Long-term disability after cerebral ischemic stroke following a Bothrops atrox snakebite in the Brazilian Amazon.

Bothrops atrox envenomations in the Brazilian Amazon are responsible for a number of local and systemic effects. Among these, stroke presents the worst prognosis for the patient since it may evolve into disabilities and/or premature death. This complication is caused by coagulation disorders and generates hemorrhagic and thrombotic conditions. This study presents a case report of a 54-year-old female patient who presented extensive cerebral ischemia after a B. atrox envenomation that occurred in the state of Amazonas, Brazil. The patient was hospitalized for 102 days, which included a stay in the intensive care unit. Clinical and laboratory findings indicated a thrombogenic coagulopathy. On discharge, the patient had no verbal response, partial motor response, and right hemiplegia. The assessment carried out four years after discharge evidenced incapacitation, global aphasia and bilateral lower and upper limbs showed hypotrophy with a global decrease in strength. Ischemic stroke is a possible complication of B. atrox snakebites even after antivenom treatment, with the potential to cause debilitating long-term consequences.

Inflammatory Profile Associated with Secondary Infection from Bothrops atrox Snakebites in the Brazilian Amazon.

Bothrops snakebite envenomation (SBE) is consider an important health problem in Brazil, where Bothrops atrox is mainly responsible in the Brazilian Amazon. Local effects represent a relevant clinical issue, in which inflammatory signs and symptoms in the bite site represent a potential risk for short and long-term disabilities. Among local complications, secondary infections (SIs) are a common clinical finding during Bothrops atrox SBE and are described by the appearance of signs such as abscess, cellulitis or necrotizing fasciitis in the affected site. However, the influence of SI in the local events is still poorly understood. Therefore, the present study describes for the first time the impact of SBE wound infection on local manifestations and inflammatory response from patients of Bothrops atrox SBE in the Brazilian Amazon. This was an observational study carried out at the Fundação de Medicina Tropical Dr. Heitor Vieira Dourado, Manaus (Brazil), involving victims of Bothrops SBE. Clinical and laboratorial data were collected along with blood samples for the quantification of circulating cytokines and chemokines before antivenom administrations (T0) and 24 h (T1), 48 h (T2), 72 h (T3) and 7 days after (T4). From the 94 patients included in this study, 42 presented SI (44.7%) and 52 were without SI (NSI, 55.3%). Patients classified as moderate envenoming presented an increased risk of developing SI (OR = 2.69; CI 95% = 1.08-6.66, p = 0.033), while patients with bites in hands showed a lower risk (OR = 0.20; CI 95% = 0.04-0.96, p = 0.045). During follow-up, SI patients presented a worsening of local temperature along with a sustained profile of edema and pain, while NSI patients showed a tendency to restore and were highlighted in patients where SI was diagnosed at T2. As for laboratorial parameters, leukocytes, erythrocyte sedimentation ratio, fibrinogen and C-reactive protein were found increased in patients with SI and more frequently in patients diagnosed with SI at T3. Higher levels of circulating IL-2, IL-10, IL-6, TNF, INF-γ and CXCL-10 were observed in SI patients along with marked correlations between these mediators and IL-4 and IL-17, showing a plurality in the profile with a mix of Th1/Th2/Th17 response. The present study reports for the first time the synergistic effects of local infection and envenoming on the inflammatory response represented by local manifestations, which reflected on laboratorial parameters and inflammatory mediators and thus help improve the clinical management of SI associated to Bothrops SBE.

Exploring the Profile of Cell Populations and Soluble Immunological Mediators in Bothrops atrox Envenomations.

Bothrops atrox envenomations are common in the Brazilian Amazon. The venom of B. atrox is highly inflammatory, which results in severe local complications, including the formation of blisters. Moreover, there is little information on the immune mechanisms associated with this condition. Thus, a longitudinal study was carried out to characterize the profile of the cell populations and soluble immunological mediators in the peripheral blood and blisters in B. atrox patients s according to their clinical manifestations (mild and severe). A similar response in both B. atrox patient groups (MILD and SEV) was observed, with an increase in inflammatory monocytes, NKT, and T and B cells, as well as CCL2, CCL5, CXCL9, CXCL10, IL-1ß and IL-10, when compared with the group of healthy blood donors. After the administration of antivenom, the participation of patrolling monocytes and IL-10 in the MILD group was observed. In the SEV group, the participation of B cells was observed, with high levels of CCL2 and IL-6. In the blister exudate, a hyperinflammatory profile was observed. In conclusion, we revealed the involvement of cell populations and soluble mediators in the immune response to B. atrox envenomation at the local and peripheral level, which is related to the onset and extent of the inflammation/clinical manifestation.

Prostatic artery embolization in people with spinal cord injury: a safe and effective technique to ease intermittent catheterization in case of concomitant benign prostatic hyperplasia.

STUDY DESIGN: Case series. OBJECTIVES: To assess the safety and efficacy of prostatic artery embolization (PAE) to reduce prostate volume (PV) and facilitate intermittent catheterization (IC) in individuals with concurrent benign prostatic hyperplasia (BPH) and spinal cord injury (SCI). SETTING: Italian tertiary referral center for people with SCI. METHODS: We retrospectively collected all data of individuals undergoing PAE from 2015 to 2020 because of BPH-related problems during IC. PAE was performed under local anesthesia with superselective catheterization of the prostatic arteries. Technical success was defined as bilateral embolization. We determined pre- and post-procedural PV by magnetic resonance imaging. RESULTS: We considered 10 cases with tetraplegia (n = 5) and paraplegia (n = 5). Nine (90%) procedures were technically successful, while we performed monolateral PAE in one case. The median pre-procedural PV was 61 mL. After 6 months, all patients experienced significant PV reduction (median: 19.6%), and 7/7 patients with indwelling urinary catheter started successfully IC. Neither intra- nor post-procedural complications occurred within a median follow-up of 33 months. CONCLUSIONS: PAE proved to be a safe and effective treatment for BPH to facilitate IC in people with SCI. Considering the minimal morbidity of PAE, further multi-center studies are mandatory to draw definitive conclusions and warrant its widespread adoption in this population.

Severe tissue complications in patients of Bothrops snakebite at a tertiary health unit in the Brazilian Amazon: clinical characteristics and associated factors.

INTRODUCTION: Snakebites in the Brazilian Amazon are caused mostly by snakes from the Bothrops genus and envenomated patients may suffer from tissue complications. METHODS: This study aimed to identify risk factors for severe tissue complications (STC) in patients with Bothrops snakebite in the Amazonas state, Brazil. RESULTS: Snakebites that were classified as severe and affected female patients with comorbidities presented greater risks of developing STCs. In addition, hospitalizations of patients with STC exceeded 5 days. CONCLUSIONS: Clinical and epidemiological characteristics can prove essential for assessing the evolution of STC and clinical prognosis of patients with Bothrops snakebites.

Endovascular approach for arteriovenous limb malformations: a single center experience.

BACKGROUND: To assess imaging findings, describe endovascular technical aspects and analyzed procedural outcomes in a population of patients underwent limb arteriovenous malformation (LAVMs). METHODS: From January 2015 to December 2018, all consecutive patients underwent an endovascular procedure for ICD-9-CM codex for arteriovenous malformation problems were retrospective reviewed. Among these, patients with LAVMs were selected. Demographic, preoperative imaging, interventional and postprocedural data were collected for each patient and procedure. The International Society of Vascular Anomalies, the Cho-Do angiographic classification and the Schobinger clinical stage were used to describe disease type, aspect and clinical severity. Angiographic and clinical outcomes were also described. RESULTS: During the study period, 76 intervention for AVMs were performed in 52 patients. Among these, 26 LAVMs were selected and analyzed in 21 patients (number of LAVMs per patient: 1.2±0.5), 14 affecting upper limbs, 17 lower limbs. Pain, discomfort and swelling were main symptoms reported (95%, 90% and 62%, respectively). Ultrasound scan, computed angiography tomography and magnetic resonance angiography were used - alone or in combination - as preoperative imaging in 67%, 62% and 48% of patients, respectively. Cho-Do class ≥3 was described in 70% of treated LAVMs and a Schobinger stage ≥2 in more than 90%. Ipsilateral femoral access with a 5F introducer was preferred. Selective embolization with glue was the most preferred technique (57%), in combination or not with microsphere embolization (19%) and/or direct nidus sclerotherapy (14%). Optimal and suboptimal results were achieved in 86% of cases. Further interventions were performed in 52% of cases, with more than two interventions in 29% of cases. CONCLUSIONS: Quality of life, clinical picture and anatomical structure are items of paramount importance during preoperative LAVMs endovascular treatment planning. Treatment must be focused on LAVMs type, minimizing invasiveness and number of interventions, although secondary intervention rate remains quite high.

Severe tissue complications in patients of Bothrops snakebite at a tertiary health unit in the Brazilian Amazon: clinical characteristics and associated factors

Abstract INTRODUCTION: Snakebites in the Brazilian Amazon are caused mostly by snakes from the Bothrops genus and envenomated patients may suffer from tissue complications. METHODS: This study aimed to identify risk factors for severe tissue complications (STC) in patients with Bothrops snakebite in the Amazonas state, Brazil. RESULTS: Snakebites that were classified as severe and affected female patients with comorbidities presented greater risks of developing STCs. In addition, hospitalizations of patients with STC exceeded 5 days. CONCLUSIONS: Clinical and epidemiological characteristics can prove essential for assessing the evolution of STC and clinical prognosis of patients with Bothrops snakebites.

Patient-Derived Xenografts and Matched Cell Lines Identify Pharmacogenomic Vulnerabilities in Colorectal Cancer.

PURPOSE: Patient-derived xenograft (PDX) models accurately recapitulate the tumor of origin in terms of histopathology, genomic landscape, and therapeutic response, but some limitations due to costs associated with their maintenance and restricted amenability for large-scale screenings still exist. To overcome these issues, we established a platform of 2D cell lines (xeno-cell lines, XL), derived from PDXs of colorectal cancer with matched patient germline gDNA available. EXPERIMENTAL DESIGN: Whole-exome and transcriptome sequencing analyses were performed. Biomarkers of response and resistance to anti-HER therapy were annotated. Dependency on the WRN helicase gene was assessed in MSS, MSI-H, and MSI-like XLs using a reverse genetics functional approach. RESULTS: XLs recapitulated the entire spectrum of colorectal cancer transcriptional subtypes. Exome and RNA-seq analyses delineated several molecular biomarkers of response and resistance to EGFR and HER2 blockade. Genotype-driven responses observed in vitro in XLs were confirmed in vivo in the matched PDXs. MSI-H models were dependent upon WRN gene expression, while loss of WRN did not affect MSS XLs growth. Interestingly, one MSS XL with transcriptional MSI-like traits was sensitive to WRN depletion. CONCLUSIONS: The XL platform represents a preclinical tool for functional gene validation and proof-of-concept studies to identify novel druggable vulnerabilities in colorectal cancer.

Hemoperitoneum in cirrhotic patients in the absence of abdominal trauma.

Introduction: Hemoperitoneum can be a life-threating condition in cirrhotic patients who have a limited compensatory reserve during hemorrhagic shock. We aim to review the literature on the different etiologies associated with non-traumatic hemoperitoneum (NTH), summarizing the most relevant conditions associated with spontaneous and iatrogenic peritoneal and retroperitoneal bleeding that may occur in cirrhotic patients and to illustrate the most relevant diagnostic strategies and optimal management. Area covered: This review encompasses the current literature in hemoperitoneum in cirrhotic patients in the absence of abdominal trauma. Established diagnostic procedures, therapeutic interventions and potential novel targets are reported and discussed. Expert opinion: To ensure the optimal management regardless of the underlying etiology of NTH, the first goal for the clinician is to obtain immediate hemodynamic stabilization with supportive measures and to control the source of bleeding. The latter can be achieved with angiographic embolization, which is usually the first choice, or with open surgery. Other therapeutic options according to specific etiologies include transjugular intrahepatic portosystemic shunt (TIPS), balloon-occluded retrograde transvenous obliteration (BRTO), balloon-occluded anterograde transvenous obliteration (BATO) or intra operative radio frequency (RF).

The Evaluation of Aortic Diameter Changes During Long-Term Follow-Up After Endovascular Treatment of Acute Blunt Traumatic Thoracic Aortic Injuries.

PURPOSE: To evaluate the variations in aortic diameters and long-term results in patients who underwent thoracic endovascular aortic repair (TEVAR) for acute blunt traumatic thoracic aortic injuries (BTTAIs). MATERIALS AND METHODS: We retrospectively evaluated 23 patients with a mean age of 39 years (range: 17-74 years) who underwent TEVAR for BTTAI between October 2000 and November 2014. All of the patients underwent computed tomography angiography (CTA) before hospital discharge as a baseline imaging for the subsequent follow-up examinations. The technical success, overall survival, and complications were evaluated. Furthermore, the aortic diameters outside of the stent-graft (1 cm proximal and 1 cm distal to the stent-graft) and the aortic diameters within the stent-graft (2 cm distal to the proximal end and 2 cm proximal to the distal end) were assessed. The diameters at baseline on CTA were compared with those of the latest available follow-up examination. RESULTS: Technical success was 100% with a mean follow-up of 65.4 months (range: 12-171 months). No death was registered, and 2 (8.7%) of 23 endograft-related complications (1 stent-graft distal infolding and 1 endoleak 2 and 4 months after the procedure, respectively) were observed. An increase in aortic diameter either proximal or distal to the stent-graft (mean value 0.7 and 0.5 mm, respectively) or within the stent-graft (mean value of 0.5 mm for both proximal and distal diameters) was registered (mean follow-up at 65.4 months, range: 12-171 months). CONCLUSION: Aortic dilatation following TEVAR for BTTAI is minimal during long-term follow-up. Endovascular treatment represents a durable and safe option in acute BTTAIs.

Depressive behavior induced by unpredictable chronic mild stress increases dentin hypersensitivity in rats.

OBJECTIVE: The present study evaluated the nociceptive response induced by dentin hypersensitivity after dental erosion in rats that were exhibited to unpredictable chronic mild stress (UCMS)-induced depressive-like behavior. DESIGN: Adult male rats were subjected to UCMS (depression [D] group) or not (no depression [ND] group) for 30days and received either acidic solution to induce dental erosion (E) or water (W), thus forming the WND, END, WD, and ED groups. After the end of treatment, depressive-like parameters (i.e., sucrose preference and immobility in the forced swim test) and dentin hypersensitivity were evaluated. Plasma tumor necrosis factor α (TNF-α) and corticosterone levels were measured, and astrocytic glial fibrillary acidic protein (GFAP) expression was evaluated in the prefrontal cortex, hippocampus, amygdala, and hypothalamus. RESULTS: Administration of the acidic solution potentiated dentin hypersensitivity and increased corticosterone levels in the ED group compared with the WD group. TNF-α levels only increased in the WD group. The ED group exhibited an increase in astrocytic GFAP expression in the hypothalamus and prefrontal cortex but decreases in the hippocampus. CONCLUSIONS: These results suggest that UCMS exacerbated the nociceptive response associated with dentin hypersensitivity, concomitant with an increase in plasma corticosterone levels. Hypothalamic and prefrontal cortex astrogliosis in the ED group may be attributable to the increase in corticosterone associated to UCMS procedure. The reduction of astrocytic GFAP expression in the hippocampus in the ED group supports the association between dentin hypersensitivity and depression.

Prostatic Artery Embolization as an Alternative to Indwelling Bladder Catheterization to Manage Benign Prostatic Hyperplasia in Poor Surgical Candidates.

PURPOSE: To prospectively assess discontinuation of indwelling bladder catheterization (IBC) and relief of lower urinary tract symptoms (LUTS) due to benign prostatic hyperplasia (BPH) following prostate artery embolization (PAE) in poor surgical candidates. METHODS: Patients ineligible for surgical intervention were offered PAE after at least 1 month of IBC for management of urinary retention secondary to BPH; exclusion criteria for PAE included eligibility for surgery, active bladder cancer or known prostate cancer. Embolization technical and clinical success were defined as bilateral prostate embolization and removal of IBC, respectively. Patients were followed for at least 6 months and evaluated for International Prostate Symptom Score, quality of life, prostate size and uroflowmetric parameters. RESULTS: A total of 43 patients were enrolled; bilateral embolization was performed in 33 (76.7%), unilateral embolization was performed in 8 (18.6%), and two patients could not be embolized due to tortuous and atherosclerotic pelvic vasculature (4.7%). Among the patients who were embolized, mean prostate size decreased from 75.6 ± 33.2 to 63.0 ± 23.2 g (sign rank p = 0.0001, mean reduction of 19.6 ± 17.3%), and IBC removal was achieved in 33 patients (80.5%). Clavien II complications were reported in nine patients (21.9%) and included urinary tract infection (three patients, 7.3%) and recurrent acute urinary retention (six patients, 14.6%). Nine patients (22.0%) experienced post-embolization syndrome. CONCLUSIONS: PAE is a safe and feasible for the relief of LUTS and IBC in highly comorbid patients without surgical treatment options.

Subependymal nodules and giant cell tumours in tuberous sclerosis complex patients: prevalence on MRI in relation to gene mutation.

PURPOSE: The purpose of this study was to estimate the association among the presence of subependymal nodules (SENs), subependymal giant cell tumours (SGCTs) and gene mutation in tuberous sclerosis complex (TSC) patients. METHODS: Clinical records and images of 81 TSC patients were retrospectively reviewed by two neuroradiologists in consensus. All patients were assessed for gene mutations and were categorized as TSC1 or TSC2 mutation carriers, or no-mutations-identified (NMI) patients. They underwent a brain magnetic resonance imaging (MRI) using 0.1 mmol/kg of gadobutrol. Any enhancing SEN ≥ 1 cm and placed near the foramen of Monro was considered SGCT. Two MRI follow-up exams for each patient with SGCT were evaluated to assess tumour growth using Wilcoxon and chi-squared tests. RESULTS: Of 81 patients, 44 (54%) were TSC2 mutation carriers, 20 (25%) TSC1 and 17 (21%) NMI. Nine (11%) had a unilateral and three (4%) a bilateral SGCT. Fifty of 81 patients (62%) showed at least one SEN. None of the 31 patients without SEN showed SGCTs, whilst 12 (24%) of the 50 patients with at least one SEN showed SGCTs (p = 0.003). The association between the presence of SGCT or SEN and gene mutation was not significant (p = 0.251 and p = 0.187, respectively). At follow-up, the median SGCT diameter increased from 14 to 15 mm (p = 0.017), whilst the median SGCT volume increased from 589 to 791 mm(3) (p = 0.006). CONCLUSIONS: TSC patients with SENs are more likely to present with SGCT than those without SENs, in particular for TSC2 mutation carriers. The SGCT growth rate may be missed if based on the diameter instead of on the volume.

Uso da angiografia rotacional 3D (3D-RA) em portadores de cardiopatias congênitas: experiência de 53 casos / Use of 3D rotational angiography (3D-RA) in congenital heart disease: experience with 53 patients

Introdução: Os avanços no diagnóstico e tratamento das cardiopatias congênitas estão relacionados com os avanços na obtenção de imagens. As representações anatômicas obtidas com tomografia computadorizada, ressonância magnética e ecocardiografia têm ajudado, mas esses métodos não conseguem obter de forma precisa os dados hemodinâmicos. A angiografia rotacional 3D (3D-RA), um novo método de reconstrução tridimensional realizado no laboratório de cateterismo, tem sido amplamente utilizada em procedimentos neurológicos e urológicos. Nosso objetivo foi avaliar o desempenho da 3D-RA na avaliação das cardiopatias congênitas. Método: Revisão dos exames dos pacientes com cardiopatia congênita encaminhados para estudo hemodinâmico e que foram submetidos a aquisição de imagens pela 3D-RA. Foi utilizado equipamento Philips Allure FD10 e contraste de baixa osmolaridade para as angiografias. Resultados: No total, foram avaliados 53 pacientes, sendo utilizados, em média, 2,2 ± 1,1 ml/kg de contraste por paciente. Foi possível observar detalhes anatômicos não revelados em angiografias planares em 23% dos pacientes. Adicionalmente, 49% dos pacientes tiveram benefícios das imagens 3D para decisões terapêuticas. A exposição à radiação não foi estatisticamente diferente das angiografias planares. Nenhum paciente apresentou complicações relacionadas ao método. Conclusão: A 3D-RA forneceu informações que algumas das angiografias convencionais não fornecem e que ajudaram no tratamento de pacientes selecionados portadores de cardiopatia congênita. O emprego desse método poderá reduzir o número de aquisições de imagem por procedimento e, como consequência, limitar a exposição do paciente à radiação e ao contraste.

Background: Advances in the diagnosis and treatment of congenital heart disease are associated to advances in imaging techniques. Anatomic images obtained by computed tomography scan, magnetic resonance imaging (MRI) and echocardiography have been useful but cannot provide accurate hemodynamic data. 3D rotational angiography (3D-RA) is a new 3D reconstruction method carried out inthe cath lab that has been widely used in neurological and urological procedures. This study was aimed at evaluating the use of 3D-RA in the diagnosis and treatment of congenitalheart disease. Method: Review of catheterization results of patients with congenital heart disease referred for diagnostic assessment in which the 3D reconstruction method was employed. Philips Allure FD 10 equipment and low osmolarity contrast medium were used for angiographies. Results: Overall,53 patients were reviewed and 2.2 ± 1.1 mL/kg of contrast medium were used per patient. Anatomic details notpreviously shown by 2D angiographies were observed in 23% of the patients. Furthermore, 3D-RA imaging was usedto make treatment decisions in 49% of the patients. Exposure to radiation was not statistically different from 2D angiography. None of the patients had complications related to the method. Conclusion: 3D-RA provided information not usually seen by conventional angiography which was useful in thetreatment of selected patients with congenital heart disease. The use of 3D-RA may reduce the number of imaging tests per procedure and as a consequence, limit patient exposureto radiation and contrast media.

Seguimento dos 74 primeiros casos de radioterapia intra-operatória (eliot) no Hospital São Lucas da PUCPRS / Follow-up of the 74 first cases of intraoperative radiotherapy (eliot) at the Hospital São Lucas PUCPRS

Este artigo tem como objetivo descrever e analisar o uso da radioterapia intraoperatória no tratamento do câncer de mama inicial. Para isso, apresentaremos os dados obtidos com o seguimento das pacientes submetidas a essa modalidade de tratamento no Hospital São Lucas da PUCRS no período de janeiro de 2004 a abril de 2009.

Patologia benigna da mama / Mammary benign patology

A patologia benigna da mama é responsável por cerca de 80% das massas palpáveis. Seu diagnóstico diferencial é amplo, envolvendo distúrbios no desenvolvimento, doenças inflamatórias, alterações fobrocísticas, tumores benignos e doença mamária proliferativa, entre outros. O fibroadenoma, tumor benigno da mama, é a neoplasia mamária mais comum em pacientes menores de 35 anos, já os cistos são mais freqüentes na perimenopausa. O diagnóstico diferencial entre nódulos sólidos ou císticos poderá ser feito por meio da punção aspirativa com agulha fina ou pela ultra-sonografia. Nos nódulos císticos a punção por agulha fina além de diagnóstica, é conduta terapêutica. Neste artigo serão revisados aspectos dos diagnósticos diferenciados entre estes tumores, as abordagens terapêuticas e a associação entre tais patologias e câncer de mama.

Gomesin, a peptide produced by the spider Acanthoscurria gomesiana, is a potent anticryptococcal agent that acts in synergism with fluconazole.

Gomesin is an 18-residue cysteine-rich antimicrobial peptide produced by hemocytes of the spider Acanthoscurria gomesiana. In the present study, the antifungal properties of gomesin against Cryptococcus neoformans, the etiologic agent of cryptococcosis, were evaluated. Gomesin bound to the cell surface of cryptococci, which resulted in cell death associated with membrane permeabilization. Antifungal concentrations of gomesin were not toxic for human brain cells. Supplementation of cryptococcal cultures with the peptide (1 microM) caused a decrease in capsule expression and rendered fungal cells more susceptible to killing by human brain phagocytes. The possible use of gomesin in combination with fluconazole, a standard antifungal drug, was also evaluated. In association with fluconazole, gomesin concentrations with low antimicrobial activity (0.1-1 microM) inhibited fungal growth and enhanced the antimicrobial activity of brain phagocytes. These results reveal the potential of gomesin to promote inhibition of cryptococcal growth directly or by enhancing the effectiveness of host defenses.

An ectophosphatase activity in Candida parapsilosis influences the interaction of fungi with epithelial cells.

This study describes the biochemical characterization of a phosphatase activity present on the cell surface of Candida parapsilosis, a common cause of candidemia. Intact yeasts hydrolyzed p-nitrophenylphosphate to p-nitrophenol at a rate of 24.30+/-2.63 nmol p-nitrophenol h(-1) 10(-7) cells. The cell wall distribution of the Ca. parapsilosis enzyme was demonstrated by transmission electron microscopy. The duration of incubation of the yeast cells with the substrate and cell density influenced enzyme activity linearly. Values of V(max) and apparent K(m) for p-nitrophenylphosphate hydrolysis were 26.80+/-1.13 nmol p-nitrophenol h(-1) 10(-7) cells and 0.47+/-0.05 mM p-nitrophenylphosphate, respectively. The ectophosphatase activity was strongly inhibited at high pH as well as by classical inhibitors of acid phosphatases, such as sodium orthovanadate, sodium molybdate, sodium fluoride, and inorganic phosphate, the final product of the reaction. Only the inhibition caused by sodium orthovanadate was irreversible. Different phophorylated amino acids were used as substrates for the Ca. parapsilosis ectoenzyme, and the highest rate of phosphate hydrolysis was achieved using phosphotyrosine. A direct relationship between ectophosphatase activity and adhesion to host cells was established. In these assays, irreversible inhibition of enzyme activity resulted in decreased levels of yeast adhesion to epithelial cells.

Binding of glucuronoxylomannan to the CD14 receptor in human A549 alveolar cells induces interleukin-8 production.

Glucuronoxylomannan (GXM) is the major capsular polysaccharide of Cryptococcus neoformans. GXM receptors have been characterized in phagocytes and endothelial cells, but epithelial molecules recognizing the polysaccharide remain unknown. In the current study, we demonstrate that GXM binds to the CD14 receptor in human type II alveolar epithelial cells, resulting in the production of the proinflammatory chemokine interleukin-8.

Glucuronoxylomannan-mediated interaction of Cryptococcus neoformans with human alveolar cells results in fungal internalization and host cell damage.

Infection by Cryptococcus neoformans begins with inhalation of infectious propagules. Fungi reach the lung tissue and interact with epithelial cells in a crucial but poorly understood process. In this study, the interaction of C. neoformans with the human alveolar epithelial cell lineage A549 was investigated, focusing on the relevance of the capsular polysaccharide in this process. The association of encapsulated strains with A549 cells was significantly inhibited by a monoclonal antibody to glucuronoxylomannan (GXM), a major component of the cryptococcal capsule. A purified preparation of GXM produced similar results, suggesting the occurrence of surface receptors for this polysaccharide on the surface of alveolar cells. A549 cells were in fact able to bind soluble GXM, as confirmed by indirect immunofluorescence analysis using the anti-polysaccharide antibody. C. neoformans is internalized after GXM-mediated interaction with A549 cells in a process that culminates with death of host cells. Our results suggest that C. neoformans can use GXM for attachment to alveolar epithelia, allowing the fungus to reach the intracellular environment and damage host cells through still uncharacterized mechanisms.