Identification of Biological Properties of Intralymphatic Tumor Related to the Development of Lymph Node Metastasis in Lung Adenocarcinoma

<div><p>Background</p><p>Intralymphatic tumors in the extratumoral area are considered to represent the preceding phase of lymph node metastasis. The aim of this study was to clarify the biological properties of intralymphatic tumors susceptible to the development of lymph node metastasis, with special reference to the expression of cancer initiating/stem cell (CIC/CSC) related markers in cancer cells and the number of infiltrating stromal cells.</p><p>Material and Methods</p><p>Primary lung adenocarcinomas with lymphatic permeation in the extratumoral area were retrospectively examined (n = 107). We examined the expression levels of CIC/CSC related markers including ALDH1, OCT4, NANOG, SOX2 and Caveolin-1 in the intralymphatic cancer cells to evaluate their relationship to lymph node metastasis. Moreover, the number of infiltrating stromal cells expressing CD34, α-smooth muscle actin, and CD204 were also evaluated.</p><p>Results</p><p>Among the intralymphatic tissues, low ALDH1 expression in cancer cells, high SOX2 expression in cancer cells, and a high number of CD204(+) macrophages were independent predictive factors for lymph node metastasis (<i>P</i> = 0.004, <i>P</i> = 0.008, and <i>P</i> = 0.028, respectively). Among these factors, only low ALDH1 expression in cancer cells was significantly correlated with the farther spreading of lymph node metastasis (mediastinal lymph node, pathological N2) (<i>P</i> = 0.046) and the metastatic lymph node ratio (metastatic/resected) (<i>P</i> = 0.028). On the other hand, in the primary tumors, ALDH1 expression in the cancer cells was not associated with lymph node metastasis. Intralymphatic cancer cells expressing low ALDH1 levels exhibited lower E-cadherin expression levels than cancer cells with high levels of ALDH1 expression (<i>P</i> = 0.015).</p><p>Conclusions</p><p>Intralymphatic cancer cells expressing low levels of ALDH1 and infiltrating macrophages expressing CD204 have a critical impact on lymph node metastasis. Our study also highlighted the significance of evaluating the biological properties of intralymphatic tumors for tumor metastasis.</p></div

Relationship between ALDH1 and E-cadherin expression.

<p>(A) intralymphatic cancer cells showing a high level of ALDH1 expression, and (B) intralymphatic cancer cells showing a high level of E-cadherin expression. The images shown in A and B were obtained from the same case. (C) Intralymphatic cancer cells with a low level of ALDH1 expression. (D) Intralymphatic cancer cells with a low level of E-cadherin expression. The images shown in C and D were also obtained from the same case. (E) Comparison of E-cadherin expression between cancer cells with low and high levels of ALDH1 expression.</p

The aggressiveness of lymph node metastasis (Metastatic lymph node ratio).

<p>*Considered to be statistically significant (p<0.05).</p

Immunohistochemical staining of intralymphatic cancer cells (A, B, C, D) and stromal cells (E, F).

<p>(A) high ALDH1 expression level, (B) low ALDH1 expression level, (C) high SOX2 expression level, (D) low SOX2 expression level, (E) higher number of CD204-positive macrophages, (F) lower number of CD204-positive macrophages.</p

Univariate analysis about relationship of immunohistochemical staining of intralymphatic tumor and LN metastasis (Cancer cells).

<p>*Considered to be statistically significant (p<0.05).</p

The aggressiveness of lymph node metastasis (Pathological N stage).

<p>*Considered to be statistically significant (p<0.05).</p

Relationship between clinicopathological characteristics and lymph node metastasis.

<p>*Considered to be statistically significant (p<0.05).</p><p>**Solid adenocarcinoma with mucin.</p

Univariate analysis about relationship of immunohistochemical staining of primary tumor and LN metastasis.

<p>*Considered to be statistically significant (p<0.05).</p

Additional file 2: of The efficacy of the Kampo medicine rikkunshito for chemotherapy-induced anorexia (RICH trial): study protocol for a randomized controlled trial

SPIRIT 2013 Checklist: recommended items to address in a clinical trial protocol and related documents*. (DOC 121 kb

Additional file 1: of The efficacy of the Kampo medicine rikkunshito for chemotherapy-induced anorexia (RICH trial): study protocol for a randomized controlled trial

Sample size calculation. (DOCX 23 kb