Deficient glucose uptake is linked to impaired Glut1 expression upon CD3/CD28 stimulation in memory T cells from pleural effusions secondary to lung cancer

Glucose and nutrient uptake is essential in supporting T cell activation and is increased upon CD3/CD28 stimulation. As T cells from pleural effusions secondary to lung cancer show impaired function, we hypothesized that these cells might have altered expression of nutrient transporters. Here, we analysed by flow cytometry the expression of the transferrin receptor CD71, amino acid transporter CD98 and glucose transporter Glut1 and glucose uptake in pleural effusion‐derived T cells from lung cancer patients, after stimulation via CD3/CD28 under normoxia or hypoxia (2% O₂). We compared the response of T cells from pleural effusions secondary to lung cancer with that of T cells from nonmalignant effusions. In memory T cells from both groups, anti‐CD3/CD28‐stimulation under normoxia upregulated CD98 and CD71 expression (measured as median fluorescence intensity, MFI) in comparison with anti‐CD3‐stimulation. Costimulation under hypoxia tended to increase CD98 expression compared to CD3‐stimulation in memory T cells from both groups. Remarkably, in the cancer group, memory T cells stimulated via CD3/CD28 under hypoxia failed to increase CD71 and Glut1 expression levels compared to the cells receiving anti‐CD3 stimulation, a phenomenon that contrasted with the behaviour of memory T cells from nonmalignant effusions. Consequently, glucose uptake by memory T cells from the cancer group was not increased after CD3/CD28 stimulation under hypoxia, implying that their glycolytic metabolism is defective. As this process is required for inducing an antitumoural response, our study suggests that memory T cells are rendered dysfunctional and are unable to eliminate lung tumour cells.     

Determination of cell expansion and surface molecule expression on anti‐CD3/28 expanded CD4+ T cells

CD4⁺ T cell immunotherapy has potential for treatment in HIV‐infected patients. A large number of expanded CD4⁺ T cells and confirmation of functional‐related phenotypes are required for ensuring the successful outcomes of treatment. Freshly isolated CD4⁺ T cells from healthy donors were activated with anti‐CD3/28‐coated magnetic beads at different bead‐to‐cell ratios and cultured in the absence and presence of IL‐2 supplementation for 3 weeks. Fold expansion, cell viability, growth kinetic and lymphocyte subset identities were determined. Data demonstrated that a 1:1 bead‐to‐cell ratio rendered the highest expansion of 1044‐fold with 88% viability and 99.5% purity followed by the 2:1 and 0.5:1 ratios. No significant difference in proliferation and phenotypes was found between non–IL‐2 and IL‐2 supplementation groups. Several specific surface molecule expressions of the expanded cells including chemokine receptors, adhesion molecules, co‐stimulatory molecules, activation molecules, maturation markers, cytokine receptors and other molecules were altered when compared to the unexpanded cells. This optimized expansion protocol using the 1:1 bead‐to‐cell ratio of anti‐CD3/28‐coated magnetic beads and culture condition without IL‐2 supplementation provided the satisfactory yield with good reproducibility. Specific surface molecule expressions of the expanded cells presented potential roles in proliferation, differentiation, homeostasis, apoptosis and organ homing.     

Interleukin-34 drives macrophage polarization to the M2 phenotype in autoimmune hepatitis

BackgroundAutoimmune hepatitis is a chronic inflammatory disease, the abnormal immunological function is the main pathogenesis. Interleukin-34 is a newly identified cytokine that shares the same receptor as colony stimulating factor-1.MethodsWe used interleukin-34 knockout and wild-type mice in a Con A-induced hepatitis model and cocultured RAW264.7 macrophage cells with interleukin-34. We then detected associated inflammatory cytokine and chemokine levels to elucidate the role of interleukin-34.ResultsIn this study, we found that the loss of interleukin-34 resulted in higher sensitivity to Con A-induced hepatitis. RAW264.7 macrophage cells were able to differentiate to the M2 phenotype upon interleukin-34 stimulation.ConclusionsWe conclude that interleukin-34 may protect the liver from Con A-mediated hepatitis by driving M2 macrophage polarization and suppressing inflammation.     

Gastrointestinal mixed adenoneuroendocrine carcinoma (MANEC): An immunohistochemistry study of 13 microsatellite stable cases

BackgroundMixed adenoneuroendocrine carcinoma (MANEC) is currently included in the category of neuroendocrine carcinomas but the therapeutically management is not yet defined.AimsTo present the immunohistochemical (IHC) features of the epithelial mesenchymal transition (EMT) of MANEC.Materials and methodsThe clinicopathological features of 13 consecutive cases of MANEC (6 gastric and 7 colorectal) were correlated with the IHC expression of the biomarkers E-cadherin, β-catenin, N-cadherin, vimentin, maspin, CD44 and S100. In all of the cases open surgery was performed.ResultsAll of the cases showed microsatellite stable status, expressed E-cadherin and membrane β-catenin in both components (neuroendocrine and adenocarcinoma) and were negative for N-cadherin, vimentin and S-100. The colorectal MANECs were negative for maspin. In gastric MANECs, maspin showed cytoplasm positivity in the neuroendocrine component and nuclear translocation in the adenocarcinoma cells. CD44 was positive in all of the cases, in both components. No tumor buddings were identified. Three of the 13 patients survived for at least 32 months, all of them showing lymphatic emboli but not lymph node metastases. Pure neuroendocrine lymph node metastases were seen in only four of the cases: one from stomach, two of the ascending colon and two cases of the upper rectum.ConclusionsGastrointestinal MANEC is a microsatellite stable tumor with nodular growth, which components might originate from a CD44-positive stem-like precursor cell. Lymph node status remains the most reliable prognostic parameter and agressivity seems to not be influenced by tumor budding degree or EMT-related features. The histologic aspect of metastatic component (neuroendocrine versus adenocarcinoma) should be included in the histopathological reports and might be used for establishing the proper-targeted therapy of MANEC.     

Expression of HBME-1 and CD56 in follicular variant of papillary carcinoma in children: An immunohistochemical study and their diagnostic utility

Papillary thyroid carcinoma (PTC) is the most common differentiated thyroid cancer in children; and the follicular variant is the second most common variant after the classic subtype. The histological appearance of follicular variant of papillary thyroid cancer (FVPTC), can be mimicked by benign follicular nodules. Pediatric pathologists encountering such lesions with FVPTC-like appearance may err on diagnosing the benign lesions as malignant. In adult patients, several immunohistochemical markers have emerged recently as a useful adjunct to distinguish differentiated thyroid carcinomas from benign follicular lesions. We undertook an inter-institutional retrospective study to establish the diagnostic utility of immunohistochemical staining for HBME-1, Galectin-3 and CD56 in differentiating FVPTC from its benign mimics, follicular adenoma and adenomatoid nodules, in children. Our specific aim of the project was to define the sensitivity and specificity of the three antibodies in FVPTC. Based on institutional diagnoses, a total of 66 cases were obtained: 32 FVPTC and 34 benign follicular nodules that comprised of 23 follicular adenoma and 11 adenomatoid nodules. Five investigators, who were blinded to the original diagnoses, independently reviewed the slides following pre-determined criteria and semi-quantitatively scoring the immunohistochemical staining. The immunohistochemical staining revealed that a combination of positive HBME-1 and negative CD56 result gave 100% specificity and positive predictive value in distinguishing FVPTC from benign follicular nodules. However, the antibody combination suffered from a lower sensitivity (50%). We used a cutoff of 25% positivity of tumor cells in determining positivity of tumor cells to an antibody. In conclusion, our study found a very high specificity and strong positive predictive value for the combination of HBME-1 and CD56 immunohistochemical stains in distinguishing FVPTC from benign follicular lesions.