Irina Mitrofanova

SOME MORPHOPHYSIOLOGICAL FEATURES OF LAVANDER CULTIVAR MICROPROPAGATED IN VITRO BY MERISTEM CULTURE

The data about some morpho-physiological peculiarities of explants of lavender (Lavandula angustifolia Mill.) the cultivars Stepnaya, Sineva, Vdala, Record, Rannyaya, introduced by meristems in aseptic conditions and under long-term micropropagation in vitro are presented. During cultured meristems on Murashige and Skoog (MS) medium with 1.0 mg L Kinetin and 0.5 mg L GA3 all cultivars have formed the multiple shoot. The maximum number of microshoots at in vitro introduction stage was in the cultivar Sineva (5.7), and minimum – in the cultivar Rannyaya (3.9). Further, microcutting of main and additional shoots was conducted every 30-35 days. Obtained shoot segments with one node on MS medium with 1.0 mg L Kinetin and 0.5 mg L GA3 were cultured. When micropropagation of lavender follow 6 subculture it was shown that the number of shoots was increased in all cultivars till 3 subculture, and then it was decreased. In the cultivar Sineva the maximal multiplication index (12.2) was noted at the third passage, and then changed from 11.5 to 7.4. The lowest multiplication index at 5-7 passages was in the cultivar Record (3.8-4.5). Analysis of water regime of obtained microshoots at 6 subculture showed that the total water content was 83-89%, and the fraction of bound and colloidabsorbed water was 40-57%. Maximum values of water-holding capacity in the cultivars Record, Rannyaya and Vdala were found. Studying the parameters of light-induction processes it has been showed that the cultivars Vdala and Sineva were characterized by maximum photosynthetic activity of leaves and shoots, and they had high viability index.

Development of Novel Monoclonal Antibodies for Evaluation of Transmembrane Prostate Androgen-Induced Protein 1 (TMEPAI) Expression Patterns in Gastric Cancer

Transmembrane prostate androgen-induced protein 1 (TMEPAI) is a single-span membrane protein, functionally involved in transforming growth factor beta signaling pathway. The particular protein presented in cells in three isoforms, which differs in the length of the soluble N-terminal extracellular domain, making it challenging for the immunochemical recognition. By using quantitative real-time polymerase chain reaction, we identified significant upregulation of PMEPA1 gene expression in malignant tissues of patients with gastric adenocarcinoma. The main part of commercially available anti-TMEPAI antibodies are having polyclonal nature or not suitable for immunocytochemical localization of target protein in tissue specimens. Hence, we decide to generate a set of novel rat monoclonal antibodies (mAb) directed against conservative C-terminal cytoplasmic epitope. Immunoblotting analysis showed that monoclonal antibodies, 2E1, 6C6, and 10A7 were able to recognize specifically target protein in transiently transfected HEK293T and CHO-K1 cells. Especially established mAb, named 10A7, showed the excellent binding ability to target protein in immunohistochemistry. By using developed antibodies, we observed pronounced expression of TMEPAI in normal gastric epithelial cells while tumor cells from gastric adenomas, and adenocarcinoma samples were mostly negative for target protein expression. Also, we found that gastric epithelium cells lose the TMEPAI expression concurrently with severe dysplasia progression, which probably caused by a mechanism involving specific microRNA.

The effect of neoadjuvant chemotherapy on the correlation of tumor-associated macrophages with CD31 and LYVE-1

Angiogenesis and lymphangiogenesis play a crucial role in tumor growth, invasion and metastasis. Tumor-associated macrophages (TAM) induce both angiogenesis and lymphangiogenesis in mouse breast cancer models and positively correlate with these processes in human breast cancer patients. Neoadjuvant chemotherapy (NAC) is a widely used therapeutic option for cancer treatment. However, the effect of NAC on the distribution of TAM within intratumoral compartments and their correlation with angiogenesis and lymphangiogenesis remained unknown. In the present study we analyzed the effect of NAC on the distribution of CD68+ and stabilin-1+ TAM in five functionally distinct areas of human breast cancer and their correlations with microvessel density (MVD) and lymphatic microvessel density (LMVD), identified by CD31 and LYVE1, respectively. We found that NAC enhances blood vessel density in soft fibrous stroma and in coarse fibrous stroma. Without NAC the amount of CD68+ TAM in gaps of ductal tumor structures positively correlate with CD31+ microvessel density in soft fibrous stroma. NAC had enhancing effect on the amount of CD68+ TAM but not stabilin-1+ TAM in soft fibrous stroma. However, no correlation between TAM and CD31+ microvessel density was identified after NAC. NAC did not enhance the lymphatic microvessel density. But after NAC stabilin-1 expressing subpopulation of TAM positively correlated with expression of LYVE-1. We hypothesized that CD68+ TAM can support tumor angiogenesis primarily before NAC, while stabilin-1+ TAM can contribute to the maintenance of lymphatic microvessel density after NAC.