Diagnostic Value of GDF10 for the Tumorigenesis and Immune Infiltration in Lung Squamous Cell Carcinoma.

OBJECTIVE: Lung squamous cell carcinoma (LUSC) is associated with a low survival rate. Evidence suggests that bone morphogenetic proteins (BMPs) and their receptors (BMPRs) play crucial roles in tumorigenesis and progression. However, a comprehensive analysis of their role in LUSC is lacking. Our study aimed to explore the relationship between BMPs/BMPRs expression levels and the tumorigenesis and prognosis of LUSC. METHODS: The "R/Limma" package was utilized to analyze the differential expression characteristics of BMPs/BMPRs in LUSC, using data from TCGA, GTEx, and GEO databases. Concurrently, the "survminer" packages were employed to investigate their prognostic value and correlation with clinical features in LUSC. The core gene associated with LUSC progression was further explored through weighted gene correlation network analysis (WGCNA). LASSO analysis was conducted to construct a prognostic risk model for LUSC. Clinical specimens were examined by immunohistochemical analysis to confirm the diagnostic value in LUSC. Furthermore, based on the tumor immune estimation resource database and tumor-immune system interaction database, the role of the core gene in the tumor microenvironment of LUSC was explored. RESULTS: GDF10 had a significant correlation only with the pathological T stage of LUSC, and the protein expression level of GDF10 decreased with the tumorigenesis of LUSC. A prognostic risk model was constructed with GDF10 as the core gene and 5 hub genes (HRASLS, HIST1H2BH, FLRT3, CHEK2, and ALPL) for LUSC. GDF10 showed a significant positive correlation with immune cell infiltration and immune checkpoint expression. CONCLUSION: GDF10 might serve as a diagnostic biomarker reflecting the tumorigenesis of LUSC and regulating the tumor immune microenvironment to guide more effective treatment for LUSC.

MRPS16 promotes lung adenocarcinoma growth via the PI3K/AKT/Frataxin signalling axis.

Although MRPS16 is involved in cancer development, its mechanisms in developing LAUD remain unclear. Herein, qRT-PCR, WB and IHC were utilized for evaluating MRPS16 expression levels, while functional assays besides animal experiments were performed to measure MRPS16 effect on LAUD progression. Using WB, the MRPS16 effect on PI3K/AKT/Frataxin signalling pathway was tested. According to our study, MRPS16 was upregulated in LAUD and was correlated to the advanced TNM stage as well as poor clinical outcomes, which represent an independent prognostic factor. Based on functional assays, MRPS16 is involved in promoting LAUD growth, migration and invasion, which was validated further in subsequent analyses through PI3K/AKT/Frataxin pathway activation. Moreover, MRPS16-knockdown-mediated Frataxin overexpression was shown to restore the reduction in tumour cells proliferation, migration and invasion. Our results revealed that MRPS16 caused an aggressive phenotype to LAUD and was a poor prognosticator; thus, targeting MRPS16 may be effectual in LAUD treatment.

Tumor-associated macrophage-derived exosomes LINC01592 induce the immune escape of esophageal cancer by decreasing MHC-I surface expression.

BACKGROUND: TAMs (tumor-associated macrophages) infiltration promotes the progression of esophageal cancer (EC). However, the underlying mechanisms remain unclear. METHODS: Abnormal expression of LINC01592 from EC microarrays of the TCGA database was analyzed. LINC01592 expression level was validated in both EC cell lines and tissues. Stable LINC01592 knockdown and overexpression of EC cell lines were established. In vitro and in vivo trials were conducted to test the impact of LINC01592 knockdown and overexpression on EC cells. RNA binding protein immunoprecipitation (RIP), RNA pulldown assays, and Immunofluorescence (IF) were used to verify the combination of E2F6 and LINC01592. The combination of E2F6 and NBR1 was verified through the utilization of ChIP and dual luciferase reporter assays. RESULTS: LINC01592 is carried and transferred by exosomes secreted by M2-TAMs to tumor cells. The molecular mechanism underlying the promotion of NBR1 transcription involves the direct binding of LINC01592 to E2F6, which facilitates the nuclear entry of E2F6. The collaborative action of LINC01592 and E2F6 results in improved NBR1 transcription. The elevation of NBR1 binding to the ubiquitinated protein MHC-I via the ubiquitin domain caused a higher degradation of MHC-I in autophagolysosomes and a reduction in MHC-I expression on the exterior of cancerous cell. Consequently, this caused cancerous cells to escape from CD8+ CTL immune attack. The tumor-promoting impacts of LINC01592, as well as the growth of M2-type macrophage-driven tumors, were significantly suppressed by the interruption of E2F6/NBR1/MHC-I signaling through the effect of siRNA or the corresponding antibody blockade. Significantly, the suppression of LINC01592 resulted in an upregulation of MHC-I expression on the tumor cell membrane, thereby enhancing the efficacy of CD8+ T cell reinfusion therapy. CONCLUSIONS: The investigation conducted has revealed a significant molecular interaction between TAMs and EC via the LINC01592/E2F6/NBR1/MHC-I axis, which facilitates the progression of malignant tumors. This suggests that a therapeutic intervention targeting this axis may hold promise for the treatment of the disease.

Clinical characteristics, prognostic factors, and survival trends in esophageal neuroendocrine carcinomas: A population-based study.

BACKGROUND: Esophageal neuroendocrine carcinoma (ENEC) is an extremely rare type of malignancy. Clinical data of ENEC are limited to case reports and case series. More information is needed on its clinical feature, management, and prognosis. METHODS: This study collected information of ENEC patients diagnosed pathologically from 2010 to 2018. Data including demographic information, clinical features, and survival trends were obtained from the Surveillance, Epidemiology, and End Results (SEER) database. Statistical analyses were performed with STATA/SE 15.1, SPSS 25.0, and GraphPad Prism 8. RESULTS: A total of 283 ENEC patients were included in this study. The small-cell and large-cell subtypes of ENEC possess similar clinical features. The lower third of the esophagus (58%) was the most common location of ENEC. At the time of diagnosis, most ENEC patients were AJCC 7th stage IV (48.1%). Metastasis occurred in more than half of the ENEC patients (53.4%), and the most common metastatic site was the liver (37.1%). Compared with poorly differentiated esophageal squamous cell carcinoma (ESCC), another aggressive malignancy of the esophagus sometimes confused with ENEC because of similar histological features, our study showed differences in tumor location and metastatic rate, but similar poor survival rates. Multivariate survival analysis showed that ENEC located at the middle third of esophagus (p = 0.013), "Brain metastasis" (p = 0.019), and "Liver metastasis" (p < 0.001) were independent predictors of worse outcomes. "Surgery" (p = 0.003), and "Chemotherapy" (p < 0.001) were associated with better survival. CONCLUSION: A significant proportion of patients with newly diagnosed ENEC presented with metastatic disease. Predictors of poor survival included tumor location, brain metastasis, and liver metastasis. ENEC and poorly differentiated ESCC share certain histological features, but differ in tumor location and metastatic rate. Yet, no standard treatment strategy has been established, but surgery and chemotherapy were related to better outcomes.

Ferroptosis-Related Long Noncoding RNAs as Prognostic Marker for Colon Adenocarcinoma.

Background: The incidence of colon adenocarcinoma (COAD) has been increasing over time. Although ferroptosis and long noncoding RNAs (lncRNAs) have been extensively reported to participate in the tumorigenesis and development of COAD, few studies have investigated the role of ferroptosis-related lncRNAs in the prognosis of COAD. Methods: Gene-sequencing and clinical data for COAD were obtained from The Cancer Genome Atlas database. The coexpression network was constructed using known ferroptosis-related genes. Cox and least absolute shrinkage and selection operator regression were used to screen ferroptosis-related lncRNAs with prognostic value and to identify a predictive model of COAD. Patients with COAD were divided into low- and high-risk groups according to their risk score. Cases of COAD in the International Cancer Genome Consortium database were included as the testing cohort. Results: In total, nine lncRNAs (LINC02381, AC105219.1, AC009283.1, LINC01011, ELFN1-AS1, EIF3J-DT, NKILA, LINC01063, and SNHG16) were considered prognostic factors for COAD. Then, a risk score model was established. The overall survival rate of COAD patients was negatively associated with the risk score. Kaplan-Meier analyses in the original and testing cohorts showed similar results. The expression of the lncRNAs in tissue was consistent with the risk score, and the relationship with tumor mutation burden, immunity, and drug sensitivity presented a marked link between the signature and COAD. A nomogram was established for clinical applications. Conclusions: Nine ferroptosis-related lncRNAs and the established signature have a certain predictive value for prognosis of COAD patients and can be used as potential research targets for exploring treatment of COAD.

Long non-coding RNA Opa interacting protein 5-antisense RNA 1 binds to micorRNA-34a to upregulate oncogenic PD-L1 in non-small cell lung cancer.

Long non-coding RNA (lncRNA) OPA-interacting protein 5 antisense transcript 1 (OIP5-AS1) plays an oncogenic role in several types of cancer, but whether it is involved in non-small-cell lung cancer (NSCLC) is unclear. Our preliminary sequencing analysis revealed the upregulation of OIP5-AS1 in NSCLC. In this study, gene expression levels were analyzed by RT-qPCR. RNA-RNA pull-down assay was applied to detect direct interactions between RNAs. Overexpression assays were performed to explore the relationship between miR-34a and OIP5-AS1. CCK-8 assay and colony formation assay were applied to evaluate cell proliferation. In NSCLC cells (H23), overexpression of OIP5-AS1 increased the expression levels of programmed death-ligand 1 (PD-L1). In addition, inhibition of OIP5-AS1 and overexpression of miR-34a decreased the expression levels of PD-L1, and miR-34a significantly blocked the role of overexpression of OIP5-AS1. Overexpression of OIP5-AS1 and PD-L1 promoted H23 and H22 cells proliferation, while silencing of miR-34a and OIP5-AS1 played opposite roles and eliminated the effects of overexpression of OIP5-AS1 on cell proliferation. Therefore, OIP5-AS1 was upregulated to enhance the expression of oncogenic PD-L1 by sponging miR-34a in NSCLC, leading to promoted NSCLC cell proliferation. Our study also demonstrated that OIP5-AS1 was upregulated while miR-34a was downregulated in NSCLC.

Identification of ABCC5 Among ATP-Binding Cassette Transporter Family as a New Biomarker for Hepatocellular Carcinoma Based on Bioinformatics Analysis.

PURPOSE: Liver cancer is the fifth most common type of cancer worldwide, and the ATP-binding cassette (ABC) transporter family has been widely accepted as a cause of multidrug resistance. This study was conducted to explore the potential value and mechanisms of the ABC transporter gene family in the liver hepatocellular carcinoma (LIHC). MATERIALS AND METHODS: Data were collected from different public databases. UALCAN, ONCOMINE, and GEPIA were used to retrieve a selection of differently expressed and pathological stage-related genes among the ABC family. Principal component analysis (PCA) was utilized for grouping, and its prognostic value was evaluated by univariate and multivariate Cox analyses. The co-expression pattern was constructed with UALCAN, and the functional analyses were carried out with DAVID. The correlation between the biomarker and immune infiltration, genetic alteration frequency, and drug sensitivity were explored with TIMER, cBioPortal, GDSC and CTRP, respectively. Finally, tSNE algorithm was used to explore the distribution of ABCC5 expressed cells. RESULTS: Among the ABC transporter family members, ABCC5 was differently expressed and strongly related to the pathological stage of LIHC. PCA divided patients of LIHC into two groups, and Cox analyses demonstrated that ABCC5 was an independent risk factor of LIHC. Functional analyses indicated that the genes were enriched in the pathways of transmembrane transporter, ATPase activity, and bile secretion. ABCC5 is also associated with immune infiltration of cells like macrophages, neutrophils, and dendritic cells. The genetic alteration frequency of ABCC5 confirmed its potential value in LIHC. In addition, several drugs were explored and found to be relevant to LIHC. The t-SNE showed that expression of ABCC5 was most concentrated in macrophages, followed by hepatocytes. CONCLUSION: ABCC5 may facilitate LIHC progression through different mechanisms and be a potential biomarker and target for diagnosis, prognosis, and therapy of LIHC.

Monocytes promote pyroptosis of endothelial cells during lung ischemia-reperfusion via IL-1R/NF-κB/NLRP3 signaling.

In our previous study, we observed that donor pulmonary intravascular nonclassical monocytes play a major role in early PGF, but the specific mechanism remained unclear. In this study, we investigated the mechanistic role of monocytes in inducing pyroptosis of human pulmonary microvascular endothelial cells (HPMECs) during IRI. A murine hilar ligation model of IRI was utilized whereby left lungs underwent 1 h of ischemia and 23 h of reperfusion. Monocyte depletion by intraperitoneal clodronate-liposome treatment on pulmonary edema and pyroptosis activation were determined. In vitro experiments, we performed the co-culture experiments under hypoxia-reoxygenation (H/R) conditions to mimic the IRI environment. We monitored the expression of NLRP3, caspase-1 and IL-1ß in co-cultures of monocytes (U937 cells) and HPMECs under H/R conditions. NLRP3, IL-1ß and IL-1R siRNA knockdown, caspase-1 and NF-κB pathway inhibitors were employed to elucidate the mechanism modulating HPMEC pyroptosis during H/R. Treatment of mice with clodronate-liposome attenuated IR-induced pulmonary edema, cytokine production and pyroptosis activation. In vitro, NLRP3 knockdown in monocytes reduced caspase-1 and IL-1ß secretion in co-cultures of monocytes and HPMECs. Reduced HPMEC pyroptosis was also observed either containing HPMECs with genetically engineered IL-1R knockdown or in co-culture treated with a Triplotide inhibitor that disrupts NF-κB signaling. Monocytes play a vital role in the development of transplant-associated ischemia-reperfusion injury. A potential role is that monocytes secrete IL-1ß to induce HPMEC pyroptosis via the IL-1R/NF-κB/NLRP3 pathway.

The Evolving Landscape of PD-1/PD-L1 Pathway in Head and Neck Cancer.

Over the past 10 years, cancer immunotherapy has made significant progress in multiple cancer types and has been gradually been applied to clinical cancer care, in which the programmed cell death protein-1 (PD-1)/programmed cell death ligand 1 (PD-L1) pathway is one of the most attractive targets. Compared with traditional therapies, the emerging PD-1/PD-L1 blockade immunotherapy exhibited more satisfactory curative effects and lower toxicity for patients with advanced head and neck squamous cell carcinoma (HNSCC). This review analyzes the expression characteristics and clinical significance of PD-1/PD-L1 in HNSCC, the immunosuppressive roles of tumor cell and stromal cell expressing PD-1/PD-L1 in this disease, and presents the development landscape of PD-1/PD-L1 inhibitors, which may provide new curative alternatives for recurrent or metastatic HNSCC.

Interplay between cancer cells and M2 macrophages is necessary for miR-550a-3-5p down-regulation-mediated HPV-positive OSCC progression.

BACKGROUND: Human papillomavirus (HPV)-positive oral squamous cell carcinoma (OSCC) is increasing worldwide with typically higher grade and stage, while better prognosis. microRNAs (miRNAs) has been shown to play a critical role in cancer, however, their role in HPV-positive OSCC progression remains unclear. METHODS: miRNA microarray was performed to identify differentially expressed miRNAs. qRT-PCR and FISH were performed to determine the relative expression of miR-550a-3-5p. CCK-8, Flow cytometry, Wound healing, Cell invasion assays and xenograft experiments were conducted to analyze the biological roles of miR-550a-3-5p. Tumor-associated macrophages (TAMs) generation, co-culturing of cancer cells with TAMs, Western blot, Dual-luciferase reporter gene assay, Immunohistochemistry and animal studies were performed to explore the mechanisms underlying the functions of miR-550a-3-5p. RESULTS: We identified 19 miRNAs differentially expressed in HPV-positive OSCC specimens and miR-550a-3-5p was down-regulated. The low expression of miR-550a-3-5p correlated with higher tumor size and nodal metastasis of HPV-positive OSCC patients. Then, we found that miR-550a-3-5p suppressed the migration, invasion and EMT of HPV-positive OSCC cells dependent on decreasing M2 macrophages polarization. Moreover, miR-550a-3-5p, down-regulated by E6 oncoprotein, inhibited M2 macrophages polarization by YAP/CCL2 signaling, which in turn abrogating EMT program in HPV-positive OSCC cells. In addition, in both xenografts and clinical HPV-positive OSCC samples, miR-550a-3-5p levels were inversely associated with YAP, CCL2 expressions and the number of M2 macrophages. CONCLUSIONS: E6/miR-550a-3-5p/YAP/CCL2 signaling induces M2 macrophages polarization to enhance EMT and progression, revealing a novel crosstalk between cancer cells and immune cells in HPV-positive OSCC microenvironment.

Prognostic relevance of UCH-L1 and α-internexin in pancreatic neuroendocrine tumors.

Prognostic biomarkers for the pancreatic neuroendocrine tumors are needed. Proteomic study on insulinoma has been rarely reported. We identified the differential expression of proteins between insulinoma and their paired tissues by proteomic analysis, and evaluated the prognostic significance of specific proteins in pancreatic neuroendocrine tumors including insulinoma. The differential expression of select proteins was validated in more than 300 tumors using immunohistochemical staining and western blot. Methylation of UCH-L1 promoter in tumors was examined by methylation specific PCR and validated by sequencing. The concurrent expression of UCH-L1 and α-internexin was correlated with the prognosis in 2 independent collectives of patients with tumors. Sixty-two and 219 proteins were significantly down-regulated and up-regulated in insulinomas, respectively. Demethylation of UCH-L1 promoter was associated with UCH-L1 expression in tumors (p = 0.002). The concurrent expression of UCH-L1 and α-internexin in pancreatic neuroendocrine tumors was significantly associated with better overall survival and disease-free survival in the combination of both cohorts (log rank p = 3.90 × 10-4 and p = 3.75 × 10-5, respectively) and in each of cohorts. The prognostic value of both proteins was also validated in patients with stage II and III tumors (p = 0.017 and p = 0.006, respectively). The proteins UCH-L1 and α-internexin could be independent prognostic biomarkers of pancreatic neuroendocrine tumors.

Apoptosis of the Tracheal Epithelium Can Increase the Number of Recipient Bone Marrow-Derived Myofibroblasts in Allografts and Exacerbate Obliterative Bronchiolitis After Tracheal Transplantation in Mice.

BACKGROUND: The long-term outcome of lung transplantation is impeded by the development of obliterative bronchiolitis (OB). We sought to investigate the relationship between the apoptosis of tracheal epithelial cells and bone marrow-derived myofibroblasts in the process of OB. METHODS: The mouse orthotopic tracheal transplant model was established. The allografts and syngrafts were harvested at 1, 2, 3, and 4 weeks after tracheal transplant. The percentage of tracheal lumen occlusion was assayed via histology and morphometry. Immunofluorescence staining was used to detect the apoptotic epithelial cells and recipient-derived myofibroblasts. The expression of SDF-1α and TGF-ß and the infiltrations of CD4 and CD8 T cells in the grafts were detected using immunohistochemical staining. RESULTS: We found that there were more apoptotic epithelia in the allograft group than in the syngraft group and that the level of tracheal lumen occlusion was higher at different time points. Moreover, the increase in the apoptosis of the tracheal epithelium occurred earlier than that of occlusion of the tracheal lumen. There were more myofibroblasts derived from the recipient's bone marrow and more CD4 and CD8 T cells in the allografts. The expression of SDF-1α and TGF-ß was higher in the epithelium from allografts than in those of the syngrafts. CONCLUSIONS: Our study indicated that the apoptotic tracheal epithelia in the OB model might increase the amount of myofibroblasts derived from the recipient's bone marrow. Therapeutic methods aimed at preventing apoptosis of the tracheal epithelium may improve the outcome of lung transplantation.

Differences and Similarities in the Clinicopathological Features of Pancreatic Neuroendocrine Tumors in China and the United States: A Multicenter Study.

The presentation, pathology, and prognosis of pancreatic neuroendocrine tumors (PNETs) in Asian patients have not been studied in large cohorts. We hypothesized that the clinicopathological features of PNETs of Chinese patients might be different from those of US patients. The objectives of this study were to address whether PNETs in Chinese patients exhibit unique clinicopathological features and natural history, and can be graded and staged using the WHO/ENETS criteria. This is a retrospective review of medical records of patients with PNETs in multiple academic medical centers in China (7) and the United States (2). Tumor grading and staging were based on WHO/ENETS criteria. The clinicopathological features of PNETs of Chinese and US patients were compared. Univariate and multivariate analyses were performed to find associations between survival and patient demographics, tumor grade and stage, and other clinicopathological characteristics. A total of 977 (527 Chinese and 450 US) patients with PNETs were studied. In general, Chinese patients were younger than US patients (median age 46 vs 56 years). In Chinese patients, insulinomas were the most common (52.2%), followed by nonfunctional tumors (39.7%), whereas the order was reversed in US patients. Tumor grade distribution was similar in the 2 countries (G1: 57.5% vs 55.0%; G2: 38.5% vs 41.3%; and G3: 4.0% vs 3.7%). However, age, primary tumor size, primary tumor location, grade, and stage of subtypes of PNETs were significantly different between the 2 countries. The Chinese nonfunctional tumors were significantly larger than US ones (median size 4 vs 3 cm) and more frequently located in the head/neck region (54.9% vs 34.8%). The Chinese and US insulinomas were similar in size (median 1.5 cm) but the Chinese insulinomas relatively more frequently located in the head/neck region (48.3% vs 26.1%). Higher grade, advanced stage, metastasis, and larger primary tumor size were significantly associated with unfavorable survival in both countries. Several clinicopathological differences are found between Chinese and US PNETs but the PNETs of both countries follow a similar natural history. The WHO tumor grading and ENETS staging criteria are applicable to both Chinese and US patients.

Chromogranin A is a reliable serum diagnostic biomarker for pancreatic neuroendocrine tumors but not for insulinomas.

BACKGROUND: Pancreatic neuroendocrine tumors (PNETs) are a group of rare tumors. Chromogranin A (CgA) was considered as the most practical and useful serum tumor marker in PNET patients. But peripheral blood levels of CgA are not routinely tested in Chinese patients with PNETs. This study was to assess the diagnostic value of CgA in Chinese patients with PNETs especially in patients with insulinomas. METHODS: Eighty-nine patients with PNETs including 57 insulinomas and 32 non-insulinoma PNETs as well as 86 healthy participants were enrolled in this study between September 2003 and June 2013. Serum levels of CgA were measured by ELISA method. Expression of CgA protein was detected in 26 PNET tissues including 14 insulinomas by immunohistochemical staining. RESULTS: Serum levels of CgA in 89 PNET patients were significantly higher than that in healthy controls (P = 7.2 × 10-9). Serum levels of CgA in 57 patients with insulinomas (median 64.8 ng/ml, range 25-164) were slightly higher than the levels in healthy controls (median 53.4 ng/ml, range 39-94) but much lower than the levels in 32 patients with non-insulinoma PNETs (median 193 ng/ml, range 27-9021), P = 0.001. The serum CgA levels were reduced in 16 of 17 patients with insulinomas after tumor resection. ROC curve showed that CgA values at 60 ng/ml distinguished patients with insulinomas from healthy controls but its sensitivity and specificity were 66.7% and 73.3%, respectively. In contrast, CgA values at 74 ng/ml distinguished patients with non-insulinoma PNETs from healthy controls, and the sensitivity and specificity were 65.6% and 91.9%, respectively. Except for two insulinomas with negative staining of CgA, 12 insulinoma tissues showed positive staining of CgA. CONCLUSION: CgA is a reliable serum diagnostic biomarker for PNETs but not for insulinomas.

Increased expression of Tim-3 and its ligand Galectin-9 in rat allografts during acute rejection episodes.

The aim of the study is to elucidate the profiles of T-cell immunoglobulin and mucin domain-3 (Tim-3) and its ligand Galecin-9 in acute pulmonary rejection by using a rat model of lung transplantation. Left lung grafts retrieved from Lewis or Fisher 344 rats were orthotopically transplanted into Lewis recipients without any immunosuppressions; the grafts were harvested at day 3, 7 or 10 after transplantation. The grade of acute rejection was histopathologically evaluated. Tim-3, Galectin-9, immune antigen and related cytokines expression were assessed with immunological techniques and real-time polymerase chain reaction (RT-PCR), respectively. Then, our results showed that Tim-3 and its ligand Galectin-9 were markedly up-regulated at protein and mRNA levels in allografts compared with syngrafts. Meanwhile, the decreased CD4/CD8 ratio was associated with acute rejection occurring and Tim-3 expression on CD4(+) and CD8(+) T cells in allografts was increased. Therefore, our study firstly described that enhanced Tim-3 and its ligand Galectin-9 in allografts might play an important role in the pathogenesis of rat lung transplant rejection, implying new valuable markers for detecting acute allograft rejection.

α-Internexin: a novel biomarker for pancreatic neuroendocrine tumor aggressiveness.

PURPOSE: We aimed to test whether α-internexin could be a molecular biomarker of tumor aggressiveness and prognosis in pancreatic neuroendocrine tumors (PNETs). PATIENTS AND METHODS: Using immunohistochemical staining and Western blot, we detected the expression of α-internexin in 350 tumors from 343 patients, of whom 257 were followed up. Methylation of α-internexin promoter was examined by bisulfite sequencing to identify the crucial region that determines gene expression. Methylation of gene promoter in tumors was quantitatively measured by denaturing high performance liquid chromatography (DHPLC). We correlated α-internexin expression with clinicopathological characteristics. RESULTS: α-Internexin was expressed in 53% of 350 PNETs. The reduced expression of α-internexin was significantly associated with advanced stage (P < .0001), metastases (P < .0001), and recurrence (P = .003). α-Internexin expression was found in 57.1% of 212 surviving patients and in 17.1% of 35 deceased patients (P < .0001). Reduced expression of α-internexin was associated with shorter overall survival in PNET patients (log rank P < .0001) as well as in patients with noninsulinoma and nonfunctional (NF)-PNETs (log rank P = 0.0073 and P = 0.010, respectively). The crucial region of α-internexin promoter (-149 to +96 nucleotides [nt]) was identified, and the hypomethylation of this area in PNETs was significantly associated with gene expression (P = .015). CONCLUSION: α-Internexin can be a useful prognostic biomarker for PNETs.

T cell immunohistochemistry refines lung transplant acute rejection diagnosis and grading.

OBJECTIVE: Lung transplant volume has been increasing. However, inaccurate and uncertain diagnosis for lung transplant rejection hurdles long-term outcome due to, in part, interobserver variability in rejection grading. Therefore, a more reliable method to facilitate diagnosing and grading rejection is warranted. METHOD: Rat lung grafts were harvested on day 3, 7, 14 and 28 post transplant for histological and immunohistochemical assessment. No immunosuppressive treatment was administered. We explored the value of interstitial T lymphocytes quantification by immunohistochemistry and compared the role of T cell immunohistochemistry with H&E staining in diagnosing and grading lung transplant rejection. RESULTS: Typical acute rejection from grade A1 to A4 was found. Rejection severity was heterogeneously distributed in one-third transplanted lungs (14/40): lesions in apex and center were more augmented than in the base and periphery of the grafts, respectively. Immunohistochemistry showed profound difference in T lymphocyte infiltration among grade A1 to A4 rejections. The coincidence rate of H&E and immunohistochemistry was 77.5%. The amount of interstitial T lymphocyte infiltration increased gradually with the upgrading of rejection. The statistical analysis demonstrated that the difference in the amount of interstitial T lymphocytes between grade A2 and A3 was not obvious. However, T lymphocytes in lung tissue of grade A4 were significantly more abundant than in other grades. CONCLUSIONS: Rejection severity was heterogeneously distributed within lung grafts. Immunohistochemistry improves the sensitivity and specificity of rejection diagnosis, and interstitial T lymphocyte quantitation has potential value in diagnosing and monitoring lung allograft rejection. VIRTUAL SLIDES: The virtual slides for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1536075282108217.

Improvements of the surgical technique on the established mouse model of orthotopic single lung transplantation.

BACKGROUND: A wide range of knockout and transgenic murine models for the study of nonimmune and immune mechanisms in lung transplants are available nowadays, but the microsurgical techniques are difficult to learn. We describe methods to simplify techniques and facilitate learning. METHODS: Traditional procedures were implemented to perform lung transplants in 30 cases (group 1). Improved techniques which included cuff without tail, broadening of the cuff diameter for bronchus, establishment of one tunnel between three structures, innovative technology of the vascular anastomosis and placement of the chest tube post-operation were used to perform lung transplants in 30 cases (group 2). RESULTS: The improved techniques considerably shorten operative times (96.75 ± 6.16 min and 85.32 ± 6.98 min in groups 1 and 2, respectively). The survival rates in the recipient animals were 86.7% and 96.7% in groups 1 and 2, respectively. Chest X-rays and macroscopic changes of transplanted recipients showed that grafts were well inflated on postoperative day 30. There was no significant difference of the arterial oxygen tension (PaO2) between two groups (115.9 ± 7.11 mm Hg and 116.3 ± 6.87 mm Hg in groups 1 and 2, respectively). Histologically, no lung injury was seen in grafts. CONCLUSIONS: We described the modified procedures of orthotopic left lung transplants in mice, which could shorten operative time and increase survival rate.

Orthotopic and heterotopic tracheal transplantation model in studying obliterative bronchiolitis.

BACKGROUND: Several animal models have been established to investigate the mechanisms of obliterative bronchiolitis after lung transplantation. In this study, we compared three prevalent murine models of obliterative bronchiolitis in terms of several basic pathologic changes in a relatively short span of time after transplantation. METHODS: Each of the recipient mice simultaneously received orthotopic, intra-omental and subcutaneous tracheal transplantation in both syngeneic and allogeneic settings. No immunosuppressive treatment was administered. Tracheal grafts were harvested on Day 14, 21 and 28 after transplantation for histological and immunohistochemical analyses. RESULTS: Syngeneic tracheal grafts from different transplant sites retained normal histologic structures, while their corresponding allografts demonstrated more occlusion of the airway lumen as well as more infiltration of CD4(+)/CD8(+) mononuclear cells and myofibroblasts, but less regenerative epithelium and neovascularized vessels at indicated times (P<0.05). Compared with two heterotopic allografts, orthotopic allografts had less occlusion of the tracheal lumen as well as less infiltration of CD4(+)/CD8(+) mononuclear cells and myofibroblasts, but more regenerative epithelium and neovascularized vessels (P<0.05). CONCLUSIONS: Orthotopic tracheal transplantation in mice can be considered as a model to study early stages of obliterative bronchiolitis, and heterotopic tracheal transplantation can be a model for late stages of obliterative bronchiolitis.