Evaluating tumor-infiltrating lymphocytes in hepatocellular carcinoma using hematoxylin and eosin-stained tumor sections.

BACKGROUND: Tumor-infiltrating lymphocytes (TILs) constitute a prognostic factor in hepatocellular carcinoma (HCC). However, different methods of assessing TILs have various pre-analytical, analytical, and post-analytical challenges. The evaluation of TILs in hematoxylin and eosin (H&E)-stained tumor sections proposed by the International Immuno-Oncology Biomarker Working Group was demonstrated to be a reproducible, affordable and easily applied method in many tumors. AIM: To evaluate the prognostic significance of TILs in H&E-stained slides of HCCs. METHODS: This was a retrospective study performed in the hospital. HCC patients who underwent liver resection between 2015 and 2017 in Zhongshan Hospital were enrolled in this study. Patients who experienced recurrence or received therapy in addition to antiviral therapy before surgery at this time were excluded. A total of 204 patients were enrolled in the study. The ILs were counted manually in tumor sections stained with H&E under an optical microscope at 400 ×. The ILs were assessed separately in the center of the tumor (TILsCT), the invasive front (TILsIF), and peritumor (PILs) areas. Univariate and multivariate survival analyses were performed using a Cox regression model. P < 0.05 was considered statistically significant and all P-values were two-sided. RESULTS: Among the 204 patients, univariate analysis indicated that macrovascular invasion (MaVI) (P = 0.001), microvascular invasion (MVI) (P = 0.012), multiple tumors (P = 0.008), large tumors (> 10 cm) (P = 0.001), absence of a tumor capsule (P = 0.026), macrotrabecular histological subtype (P = 0.001), low density of TILsCT (P = 0.039), TILsIF (P = 0.014), and PILs (P = 0.010) were predictors of progression-free survival (PFS). Cox multivariate analysis indicated that MaVI (P = 0.009), absence of a tumor capsule (P = 0.031), low-density of TILsIF (P = 0.047) and PILs (P = 0.0495) were independent predictors of PFS. A three-category analysis was carried out by combining TILsCT, TILsIF, and PILs, after which HCCs were classified into immunehigh [(TILsCT)high, (TILsIF)high, and PILshigh, 83 cases], immunemod (tumors other than immunehigh and immunelow subtypes, 94 cases), and immunelow [(TILsCT)low, (TILsIF)low, and PILslow, 27 cases)] subtypes. The immunehigh subtype had a lower rate of MVI (40.96%) than the immunemod (61.70%, P = 0.017) and immunelow (66.67%, P = 0.020) subtypes. The recurrence rates of the immunehigh, immunemod and immunelow subtypes were 10.8%, 25.5% and 33.3%, respectively. CONCLUSION: HCC patients with high infiltrating lymphocytes tend to have a lower recurrence rate and less MVI. The evaluation of TILs in H&E-stained specimens could be a prognostic parameter for HCC.

Caspase Recruitment Domain Containing Protein 9 Suppresses Non-Small Cell Lung Cancer Proliferation and Invasion via Inhibiting MAPK/p38 Pathway.

PURPOSE: Caspase recruitment domain containing protein 9 (CARD9) has been demonstrated to be a pro-tumor factor in various cancers. However, our previous study found a significant decrease of CARD9 in malignant pleural effusion compared with benign pleural effusion. So we investigated the role of CARD9 in non-small cell lung cancer (NSCLC) and its working mechanism. MATERIALS AND METHODS: Immunohistochemistry, western blot, and quantitative real-time polymerase chain reaction were used to detect the expression of CARD9 in specimens of NSCLC patients. The Cancer Genome Atlas (TCGA) databasewas also used to analyze the expression of CARD9 in NSCLC and its predicting value for prognosis. Immunofluorescence was used for CARD9 cellular location. Cell growth assay, clonal formation assay, wound healing assay, matrigel invasion assay, and flow cytometry were used to test cell proliferation, migration, invasion, apoptosis, and cycle progression of NSCLC cells with CARD9 knockdown or CARD9 overexpression. Co-immunoprecipitation was used to identify the interaction between CARD9 and B-cell lymphoma 10 (BCL10). SB203580 was used to inhibit p38 activation. RESULTS: CARD9 was decreased in NSCLC tissues compared with normal tissues; low CARD9 expression was associated with poor survival. CARD9 was expressed both in tumor cells and macrophages. Downregulation of CARD9 in NSCLC cells enhanced the abilities of proliferation, invasion and migration via activated MAPK/p38 signaling, while overexpression of CARD9 presented antitumor effects. BCL10 was identified to interact with CARD9. CONCLUSION: We demonstrate that CARD9 is an independent prognostic factor in NSCLC patients and inhibits proliferation, migration, and invasion by suppressing MAPK/p38 pathway in NSCLC cells.

Self-activation of biochar from furfural residues by recycled pyrolysis gas.

Biochar samples with controllable specific surface area and mesopore ratio were self-activated from furfural residues by recycled pyrolysis gas. The objective of this study was to develop a new cyclic utilization method for the gas produced by pyrolysis. The influences of preparation parameters on the resulting biochar were studied by varying the pyrolysis-gas flow rate, activation time and temperature. Structural characterization of the produced biochar was performed by analysis of nitrogen adsorption isotherms at 77 K and scanning electron microscope (SEM). The pyrolysis gas compositions before and after activation were determined by a gas chromatograph. The results indicated that the surface area of the biochar was increased from 167 m2/g to 567 m2/g, the total pore volume increased from 0.121 cm3/g to 0.380 cm3/g, and the ratio of the mesopore pore volume to the total pore volume increased 17-39.7%. The CO volume fraction of the pyrolysis gas changed from 34.66 to 62.29% and the CO2 volume fraction decreased from 48.26% to 12.17% under different conditions of pyrolysis-gas flow rate, activation time and temperature. The calorific values of pyrolysis gas changed from 8.82 J/cm3 to 14.00 J/cm3, which were higher than those of conventional pyrolysis gases. The slower pyrolysis-gas flow rate and higher activation time increased the efficiency of the reaction between carbon and pyrolysis gas. These results demonstrated the feasibility of treatment of the furfural residues to produce microporous and mesoporous biochar. The pyrolysis gas that results from the activation process could be used as fuel. Overall, this new self-activation method meets the development requirements of cyclic economy and cleaner production.

Hepatitis C virus infection and risk of thyroid cancer: A systematic review and meta-analysis.

PURPOSE: Several epidemiological studies investigated the relationship between hepatitis C virus (HCV) infection and risk of thyroid cancer, but the results were not consistent. A systematic review and meta-analysis was conducted to assess the impact of HCV infection on thyroid cancer risk. METHODS: The literature was searched up to March 15, 2016 for case-control or cohort studies on the association between HCV infection and thyroid cancer risk. The summary relative risks (RR) and 95% confidence intervals (CI) were calculated. RESULTS: Five studies (two case-control studies and three cohort studies) were included in the meta-analysis, with a total of 751,551 participants and 367 cases of thyroid cancer. Meta-analysis of those 5 studies found that there was no statistically significant association between HCV infection and thyroid cancer risk (summary RR=2.09, 95%CI 0.78-5.64, p=0.145; I2=81.2%). However, HCV infection was significantly associated with increased risk of thyroid cancer (summary RR=2.86, 95%CI 1.63-5.03, p=0.003; I2=24.9%) after adjusting the heterogeneity. CONCLUSION: There is a possible association between HCV infection and increased risk of thyroid cancer, and more cohort studies are needed to validate the possible association.

Low-dose volume perfusion computed tomography (VPCT) for diagnosis of solitary pulmonary nodules.

OBJECTIVE: To characterize solitary pulmonary nodules (SPN) in terms of perfusion parameters using low-dose volume perfusion computed tomography (VPCT) and two different calculation methods, compare their results, look for interobserver agreement of measurements and investigate the feasibility of low-dose VPCT. MATERIALS AND METHODS: This study was approved by the local Institutional Review Board and all patients provided written informed consent. Seventy-one patients (mean age 60.8 years ±9.6) with solitary pulmonary nodules were enrolled. Low-dose VPCT was performed for 38.63s covering the involved lung (70kV, 120mAs, 22 consecutive volume measurements, 50mL iodinated contrast, flow rate 4mL/s). Mean blood flow (BF), blood volume (BV) and k-trans were determined both with the maximum slope+Patlak vs. deconvolution method. Additionally, the difference of VPCT parameters between different type lesions and normal tissue was analyzed. Interobserver agreement for all perfusion parameters was calculated using intraclass correlation coefficients (ICC). The effective radiation dose of the VPCT and the total CT scan protocol were recorded. All CT findings were histologically confirmed by surgical intervention. RESULTS: The mean lesion size was 18.6mm. Interobserver agreement measure with ICC shows high agreement between the measurements (κ=0.85). The effective radiation dose of the VPCT was 9.3mSv. The mean perfusion values for BF, BV and k-trans of 120.6mL/100g tissue/', 11.6mL/100g tissue/', and 18.5mL/100g tissue/' for the deconvolution method, and 50.1mL/100g tissue/', 11.6mL/100g tissue/' and 24.3mL/100g tissue/' for the maximum slope+Patlak method, which were significantly higher than those of normal muscle (20.7mL/100g tissue/', 2.6mL/100g tissue/', and 7.6mL/100g tissue/' for the deconvolution method and 10.9mL/100g tissue/', 3.1mL/100g tissue/' and 8.1mL/100g tissue/' for the maximum slope method). The best overall correlation between calculation methods was achieved for measurements of BF. CONCLUSION: The low-dose volume perfusion CT of the solitary pulmonary nodules can effectively reduce the radiation dose and non-invasively assess perfusion of SPN within the entire lesion volume.

The recycling of incinerated sewage sludge ash as a raw material for CaO-Al2O3-SiO2-P2O5 glass-ceramic production.

In this paper, the recycling of incinerated sewage sludge ash (ISSA) into glass-ceramic materials by a two-stage sintering cycle of nucleation stage and crystallization stage without any pressure and binder is presented. The parent glasses were subjected to the following nucleation/crystallization temperature and time level: (A) 790°C, 1.0 h/870°C, 1.0-3.0 h; (B) 790°C, 1.0 h/945°C, 1.0-3.0 h and (C) 790°C, 1.0 h/1065°C, 1.0-3.0 h. X-ray power diffraction analysis results revealed that multiple crystalline phases coexisted in the glass-ceramic materials and the crystalline phase compositions were more affected by crystallization temperature than crystallization time. Scanning electron microscopy analysis showed an interlocking microstructure of glass phases and crystals with different sizes and spatial distribution. The glass-ceramics crystallized at 945°C for 2.0 h exhibited optimal properties of density of 2.88±0.08 g/cm3, compression strength of 247±12 MPa, bending strength of 118±14 MPa and water absorption of 0.42±0.04. The leaching concentrations of heavy metals were far lower than the limits required by the regulatory standard of EPA. This paper provides a feasible, low-cost and promising method to produce ISSA-based glass-ceramics and highlights the principal characteristics that must be taken into account to use ISSA correctly in glass-ceramics.

[Immunohistochemical classification and prognosis of diffuse large B-cell lymphoma in China].

OBJECTIVE: To study the immunohistochemical classification and prognosis of diffuse large B-cell lymphoma (DLBCL). METHODS: A total of 148 cases of DLBCL were classified into germinal center B-cell-like (GCB) and non-GCB/activated B-cell-like (ABC) subtypes by Hans, Choi and Tally immunohistochemical stain algorithms. The clinical features and survival data of GCB and non-GCB/ABC subtypes were compared. Multivariate analysis about clinical features and results of immunohistochemical stain algorithms was carried out by using Cox regression, with overall survival as the outcome. RESULTS: The prevalence of GCB subtype was significantly lower than that of non-GCB/ABC subtype, as classified by whichever algorithms in the 148 DLBCL cases studied. The prevalence of GCB subtype by Tally algorithm was lowest. The prevalence of GCB subtype (19 cases, 16.7%) was also significantly lower than non-GCB/ABC subtype (95 cases, 83.3%; P = 0.000 1) in the 114 (77.0%) concordant cases by the three algorithms. There was no difference between GCB and non-GCB/ABC subtypes by the three algorithms in five-year overall survival rate and survival curve of the 80 DLBCL patients with follow-up data available (P > 0.05). Primary gastric DLBCL tended to show a higher prevalence of GCB subtype, a better five-year overall survival rate and survival curve than the other groups. Multivariate analysis showed that patient age (HR = 1.036, P = 0.001) and tumor stage (HR = 1.997, P = 0) were also significantly adverse predictors of overall survival. CONCLUSION: The Hans, Choi and Tally immunohistochemical stain algorithms cannot effectively classify Chinese DLBCL into different prognostic subtypes. Primary gastric DLBCL has different immunophenotype and outcome, as compared with DLCBL in other sites.

Relationship between programmed death-ligand 1 and clinicopathological characteristics in non-small cell lung cancer patients.

OBJECTIVE: To evaluate the correlation between programmed death-ligand 1 (PD-L1) expression in primary lung cancer cells, tumor associated macrophages (TAM) and patients' clinicopathological characteristics. METHODS: From 2008 to 2010, 208 non-small cell lung cancer patients who underwent surgery or CT-guided biopsy were recruited from Huadong Hospital, Fudan University. Immunohistochemistry staining was performed to evaluate the PD-L1 expression in both primary lung cancer cells and CD68 positive TAM. The relationship between PD-L1 expression and the clinical pathology was evaluated using χ(2) test. Spearman's rank correlations were used to determine the correlation between PD-L1 expression in tumor cells and macrophages. RESULTS: Positive PD-L1 expression in primary cancer cells was found in 136 (65.3%) patients, which were negatively correlated with lymph node metastasis (P=0.009) and smoking history (P=0.036). Besides, TAM with PD-L1 expression (found in 116 patients) was positively associated with smoking history (P=0.034), well-differentiation (P=0.029) and negative lymph node metastasis (P=0.0096). A correlation between PD-L1 expression in primary tumor cells and non-small cell lung cancer associated macrophages was found (r=0.228, P=0.021). CONCLUSION: PD-L1, secreted from TAM, might induce cancer cells apoptosis, and decrease lymph node metastasis.

Expression of monocyte chemoattractant protein-1 and CC chemokine receptor 2 in non-small cell lung cancer and its significance.

Expression of monocyte chemoattractant protein-1 (MCP-1) and CC chemokine receptor 2 (CCR2) and its significance has been demonstrated in some cancer cells in recent clinical studies. However, the role of tumor MCP-1 and CCR2 expression in non-small cell lung cancer (NSCLC) remains unknown. The aim of the present study was to investigate the prognostic significance of MCP-1 and CCR2 expression in NSCLC cells. The relationship between MCP-1 and CCR2 expression in NSCLC cancer cells was examined by immunohistochemical staining of surgical specimens from 134 patients. Sixty-five of these patients had follow-up records. Kaplan-Meier analysis and Cox regression model were used to assess overall survival according to the presence or absence of MCP-1 and CCR2 expression in tumor cells. MCP-1 was detected in cancer cells of 107 NSCLC (79.9 %) and CCR2 was detected in cancer cells of 39 NSCLC (29.1 %). MCP-1 expression was correlated with sex, smoking habits, histology, and tumor size. Presence of MCP-1 in tumor cells was associated with better overall survival (P = 0.018). By multivariate analysis, MCP-1 expression in cancer cells showed an independent prognostic factor for overall survival (P = 0.002, hazard ratio [HR] = 0.256, 95 % confidence interval [CI] = 0.106-0.616). There was no significant relationship between CCR2 expression in tumor cells and clinical and pathological characteristics. Also, no significant positive correlation between MCP-1 and CCR2 expression was revealed by Spearman correlation analysis. Our data indicate that MCP-1 is overexpressed in NSCLC cells. Its expression in cancer cells is associated with better survival in NSCLC patients.

Expression of the adipocytokine resistin and its association with the clinicopathological features and prognosis of pancreatic ductal adenocarcinoma.

Fat tissue is viewed as an active endocrine organ that secretes a variety of bioactive substances. Resistin, an adipocyte-secreted factor, is thought to be closely related to obesity, insulin resistance and inflammation, the three most significant risk factors for the progression of pancreatic cancer. However, the association between resistin and pancreatic cancer is still unknown. In this study, pancreatic tumor samples from 45 patients with pancreatic ductal adenocarcinoma were analyzed with immunohistochemistry for the expression of resistin. The correlation between resistin expression and clinicopathological features and prognosis were evaluated. Resistin staining was observed in 48.9% (22 of 45) of the cases. Resistin expression was more frequent in poorly differentiated tumors (9 of 9, 100%) compared to moderately differentiated tumors (11 of 28, 39.3%) and well-differentiated tumors (2 of 8, 25%) (p<0.01). The incidence of resistin expression in patients with Japan Pancreas Society stages III-IV (18 of 27, 66.7%) was significantly higher than in subjects with stages I-II (4 of 18, 22.2%) (p<0.01). Patients with resistin-stained tumors had significantly shorter relapse-free survival times (median, 9 months) than patients with negative tumors (median, 18 months) (p<0.05). In addition, multivariate analysis showed that resistin expression was an independent prognostic factor for relapse-free survival of patients with pancreatic ductal adenocarcinoma (p<0.05). These results demonstrate that resistin may influence the progression of pancreatic tumors and may be a useful predictor of relapse-free survival in patients with pancreatic ductal adenocarcinoma.

[Cytogenetic profiles of follicular lymphoma].

OBJECTIVE: To study the cytogenetic profiles of follicular lymphoma (FL) in Chinese patients. METHODS: Conventional karyotype in 57 FL patients from Shanghai area was analyzed. Fluorescence in-situ hybridization (FISH) for t(14;18) and Bcl-6 and IgH gene rearrangement was performed in these cases. RESULTS: The most frequent breakpoints (frequency > or = 10% ) of the 57 FL cases were at band 14q32 (68.4%), 18q21 (38.6%), 3q27 (21.1%), 1q10 (15.8%) and 1q21 (12.3%). Nineteen (33.3%) of the 57 cases had t(14;18). The breakpoint of 18q21 and t(14;18) were more frequent in FL grade 1-2 and less frequent in FL grade 3 (57.6% vs. 12.5%; 54.5% vs. 4.2%, P < 0.05), whereas the 3q27/Bcl-6 rearrangement was more frequent in FL grade 3 and less frequent in FL grade 1-2 (37.5% vs. 9.1% , P < 0.05). The cohort of FL was more frequent in gains of chromosomes X, 1q, 5, 6p, 7 and 12q and losses of chromosomes 1p, 6p and 14q32. Gain of 18q was more frequent in FL grade 1-2 than in FL grade 3 (P < 0.05). Loss of 14q32 was more frequent in t(14;18) negative FL than in t(14;18) positive FL (P < 0.05). CONCLUSIONS: Compared with the data of Western patients reported in the literature, Chinese FL cases had distinct cytogenetic profiles from Western FL cases that the t(14;18) is less frequent and the gain of 1q is more frequent in Chinese FL cases, which are more significant in high grade FL.

[Urothelial hyperplastic lesion with endophytic growth pattern: a clinicopathologic study].

OBJECTIVE: To study the clinicopathologic features of urothelial hyperplastic lesion with an endophytic growth pattern and the role of immunohistochemistry and multitargeted fluorescence in situ hybridization (FISH) in the differential diagnosis. METHODS: Forty-one cases of urothelial lesions exhibiting endophytic growth patterns were reviewed and reclassified as inverted papilloma, urothelial carcinoma with an endophytic growth pattern, and florid von Brunn nest. The gains of chromosomes 3, 7, and 17 and loss of 9p21 was detected by FISH, and performed immunohistochemical staining for CK20, p53, and Ki-67. Follow-up data of 12 cases were obtained. RESULTS: (1) Twelve inverted papillomas sized 1.2 cm in average, consisted of anastomosing cords and nests with uniform width distribution involving the lamina propria, the central portion contained streaming cells with squamous metaplasia, and the periphery showed palisading. No or rare atypia and mitosis were found. Focal exophytic papillary component lined by less than 6 layers of normal urothelium were observed in 4 cases. (2) Twenty-four urothelial carcinomas with an endophytic growth pattern sized 2.1 cm in average, demonstrated the similar architecture with inverted papilloma, but exhibited thick columns and variable thickness of the cords, irregular size and shape of large nests with transition into solids. Mild to moderate cytologic atypia was shown, and mitotic figures ranged 1 to 8 per 10 HPFs. Exophytic papillary component was not observed in 3 cases, but the superficial urothelium showed dysplasia, while coexisted exophytic component in other cases was associated with low malignant potential or low grade tumor. (3) Five florid von Brunn nests sized 0.9 cm in average, had normal or hyperplastic urothelium, variable nests with cysts compacted in lamina propria, no cytologic atypia and mitosis. Twenty-one of 24 (79.1%) urothelial carcinomas with an endophytic growth pattern displayed abnormally positive results by multitargeted FISH, whereas all inverted papillomas and florid von Brunn nests were negative. Immunohistochemically, CK20 was weakly positive in 2 cases of urothelial carcinoma with an endophytic growth pattern, and negative in all inverted papillomas and florid von Brunn nests. p53 weakly stained 5% to 50% nuclei of the tumor cells in 16 cases of urothelial carcinomas with an endophytic growth pattern and 1 inverted papilloma. 1%-5% tumor cells expressed Ki-67 in urothelial carcinoma with an endophytic growth pattern, and less than 1% in inverted papilloma and florid von Brunn nests. Follow-up study revealed that 2 cases of urothelial carcinoma with an endophytic growth pattern had developed invasive carcinoma, underwent cystectomy, and metastasized remotely. No recurrence occurred in cases of inverted papilloma. CONCLUSIONS: Benign and malignant urothelial lesions with an endophytic growth pattern present histologic overlapping. Urothelial carcinoma with an endophytic growth pattern displays unique characteristics in morphology and immunohistochemistry. Multitargeted FISH analysis is helpful in the differential diagnosis.

[Value of spiral CT in diagnosis of cystic renal cell carcinoma].

OBJECTIVE: To investigate the image features and the diagnostic value of spiral CT for cystic renal cell carcinoma. METHODS: The clinical data and CT manifestations of 17 operated and pathologically proven cystic renal cell carcinoma were retrospectively analyzed. There were 12 males and 5 females with an average age of 47.3 years (33 - 82 years). Plain and contrast CT scan (Siemens somatom) single layer sensation 16 layer spiral CT had been performed before operation. The image of artery phase (30 - 40 s), venous (60 - 70 s) and excretory (120 - 180 s) were respectively obtained after contrast administration. Various image reconstructions were done using Siemens Wizard workstation based on the raw images. RESULTS: It was found that 5 cystic renal cell cancers located in the right kidney and 12 in the left kidney. The long dimension of the tumor arranged from 21 - 100 mm with an average of 57 mm. The tumor looked like a round or round-like shape with density similar to fluid on plain CT scan. Some cystic renal carcinomas had a thick wall. Some had single or multiple cystic spaces filled with fluid of different densities. Some had infiltrated out of kidney surface or into renal sinus. Some showed enhanced nodules on the wall. CONCLUSION: Cystic renal cell carcinoma has its own specific morphologic features in spiral CT scan. Spiral CT may be very helpful in the diagnosis of cystic renal cell carcinoma before operation.