| Objective:To investigate the proliferation status of helper T lymphocytes,dendritic cells,monocytes,and B lymphocytes in focal and paracancerous tissues of papillary thyroid carcinoma(PTC)complicated with Hashimoto’s thyroiditis(HT),and to analyze the clinical characteristics of the patients,to explore the changes of local immune microenvironment in PTC+HT patients,so as to provide a new idea for the study of the pathogenesis of PTC+HT.Methods:1.Sample collection and inclusion criteria(1)Sample collection: 54 patients with papillary thyroid carcinoma who were hospitalized in the Department of Basic Surgery of the Affiliated Hospital of Hebei University from May 2019 to July 2020 were selected,including 17 male patients and 37 female patients.All patients underwent total thyroidectomy or subtotal thyroidectomy and were diagnosed as papillary thyroid carcinoma.The age,gender,tumor size,tumor stage and other clinical characteristics of patients were recorded.(2)Inclusion criteria:(1)The samples were pathologically diagnosed as PTC or PTC+HT in both intraoperative frozen sections and postoperative paraffin sections;(2)The patient did not receive any preoperative chemoradiotherapy;(3)Patients did not have hyperthyroidism and other thyroid diseases;(4)All patients were newly diagnosed with PTC;(5)Diagnostic criteria for HT: diffuse thyroid enlargement,significantly increased serum TPOAb and TGAB;lymphocytic infiltration was found in thyroid fine needle biopsy;a large number of lymphocytes were infiltrated in paraffin section after operation,which was confirmed as HT.(6)No other tumor-related diseases;(7)Complete clinical and pathological data.(3)Exclusion criteria:(1)Unclear diagnosis or other types of thyroid cancer;(2)A history of thyroid or lymph node related;(3)The head and neck underwent radiotherapy and chemotherapy;(4)Have a history of other malignant tumors.2.Experiment groupingThis study was divided into 2 groups: PTC+HT group,13 cases;PTC group,41 cases.All patients ranged in age from 21 to 68 years,with a median age of 55 years and a mean age of 53±11.3 years.3.Treatment and HE staining of tissue samples(1)Fresh cancer tissues were collected during the operation,and the adjacent tissues were cut at the farthest distance from the tumor as the control.The removed tissue was first put into a 50 ml centrifugal tube,which was quickly cooled in liquid nitrogen,and then wrapped with aluminum foil or plastic film and transferred to a-80 ℃ refrigerator for preservation for subsequent experiments.(2)HE staining techniques were used to observe PTC + HT with PTC group of cancer tissue and tissue adjacent to carcinoma pathology morphology,and randomly selected from three different area of the same field of vision.The total number of lymphocytes and the total number of cells were counted and recorded,and the ratio of total number of lymphocytes(%)= total number of lymphocytes in cancer(paracancer)tissue/total number of cells in cancer(paracancer)tissue ×100.The proliferation status of lymphocytes was observed and the differences were compared.4.The proliferation status of helper T lymphocytes,monocytes,B lymphocytes and dendritic cells in tumor focal tissues and adjacent tissues of PTC+HT group and PTC group was observed by immunofluorescence stainingCD4+T lymphocytes,CD14+monocytes,CD19+B lymphocytes and CD83+ dendritic cells were detected respectively.Their proliferation status in the local immune microenvironment of PTC+HT and PTC patients was analyzed by calculating their average optical density and positive rate,and the differences were compared.Result:1.The clinical characteristics of PTC+HT and PTC patients were analyzed.Compared with the PTC group,the tumor diameter of PTC+HT patients was smaller,but there was no significant difference in age,gender,TNM stage and lymph node metastasis between the two groups.2.HE staining method was used to analyze the PTC + HT and PTC groups overall state of lymphocyte proliferation.Lymphocyte proliferation in PTC+HT paracancerous tissues was significantly more than that in PTC paracancerous tissues,lymphatic follicle formation and part of the lymphogenic centers were observed.The proliferation of lymphocytes in PTC carcinoma tissues was significantly higher than that in paracancerous tissues.Lymphocyte proliferation in PTC+HT carcinoma tissues was significantly higher than that in PTC carcinoma tissues.3.The proliferation status of CD4+T lymphocytes,CD14+ monocytes,CD19+B lymphocytes and CD83+ dendritic cells in PTC+HT and PTC cancer tissues and paracancerous tissues were detected by immunofluorescence staining.Multiple analysis showed that:(1)CD4+T lymphocyte proliferation status: the average optical density and positive rate in PTC cancer tissues were significantly higher than those in adjacent tissues(P<0.001,P<0.05).The mean optical density in PTC+HT paracancerous tissue was significantly higher than that in PTC group(P<0.001).The average optical density and positive rate in PTC + HT cancer tissues were significantly higher than those in PTC group(P < 0.001,P < 0.05).(2)CD14+ mononuclear cells: the average optical density and positive rate in PTC cancer tissues were significantly higher than those in adjacent tissues(P<0.001,P<0.01);The positive rate in PTC+HT paracancerous tissue was significantly higher than that in cancer tissues(P<0.001).The positive rate in PTC cancer tissues was significantly higher than that in PTC+HT group(P<0.001).The mean optical density in PTC+HT paracancerous tissue was significantly higher than that in PTC group(P<0.001).(3)CD19+B lymphocytes: the average optical density in the PTC+HT cancer tissues was significantly higher than that in the paracancer tissue,and the positive rate was lower than that in the paracancer tissue(P<0.05,P<0.001).The average optical density and positive rate in PTC+HT cancer tissues were significantly higher than those in PTC group(P<0.001,P<0.001).The average optical density and positive rate in PTC+HT paracancer tissues were significantly higher than those in PTC group(P<0.001,P<0.001).(4)CD83+ dendritic cells:the average optical density and positive rate in PTC+HT cancer tissues were significantly higher than those in adjacent tissues(P<0.001,P<0.05).The average optical density and positive rate in PTC cancer tissues were significantly higher than those in adjacent tissues(P<0.001,P<0.01).The average optical density in PTC+HT cancer tissue was significantly higher than that in PTC cancer tissue(P<0.001).The mean optical density in PTC+HT paracancerous tissues was significantly higher than that in PTC(P<0.001).Conclusion:1.The diameter of tumor focus in PTC+HT patients is generally smaller,which may be affected by immune function in the process of PTC growth.2.A large number of lymphocyte proliferation,lymphatic follicle formation and lymphatic germinal center were observed in the carcinoma and paracancerous tissues of PTC+HT patients,indicating that PTC+HT patients had an obvious local immune response,which may be the process of developing from HT to PTC,or PTC stimulated obvious immune response;At the same time,the proliferation of CD4+T lymphocytes,CD14+monocytes,CD19+B lymphocytes and CD83+ dendritic cells increased significantly,indicating that the local immune response occurred in PTC+HT,the local immune microenvironment of PTC+HT was changed,and the obvious immune response occurred in the local immune microenvironment of PTC.3.The proliferation of CD4+T lymphocytes,CD14+ monocytes and CD83+ dendritic cells,which are the main types of inflammation,was significantly increased in PTC+HT patients,indicating that they were involved in tissue destruction.While the proliferation of CD19+B lymphocytes was large,indicating that PTC may stimulate the human autoimmune response.Under the action of the above immune cells,it may influence the development of PTC+HT local tumors. |
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