A Study Focused On Tumor Immune Microenvironment During Gastric Cancer Invasion And Metastasis

Part I:The tumor stromal ratio as a strong prognosticator for advanced gastric cancer patientsObjective Insufficient attention is paid to the underlying tumor microenvironment(TME)evolution,that resulting in tumor heterogeneity and driving differences in cancer aggressiveness and treatment outcomes.The morphological evaluation of the proportion of the stroma at the most invasive part of primary tumor(tumor stromal ratio,TSR)in cancer is gaining momentum as evidence strengthens for the clinical relevance.Methods Tissue samples from the most invasive part of the primary gastric cancer(GC)of 494 patients were analyzed for their TSR,and correlations between TSR and major clinic-pathological features and overall survival(OS)of GC patients were established and assessed.Results TSR is a new and strong independent prognostic factor for GC patients.The likelihood of tumor invasion is increased significantly for patients in the stromal-high subgroup compared to those in the stromal-low subgroup(P = 0.011).The discrimination ability of TSR was not less than the TNM staging system and was better in patients with stages I and II GC.Conclusion This study confirms the TSR as a TME prognostic factor for GC.TSR is a candidate TME parameter that could easily be implemented in routine pathology diagnostics.Part Ⅱ:Morphological study and comprehensive cellular constituents of milky spots in the human omentumObjective To analyze morphological features of omental milky spots(MS).Methods Hematoxylin-eosin staining and immunohistochemistry technique were used to study the omental MS of gastric cancer(GC)patients and rectal cancer(RC)patients.We focused on morphological features of MS and conducted quantitative analysis on the cells number and cellular constituents.Differences in MS parameters between GC and RC were also analyzed.Results Various shapes of MS were mainly round,oval,irregular form in the adipose and perivascular annular.The median MS perimeter was 2752(range 817～7753)computer-based pixels.The median value of immune cells in one MS was 141(43～650),comprising T lymphocytes(46.1%),B lymphocytes(28.4%),macrophages(12.4%)and other immune cells(13.1%).Relatively high density of vessels in MS could be calculated by microvessel density(MVD)as 4(0～13).The median value of mesothelial cells loosely arranged in the surface layer was 5(0～51).There were no signifcant differences in MS perimeter,MVD,the number of mesothelial cells,total immune cells,T lymphocytes and macrophages between GC and RC(P>0.05),while the number of MS B lymphocytes in RC was signifcantly higher than that in GC(P<0.001).Conclusion MS are structural bases for development and progression of peritoneal dissemination of GC and RC.Analyzing the morphology and cellular constituents could help understanding the mechanism of peritoneal metastasis.Tumor immune microenvironment within MS are composed of lymphocytes and macrophages.Part Ⅲ:The percentage of stromal tumor-infiltrating lymphocytes correlates with prognosis in patients with gastric cancerObjective Tumor-infiltrating lymphocytes(TIL)become increasingly relevant with tumor progression.The present study aimed to investigate the impact of TIL on the prognosis of patients with gastric cancer(GC).Methods The percentage of stromal TIL(psTEL)was reported semi-quantitatively by hematoxylin and eosin(H&E)staining-based evaluation.Herein,we screened one independent cohort of GC(n = 481)for psTIL characterization.Correlations between psTIL and clinic-pathological features,as well as overall survival(OS)were further explored.Additionally,psTIL was evaluated by receiver operating characteristic curve(ROC).Results A two-grade scale for psTIL was appropriate to be divided into-low and-high subgroups,using the median value 10.0%as the threshold.High psTIL was correlated with no serosa invasion,earlier TNM stage and better survival state(P<0.05 for all),and identified as a favorable prognostic factor both by uni-(HR:0.734,P = 0.047)and multivariate analysis(HR:0.722,P = 0.030).A beneficial OS of high psTIL was found in a linear manner with increasing TIL infiltrates associated with improved survival by survival curve(P = 0.030)and receiver operating characteristic curve(ROC)analysis(Area under the curve:0.432[95%CI:0.380-0.484],P=0.012).Conclusion The parameter psTIL by H&E staining-based evaluation could be served as an independent,favorable prognostic factor of GC.Part IV:Distribution pattern of tumor associated macrophages predicts the prognosis of gastric cancerObjective As mayor biomarkers in tumor microenvironment(TME),tumor associated macrophages(TAM)of gastric cancer(GC)still needs further studies in terms of the number and distribution pattern.Methods Herein,tissue microarrays(TMA)incorporating 494 GC surgical samples in duplicate were stained for TAM infiltration analysis.TAM number was counted according to the locations,including infiltrating macrophages in cancer nest(MC),in invasive front(MF)and in stroma(MS).Correlations between TAM number,distribution pattern and clinic-pathological features and survival analyses were performed.Results Infiltrating macrophages number in GC tissues was much higher than that in peritumoral tissues.TAM number was not significantly correlated with the overall survival(OS).TAM distribution pattern could be categorized into MC or MF/MS dominant pattern,and correlated with histological grade(P = 0.001).The median OS of MF/MS dominant pattern(46.7,95%CI:40.1～53.3)was significantly shorter than that of MC dominant pattern(66.6,95%CI:58.3～74.9)(P = 0.002).By receiver operating characteristic curve(ROC)analysis,the predictive value of TAM distribution pattern was superior to histological grade and pM stage,but inferior to pN and TNM stage.Conclusion TAM distribution pattern could be an independent prognostic factor for the OS of GC patients,and patients with MF/MS dominant pattern had worse outcomes.