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Introduction: Understanding immune cell dynamics in kidney transplantation may provide insight into the mechanisms of rejection and improve patient management. B cells have gained interest with a special relevance of the "regulatory" subsets and their graft outcome prognostic value. In this study, we aimed to prove that the direct immunophenotyping and target gene expression analysis of kidney transplant patients' fresh whole blood will help to identify graft rejection risk and assist in the monitoring of kidney transplanted patients. Methods: We employed flow cytometry and qPCR techniques to characterize B and T cell subsets within fresh whole blood samples, with particular emphasis on transitional B cells (TrB) identified as CD19+CD24hiCD38hi. TrB are a relevant population in the context of kidney transplantation and are closely associated with regulatory B cells (Bregs) in humans. Patients were monitored, tracking pertinent clinical parameters and kidney-related events, including alterations in graft function and episodes of biopsy proven rejection. Results: Higher percentages of TrB cells at 3 months after transplantation were positively associated with better graft outcomes and lower biopsy-proven acute rejection risk. Furthermore, a novel panel of B cell regulatory associated genes was validated at 3 months post-transplantation by qPCR analysis of peripheral blood mononuclear cell (PBMC) mRNA, showing high predictive power of graft events and prognostic value. Discussion: These findings suggest that monitoring TrB may provide interesting patient management information, improve transplant outcomes, and allow for personalized drug regimens to minimize clinical complications.
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Rejeição de Enxerto , Transplante de Rim , Humanos , Transplante de Rim/efeitos adversos , Rejeição de Enxerto/imunologia , Rejeição de Enxerto/diagnóstico , Masculino , Feminino , Pessoa de Meia-Idade , Prognóstico , Adulto , Imunofenotipagem , Biomarcadores , Idoso , Sobrevivência de Enxerto/imunologia , Resultado do Tratamento , Linfócitos B Reguladores/imunologiaRESUMO
This study aimed to investigate obesity-related glomerulopathy (ORG) at cellular, structural, and transcriptomic levels. Thirty Wistar rats were randomized into two groups: 15 rats were fed with a standard diet (SD-rats), and 15 rats were fed with a high-fat diet (HFD-rats). After 10 weeks, the weight, kidney function, histological features, and transcriptomic changes were assessed. HFD-rats gained significantly more weight (55.8% vs. 29.2%; p < 0.001) and albuminuria (10,384.04 ng/mL vs. 5845.45 ng/mL; p < 0.001) compared to SD-rats. HFD-rats exhibited early stages of ORG, with predominant mesangial matrix increase and podocyte hypertrophy (PH). These lesions correlated with differentially expressed (DE) genes and miRNAs. Functional analysis showed that miR-205, which was DE in both the kidneys and urine of HFD-rats, negatively regulated the PTEN gene, promoting lipid endocytosis in podocytes. The downregulation of PTEN was proved through a higher PTEN/nephrin ratio in the SD-rats and the presence of lipid vacuoles in HFD-podocytes. This study has found a specific targetome of miRNAs and gene expression in early stages of ORG. Also, it emphasizes the potential value of miR-205 as a urinary biomarker for detecting podocyte injury in ORG, offering a tool for early diagnosis, and opening new avenues for future therapeutic research of obesity-related glomerulopathy.
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Dieta Hiperlipídica , MicroRNAs , Obesidade , Podócitos , RNA Mensageiro , Ratos Wistar , Animais , MicroRNAs/genética , Obesidade/complicações , Obesidade/genética , Obesidade/metabolismo , Ratos , Dieta Hiperlipídica/efeitos adversos , Masculino , Podócitos/metabolismo , Podócitos/patologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Nefropatias/etiologia , Nefropatias/genética , Nefropatias/metabolismo , Nefropatias/patologia , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica , Transcriptoma , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismoRESUMO
Neoadjuvant treatment (NAT) is one of the most widely used options for HER2+ and triple negative (TN) early breast cancer (BC). Since around half of the patients treated with NAT do not achieve a pathologically complete response (pCR), biomarkers to predict resistance are urgently needed. The correlation of clinicopathological factors with pCR was studied in 150 patients (HER2 = 81; TN = 69) and pre- and post-NAT differences in tumour biomarkers were compared. Low estrogen receptor (ER) expression, high tumour-infiltrating lymphocytes (TILs) and low cT-stage were associated with pCR in HER2+ tumours (p = 0.022; p = 0.032 and p = 0.005, respectively). Furthermore, ER expression was also associated with residual cancer burden (RCB; p = 0.046) in the HER2+ subtype. Similarly, pre-NAT, low progesterone receptor expression (PR; 1-10%) was associated with higher RCB (p < 0.001) in TN tumours. Only clinical and pathological T-stage (cpT-stage) had prognostic capacity in HER2+ tumours, whereas pre-NAT cpT-stage and post-NAT TILs had this capacity for the prognosis of TN tumours. We conclude that ER and PR expression may help predict response to NAT in HER2 and TN BC and should be taken into account in residual tumours. Also, changes observed in the phenotype after NAT suggest the need to reevaluate biomarkers in surviving residual tumour cells.
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Invasive lobular carcinoma (ILC) represents the second most common subtype of breast cancer (BC), accounting for up to 15% of all invasive BC. Loss of cell adhesion due to functional inactivation of E-cadherin is the hallmark of ILC. Although the current world health organization (WHO) classification for diagnosing ILC requires the recognition of the dispersed or linear non-cohesive growth pattern, it is not mandatory to demonstrate E-cadherin loss by immunohistochemistry (IHC). Recent results of central pathology review of two large randomized clinical trials have demonstrated relative overdiagnosis of ILC, as only ~60% of the locally diagnosed ILCs were confirmed by central pathology. To understand the possible underlying reasons of this discrepancy, we undertook a worldwide survey on the current practice of diagnosing BC as ILC. A survey was drafted by a panel of pathologists and researchers from the European lobular breast cancer consortium (ELBCC) using the online tool SurveyMonkey®. Various parameters such as indications for IHC staining, IHC clones, and IHC staining procedures were questioned. Finally, systematic reporting of non-classical ILC variants were also interrogated. This survey was sent out to pathologists worldwide and circulated from December 14, 2020 until July, 1 2021. The results demonstrate that approximately half of the institutions use E-cadherin expression loss by IHC as an ancillary test to diagnose ILC and that there is a great variability in immunostaining protocols. This might cause different staining results and discordant interpretations. As ILC-specific therapeutic and diagnostic avenues are currently explored in the context of clinical trials, it is of importance to improve standardization of histopathologic diagnosis of ILC diagnosis.
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Neoplasias da Mama , Carcinoma in Situ , Carcinoma Ductal de Mama , Carcinoma Lobular , Feminino , Humanos , Neoplasias da Mama/patologia , Caderinas/metabolismo , Carcinoma Ductal de Mama/patologia , Carcinoma Lobular/patologia , Imuno-Histoquímica , Ensaios Clínicos Controlados Aleatórios como AssuntoRESUMO
Digital counting methods were developed to decrease the high intra- and inter-observer variability of immunohistochemical markers such as Ki67, with most presenting a good correlation coefficient (CC). Since Ki67 is one of the major contributors to Oncotype DX, it is conceivable that Ki67 expression and the recurrence score (RS) obtained by the multigene panel are positively correlated. We decided first to test to what extent conventional and digital Ki67 quantification methods correlate in daily practice and, second, to determine which of these methods correlates better with the prognostic capacity of the Oncotype DX test. Both Ki67 evaluations were performed in 89 core biopsies with a diagnosis of estrogen receptor (ER) positive HER2-negative breast cancer (BC). Cases were, thus, classified twice for surrogate subtype: first by conventional analysis and then by digital evaluation. The Oncotype RS was obtained in 55 cases that were subsequently correlated to Ki67 evaluation by both methods. Conventional and digital Ki67 evaluation showed good concordance and correlation (CC = 0.81 (95% CI 0.73-0.89)). The correlation of Oncotype DX risk groups and surrogate derived subtypes was slightly higher for the digital technique (rs = 0.46, p < 0.01) compared to the conventional method (rs = 0.39, p < 0.01), even though both were statistically significant. In conclusion, we show that digital evaluation could be an alternative to conventional counting, and also has advantages for predicting the risk established by the Oncotype DX test in ER-positive BC. This study also supports the importance of an accurate Ki67 analysis which can influence the decision to submit ER-positive HER2-negative BC to prognostic molecular platforms.
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Neoplasias da Mama , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Feminino , Humanos , Antígeno Ki-67/genética , Antígeno Ki-67/metabolismo , Recidiva Local de Neoplasia/patologia , Prognóstico , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Receptores de Estrogênio/metabolismoRESUMO
IMPORTANCE: Reactions to tattoo may simulate common dermatosis or skin neoplasms. Histopathology allows diagnosis and helps determining the level and degree of inflammation associated, consequently orientating treatment. OBJECTIVE: To describe the histological features found in biopsies of cutaneous reactions to tattoo. DESIGN: This study was designed as a multicenter case series. SETTING: All consecutive histopathological samples of tattoos referred from 1992 to 2019 to the Hospital General de Catalunya, Hospital Germans Trias i Pujol, and a private practice, all in Barcelona, Spain, and from the Kempf und Pfaltz Histologische Diagnostik in Zurich, Switzerland were retrieved from the files. PARTICIPANTS AND EXPOSURE: The inclusion criteria were all cosmetic/permanent makeup, artistic/professional, and traumatic tattoos associated with either inflammatory reactions alone and/or with tumors and/or infections. Exclusion criteria were cases without any associated pathologic finding in the place of the ink, amalgam tattoos, and medical or temporary tattoos. MAIN OUTCOMES AND MEASURES: In all patients, clinical features (age, sex, location, tattoo color, and presentation) were recorded. Histological features evaluated included ink color, associated tumors or infections, and inflammatory reaction pattern. Inflammation was graded in low to moderate or severe. RESULTS: From 477 biopsies diagnosed as tattoos, 230 cases from 226 patients met the inclusion criteria. Samples corresponded to 107 male and 120 female subjects and 3 of unknown gender. Median age was 39 years (ranging from 9 to 84 years). Fifty-three samples were referred from centers in Spain and 177 from the center in Switzerland. The series was analyzed in 2 parts: tattoos associated only with inflammatory reactions (117/230) and tattoos associated with tumors or infections (113/230). The most common form of inflammatory pattern associated with tattoo was the fibrosing reaction (79/117, 68%), followed by granulomatous reaction (56/117, 48%), lichenoid reaction (33/117, 28%), epithelial hyperplasia (28/117, 24%), pseudolymphoma (27/117, 23%) and spongiotic reaction (27/117, 23%). Combined features of 2 or more types of inflammatory patterns were seen in 64% cases. CONCLUSIONS AND RELEVANCE: Our series confirms that cutaneous reactions to tattoos are polymorphous. Inflammation tends to present with combined patterns. Infections are tending to decline, and pathologic findings are not specific to ink color or clinical features.
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Dermatite/patologia , Dermatopatias Infecciosas/patologia , Neoplasias Cutâneas/patologia , Pele/patologia , Tatuagem/efeitos adversos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Biópsia , Criança , Cor , Corantes/efeitos adversos , Dermatite/etiologia , Feminino , Granuloma/etiologia , Granuloma/patologia , Humanos , Tinta , Erupções Liquenoides/etiologia , Erupções Liquenoides/patologia , Masculino , Pessoa de Meia-Idade , Pseudolinfoma/etiologia , Pseudolinfoma/patologia , Dermatopatias Infecciosas/etiologia , Adulto JovemRESUMO
PURPOSE: To assess the safety and effectiveness of magnetic seeds in preoperative localization and surgical dissection of metastatic axillary lymph nodes (LN+) in breast cancer patients with axillary involvement, after neoadjuvant chemotherapy (NAC). In addition, to assess the impact of targeted axillary dissection (TAD) in reducing the rate of false negatives (FN) in sentinel lymph node biopsy (SLNB). MATERIALS AND METHODS: A cross-sectional prospective cohort study was conducted from April 2017 to September 2019, including breast cancer patients with axillary lymph node involvement treated with NAC. Prior to NAC, the LN+ were marked by ultrasound-guided clip insertion. After NAC, a magnetic seed (Magseed®) was inserted in the clip-marked lymph node (MLN). During surgery, the MLN was located and removed with the aid of a magnetic detection probe (Sentimag®) and the sentinel lymph node was removed. Axillary lymph node dissection (ALND) was used to determine the rate of FN for SLNB alone and the combination of SLNB and MLN dissection, called TAD. RESULTS: The study included 29 patients (mean age, 55; range, 30-78 years). Selective preoperative localization and surgical dissection were successful for all 30 MLNs (100%). The MLN corresponded to the SLN in 50% of cases. After ALND, there were 21.4% (3/14) FN with SLNB alone and 5.9% (1/17) with TAD. CONCLUSIONS: Following NAC, selective surgical removal of MLN by preoperative localization using magnetic seeds is a safe and effective procedure with a success rate of 100%. Adding TAD reduces the rate of FN associated with SLNB alone.
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Neoplasias da Mama/cirurgia , Quimioterapia Adjuvante/métodos , Excisão de Linfonodo/métodos , Magnetismo , Mastectomia/métodos , Terapia Neoadjuvante/métodos , Biópsia de Linfonodo Sentinela/métodos , Adulto , Idoso , Axila , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Estudos Transversais , Feminino , Seguimentos , Humanos , Metástase Linfática , Pessoa de Meia-Idade , Cuidados Pré-Operatórios , Prognóstico , Estudos ProspectivosRESUMO
Tuberculosis (TB) is a leading cause of mortality due to infectious disease, but the factors determining disease progression are unclear. Transcriptional signatures associated with type I IFN signalling and neutrophilic inflammation were shown to correlate with disease severity in mouse models of TB. Here we show that similar transcriptional signatures correlate with increased bacterial loads and exacerbate pathology during Mycobacterium tuberculosis infection upon GM-CSF blockade. Loss of GM-CSF signalling or genetic susceptibility to TB (C3HeB/FeJ mice) result in type I IFN-induced neutrophil extracellular trap (NET) formation that promotes bacterial growth and promotes disease severity. Consistently, NETs are present in necrotic lung lesions of TB patients responding poorly to antibiotic therapy, supporting the role of NETs in a late stage of TB pathogenesis. Our findings reveal an important cytokine-based innate immune effector network with a central role in determining the outcome of M. tuberculosis infection.
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Armadilhas Extracelulares/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Interferon Tipo I/metabolismo , Pulmão/microbiologia , Mycobacterium tuberculosis/imunologia , Neutrófilos/imunologia , Pneumonia/imunologia , Tuberculose Pulmonar/imunologia , Animais , Bases de Dados Genéticas , Progressão da Doença , Perfilação da Expressão Gênica , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Humanos , Interferon Tipo I/genética , Interferon gama/genética , Interferon gama/metabolismo , Pulmão/imunologia , Pulmão/metabolismo , Pulmão/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mycobacterium tuberculosis/patogenicidade , Pneumonia/genética , Pneumonia/metabolismo , Pneumonia/patologia , RNA-Seq , Receptor de Interferon alfa e beta/genética , Receptor de Interferon alfa e beta/metabolismo , Tuberculose Pulmonar/sangue , Tuberculose Pulmonar/genética , Tuberculose Pulmonar/microbiologiaRESUMO
Modulation of immunity and disease by glycans is increasingly recognized. However, how host glycosylation shapes and is shaped by tuberculosis remains poorly understood. We show that deficiency in the glucosaminyl (N-acetyl) transferase 1 (Gcnt1), a key enzyme for core-2 O-glycans biosynthesis, drives susceptibility to Mycobacterium tuberculosis infection. The increased susceptibility of Gcnt1 deficient mice was characterized by extensive lung immune pathology, mechanistically related to neutrophils. Uninfected Gcnt1 deficient mice presented bone marrow, blood and lung neutrophilia, which further increased with infection. Blood neutrophilia required Gcnt1 deficiency in the hematopoietic compartment, relating with enhanced granulopoiesis, but normal cellular egress from the bone marrow. Interestingly, for the blood neutrophilia to translate into susceptibility to M. tuberculosis infection, Gnct1 deficiency in the stroma was also necessary. Complete Gcnt1 deficiency associated with increased lung expression of the neutrophil chemoattractant CXCL2. Lastly, we demonstrate that the transcript levels of various glycosyltransferase-encoding genes were altered in whole blood of active tuberculosis patients and that sialyl Lewis x, a glycan widely present in human neutrophils, was detected in the lung of tuberculosis patients. Our findings reveal a previously unappreciated link between Gcnt1, neutrophilia and susceptibility to M. tuberculosis infection, uncovering new players balancing the immune response in tuberculosis.
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Predisposição Genética para Doença , Mycobacterium tuberculosis , N-Acetilglucosaminiltransferases/deficiência , Neutrófilos/imunologia , Neutrófilos/metabolismo , Tuberculose/etiologia , Tuberculose/metabolismo , Animais , Carga Bacteriana , Biomarcadores , Modelos Animais de Doenças , Ativação Enzimática , Regulação da Expressão Gênica , Glicosilação , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Camundongos , Camundongos Knockout , Mycobacterium tuberculosis/imunologia , Neutrófilos/patologia , Taxa de Sobrevida , Tuberculose/diagnóstico , Tuberculose/mortalidadeRESUMO
BACKGROUND: Colonic cytomegalovirus [CMV] reactivation has been involved in steroid refractoriness in patients with active ulcerative colitis [UC]. The benefits of antiviral therapy in this clinical setting are still under debate, but the burden of viral reactivation has been associated with a poorer outcome in some studies. Our aim was to assess whether the burden of CMV reactivation measured by the number of viral inclusions by immunohistochemistry [IHC-CMV] is associated with a risk of colectomy. METHODS: Biopsy sets of UC patients with positive IHC-CMV were identified from the Pathology departments of three university hospitals. All biopsies were reviewed by expert pathologists, and the maximum number of IHC-CMV-positive cells in each biopsy set was re-assessed. Epidemiological and clinical features and clinical outcomes were recorded. RESULTS: Forty-six positive IHC-CMV cases with UC were included. At the time of CMV reactivation, 70% were receiving corticosteroids, 33% azathioprine, and 24% anti-tumour necrosis factor [TNF] agents. Thirty-two patients [70%] were treated with antiviral therapy. The median number of IHC-CMV-positive cells was 2 cells/biopsy [IQR 1-4]. Fourteen patients [30%] underwent colectomy, and 4 of them [29%] showed persistence of CMV in the surgical specimen. In the multivariate analysis, colectomy was only associated with >2 positive cells/biopsy [p = 0.048] and younger age [p = 0.023]. CONCLUSIONS: The burden of CMV colonic reactivation in patients with active UC, as measured by IHC, seems to be related to the risk of colectomy, and more data is needed to understand whether antiviral therapy guided by CMV burden will alter the clinical outcome.
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Colectomia , Colite Ulcerativa/cirurgia , Colo/virologia , Infecções por Citomegalovirus/tratamento farmacológico , Citomegalovirus/fisiologia , Ativação Viral , Corticosteroides/uso terapêutico , Adulto , Fatores Etários , Antivirais/uso terapêutico , Azatioprina/uso terapêutico , Colite Ulcerativa/complicações , Colite Ulcerativa/tratamento farmacológico , Colo/patologia , Infecções por Citomegalovirus/complicações , Feminino , Humanos , Imuno-Histoquímica , Imunossupressores/uso terapêutico , Masculino , Pessoa de Meia-Idade , Prognóstico , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Valganciclovir/uso terapêuticoRESUMO
An excessive, non-productive host-immune response is detrimental in active, chronic tuberculosis (TB) disease as it typically leads to tissue damage. Given their anti-inflammatory effect, non-steroidal anti-inflammatory drugs can potentially attenuate excessive inflammation in active TB disease. As such, we investigated the prophylactic and therapeutic effect of low-dose aspirin (LDA) (3 mg/kg/day), either alone or in combination with common anti-TB treatment or BCG vaccination, on disease outcome in an experimental murine model of active TB. Survival rate, bacillary load (BL) in lungs, and lung pathology were measured. The possible mechanism of action of LDA on the host's immune response was also evaluated by measuring levels of CD5L/AIM, selected cytokines/chemokines and other inflammatory markers in serum and lung tissue. LDA increased survival, had anti-inflammatory effects, reduced lung pathology, and decreased bacillary load in late-stage TB disease. Moreover, in combination with common anti-TB treatment, LDA enhanced survival and reduced lung pathology. Results from the immunological studies suggest the anti-inflammatory action of LDA at both a local and a systemic level. Our results showed a systemic decrease in neutrophilic recruitment, decreased levels of acute-phase reaction cytokines (IL-6, IL-1ß, and TNF-α) at late stage and a delay in the decrease in T cell response (in terms of IFN-γ, IL-2, and IL-10 serum levels) that occurs during the course of Mycobacterium tuberculosis infection. An anti-inflammatory milieu was detected in the lung, with less neutrophil recruitment and lower levels of tissue factor. In conclusion, LDA may be beneficial as an adjunct to standard anti-TB treatment in the later stage of active TB by reducing excess, non-productive inflammation, while enhancing Th1-cell responses for elimination of the bacilli.