Tyndallized bacteria prime bronchial epithelial cells to mount an effective innate immune response against infections.

Airway epithelium represents a physical barrier against toxic substances and pathogens but also presents pattern recognition receptors on the epithelial cells that detect pathogens leading to molecule release and sending signals that activate both the innate and adaptive immune responses. Thus, impaired airway epithelial function and poor integrity may increase the recurrence of infections. Probiotic use in respiratory diseases as adjuvant of traditional therapy is increasingly widespread. There is growing interest in the use of non-viable heat-killed bacteria, such as tyndallized bacteria (TB), due to safety concerns and to their immunomodulatory properties. This study explores in vitro the effects of a TB blend on the immune activation of airway epithelium. 16HBE bronchial epithelial cells were exposed to different concentrations of TB. Cell viability, TB internalization, TLR2 expression, IL-6, IL-8 and TGF-ßl expression/release, E-cadherin expression and wound healing were assessed. We found that TB were tolerated, internalized, increased TLR2, E-cadherin expression, IL-6 release and wound healing but decreased both IL-8 and TGF-ßl release. In conclusion, TB activate TLR2 pathway without inducing a relevant pro-inflammatory response and improve barrier function, leading to the concept that TB preserve epithelial homeostasis and could be used as strategy to prevent and to manage respiratory infection, exacerbations included.

Formoterol Exerts Anti-Cancer Effects Modulating Oxidative Stress and Epithelial-Mesenchymal Transition Processes in Cigarette Smoke Extract Exposed Lung Adenocarcinoma Cells.

Lung cancer frequently affects patients with Chronic Obstructive Pulmonary Disease (COPD). Cigarette smoke (CS) fosters cancer progression by increasing oxidative stress and by modulating epithelial-mesenchymal transition (EMT) processes in cancer cells. Formoterol (FO), a long-acting ß2-agonist widely used for the treatment of COPD, exerts antioxidant activities. This study explored in a lung adenocarcinoma cell line (A549) whether FO counteracted the effects of cigarette smoke extract (CSE) relative to oxidative stress, inflammation, EMT processes, and cell migration and proliferation. A549 was stimulated with CSE and FO, ROS were evaluated by flow-cytometry and by nanostructured electrochemical sensor, EMT markers were evaluated by flow-cytometry and Real-Time PCR, IL-8 was evaluated by ELISA, cell migration was assessed by scratch and phalloidin test, and cell proliferation was assessed by clonogenic assay. CSE significantly increased the production of ROS, IL-8 release, cell migration and proliferation, and SNAIL1 expression but significantly decreased E-cadherin expression. FO reverted all these phenomena in CSE-stimulated A549 cells. The present study provides intriguing evidence that FO may exert anti-cancer effects by reverting oxidative stress, inflammation, and EMT markers induced by CS. These findings must be validated in future clinical studies to support FO as a valuable add-on treatment for lung cancer management.

Effects of condensates from volcanic fumaroles and cigarette smoke extracts on airway epithelial cells.

The impact of volcanic airborne products on airway epithelium homeostasis is largely unknown. This study assessed the effects of volcanic Fumarole Condensates (FC) alone or combined with Cigarette Smoke Extracts (CSE) on airway epithelial cells (16HBE and A549). Chemical composition of FC was analyzed by gas chromatography and HPLC. Cells were exposed to FC and IL-33 and IL-8 were assessed. The effects of FC and CSE on cell injury were evaluated assessing cell metabolism/cell viability, mitochondrial stress, cell apoptosis/cell necrosis, and cell proliferation. FC contained: water vapor (70-97%), CO2 (3-30%), acid gases (H2S, SO2, HCl, HF) around 1%. FC increased the intracellular IL-33 but differently modulated IL-33 and IL-8 gene expression and IL-8 release in the tested cell lines. FC without/with CSE: (a) increased cell metabolism/cell viability in 16HBE, while decreased it in A549; (b) increased mitochondrial stress in both cell types. FC with CSE increased cell necrosis in A549 in comparison to CSE alone. CSE reduced cell proliferation in 16HB,E while increased it in A549 and FC counteracted these effects in both cell types. Overall, FC induce a pro-inflammatory profile associated to a metabolic reprogramming without a relevant toxicity also in presence of CSE in airway epithelial cells.

Tyndallized Bacteria Preferentially Induce Human Macrophage M1 Polarization: An Effect Useful to Balance Allergic Immune Responses and to Control Infections.

Macrophage polarization is a dynamic process through which macrophages acquire specific features whose extremes are represented by M1 and M2 polarization. Interleukin (IL)-6, IL-1ß, IL-12 and IL-8 belong to M1 macrophages while transforming growth factor-beta (TGF-ß belongs to M2 cytokines. M2 polarization prevalence is observed in allergic diseases. Tyndallization is a thermal process able to inactivate microorganisms and to allow their use for chronic respiratory disease treatment via immune response modulation. The present study explores the effects of a blend of tyndallized bacteria (TB) on macrophage polarization. THP-1-derived macrophages were exposed to different concentrations of TB (106, 5 × 106, 107, 5 × 107, 108 CFU/mL) and then cell viability and TB phagocytosis, and IL-8, IL-1ß, IL-6, IL-12 and TGF-ß1 gene expression and release were assessed. TB were tolerated, phagocyted and able to increase IL-8, IL-1ß and IL-6 gene expression and release IL-12 gene expression, as well as decrease TGF-ß1 gene expression and release. The effects on IL-8, IL-6 and TGF-ß1 release were confirmed in human monocyte-derived macrophages (hMDMs) exposed to TB. In conclusion, TB promote M1 polarization, and this mechanism might have valuable potential in controlling allergic diseases and infections, possibly preventing disease exacerbations.

Disease Behavior and Treatment Response of Special Histological Types of Triple-Negative Breast Cancer.

BACKGROUND: Special histological types (SHT) of triple-negative breast cancer (TNBC) are a heterogeneous group of rare poorly understood diseases. We aimed to evaluate the clinical features, treatment, and outcomes of patients with SHT of TNBC. METHODS: We evaluated patients with a SHT of TNBC treated in a cancer center between 2009 and 2020. The endpoints were characterization of clinical and pathological features, pathologic complete response (PCR) rate after neoadjuvant chemotherapy, disease-free survival (DFS), progression-free survival, and overall survival (OS). RESULTS: The 132 patients included had the following histologies: metaplastic (n=71), medullary pattern (n=14), lobular (n=12), adenoid cystic (n=12), apocrine (n=10), and others (n=13). Metaplastic, lobular, and medullary pattern tumors had higher grade (66.6-85.7% grade 3); adenoid cystic and apocrine had mainly grade 1-2 (70-83.3%). Metaplastic and lobular carcinomas had higher disease stages (47.8% and 58.2% stages III-IV). PCR rates were 10.3% for metaplastic and 33.3% for lobular carcinomas, with 5-year DFS rates of 56% and 51.4%. Medullary pattern carcinomas had a great response to treatment, with PCR rate of 100%, and 5-year DFS rate of 92.8%. Apocrine carcinomas also had favorable prognosis, with no recurrence after early disease treatment, and 5-year DFS rate of 83.3%. Adenoid cystic carcinomas had intermediate prognosis, with 5-year DFS rate of 66.6%. CONCLUSION: SHT of TNBC encompasses heterogeneous malignancies with distinct behaviors. Lobular and metaplastic carcinomas showed high aggressiveness and poor treatment response, while medullary pattern and apocrine carcinomas had favorable outcomes. Treatment strategies focus on molecular features of each of these diseases are warranted.

Haemolymphatic Parameters in Two Aquaculture Crustacean Species Cherax destructor (Clark, 1836) and Cherax quadricarinatus (Von Martens, 1868).

In the last few years, there has been a notable development in the breeding of freshwater shrimp (astaciculture), which involved various species and in particular, the two Australian Parastacidae species, Cherax destructor and Cherax quadricarinatus. Information about the haemolymphatic parameters of these two species is fragmentary, and filling these gaps becomes important given their importance in aquaculture. Cellular and biochemical parameters were analyzed in both species to create a reference baseline for these parameters to identify the state of welfare or suffering of these animals. The results showed that the total haemocyte count, haemocyte subpopulations, enzymatic activities and pH are similar between the two species, while total protein and osmolality are higher in C. destructor than C. quadricarinatus. Knowledge of these parameters could assist in evaluating the good health status of these species kept in aquaculture facilities.

ANGEL2 is a member of the CCR4 family of deadenylases with 2',3'-cyclic phosphatase activity.

RNA molecules are frequently modified with a terminal 2',3'-cyclic phosphate group as a result of endonuclease cleavage, exonuclease trimming, or de novo synthesis. During pre-transfer RNA (tRNA) and unconventional messenger RNA (mRNA) splicing, 2',3'-cyclic phosphates are substrates of the tRNA ligase complex, and their removal is critical for recycling of tRNAs upon ribosome stalling. We identified the predicted deadenylase angel homolog 2 (ANGEL2) as a human phosphatase that converts 2',3'-cyclic phosphates into 2',3'-OH nucleotides. We analyzed ANGEL2's substrate preference, structure, and reaction mechanism. Perturbing ANGEL2 expression affected the efficiency of pre-tRNA processing, X-box-binding protein 1 (XBP1) mRNA splicing during the unfolded protein response, and tRNA nucleotidyltransferase 1 (TRNT1)-mediated CCA addition onto tRNAs. Our results indicate that ANGEL2 is involved in RNA pathways that rely on the ligation or hydrolysis of 2',3'-cyclic phosphates.

Crecimiento físico de niños con hiperplasia suprarrenal congénita perdedora de sal durante los dos primeros años de vida en el Instituto Nacional de Salud del Niño / Physical growth in children with salt-wasting adrenal hyperplasia during their first two years of life in the Peruvian National Children's Health Institute

Objetivo: Describir el crecimiento físico de niños con hiperplasia suprarrenal congénita (HSC) perdedora de sal durante los dos primeros años de vida, evaluados en el Servicio de Endocrinología del Instituto Nacional de Salud del Niño (INSN), Lima-Perú. Materiales y métodos: Es un estudio observacional, descriptivo y retrospectivo. Se incluyeron 55 niños con HSC que tuvieron seguimiento en el Servicio de Endocrinología del INSN del 2000 al 2016; se recopilaron datos como peso y talla al nacer y cada 6 meses, edad al diagnóstico, dosis de hidrocortisona y fludrocortisona y velocidad de crecimiento. Resultados: Se analizaron 55 niños con HSC perdedora de sal siendo el diagnóstico más precoz en mujeres (mediana: 21 días) que en varones (mediana: 52 días). Al nacimiento, su longitud promedio fue 0,07 + 0,75 DE; a los 6 meses fue -1,67 + 1,33 DE con mayor compromiso en varones (-2,29 + 1,18 DE; p=0,022); a los 12 meses fue -1,84 + 1,27 DE, con recuperación a los 24 meses (-1,51 + 1,10 DE). La velocidad de crecimiento fue -1,03 + 1,62 DE y -0,89 + 1,06 DE en el en el primer y segundo año, respectivamente. Conclusiones: Los niños con HSC perdedora de sal que tuvieron seguimiento en el INSN, tuvieron menor longitud a los 6 y 12 meses de edad y lograron recuperarse a los 24 meses.

Objective: To describe physical growth features in children with salt-wasting congenital adrenal hyperplasia (CAH) during their first two years of life, in the Endocrinology Service of the Peruvian National Children's Health Institute (CHI) in Lima, Peru. Materials and Methods: This is a retrospective, observational, and descriptive study. Fifty-five children with CAH underwent follow-up in the Endocrinology Service of the CHI from 2000 to 2016; data such as body weight and height were collected at birth time and every six months, as well as age at diagnosis, hydrocortisone and fludrocortisone doses, and growth velocity. Results: Fifty-five children with salt-wasting CAH were assessed, the diagnosis was more promptly made in females (median: 21 days) compared to males (median: 52 days). At birth, their average length was 0.07 ± 0.75 SD; at 6 months it was -1.67 ± 1.33 SD, and male subjects were more affected (-2.29 ± 1.18 SD; p= 0.022); at 12 months, this parameter was -1.84 ± 1.27 SD, and recovery was observed at 24 months (-1.51 ± 1.10 SD). Growth velocity values were -1.03 ± 1.62 SD and -0.89 ± 1.06 SD in the first and second years of life, respectively. Conclusions: Children with salt-wasting CAH who were followed-up at CHI had shorter length at 6- and 12- months of age, and their recovered at 24-monbths.

Estudio de la interferencia por hemólisis en la determinación de bilirrubina total / Hemolysis interference study in determination of total bilirubin / Estudo da interferência por hemólise na determinação de bilirrubina total

La hemólisis es una causa frecuente de rechazo de muestras que genera un sesgo negativo en la determinación de bilirrubina total (BT). Los objetivos de este trabajo fueron determinar el grado de hemólisis (GH) para reemplazar la inspección visual subjetiva, establecer la magnitud de su interferencia y conocer la concentración inicial de hemoglobina a partir de la cual se produce interferencia clínicamente relevante (ICR). Se recolectaron sueros con diferentes concentraciones de BT. Se preparó un hemolizado para generar distintas concentraciones de interferente. Se cuantificó la BT mediante el método de diclorofenildiazonio en los sueros con las diferentes concentraciones de hemoglobina y se determinó GH midiendo absorbancia a 578 nm. Se consideró ICR cuando se superó la máxima inexactitud deseable. Para cada nivel de BT se determinó la concentración de hemoglobina máxima aceptable (CHbmáx aceptable) por sobre la cual se considera ICR. La Abs578 permitió definir cuatro GH (GH1, GH2, GH3 y GH4). A partir de los interferogramas se determinó la CHbmáx aceptable para cada nivel de BT; ésta mostró un aumento lineal con la BT. A partir de 14,8 mg/dL de BT no se encontró ICR en los rangos estudiados. El GH permite tomar conductas objetivas ante muestras hemolizadas según el algoritmo propuesto.

Hemolysis is a common cause for rejection of samples that generates a negative bias in total bilirubin (TB) determination. The objectives of this study were to determine the hemolysis degree (HD) to replace the subjective visual inspection, to establish the extent of their interference and to meet the initial hemoglobin concentration from which clinically relevant interference (CRI) occurs. Sera was collected at different TB concentrations.Hemolysate was prepared to generate different concentrations of interfering. TB was quantified by dichlorophenyldiazonium method in sera with several hemoglobin concentrations, and DH was determined by measuring absorbance at 578 nm (Abs578). CRI was considered when it exceeded the maximum desirable inaccuracy. For each TB level, the maximum acceptable concentration of hemoglobin (CHbmáx acceptable) above which ICR is considered was determined.The Abs578 made it possible to define four DH (DH1, DH2, DH3 and DH4). The CHbmax acceptable was determined for each level of TB from the interferograms; it showed a linear increase with TB. From 14,8 mg/dL BT, CRI was not found in the ranges studied. The DH makes it possible to have objective behaviours in hemolyzed samples according to the proposed algorithm.

A hemólise é uma causa frequente de rejeição de amostras que gera um viés negativo na determinação da bilirrubina total (BT). Os objetivos deste estudo foram: determinar o grau de hemólise (GH) para substituir a inspeção visual subjetiva, estabelecer a extensão da sua interferência e conhecer a concentração inicial de hemoglobina a partir da qual ocorre a interferência clinicamente relevante (ICR). Foram coletados soros com diferentes concentrações de BT. Um hemolisado foi preparado para gerar diferentes concentrações de interferente. Foi quantificada a BT pelo método de diclorofenil diazônio nos soros com as diferentes concentrações de hemoglobina e foi determinado o GH medindo a absorvância a 578 nm (Abs578). ICR foi considerada quando se superou a máxima inexatidão desejável. Para cada nível de BT foi determinada a concentração de hemoglobina máxima aceitável (CHbMax aceitável) acima da qual é considerada ICR. A Abs578 permitiu definir quatro GH (GH1, GH2, GH3 e GH4). A partir dos interferogramas se determinou a CHbMax aceitável para cada nível de BT; ela apresentou um aumento linear com a BT. A partir de 14,8 mg/dL de BT, a ICR não foi encontrada nos intervalos estudados. O GH permite ter condutas objetivas diante de amostras hemolisadas de acordo com o algoritmo proposto.

Central diabetes insipidus: clinical profile that suggests organicity in Peruvian children: Lima - Peru 2001-2013.

BACKGROUND: Central diabetes insipidus (CDI) is a heterogeneous disease caused by arginine vasopressin deficiency; its management implies a profound understanding of the pathophysiology and the clinical spectrum. The aim of the study was to describe the clinical characteristics that indicate organicity in children and adolescents with central diabetes insipidus treated at the Department of Endocrinology from The Child Health's Institute during 2001 to 2013. METHODS: Cross-sectional, retrospective study. 79 cases of patients diagnosed with CDI (51 males and 28 females) from 1 month to 16 years of age were reviewed. For the descriptive analysis, measures of central tendency and dispersion were used; groups of organic and idiopathic CDI were compared using χ2-test and t-test. A p-value<0.05 was considered significant. RESULTS: The average age of patients was 8.1±4.2 years. Organic causes were intracranial tumors, 44 (55.7%), Langerhans cell histiocytosis (LCH), 11 (13.9%) and cerebral malformations in 7 (8.9%) patients, while the idiopathic group was 14 (17.7%) patients. Regarding clinical characteristics suggestive of organicity, headache (p=0.02) and visual disturbances (p=0.01) were found statistically significant. The anterior pituitary hormonal abnormalities were documented in 34 (52.3%) organic CDI patients. Furthermore, we did not find a significant difference in the average daily dose of desmopressin between patients with permanent vs. transitory CDI (0.81±0.65 vs. 0.59±0.62; p=0.363). CONCLUSIONS: The main clinical features suggestive of organicity in pediatric patients with central diabetes insipidus were headache and visual disturbances; furthermore, anterior pituitary hormonal abnormalities suggest an underlying organic etiology.

[Early puberty and thoracic mass: presentation of a case]. / Pubertad precoz y tumoración torácica: presentación de un caso.

A case is reported of a 7 year old patient with progressive dyspnea and genital development according to Marshall and Tanner criteria at the G2 VP3 stage. A mass of regular and well-defined borders was found in the chest, with calcifications and cystic areas of heterogeneous content. Basal measurements of FSH, LH and testosterone were <10 mIU/mL, 12.2 mIU/mL and 724 ng/dL, respectively. α-fetoprotein was 145.1 ng/mL. Excising the tumor improved respiratory symptoms, diminished hormone levels and stopped the process of precocious puberty. Pathology of the mediastinal tumor corresponded to a germ cell tumor with a predominant component of mature teratoma and choriocarcinoma. Neoplastic causes of precocious puberty are uncommon; its diagnosis is essential in order to provide timely treatment.

Pubertad precoz y tumoración torácica: presentación de un caso / Early puberty and thoracic mass: presentation of a case

Se reporta el caso de un paciente de 7 años de edad, que inicia su enfermedad con disnea progresiva, el desarrollo genital, de acuerdo con los criterios de Marshall y Tanner, muestra un estadio G2 VP3. En el tórax se halló una tumoración de bordes regulares y definidos, con presencia de calcificaciones y áreas quísticas de contenido heterogéneo. Las determinaciones basales de FSH, LH y testosterona fueron de < 10 mUI/mL, 12,2 mUI/mL y 724 ng/dL, respectivamente. La determinación de la α-feto proteína fue de 145,1 ng/mL. La exéresis del tumor mejoró el cuadro respiratorio, la remisión de los niveles hormonales y la detención del proceso de pubertad precoz. El estudio anatomopatológico de la tumoración mediastinal correspondió a un tumor de células germinales con componente predominante de teratoma maduro y coriocarcinoma. La pubertad precoz de etiología neoplásica es una causa rara, su diagnóstico es primordial para aplicar un tratamiento oportuno...

A case is reported of a 7 year old patient with progressive dyspnea and genital development according to Marshall and Tanner criteria at the G2 VP3 stage. A mass of regular and well-defined borders was found in the chest, with calcifications and cystic areas of heterogeneous content. Basal measurements of FSH, LH and testosterone were <10 mIU/mL, 12.2 mIU/mL and 724 ng/dL, respectively. α-fetoprotein was 145.1 ng/mL. Excising the tumor improved respiratory symptoms, diminished hormone levels and stopped the process of precocious puberty. Pathology of the mediastinal tumor corresponded to a germ cell tumor with a predominant component of mature teratoma and choriocarcinoma. Neoplastic causes of precocious puberty are uncommon; its diagnosis is essential in order to provide timely treatment...

The p38 MAPK-MK2 axis regulates E2F1 and FOXM1 expression after epirubicin treatment.

E2F1 is responsible for the regulation of FOXM1 expression, which plays a key role in epirubicin resistance. Here, we examined the role and regulation of E2F1 in response to epirubicin in cancer cells. We first showed that E2F1 plays a key role in promoting FOXM1 expression, cell survival, and epirubicin resistance as its depletion by siRNA attenuated FOXM1 induction and cell viability in response to epirubicin. We also found that the p38-MAPK activity mirrors the expression patterns of E2F1 and FOXM1 in both epirubicin-sensitive and -resistant MCF-7 breast cancer cells, suggesting that p38 has a role in regulating E2F1 expression and epirubicin resistance. Consistently, studies using pharmacologic inhibitors, siRNA knockdown, and knockout mouse embryonic fibroblasts (MEF) revealed that p38 mediates the E2F1 induction by epirubicin and that the induction of E2F1 by p38 is, in turn, mediated through its downstream kinase MK2 [mitogen-activated protein kinase (MAPK)-activated protein kinase 2; MAPKAPK2]. In agreement, in vitro phosphorylation assays showed that MK2 can directly phosphorylate E2F1 at Ser-364. Transfection assays also showed that E2F1 phosphorylation at Ser-364 participates in its induction by epirubicin but also suggests that other phosphorylation events are also involved. In addition, the p38-MK2 axis can also limit c-jun-NH(2)-kinase (JNK) induction by epirubicin and, notably, JNK represses FOXM1 expression. Collectively, these findings underscore the importance of p38-MK2 signaling in the control of E2F1 and FOXM1 expression as well as epirubicin sensitivity.

Metastatic mucinous carcinomas in the ovary: a practical approach to diagnosis related to gross aspects and to immunohistochemical evaluation.

Ovarian mucinous carcinomas are uncommon, and the differential diagnosis is metastatic carcinoma mainly from the gastrointestinal tract. The aim was to verify the importance of immunohistochemical reactions and the algorithm described in literature on the basis of laterality and tumor size. Twenty-five cases identified as metastatic mucinous adenocarcinomas were reviewed, along with clinical records; a tissue microarray was created, and immunohistochemical reactions for CK7, CK20, Ca125, hormonal receptors, WT1, DPC4, ß-catenin, and Cdx2 were determined. The median age was 51, and only 9 patients had a history of cancer. Sixteen patients (64%) had bilateral tumors, with sizes ranging from 5 to 36 cm (average, 20.5 cm); 9 (36%) had unilateral tumors varying from 5.5 to 38 cm (average, 21.8 cm). Algorithm agreement was 76%; most unilateral tumors were >13 cm. Common positive markers were Dpc4 (88%), Cdx2 (68%), CK20 (60%), and CK7 (44%). The useful markers were CK7, CK20, and Cdx2, although there were cases with overlapping results. The most common primary tumor was of colorectal origin (14 cases). The mean survival age was 32.6 mo. Although the proposed algorithm and immunohistochemical reactions are useful tools for diagnosis, some mucinous tumors cannot be definitively classified as primary or metastatic without further clinical evaluation, emphasizing the limits of this challenging diagnosis.