Olesya N. Tomchuk Postgraduate Student, Orenburg State Medical University, Russia

1. Cancer Middle East 2015 Morphological features of lymphoid tissue and microvessels in regional lymph nodes in gastric cancer Olesya N. Tomchuk Postgraduate Student, Orenburg State Medical University, Russia

2. Gastric cancer is one of the most prevalent malignant tumors in the world. High level of gastric cancer morbidity and mortality determines have undying interest in this problem. Thereby, searching of the gastric cancer factors connected with aggressiveness and prognosis of the disease is still actual. • One of the most important factors connected with progression of malignant tumors is angiogenesis. A number of authors noted to the close connection of angiogenesis activity with the depth of tumor invasion, presence of metastases in regional lymph nodes and prognosis of disease, including the gastric cancer.

3. Characteristics of patients 44 patients with gastric cancer 26 men (58,9%) and 18 women (41,1%). The average age – 61.6±7,3 years (from 34 to 75 years) The median – 61 years. The patients with decompensation of chronic diseases, acute infection pathology, severe allergic processes were not included in the study as well as the ones who received corticosteroids, antihistamines, non-steroidal anti-inflammatory drugs and neoadjuvant chemotherapy.

4. The distribution of patients according to the tumor location, histology, grade Upper third – 13 (29.5%) Middle third – 9 (20.5%) Lower third – 21 (47.7%) Total – 1 (2.3%). Tumors of intestinal type – 19 (43.2%) Tumors of diffuse type – 25 (56.8%) G1 – 12 (27,3%) G2 – 7 (15,9%) G3-G4 - 5 (11,4%) SRCC* – 20 (45,4%) *Signet ring cell carcinoma

5. The distribution of patients according to the stage T1-2N0M0 – 17(42,1%) T3N0M0 – 6 (13,2%) T3-4N1M0 – 7(15,9%) T3-4N2M0 – 14(28,8%)

6. Surgery Subtotal distal resection – 32 patients (72.6%) Subtotal proximal resection – 7 patients (15.9%) Gastrectomy – 5 patients (11.5%) D2 lymphadenectomy – 23 patients (52.3%) D2 lymphadenectomy – 21 patients (47.7%)

7. METHODS The specimens of lymph node without metastasis were stained with Mayer’s Hematoxylin and eosin and immunohistochemically with antibodies to CD34. The visualization system has included DAB and Hematoxylincounterstaining. For negative control sections, primary antibody was replaced with phosphate-buffered saline and processed in the same manner.

8. METHODS It was evaluated: • - Area of lymph node, • - Area of lymphoid follicles and germinal centers • - Average size of lymphoid follicles

9. On the relative unit area of lymph node equal to 1 cm2 we counted: • The density of lymphoid follicles. Depending on the density of lymphoid follicles, patients were divided in 2 groups: in the first group there were patients with low density of lymphoid follicles (<3.5 on unit area) in the second group – the patients with normal and high density of lymphoid follicles (>3.5 on unit area, 34 pertsintil). • Microvessel density in the lymphoid follicles and medulla of lymph node. By visual analog way using low magnification (x100) were evaluated: • The number of dilated capillaries with atypia (none, single- no more than two vessels in the field of view, multiple – more than two vessels in the field of view) • The presence of atypical vessels with blast phenotype of endothelial cells (absent, present) The obtained data were compared with the clinical features of gastric cancer: stage, size, localization, histology, tumor grade, 3-year overall survival and relapse-free survival.

10. Statistical methods • Spearman’s rank correlation or gamma correlation (for the evaluation of the correlations between different data); • Chi-square test (it was carried out to analyze the difference of distribution among the categorized data). • The survival was analyzed by the Kaplan-Meier method. • Log-rank test (it was used to compare survival curves between subgroups of patients). • Cox's proportional hazards model (it was used for multivariate analysis of prognostic factors). • Odds ratio (OR) and a 95% confidence interval (CI) (this methods were used for the estimated the association between the 3-year survival and different types of vessels). A value of P< 0.05 was considered statistically significant.

11. I. The normal capillaries in medulla of lymph node Staining with CD34, x200 μm

12. II. The dilated capillaries Staining with CD34, x200 μm

13. III. Atypical vessels with blast phenotype of endothelial cells Staining with CD34, x200 μm

14. III. Atypical vessels with blast phenotype of endothelial cells Staining with CD34, x200 μm

15. The clinical features of lymphoproliferativeprocesses and angiogenesis in regional lymph node in gastric cancer

16. The area of lymph nodecorrelates with: Area of lymph follicles(r=0,451, p=0,05) Area of germinal centers (r=0,394, p=0,014) Size of lymph follicles(ρ = 0,464, p = 0,004) Microvessel densityin lymph follicles (ρ=0,389, p=0,03) Density of lymph follicles (r=-0,641, p=0,001). The number of lymph follicles at the bondary of tumor and gastric mucosa(ρ=-0,714, p=0,006) The dilated capillaries in the gastric submucosa(ρ =0,027, p=0,04)

17. The area of lymph nodedepending on the area of lymph follicles

18. The area of lymph nodedepending on the area of germinal centers.

19. The area of lymph nodedepending on size of lymph follicles

20. The area of lymph nodedepending on microvessel densityin lymph follicles

21. The area of lymph nodedepending on the density of lymph follicles

22. The area of lymph nodedepending on the number of nodal in lymph follicles area in the border of the tumor-mucosa

23. The area of lymph nodedepending on the dilated capillaries in the gastric submucosa

24. The area of lymph folliclescorrelates with: Thedensity of lymph follicles (ρ=-0,469, p=0,004). Microvessel densityin lymph follicles (r=0,663, p=0,004) The area of germinal centers (r=0,721, p=0,001)

25. The area of lymph folliclesdepending on the density of lymph follicles

26. The area of lymph folliclesdepending on the microvessel densityin lymph follicles

27. The area of lymph folliclesdepending on the area of germinal centers

28. The size of lymph folliclescorrelates with: T stage (ρ=-0,464, p=0,003) Number of the cavitary structure type-1 in tumor (ρ=-0,361, p=0,003) 3-year disease-free survival rate (ρ=-0,806, p=0,00003) Log-Rank Test p=0,005 3-year overal survival rate (ρ=-0,629, p=0,02) Cox’s F-test p=0,05

29. The size of lymph follicles depending on the T stage gastric cancer

30. The size of lymph follicles depending on the number of abdominal vascular of the first type

31. The size of lymph follicles depending on the 3-year disease-free survival rate(p=0,005)

32. The size of lymph follicles depending on the 3-year overal survival rate (p=0,05)

33. The density of lymph follicles in lymph node correlates with: The tumor size(ρ=-0,632, p=0,01) N stage (ρ=-0,527, p=0,07) T stage (ρ=-0,598, p=0,001) The presence of the cavitary structure type-1 in tumor (ρ=-0,600, p=0,04). 3-year overal survival rate (ρ=0,629, p=0,02)

34. The density of lymph follicles of lymph node depending on the tumor size

35. The density of lymph follicles of lymph node depending on the N stage gastric cancer

36. The density of lymph follicles of lymph node depending on the T stage gastric cancer

37. The density of lymph follicles of lymph node depending on the presence of thepresence of the cavitary structure type-1 in tumor

38. The density of lymph follicles of lymph node depending on the 3-year overal survival rate (p=0,11)

39. The presence of atypical phenotype with blast capillary endothelial cells correlates with: The degree of tumor growth(ρ=0,516, p=0,002) The type of differentiation of tumor growth (ρ=0,739, p=0,001) Presence of the cavitary structure type-1 in tumor (ρ=0,5, p=0,01). The number of lymph follicles at the bondary of tumor and gastric mucosa(ρ=-0,714, p=0,02)

40. The size of presence of atypical phenotype with blast capillary endothelial cells depending on the degree of tumor growth

41. The size of presence of atypical phenotype with blast capillary endothelial cells depending on the type of differentiation of tumor growth

42. The size of presence of atypical phenotype with blast capillary endothelial cells depending on the presence of the abdominal vascular type 1 in tumor

43. The size of presence of atypical phenotype with blast capillary endothelial cells depending on thenumber of lymph follicles at the bondary of tumor and gastric mucosa

44. The presence of dilated capillaries was not connected with clinical and morphological characteristics of the gastric cancer.

45. Lymphoproliferative processes and features of angiogenesis in a tissue of regional lymph nodes (without metastases) and tumors are interconnected, as well as the clinical characteristics and long-term results of treatment of gastric cancer. • The most important indicators are the size and density of lymphoid follicles in the lymph node tissue, and the presence of atypical vessels with blastic phenotype of endothelial cells • The data obtained can be used for prognosis of gastric cancer • We consider promising, further research in this direction

46. Thanks Prof. Alexander A. Stadnikov – Head, the Department of Histology, Cytology and Embryology of Orenburg State Medical University, Russia. Dr. Marina A. Senchukova, Associate Professor, Department of Oncology, Orenburg State Medical University, Russia

47. Thank for your attention!