral blood, while TLR-2, TLR-4 expression, and median fluorescence intensity (MFI) were determined by immunophenotyping peripheral whole blood monocytes, and were analyzed with a BD FACSDiva flow cytometer (Becton, Dickinson and Company, USA). A comparison was performed with healthy adults. Results: Microorganisms were identified in 37% of these septic newborns, and all of them had high levels of pro-inflammatory cytokines (IL-8, IL-6, IL-1β) and anti-inflammatory cytokine (IL-10) corroborating the inflammatory/septic process. In monocytes, the frequency of TLR-4 expression was higher in infected newborns (p = 0.01). Conclusion: This study investigated the innate immune response in septic newborns. Septic newborns that relied almost exclusively on the innate immune system showed little in vivo response at monocyte activation, suggesting impaired immune response and increased susceptibility to infection. .

Objetivos: Analisar a expressão dos TLR-2 e TLR-4 em monócitos de recém-nascidos com sepse tardia. Métodos: Trata-se de um estudo prospectivo com 27 recém-nascidos a termo entre 8 e 29 dias de vida com diagnóstico clínico e laboratorial de sepse tardia dos quais dez (37%) apresentaram cultura positiva. As citocinas foram determinadas por teste de CBA em sangue periférico enquanto que a expressão e MFI (mediana de intensidade de fluorescência) dos TLR-2 e TLR-4 foi determinado por imunofenotipagem em monócitos de sangue periférico total através de análise pelo citômetro de fluxo BD FACSDiva. O grupo usado para comparação foi de adultos saudáveis. Resultados: Microrganismos foram identificados em 37% dos pacientes e estes juntamente com os pacientes com sepse clínica tiveram níveis elevados de citocinas pró-inflamatórias (IL-8, IL-6, IL-1β) e de citocina anti-inflamatória (IL-10) corroborando o processo inflamatório/infeccioso. No monócito, a frequência de expressão do TLR-4 foi mais elevada (p = 0,01). Conclusões: Este estudo analisou a resposta imune inata no recém-nascido com sepse. Recémnascidos sépticos que dependem quase exclusivamente do sistema imune inato apresentaram pouca resposta in vivo na ativação de monócitos o que sugere uma resposta imune deficiente e maior susceptibilidade à infecção. .

Gremlin activates the Smad pathway linked to epithelial mesenchymal transdifferentiation in cultured tubular epithelial cells.

Gremlin is a developmental gene upregulated in human chronic kidney disease and in renal cells in response to transforming growth factor-ß (TGF-ß). Epithelial mesenchymal transition (EMT) is one process involved in renal fibrosis. In tubular epithelial cells we have recently described that Gremlin induces EMT and acts as a downstream TGF-ß mediator. Our aim was to investigate whether Gremlin participates in EMT by the regulation of the Smad pathway. Stimulation of human tubular epithelial cells (HK2) with Gremlin caused an early activation of the Smad signaling pathway (Smad 2/3 phosphorylation, nuclear translocation, and Smad-dependent gene transcription). The blockade of TGF-ß, by a neutralizing antibody against active TGF-ß, did not modify Gremlin-induced early Smad activation. These data show that Gremlin directly, by a TGF-ß independent process, activates the Smad pathway. In tubular epithelial cells long-term incubation with Gremlin increased TGF-ß production and caused a sustained Smad activation and a phenotype conversion into myofibroblasts-like cells. Smad 7 overexpression, which blocks Smad 2/3 activation, diminished EMT changes observed in Gremlin-transfected tubuloepithelial cells. TGF-ß neutralization also diminished Gremlin-induced EMT changes. In conclusion, we propose that Gremlin could participate in renal fibrosis by inducing EMT in tubular epithelial cells through activation of Smad pathway and induction of TGF-ß.