CORRELAÇÃO DA EXPRESSÃO IMUNOISTOQUíMICA DE BETA-CATENINA E C-MYC COM A AGRESSIVIDADE EM TUMORES DO ESTÔMAGO

– Background: Biomarkers are macromolecules present in the body that may be related to neoplastic cells. None of them for diagnosis for secondary prevention were defined for gastric cancer. Objective: To investigate the immunohistochemical expression of beta-catenin and c-MYC proteins in gastric cancer and to correlate them with the aggressiveness of gastric tumors. Method: The sample consisted of histopathological slides, stained by H&E and paraffin blocks, and immunostained. Retrospective clinical data were collected. Clinical and epidemiological information was cross-referenced with the result obtained by immunostaining and its statistical analysis. Results: There was a predominance of men (69.1%), with a mean age of 63.8 years, with a predominance of lesions located in the antrum (54.5%), poorly differentiated (49.1%) with signet ring cells in 30 % of cases. On average, 18 lymph nodes were resected and in 30% no affected lymph nodes were detected. In 42.7% there was already distant metastasis, predominantly liver (61.7%). Stage IV was the classification of 43.6%, not being detected angiolymphatic invasion in 77.3% and perineural in 65.5%. Treatment was surgical and chemotherapy in 87.3%, with R0 resection in 79.1%. c-MYC was negative in 99.1% and beta-catenin was not expressed in 90.9%, being inconclusive in 6 cases. Conclusion: The immunohistochemical expression of these proteins in tissues with gastric cancer was not observed. The analyzed biomarkers, c-MYC and beta-catenin, showed no association with tumor aggressiveness in this cancerRacional: Biomarcadores são macromoléculas presentes no organismo que podem estar relacionadas com células neoplásicas. Nenhum deles para diagnóstico para prevenção secundária foi definido para o câncer gástrico. Objetivo: Investigar a expressão imunoistoquímica das proteínas beta-catenina e c-MYC no câncer gástrico e correlacioná-las com a agressividade dos tumores do estômago. Método: A amostra consistiu em lâminas histopatológicas, coradas por H&E e blocos de parafina, e realizada imunomarcação. Dados clínicos retrospectivos foram coletados. As informações clinicoepidemiológicas foram cruzadas com o resultado obtido pela imunomarcação e sua análise estatística. Resultado: Houve predomínio de homens (69,1%), com idade média de 63,8 anos, predominando lesão localizad

ANÁLISE IMUNOISTOQUÍMICA DA EXPRESSÃO DOS MARCADORES ALCAM E ALDH1 EM PACIENTES COM ADENOCARCINOMA COLORRETAL E ASSOCIAÇÃO COM DESFECHOS CLINICOPATOLÓGICOS

Background: Colorectal cancer has a high global mortality and tumor markers have emerged as diagnostic, management and prognostic indicators. New markers are being studied. Objective: To verify if there is a correlation between the immunohistochemical expression of ALCAM and ALDH1 proteins in colorectal adenocarcinoma tissue with epidemiological and clinicopathological characteristics, in particular their impact on disease progression and death. Method: Observational, single-center, analytical, retrospective study, through the investigation of patients undergoing surgical resection for colorectal cancer. 122 patients were evaluated. Regarding progression, it was shown that in individuals with positive ALCAM (n=40), 14/40 (35%) had progression, and for positive ALDH (n=54), 22/54 (40.7%). For death, the analysis of ALCAM positive (n=40), 24/40 (60%) died, and ALDH1 positive (n=54), 33/54 (61.1%). Conclusion: The immunohistochemical expression of ALCAM and ALDH1 markers was not associated with disease progression and death; it was also not possible to observe a correspondence relationship with the evaluated markers.Racional: O câncer colorretal apresenta alta mortalidade global e marcadores tumorais têm surgido como sinalizadores de diagnóstico, manejo e prognóstico. Novos marcadores estão sendo estudados. Objetivo: Verificar se há correlação da expressão por imunoistoquímica das proteínas ALCAM e ALDH1 em tecido com adenocarcinoma colorretal com as características epidemiológicas e clinicopatológicas, em particular o seu impacto na progressão de doença e no óbito. Método: Estudo observacional, unicêntrico, analítico, retrospectivo, através da investigação de pacientes submetidos à ressecção cirúrgica por câncer colorretal. Foram avaliados 122 pacientes. Em relação a progressão, mostrou-se que nos indivíduos com ALCAM positiva (n=40), 14/40 (35%) tiveram progressão, e para ALDH positiva (n=54), 22/54 (40,7%). Para óbito, a análise da ALCAM positiva (n=40), 24/40 (60%) morreram, e ALDH1 positivo (n=54), 33/54 (61,1%). Conclusão: A expressão imunoistoquímica dos marcadores ALCAM e ALDH1 não apresentou associação com a progressão de doença e óbito; também não foi possível observar relação de correspondência com os marcadores avaliados

ANÁLISE IMUNOISTOQUÍMICA DO BIOMARCADOR CICLINA D1 NOS CARCINOMAS PAPILÍFEROS DE TIREOIDE E BÓCIOS MULTINODULARES

Background: Papillary thyroid carcinomas (PTC) are the most prevalent and least aggressive thyroid carcinomas. In some cases, the diagnosis is doubtfull and the prognosis is poor. The search for tissue biomarkers that make it possible to ensure both the diagnosis for indeterminate cases and the prognosis, identifying the most aggressive cases, has been studied in recent decades. Objective: To analyze the molecular marker cyclin D1 in PTC and multinodular goiter (BMN) and to verify the correlation of the marking with the clinical-pathological characteristics in carcinomas. Methods: 118 tissues from adult patients submitted to PTC thyroidectomy and 40 BMN were selected as a control group. Tissue immunostaining was performed with cyclin D1 with subsequent immunohistochemical analysis in both groups, evaluating the expression of the marker (intensity and distribution). In the PTC group, immunostaining data were also crossed with clinical and pathological data. Results: The majority (93.3%) expressed the staining of cyclin D1 with varying intensities (weak, moderate 3 and strong) and predominantly diffuse distribution (71.2%). The BMN control group expressed staining for cyclin D1 in 57.5%, with weak intensity (47.5%) and sparse distribution (37.5%). The difference between the groups (study and control) was statistically significant (p<0.001). In the CPT group, the clinical-pathological crossings did not show differences regarding age, sex, tumor type and size, lymph node status, focus, angiolymphatic invasion. Conclusion: Cyclin D1 was expressed in the vast majority of PTC with the predominant diffuse distribution. There was no correlation between the expression of cyclin D1 and any clinical-pathological characteristic of PTC.Racional - Os carcinomas papilíferos são os mais prevalentes e menos agressivos de tireoide (CPT). Em alguns casos, o diagnóstico é duvidoso e o prognóstico ruim. A busca de biomarcadores teciduais que permitam assegurar tanto o diagnóstico para casos indeterminados, quanto o prognóstico, identificando os casos de maior agressividade, têm sido estudadas nas últimas décadas. Objetivo: Analisar a ciclina D1 nos CPT e nos bócios multinodulares (BMN) e verificar a correlação da marcação com as características clinicopatológicas. Métodos: Foram selecionados 118 tecidos de pacientes adultos submetidos àa tireoidectomia por CPT e 40 BMN como grupo controle. Realizou-se imunocoloração tecidual com ciclina D1 com subsequente análise imunoistoquímica em ambos grupos, avaliando-se a expressão do marcador (intensidade e distribuição). No grupo dos CPT os dados da imunocoloração foram também cruzados com os dados clinicopatológicos. Resultados: A maioria (93,3%) expressou a coloração da ciclina D1 com intensidades variadas (fraca, moderada e forte) e distribuição predominantemente difusa (71,2%). O grupo controle dos BMN, expressou coloração para ciclina D1 em 57,5%, com intensidade fraca (47,5%) e distribuição esparsa (37,5%). A diferença entre os grupos (estudo e controle) foi estatisticamente significante (p<0,001). No grupo dos CPT, os cruzamentos clinicopatológicos não evidenciaram diferenças quanto à idade, sexo, tipo e tamanho tumoral, estado linfonodal, focalidade e invasão angiolinfática. Conclusão: A ciclina D1 foi expressa na grande maioria dos CPT sendo a distribuição difusa predominante. Não houve correlação entre a expressão delacom qualquer característica clinicopatológica dos CPT

Perinatal Ureaplasma Exposure Is Associated With Increased Risk of Late Onset Sepsis and Imbalanced Inflammation in Preterm Infants and May Add to Lung Injury

Background: Controversy remains concerning the impact of Ureaplasma on preterm neonatal morbidity.Methods: Prospective single-center study in very low birth weight infants <30 weeks' gestation. Cord blood and initial nasopharyngeal swabs were screened for Ureaplasma parvum and U. urealyticum using culture technique and polymerase chain reaction. Neonatal outcomes were followed until death or discharge. Multi-analyte immunoassay provided cord blood levels of inflammatory markers. Using multivariate regression analyses, perinatal Ureaplasma exposure was evaluated as risk factor for the development of bronchopulmonary dysplasia (BPD), other neonatal morbidities until discharge and systemic inflammation at admission.Results: 40/103 (39%) infants were positive for Ureaplasma in one or both specimens, with U. parvum being the predominant species. While exposure to Ureaplasma alone was not associated with BPD, we found an increased risk of BPD in Ureaplasma-positive infants ventilated ≥5 days (OR 1.64; 95% CI 0.12–22.98; p = 0.009). Presence of Ureaplasma was associated with a 7-fold risk of late onset sepsis (LOS) (95% CI 1.80–27.39; p = 0.014). Moreover, Ureaplasma-positive infants had higher I/T ratios (b 0.39; 95% CI 0.08–0.71; p = 0.014), increased levels of interleukin (IL)-17 (b 0.16; 95% CI 0.02–0.30; p = 0.025) and matrix metalloproteinase 8 (b 0.77; 95% CI 0.10–1.44; p = 0.020), decreased levels of IL-10 (b −0.77; 95% CI −1.58 to −0.01; p = 0.043) and increased ratios of Tumor necrosis factor-α, IL-8, and IL-17 to anti-inflammatory IL-10 (p = 0.003, p = 0.012, p < 0.001).Conclusions: Positive Ureaplasma screening was not associated with BPD. However, exposure contributed to BPD in infants ventilated ≥5 days and conferred an increased risk of LOS and imbalanced inflammatory cytokine responses

COMO A CICLINA D1 SE COMPORTA COMO BIOMARCADOR NOS CARCINOMAS PAPILÍFEROS DE TIREOIDE E BÓCIOS MULTINODULARES?

Introduction: Papillary carcinomas are the most prevalent and least aggressive thyroid carcinomas (PTC). In some cases, the diagnosis is doubtfull and the prognosis is poor. The search for tissue biomarkers that ensure both the diagnosis for indeterminate cases and the prognosis, identifying the most aggressive cases, has been studied in recent decades. Objective: To review the literature in search of cyclin D1 as a marker of papillary thyroid carcinomas and multinodular goiters and evaluate whether its expression correlates with the clinicopathological characteristics of papillary thyroid carcinomas. Methods: Narrative review carried out by collecting information for reading and analysis from online research on virtual platforms. Initially, a search was carried out from MESH descriptors related to the topic, using the following terms: “papillary thyroid carcinoma, cyclin D1, immunohistochemistry, diagnosis, prognosis.” with AND or OR search, considering the title and/or abstract and those chosen were read in full. Results: The search included 77 articles that were compiled in this review. Conclusion: Cyclin D1 was expressed in the vast majority of PTCs, with diffuse distribution being predominant. There was no correlation between its expression and any clinicopathological characteristic of PTC.Racional - Os carcinomas papilíferos são os mais prevalentes e menos agressivos de tireoide (CPT). Em alguns casos, o diagnóstico é duvidoso e o prognóstico ruim. A busca de biomarcadores teciduais que permitam assegurar tanto o diagnóstico para casos indeterminados, quanto o prognóstico, identificando os casos de maior agressividade, têm sido estudadas nas últimas décadas. Objetivo: Revisar na literatura na busca da ciclina D1 como marcador dos carcinomas papilíferos de tireoide e nos bócios multinodulares, e avaliar se a expressão dela apresenta correlação com as características clínico-patológicas dos carcinomas papilíferos de tireoide. Métodos: Revisão narrativa feita colhendo informações para leitura e análise a partir de pesquisa online em platoformas virtuais. Inicialmente foi realizada busca por descritores DECs relacionados ao tema, utilizando os seguintes termos: “carcinoma papilífero de tireoide, ciclina D1, imunoistoquímica, diagnóstico, prognóstico.” com busca AND ou OR, considerando o título e/ou resumo e os escolhidos foram lidos na íntegra. Resultados: A busca incluiu 77 artigos que foram compilados nesta revisão.  Conclusão: A ciclina D1 foi expressa na grande maioria dos CPT sendo a distribuição difusa predominante. Não houve correlação entre a expressão dela com qualquer característica clinicopatológica dos CPT

AVALIAÇÃO DO OMENTO COMO DOADOR DE CÉLULAS TRONCO NO MIOCÁRDIO ISQUÊMICO ATRAVÉS DA ANÁLISE IMUNOHISTOQUÍMICA DO CD34

Background: Cardiovascular diseases are the leading cause of death in the world. The use of hematopoietic precursor cells and recent advances made in heart graft bioengineering offer a new therapeutic modality for post-myocardial infarction (MI)  and cardiac tissue regeneration. CD34 is a marker expressed on all hematopoietic and endothelial precursor cells, and functions as a cell adhesion factor. The antibody corresponding to this marker is used in immunohistochemistry to assess the formation of new vessels and the presence of stem cells. Aim: To evaluate the efficacy of omentopexy as stem cell donor, on previously infarcted myocardium, using immunohistochemically analysis of CD34. Method: Myocardial infarction was generated in four pigs, by ligature of the 1st and 2nd marginal branches of the circumflex artery. In three animals, abrasion of the infarcted epicardium was performed followed by multiple myocardial perforations and the mobilization of the omentum from the abdominal cavity to the mediastinum, sutured on the infarcted area. In the fourth animal, omentopexy was not performed and only the abrasion and perforation of the infarcted area were performed. All hearts were removed for CD34 immunohistochemically evaluation. Results:  In the samples from the group submitted to omentopexy, there was a 60% increase in angiogenesis, and in the samples from the control animal there was minimal staining.  Four samples from different sites of each animal, totaling 16 histopathological samples were evaluated. All samples were immunolabelled for CD34. Conclusions: Omentopexy proved to be effective in seeding previously infarcted myocardium with stem angiogenic cells, seen through immunohistochemistry, using CD34 marker.Racional: As doenças cardiovasculares são a principal causa de morte no mundo. O uso de células precursoras hematopoiéticas e os recentes progressos feitos na bioengenharia de enxertos cardíacos oferecem uma nova modalidade terapêutica para a regeneração do tecido cardíaco pós-infarto do miocárdio (IM). O CD34 é um marcador expresso em todas as células precursoras hematopoiéticas e endoteliais, e funciona como fator de adesão celular. O anticorpo que correspondente a este marcador é utilizado na imunohistoquímica para avaliar a formação de novos vasos e a presença de células-tronco. Objetivo: O estudo teve por objetivo avaliar a eficácia da omentopexia na neovascularização e na doação de células tronco de corações suínos previamente infartados, a partir da análise imunohistoquímica do CD34. Método: O infarto do miocárdio foi gerado em 4 suínos, por ligadura do 1°e 2° ramos marginais da artéria circunflexa. Em 3 animais realizou-se abrasão cuidadosa do epicárdio infartado seguido de múltiplas perfurações miocárdicas e a mobilização do omento da cavidade abdominal para o mediastino, envolvendo a área infartada e as perfurações. No quarto animal não foi realizado a omentopexia sendo realizado apenas a abrasão e perfuração da área infartada. Todos os animais foram eutanasiados ao 30º dia pós operatório e os corações retirados para avaliação macroscópica, microscópica e Imunohistoquímica do CD34. Resultados: Nas amostras do grupo submetido a omentopexia, ocorreu um aumento de 60% da angiogênese, sendo que nas amostras do animal controle houve marcação mínima. Foram avaliadas quatro amostras de diferentes sítios de cada coração dos animais, totalizando 16 amostras histopatológicas. Todas as amostras foram imunomarcadas para CD-34. Conclusões: O omento mostrou-se eficiente na indução de neovascularização pela presença de células tronco, vista através da marcação do CD34, demonstrando grande potencial como futura terapêutica para restaurar áreas de miocárdio isquêmico

Comparative Analysis of Inflammatory Cytokine Release and Alveolar Epithelial Barrier Invasion in a Transwell® Bilayer Model of Mucormycosis

Understanding the mechanisms of early invasion and epithelial defense in opportunistic mold infections is crucial for the evaluation of diagnostic biomarkers and novel treatment strategies. Recent studies revealed unique characteristics of the immunopathology of mucormycoses. We therefore adapted an alveolar Transwell® A549/HPAEC bilayer model for the assessment of epithelial barrier integrity and cytokine response to Rhizopus arrhizus, Rhizomucor pusillus, and Cunninghamella bertholletiae. Hyphal penetration of the alveolar barrier was validated by 18S ribosomal DNA detection in the endothelial compartment. Addition of dendritic cells (moDCs) to the alveolar compartment led to reduced fungal invasion and strongly enhanced pro-inflammatory cytokine response, whereas epithelial CCL2 and CCL5 release was reduced. Despite their phenotypic heterogeneity, the studied Mucorales species elicited the release of similar cytokine patterns by epithelial and dendritic cells. There were significantly elevated lactate dehydrogenase concentrations in the alveolar compartment and epithelial barrier permeability for dextran blue of different molecular weights in Mucorales-infected samples compared to Aspergillus fumigatus infection. Addition of monocyte-derived dendritic cells further aggravated LDH release and epithelial barrier permeability, highlighting the influence of the inflammatory response in mucormycosis-associated tissue damage. An important focus of this study was the evaluation of the reproducibility of readout parameters in independent experimental runs. Our results revealed consistently low coefficients of variation for cytokine concentrations and transcriptional levels of cytokine genes and cell integrity markers. As additional means of model validation, we confirmed that our bilayer model captures key principles of Mucorales biology such as accelerated growth in a hyperglycemic or ketoacidotic environment or reduced epithelial barrier invasion upon epithelial growth factor receptor blockade by gefitinib. Our findings indicate that the Transwell® bilayer model provides a reliable and reproducible tool for assessing host response in mucormycosis

Ex vivo-expanded highly pure ABCB5+ mesenchymal stromal cells as good manufacturing practice-compliant autologous advanced therapy medicinal product for clinical use: Process validation and first in-human data

© 2020 International Society for Cell & Gene Therapy Background aim: Mesenchymal stromal cells (MSCs) hold promise for the treatment of tissue damage and injury. However, MSCs comprise multiple subpopulations with diverse properties, which could explain inconsistent therapeutic outcomes seen among therapeutic attempts. Recently, the adenosine triphosphate-binding cassette transporter ABCB5 has been shown to identify a novel dermal immunomodulatory MSC subpopulation. Methods: The authors have established a validated Good Manufacturing Practice (GMP)-compliant expansion and manufacturing process by which ABCB5+ MSCs can be isolated from skin tissue and processed to generate a highly functional homogeneous cell population manufactured as an advanced therapy medicinal product (ATMP). This product has been approved by the German competent regulatory authority to be tested in a clinical trial to treat therapy-resistant chronic venous ulcers. Results: As of now, 12 wounds in nine patients have been treated with 5 × 105 autologous ABCB5+ MSCs per cm2 wound area, eliciting a median wound size reduction of 63% (range, 32–100%) at 12 weeks and early relief of pain. Conclusions: The authors describe here their GMP- and European Pharmacopoeia-compliant production and quality control process, report on a pre-clinical dose selection study and present the first in-human results. Together, these data substantiate the idea that ABCB5+ MSCs manufactured as ATMPs could deliver a clinically relevant wound closure strategy for patients with chronic therapy-resistant wounds

Upregulated Heat Shock Proteins After Hyperthermic Chemotherapy Point to Induced Cell Survival Mechanisms in Affected Tumor Cells From Peritoneal Carcinomatosis

In patients with peritoneal carcinomatosis cytoreductive surgery combined with hyperthermic intraperitoneal chemotherapy (HIPEC) represents a promising treatment strategy. Here, we studied the role of hyperthermic chemotherapy on heat shock protein (HSP) expression and induction of tumor cell death and survival. HSP27, HSP70, and HSP90 combined with effects on tumor cell proliferation and chemosensitivity were analyzed in human colon cancer. Hyperthermic chemotherapy resulted in significant HSP27/HSP70 and HSP90 gene/protein overexpression in analyzed HT-29/SW480/SW620 colon cancer cells and peritoneal metastases from patients displaying amplified expression of proliferation markers, proliferating cell nuclear antigen and antiapoptotic protein Bcl-xL. Moreover, functionally increased chemoresistance against 5-fluorouracil/mitomycin C and oxaliplatin after hyperthermic chemotherapy points to induced survival mechanisms in cancer cells. In conclusion, the results indicate that intracellular HSP-associated antiapoptotic and proliferative effects after hyperthermic chemotherapy negatively influence beneficial effects of hyperthermic chemotherapy-induced cell death. Therefore, blocking HSPs could be a promising strategy to further improve the rate of tumor cell death and outcome of patients undergoing HIPEC therapy