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INSTITUTE OF CYTOLOGY OF THE RUSSIAN ACADEMY OF SCIENCES

"CELL CULTURES"
Information Bulletin. Issue 21 (2006), St. Petersburg. 63 p.

Stem cells: their use in fundamental cell biology and medicine
CELL TECHNOLOGIES BASED ON STEM CELLS.
N. N. Nikolsky.
Institute of cytology RAS, St. Petersburg, nnnik@mail.cytspb.rssi.ru

PARTICIPATION OF BONE MARROW'S HAEMOPOIETIC STEM CELLS IN REGENERATION OF MOUSE CORNEAL STROMA.
K. K. Davitaia, T. V. Vasilieva, G. S. Kvinikhidze, T. V. Mitchurina, N. G. Khrushchov.
Institute of Zoology, Georgian Academy of Sciences, Tbilisi; N.K. Koltsov Institute of Developmental biology, Russian Academy of Sciences, Moscow; M.V. Lomonosov State University, Moscow; dato_mfa@hotmail.com
     The source of fibroblast of the post-traumatic regenerating corneal stroma is still a subject of study. It is not clear if the new formation of corneal stroma is the product of local cells, or elements of haematogeneous nature. The resent article describes our attempt to clear up this problem. In the recent work by using morphological, radio autographic and immunofluorescence methods it was shown for the first time on the Rat-mice xenogenic Radiation chimeras, that fibroblast-like cells participate in mice corneal stroma regeneretion are originated from the red bone marrow, of rat.

GENE-CELLS THERAPY OF MDX MICE STRIATED MUSCLES.
V. M. Mikhailov, A. V. Karmanova, V. V. Zenin, V. B. Serikov.
Institute of Cytology RAS, St. Petersburg; vmikhailov@mail.cytspb.rssi.ru
     Three sets of experiments were carried out. The first one involved chimeric mice, obtained by intravenously injections of bone marrow derived cells taken from transgenic C57BL/6 mice expressing GFP, to 5 Gy X-ray irradiated mdx or C57BL/6 mice. In 2 months M. quadriceps femoris of chimeric mice were destroyed by surgical clamp. Following the next 4-5 weeks, the same muscles were studied for the presence of GFP-positive striated muscle fibres. In the case of chimeric C57BL/6 mice GFP-positive striated muscle fibres were observed in 0.3±0.5 % and in 0.2±0.3 % of destroyed muscle and of lateral muscle, consequently. In the case of chimeric mdx mice, positive results were observed in 1.7±0.4 % and in 0.5±0.3 % of destroyed and control muscle, respectively. In the second set of experiments, the GFP-positive bone marrow stem cells were used for multiple intramuscular injections to M. quadriceps femoris of C57BL/6 or mdx mice in a dose of 2-5 x 105 cells per mouse. Before injection, GFP-positive bone marrow cells were fractionated in a 63% Percoll solution and then were exhausted from differentiated cells by magnetic manner using CD4, CD8, CD38, CD45R, CD119, Ly-6G, and F4/80 antibodies. After 2-3 weeks, as many as 0.15±0.4 % и 0.1±0.2 % of GFP-positive muscle fibres were found in injected and control muscles of C57BL/6 mice, respectively. In the case of mdx mice, the frequency of GFP-positive striated muscle fibres was 2±0.8 % and 1.2±0.6 % for injected and control muscles, respectively. There is conclusion that bone marrow stem cells can take part in differentiation of mdx mouse muscles after their delivery by needle injections. We also transfected C57BL/6 bone marrow stem cells by plasmids pCMV-GFP-nPRX6-L. Construction was prepared and put by Prof. N. V. Tomilin. Cell transfections was made by electroporation, cell delivery to muscle tissue was made by multiple injections. After week the part of GFP-positive striated muscle fibres were 3±1.3%. The result showed the principal possibility of transfection of bone marrow stem cells for next paticipation in differentiation of mdx mice striated muscle fibres.

Cell Biotechnology and Tissue Engineering
ENCAPSULATION OF CELLS AND CELL CULTURE BY LAYER-BY LAYER ADSORPTION OF POLYELECTROLYTES.
I. G. Belyaeva ^{1, 2}, O. V. Galibin ¹, A. D. Vilesov ³, G. B. Sukhorukov ^{2, 4}.
¹ I. P. Pavlov St-Petersburg State Medical University, irinbel1@yandex.ru; ² Max Plank Institute of Colloids and Interfaces, Germany; ³ Institute of Macromolecular Compounds RAS, St-Petersburg, Russia; ⁴ Department of Materials, Queen Mary University, London, UK.
     A principle possibility of live cell systems immunoprotection by shell fabrication made of layer-by layer adsorbtion of polyelectrolytes has been shown. This protection of the cells is expected to exclude penetration of immunocytes, antibodies and other transplant rejection mechanisms, whereas low molecular weight substances such as nutrients, electrolytes, oxygen, and bioactive secretory products are exchangeable across the capsules wall maintaining the cells to remain alive.

CULTURED HUMAN FETUS FIBROBLASTS IN THE COMPLEX TREATMENT OF LEG ULCERS.
Sedov V. M. ¹, Andreev D. Y. ¹, Smirnova T. D. ², Paramonov B. A. ³, Enkina T. N. ⁴, Sominina A. A. ², Kiselev O. I. ², Karpova N. G. ², Kuznetsova I. K. ², Lebedev L.V. ¹.
¹ I. P. Pavlov St-Petersburg State Medical University, ² Institute of Influenza of RAMS, ²³ Plastic and Esthetic Surgery Department of Saint-Petersburg Postgraduate Medical Academy, ⁴ Central Medical Unit № 122.
     Results of cell therapy as part of complex treatment of 23 patients with venous leg ulcers (due to varicose disease and post-thrombotic syndrome) are analyzed in the article. In the control group (25 patients) modern wound dressings Suprasorb® were used.
     It has been demonstrated that cell therapy using fibroblasts of the line 11 00/14 significantly increased efficacy of the treatment. Ulcers healing rate was 100% in the basic group. Epithelialization time averaged 1.5 weeks for varicose ulcers and 3.2 weeks for ulcers due to post-thrombotic syndrome. In the control group, ulcers healing rate was 86% for varicose disease and 78% for post-thrombotic syndrome. Healing time averaged 3.6 and 3.9 months, correspondingly.
     Cell therapy as part of complex treatment of venous leg ulcers excels modern wound dressings in efficacy.

Studies on cultured plant cells and tissues
PECULIARITIES OF MORPHOGENIS PATHWAYS INDUCTION IN ANTHER CULTURE IN VITRO OF WHEAT.
D. Yu. Zaytsev, O. A. Seldimirova, N. N. Kruglova.
Institute of Biology of Ufa Scientific Center, Russian Academy of Sciences, Ufa, Russia, Kruglova@anrb.ru.
     The cytological and histological analysis of morphogenis pathways in wheat anther culture in vitro on induction medium containing different concentrations of exogenic 2.4-D was conducted. On the basis of the data of immunofermentative analysis and light microscopy the dependence of the concrete morphogenis pathway induction from the balance of endogenic and exogenic phytohormones content are shown.

Development of Problem
J-CHAIN GENE EXPRESSION IN MURINE MYELOMA 653A AND MYELOMA-DERIVED HYBRYDOMAS.
Klimovich B. V., Samoylovich M. P., Klimovich B. V..
Central Research institute for Rentgeno-Radiology, St. Petersburg, Russia, vklim@rednet.ru, borrismorris@mail.ru
     J-chain gene expression in murine myeloma 653A and three hybridomas derived as a result of fusion myeloma cells and splenocytes of immunized mice was studied by RT-PCR. Immunoglobulins non-secreting myeloma cells were J-chain negative.Two hybrydoma strains secreted IgG, one produced IgM. All hybridomas under investigation showed J-chain expression. This data provide a possibility of using hybridomas and myelomas as models for J-chain gene expression regulation studies.

Methods of investigations

Technical information

Chronicle

Information of the Cell Culture Association

Instructions for authors
     The paper should not exceed 10 manuscript pages. The manuscript is to be accompanied by recommendation addressed to the Editorial Board of the Information Bulletin from the institution in which the work has been carried out. The paper should contain a brief resume, with the title of the work, names of the authors, institution, city.
     The manuscript is to be accompanied by its electron variant: one file in MS Word for Windows. The file should have the font Arial Narrow, 13; interline difference - 1.5; the indented line - 0.5 cm; the upper margin - 2.5 cm, other margins - 2.1 cm each. The file is to be e-mailed to the address of the Editorial Board.

Address of the Editorial Board:

Dr. M. Bogdanova, Institute of Cytology, Russian Academy of Sciences, Tikhoretsky Prosp. 4, St. Petersburg 194064, Russia.
Tel.: (812) 297-53-10, fax: (812) 297-03-41
e-mail: msb@mail.cytspb.rssi.ru

"Cell Cultures" ETCS Home

Stem cells: their use in fundamental cell biology and medicine CELL TECHNOLOGIES BASED ON STEM CELLS. N. N. Nikolsky. Institute of cytology RAS, St. Petersburg, nnnik@mail.cytspb.rssi.ru
PARTICIPATION OF BONE MARROW'S HAEMOPOIETIC STEM CELLS IN REGENERATION OF MOUSE CORNEAL STROMA. K. K. Davitaia, T. V. Vasilieva, G. S. Kvinikhidze, T. V. Mitchurina, N. G. Khrushchov. Institute of Zoology, Georgian Academy of Sciences, Tbilisi; N.K. Koltsov Institute of Developmental biology, Russian Academy of Sciences, Moscow; M.V. Lomonosov State University, Moscow; dato_mfa@hotmail.com The source of fibroblast of the post-traumatic regenerating corneal stroma is still a subject of study. It is not clear if the new formation of corneal stroma is the product of local cells, or elements of haematogeneous nature. The resent article describes our attempt to clear up this problem. In the recent work by using morphological, radio autographic and immunofluorescence methods it was shown for the first time on the Rat-mice xenogenic Radiation chimeras, that fibroblast-like cells participate in mice corneal stroma regeneretion are originated from the red bone marrow, of rat.
GENE-CELLS THERAPY OF MDX MICE STRIATED MUSCLES. V. M. Mikhailov, A. V. Karmanova, V. V. Zenin, V. B. Serikov. Institute of Cytology RAS, St. Petersburg; vmikhailov@mail.cytspb.rssi.ru Three sets of experiments were carried out. The first one involved chimeric mice, obtained by intravenously injections of bone marrow derived cells taken from transgenic C57BL/6 mice expressing GFP, to 5 Gy X-ray irradiated mdx or C57BL/6 mice. In 2 months M. quadriceps femoris of chimeric mice were destroyed by surgical clamp. Following the next 4-5 weeks, the same muscles were studied for the presence of GFP-positive striated muscle fibres. In the case of chimeric C57BL/6 mice GFP-positive striated muscle fibres were observed in 0.3±0.5 % and in 0.2±0.3 % of destroyed muscle and of lateral muscle, consequently. In the case of chimeric mdx mice, positive results were observed in 1.7±0.4 % and in 0.5±0.3 % of destroyed and control muscle, respectively. In the second set of experiments, the GFP-positive bone marrow stem cells were used for multiple intramuscular injections to M. quadriceps femoris of C57BL/6 or mdx mice in a dose of 2-5 x 105 cells per mouse. Before injection, GFP-positive bone marrow cells were fractionated in a 63% Percoll solution and then were exhausted from differentiated cells by magnetic manner using CD4, CD8, CD38, CD45R, CD119, Ly-6G, and F4/80 antibodies. After 2-3 weeks, as many as 0.15±0.4 % и 0.1±0.2 % of GFP-positive muscle fibres were found in injected and control muscles of C57BL/6 mice, respectively. In the case of mdx mice, the frequency of GFP-positive striated muscle fibres was 2±0.8 % and 1.2±0.6 % for injected and control muscles, respectively. There is conclusion that bone marrow stem cells can take part in differentiation of mdx mouse muscles after their delivery by needle injections. We also transfected C57BL/6 bone marrow stem cells by plasmids pCMV-GFP-nPRX6-L. Construction was prepared and put by Prof. N. V. Tomilin. Cell transfections was made by electroporation, cell delivery to muscle tissue was made by multiple injections. After week the part of GFP-positive striated muscle fibres were 3±1.3%. The result showed the principal possibility of transfection of bone marrow stem cells for next paticipation in differentiation of mdx mice striated muscle fibres.
Cell Biotechnology and Tissue Engineering ENCAPSULATION OF CELLS AND CELL CULTURE BY LAYER-BY LAYER ADSORPTION OF POLYELECTROLYTES. I. G. Belyaeva ^{1, 2}, O. V. Galibin ¹, A. D. Vilesov ³, G. B. Sukhorukov ^{2, 4}. ¹ I. P. Pavlov St-Petersburg State Medical University, irinbel1@yandex.ru; ² Max Plank Institute of Colloids and Interfaces, Germany; ³ Institute of Macromolecular Compounds RAS, St-Petersburg, Russia; ⁴ Department of Materials, Queen Mary University, London, UK. A principle possibility of live cell systems immunoprotection by shell fabrication made of layer-by layer adsorbtion of polyelectrolytes has been shown. This protection of the cells is expected to exclude penetration of immunocytes, antibodies and other transplant rejection mechanisms, whereas low molecular weight substances such as nutrients, electrolytes, oxygen, and bioactive secretory products are exchangeable across the capsules wall maintaining the cells to remain alive.
CULTURED HUMAN FETUS FIBROBLASTS IN THE COMPLEX TREATMENT OF LEG ULCERS. Sedov V. M. ¹, Andreev D. Y. ¹, Smirnova T. D. ², Paramonov B. A. ³, Enkina T. N. ⁴, Sominina A. A. ², Kiselev O. I. ², Karpova N. G. ², Kuznetsova I. K. ², Lebedev L.V. ¹. ¹ I. P. Pavlov St-Petersburg State Medical University, ² Institute of Influenza of RAMS, ²³ Plastic and Esthetic Surgery Department of Saint-Petersburg Postgraduate Medical Academy, ⁴ Central Medical Unit № 122. Results of cell therapy as part of complex treatment of 23 patients with venous leg ulcers (due to varicose disease and post-thrombotic syndrome) are analyzed in the article. In the control group (25 patients) modern wound dressings Suprasorb® were used. It has been demonstrated that cell therapy using fibroblasts of the line 11 00/14 significantly increased efficacy of the treatment. Ulcers healing rate was 100% in the basic group. Epithelialization time averaged 1.5 weeks for varicose ulcers and 3.2 weeks for ulcers due to post-thrombotic syndrome. In the control group, ulcers healing rate was 86% for varicose disease and 78% for post-thrombotic syndrome. Healing time averaged 3.6 and 3.9 months, correspondingly. Cell therapy as part of complex treatment of venous leg ulcers excels modern wound dressings in efficacy.
Studies on cultured plant cells and tissues PECULIARITIES OF MORPHOGENIS PATHWAYS INDUCTION IN ANTHER CULTURE IN VITRO OF WHEAT. D. Yu. Zaytsev, O. A. Seldimirova, N. N. Kruglova. Institute of Biology of Ufa Scientific Center, Russian Academy of Sciences, Ufa, Russia, Kruglova@anrb.ru. The cytological and histological analysis of morphogenis pathways in wheat anther culture in vitro on induction medium containing different concentrations of exogenic 2.4-D was conducted. On the basis of the data of immunofermentative analysis and light microscopy the dependence of the concrete morphogenis pathway induction from the balance of endogenic and exogenic phytohormones content are shown.
Development of Problem J-CHAIN GENE EXPRESSION IN MURINE MYELOMA 653A AND MYELOMA-DERIVED HYBRYDOMAS. Klimovich B. V., Samoylovich M. P., Klimovich B. V.. Central Research institute for Rentgeno-Radiology, St. Petersburg, Russia, vklim@rednet.ru, borrismorris@mail.ru J-chain gene expression in murine myeloma 653A and three hybridomas derived as a result of fusion myeloma cells and splenocytes of immunized mice was studied by RT-PCR. Immunoglobulins non-secreting myeloma cells were J-chain negative.Two hybrydoma strains secreted IgG, one produced IgM. All hybridomas under investigation showed J-chain expression. This data provide a possibility of using hybridomas and myelomas as models for J-chain gene expression regulation studies.
Methods of investigations
Technical information
Chronicle
Information of the Cell Culture Association
Instructions for authors The paper should not exceed 10 manuscript pages. The manuscript is to be accompanied by recommendation addressed to the Editorial Board of the Information Bulletin from the institution in which the work has been carried out. The paper should contain a brief resume, with the title of the work, names of the authors, institution, city. The manuscript is to be accompanied by its electron variant: one file in MS Word for Windows. The file should have the font Arial Narrow, 13; interline difference - 1.5; the indented line - 0.5 cm; the upper margin - 2.5 cm, other margins - 2.1 cm each. The file is to be e-mailed to the address of the Editorial Board.