Boldine prevents the inflammatory response of cardiac fibroblasts induced by SGK1-NFκB signaling pathway activation.

Cardiac fibroblasts (CF) are mesenchymal-type cells responsible for maintaining the homeostasis of the heart's extracellular matrix (ECM). Their dysfunction leads to excessive secretion of ECM proteins, tissue stiffening, impaired nutrient and oxygen exchange, and electrical abnormalities in the heart. Additionally, CF act as sentinel cells in the cardiac tissue microenvironment, responding to various stimuli that may affect heart function. Deleterious stimuli induce an inflammatory response in CF, increasing the secretion of cytokines such as IL-1ß and TNF-α and the expression of cell adhesion molecules like ICAM1 and VCAM1, initially promoting damage resolution by recruiting immune cells. However, constant harmful stimuli lead to a chronic inflammatory process and heart dysfunction. Therefore, it is necessary to study the mechanisms that govern CF inflammation. NFκB is a key regulator of the cardiac inflammatory process, making the search for mechanisms of NFκB regulation and CF inflammatory response crucial for developing new treatment options for cardiovascular diseases. SGK1, a serine-threonine protein kinase, is one of the regulators of NFκB and is involved in the fibrotic effects of angiotensin II and aldosterone, as well as in CF differentiation. However, its role in the CF inflammatory response is unknown. On the other hand, many bioactive natural products have demonstrated anti-inflammatory effects, but their role in CF inflammation is unknown. One such molecule is boldine, an alkaloid obtained from Boldo (Peumus boldus), a Chilean endemic tree with proven cytoprotective effects. However, its involvement in the regulation of SGK1 and CF inflammation is unknown. In this study, we evaluated the role of SGK1 and boldine in the inflammatory response in CF isolated from neonatal Sprague-Dawley rats. The involvement of SGK1 was analyzed using GSK650394, a specific SGK1 inhibitor. Our results demonstrate that SGK1 is crucial for LPS- and IFN-γ-induced inflammatory responses in CF (cytokine expression, cell adhesion molecule expression, and leukocyte adhesion). Furthermore, a conditioned medium (intracellular content of CF subject to freeze/thaw cycles) was used to simulate a sterile inflammation condition. The conditioned medium induced a potent inflammatory response in CF, which was completely prevented by the SGK1 inhibitor. Finally, our results indicate that boldine inhibits both SGK1 activation and the CF inflammatory response induced by LPS, IFN-γ, and CF-conditioned medium. Taken together, our results position SGK1 as an important regulator of the CF inflammatory response and boldine as a promising anti-inflammatory drug in the context of cardiovascular diseases.

In cardiac fibroblasts, interferon-beta attenuates differentiation, collagen synthesis, and TGF-ß1-induced collagen gel contraction.

Cardiac fibroblasts (CF) play a key role in the homeostasis of the extracellular matrix in cardiac tissue and are newly recognized as inflammatory supporter cells. Besides, CF-to-Cardiac myofibroblast differentiation is commanded by TGF-b, through SMAD signaling pathways, and these last cells are strongly implicated in cardiac fibrosis. In the heart IFN-ß is produced by CF; however, the role of IFN-ß, STAT proteins, and STAT-homo or heterodimers in the regulation of CF function with or without a fibrotic environment is unknown. CF were isolated from hearts of adult rats, and by western blot analysis we studied STAT1, STAT2, and STAT3 phosphorylation and through specific siRNA against these proteins we analyzed their role in CF functions such as differentiation (α-SMA expression); and pro-collagen type-I synthesis and secretion expression levels; collagen gels contraction and CF migration. In cultured adult rats CF, IFN-ß increases phosphorylation of STAT1, STAT2, and STAT3. Both STAT1 and STAT2 were involved in decreasing α-SMA and CF migration induced by TGF-ß1. Also, IFN-ß through STAT1 regulated pro-collagen type-I protein expression levels, and collagen gels contraction induced by TGF-ß1. STAT3 was not involved in any effects of IFN-ß studied. In conclusion, IFN-ß through STAT1 and STAT2 shows antifibrotic effects on CF TGF-ß1-treated, whereas STAT3 did not participate in such effect.

TGF-ß1 decreases CHOP expression and prevents cardiac fibroblast apoptosis induced by endoplasmic reticulum stress.

Transforming growth factor-beta 1 (TGF-ß1) is a cytokine with marked pro-fibrotic action on cardiac fibroblasts (CF). TGF-ß1 induces CF-to-cardiac myofibroblast (CMF) differentiation, defined by an increase in α-smooth muscle cells (α-SMA), collagen secretion and it has a cytoprotective effect against stimuli that induce apoptosis. In the Endoplasmic Reticulum (ER) lumen, misfolded protein accumulation triggers ER stress and induces apoptosis, and this process plays a critical role in cell death mediated by Ischemia/Reperfusion (I/R) injury and by ER stress inducers, such as Tunicamycin (Tn). Here, we studied the regulation of CHOP, a proapoptotic ER-stress-related transcription factor in CF under simulated I/R (sI/R) or exposed to Tn. Even though TGF-ß1 has been shown to participate in ER stress, its regulatory effect on CF apoptosis and ER stress-induced by sI/R or TN has not been evaluated yet. CF from neonatal rats were exposed to sI/R, and cell death was evaluated by cell count and apoptosis by flow cytometry. ER stress was assessed by western blot against CHOP. Our results evidenced that sI/R (8/24) h or Tn triggers CF apoptosis and an increase in CHOP protein levels. TGF-ß1 pre-treatment partially prevented apoptosis induced by sI/R or Tn. Furthermore, TGF-ß1 pre-treatment completely prevented CHOP increase by sI/R or Tn. Additionally, we found a decrease in α-SMA expression induced by sI/R and in collagen secretion induced by Tn, which were not prevented by TGF-ß1 treatment. In conclusion, TGF-ß1 partially protects CF apoptosis induced by sI/R or Tn, through a mechanism that would involve ER stress.

Evaluation of a novel infra-red endoscopy system in the assessment of early neoplasia in Barretts esophagus: pilot study from a single center.

Infrared endoscopy (IRE) has been shown to be useful in detecting submucosal (SM) invasion in early gastric cancer. Its role in the endoscopic assessment of Barrett's neoplasia has not been reported to date. We aimed in this study to evaluate the role of IRE in the detection and characterization of early neoplastic lesions within Barrett's esophagus (BE). The secondary aim was to explore its usefulness for the assessment of the presence of submucosal invasion in these early neoplastic Barrett's lesions. We included in the study patients with dysplastic BE who were referred to our institution for endoscopic therapy of a previously diagnosed early Barrett's neoplasia. An examination with white light high resolution endoscopy (HRE) and near IRE after intravenous injection of indocyanine green was performed for all patients using an infrared endoscope prototype. Staining on IRE and correlation with final histological staging by endoscopic mucosal resection/surgery or histological diagnosis on mapping biopsies was analyzed. A total of 23 patients were enrolled in our study: 17 of them with 19 visible lesions and 6 patients with flat BE and no lesions. Staining on IRE was noted in 18 cases: 17 (94%) had at least high grade dysplasia (HGD). No stain was noted in 7 cases: final histology was

Incidence of metachronous visible lesions in patients referred for radiofrequency ablation (RFA) therapy for early Barrett's neoplasia: a single-centre experience.

OBJECTIVE: Evaluate the incidence of metachronous visible lesions (VLs) in patients referred for radiofrequency ablation (RFA) for early Barrett's neoplasia. DESIGN: This study was conducted as part of the service evaluation audit. SETTING: Tertiary referral centre. PATIENTS: All patients with dysplastic Barrett's oesophagus referred for RFA were included for analysis. White light high-resolution endoscopy (HRE), autofluorescence imaging and narrow band imaging were sequentially performed. Endoscopic mucosal resection (EMR) was performed for all VL. Three to six months after EMR, all patients underwent initial RFA and then repeat RFA procedures at three monthly intervals. INTERVENTIONS: All endoscopy reports and final staging by EMR/surgery were evaluated and included for analysis. RESULTS: Fifty patients were analysed; median age 73 years, 84% men. 38/50 patients (76%) had a previous EMR due to the presence of VL before referred for ablation; twelve patients had no previous treatment. In total, 151 ablation procedures were performed, median per patient 2.68. Twenty metachronous VL were identified in 14 patients before the first ablation or during the RFA protocol; incidence was 28%. All metachronous lesions were successfully resected by EMR. Upstaging after rescue EMR compared with the initial histology was observed in four patients (28%). CONCLUSIONS: In total, 28% of patients enrolled in the RFA programme were diagnosed to have metachronous lesions. This high-incidence rate highlights the importance of a meticulous examination to identify and resect any VL before every ablation session. RFA treatment for early Barrett's neoplasia should be performed in tertiary referral centres with HRE and EMR facilities and expertise.

Cardiac fibroblasts as sentinel cells in cardiac tissue: Receptors, signaling pathways and cellular functions.

Cardiac fibroblasts (CF) not only modulate extracellular matrix (ECM) proteins homeostasis, but also respond to chemical and mechanical signals. CF express a variety of receptors through which they modulate the proliferation/cell death, autophagy, adhesion, migration, turnover of ECM, expression of cytokines, chemokines, growth factors and differentiation into cardiac myofibroblasts (CMF). Differentiation of CF to CMF involves changes in the expression levels of various receptors, as well as, changes in cell phenotype and their associated functions. CF and CMF express the ß2-adrenergic receptor, and its stimulation activates PKA and EPAC proteins, which differentially modulate the CF and CMF functions mentioned above. CF and CMF also express different levels of Angiotensin II receptors, in particular, AT1R activation increases collagen synthesis and cell proliferation, but its overexpression activates apoptosis. CF and CMF express different levels of B1 and B2 kinin receptors, whose stimulation by their respective agonists activates common signaling transduction pathways that decrease the synthesis and secretion of collagen through nitric oxide and prostacyclin I2 secretion. Besides these classical functions, CF can also participate in the inflammatory response of cardiac repair, through the expression of receptors commonly associated to immune cells such as Toll like receptor 4, NLRP3 and interferon receptor. The activation by their respective agonists modulates the cellular functions already described and the release of cytokines and chemokines. Thus, CF and CMF act as sentinel cells responding to a plethora of stimulus, modifying their own behavior, and that of neighboring cells.

4-Phenylbutyric acid prevent cytotoxicity induced by thapsigargin in rat cardiac fibroblast.

Cardiac fibroblast (CF) survival is important for the maintenance of the extracellular matrix homeostasis in the heart; providing a functional support to cardiomyocytes necessary for the correct myocardial function. Endoplasmic reticulum (ER) stress causes cellular dysfunction and cell death by apoptosis; and thapsigargin is a well-known ER stress inducer. On the other hand, the chemical chaperone, 4-phenylbutyric acid (4-PBA) had showed to prevent ER stress; however, in cardiac fibroblast both the ER stress induced by thapsigargin and prevention by 4-PBA, have not been studied in detail. Neonate rat CF were treated with thapsigargin in presence or absence of 4-PBA, and cell viability was evaluated by trypan blue exclusion and apoptosis by flow cytometry; whereas CHOP, BIP, PDI, ATF4 and procollagen protein levels were assessed by western blot. In CF, thapsigargin triggered the unfolded protein response detected by early increases in ATF4, CHOP, PDI and BIP protein levels as well as, the accumulation of intracellular procollagen. Thapsigargin also stimulated CF death in a time and concentration-dependent manner. ER stress, CF death and apoptosis induced by thapsigargin were prevented by 4-PBA. Conclusion our data suggest that 4-PBA prevent ER stress, intracellular procollagen accumulation, CF death and apoptosis induced by thapsigargin.

Effect of awake Carotid Endarterectomy under local anaesthesia on peri-operative blood pressure: blood pressure is normalised when carotid stenosis is treated under local anaesthesia.

BACKGROUND: Carotid Endarterectomy can be performed under local, regional or general anaesthesia. One of the most important effects of the type of anaesthetic used is on the systemic blood pressure. Although variations in blood pressure during and following carotid endarterectomy have been studied previously, the effects of awake carotid endarterectomy under local anaesthesia on blood pressure and its comparison with similar procedures under similar types of anaesthesia have not. METHODS: Peri-operative blood pressure measurements were collected from the records of 25 consecutive patients for each of the following five procedures; Carotid Endarterectomy under general anaesthesia (CEAGA), Anterior Cervical Discectomy and Fusion under general anaesthesia (ACDF), Cerebral Angiography under local anaesthesia (ANG), Carotid Endarterectomy patients under local anaesthesia who were symptomatic (CEALAS) and Carotid Endarterectomy patients under local anaesthesia who were asymptomatic (CEALAA). The recordings were then analysed to find out if there were any clinically significant variations in peri-operative blood pressure. FINDINGS: There is a significant and consistent difference when the pre-operative value was compared with the 4 hour and 24 hour post-operative recordings between the local and general anaesthetic groups for carotid endarterectomy. Carotid endarterectomy reduces the systolic and diastolic blood pressures post-operatively when performed under local anaesthesia and only the diastolic pressure was reduced when performed under general anaesthesia. CONCLUSION: The study provides evidence about the effect of carotid endarterectomy on the systemic blood pressure and its variations when performed under different types of anaesthesia. There is significant post-operative reduction in both the systolic and diastolic blood pressure values and the intraoperative fluctuation is minimal when local anaesthesia is used. Further studies are required to find out how this affects the long-term blood pressure and clinical outcome of the patient.

Bispectral index monitoring may not reliably indicate cerebral ischaemia during awake carotid endarterectomy.

BACKGROUND: Intraoperative ischaemia during carotid cross-clamping in patients undergoing carotid endarterectomy (CEA) is a major complication and prompt recognition of insufficient collateral blood supply is crucial. Electroencephalogram (EEG) is believed to be one of the useful forms of monitoring cerebrovascular insufficiency during CEA. The aim of this study was to evaluate the utility of bispectral index (BIS) monitoring, a processed EEG parameter, for the reliable detection of intraoperative cerebral ischaemia during awake CEA. METHODS: We monitored 52 patients continuously with the BIS monitor together with assessment of neurological function (contralateral upper and lower limb strength and the verbal component of the Glasgow Coma Scale for speech) in patients undergoing awake CEA. RESULTS: Overall mean BIS value in all patients was 96 (SD 2.9). In five patients who showed clinical evidence of cortical ischaemia during carotid cross-clamping, there was no change in the original range of BIS values throughout the procedure (96.7 [3.2]). In one patient BIS values decreased to 38 about 5 min after the incision and recovered within the next 10 min. The mean BIS value in the remaining 46 patients who did not develop clinical signs of ischaemia was 95.4 (2.6). Three cases are presented which demonstrate the inability of the BIS monitor to detect cerebral ischaemia. CONCLUSIONS: Lack of correlation of BIS with the signs of cerebral ischaemia during CEA makes it unreliable for detection of cerebrovascular insufficiency. We conclude that awake neurological testing is the preferred method of monitoring in these patients.

[Validation of a version in Spanish of the Yale-Brown Obsessive-Compulsive Scale]. / Validación de una versión en español de la Escala Yale-Brown para el Trastorno Obsesivo-Compulsivo.

INTRODUCTION: The objective of this study is to validate a version in Spanish of the Yale-Brown Obsessive-Compulsive Scale (Y-BOCS). METHOD: We administered to 20 patients with obsessive-compulsive disorder (OCD) the Maudsley Obsessive-Compulsive Inventory (MOI) and a version in Spanish of the Y-BOCS. The application of the last one was performed by an interviewer, filmed, and then watched and rated by other 4 observers. In addition, the opinion of the treating psychiatrist about the severity of the patient's symptoms was registered. The interviewer and the observers were named raters. RESULTS: The mean scores of the Y-BOCS assigned by the 4 observers were from 18.35 to 19.55; the maximum scores, from 34 to 35; and the minimum scores, from 1 to 5. The Pearson's correlations between raters were significant for all cases (r>0.86 and p<0.001). For all raters, Y-BOCS scores significantly correlated with MOI scores (p<0.001) and the opinion of the treating psychiatrist (p<0.05). We did not find significant correlation between the last one and MOI scores. The Cronbach's coefficients ranged from 0.8789 to 0.9488. CONCLUSIONS: The Spanish version of the Y-BOCS is reliable, homogeneous and valid for quantification of the obsessive-compulsive symptoms, and has more correlation with the opinion of the treating psychiatrist than the MOI.

Comparative in vitro activity of moxifloxacin by E-test against Streptococcus pyogenes.

Macrolides are currently used to treat Streptococcus pyogenes infections where allergy or resistance prevents the use of penicillin. However, growing macrolide resistance is now seen worldwide, with rates of 5%-40% being reported. In this context it is therefore important to have other therapeutic options. The aim of this study was to ascertain the potential role of moxifloxacin, a third-generation fluoroquinolone, in the treatment of infections caused by group A S. pyogenes. The antimicrobial susceptibilities of S. pyogenes isolated from 197 adult patients with pharyngotonsillitis were analyzed by the E-test. Twelve percent of the isolates were resistant to macrolides, and 5% showed diminished susceptibility toward penicillin; none of the strains were resistant to cefotaxime or to moxifloxacin (90% minimum inhibitory concentration, 0.25 microg/mL). Therefore, moxifloxacin may be a therapeutic option in the management of S. pyogenes infections when penicillin cannot be used or when macrolide resistance may be a local issue. Clinical studies of moxifloxacin in pharyngotonsillitis are warranted.