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Cell Biology International (2005) 29, 10471049 (Printed in Great Britain)
Possible role of endogenous growth inhibitors in regeneration of organs: Searching for new approaches
N. Giorgobiania*, D. Dzidziguria, M. Rukhadzeb, L. Rusishvilia and G. Tumanishvilia
aLaboratory of Developmental Biology, Tbilisi State University, 1 Chavchavadze Avenue, 0108 Tbilisi, Georgia, USA
bLaboratory of Neurobiology, Tbilisi State University, Tbilisi, Georgia, USA
A hydrophilic protein component (12–17
Keywords: Endogenic growth inhibitor, Growth regulation.
Endogenous regulators of proliferation are protein molecules, encoded by regulating genes. They send specific signals to target cells to induce proliferation. In our opinion, one of the topical trends in the field of regulation of cell proliferation is investigation of endogenous growth inhibitory factors. Growth regulatory protein factors have been obtained from different organs of adult animals (Balazs and Blazsek, 1979; Kusen and Stoika, 1985; Amano and Iseki, 2001); at those times, comparatively scanty data existed about growth inhibitory factors of the myocardium. Most investigations included non-specific factors that affect: (a) the expression of synthesis of the different myocardium proteins; (b) the hypertrophy of cardiomyocytes; and (c) a directed action against death of cardiomyocytes after infarction (Kardami, 1990; Detillieux et al., 2003; Nishida et al., 2003; Kardami et al., 2004).
In general, our interest is mainly in specific growth inhibitory factors of myocardium and identification of similar factors in other organs. The objective of the present work was growth inhibitory factors obtained from: (i) the myocardium of animals of different phylogenetic stages; and (ii) different organs of the same animal.
2 Materials and methods
Myocardium, liver, kidney, and brain of white adult rats, as well as myocardium of adult snail, pigeon, and pig, were used as a material for investigation. Thermostable protein fractions were obtained by the method of alcohol precipitation of Balazs and Blazsek (1979), with modification. Animals were decapitated under diethyl ether. Organs were removed quickly, separated from capsules of connective tissues and vessels, rinsed with the physiological solution, and crushed. Aqueous homogenates were prepared in a tissue/distilled water ratio of 1:8. The homogenates were saturated step-wise with 96% ethanol to obtain 81% ethanol fraction, which was heated in a water bath (100
Hydrophobic interaction chromatography (HIC) was used for comparative analysis of TSPC (Queiroz et al., 2001). A hydrophilic polymeric sorbent, HEMA BIO Phenyl-1000 (particle size 10
3 Results and discussion
The chromatograms in Fig. 1 show that TSPC extracted from the hearts of the animals (snail, pigeon, rat, pig) contains a hydrophilic component with a retention time of 5.5
Chromatograms of TSPC extracted from the heart of snail, pigeon, pig and rat.
The hydrophilic (1) and hydrophobic (2) components of TSPC.
In previous work, we found that TSPC extracted from heart, liver, kidney, and brain of adult white rats inhibits both proliferation and transcription on average by 40–50% in cells of homologous organs of 7-day-old rats (Salakaya et al., 2000; Giorgobiani et al., 2002, 2003; Rusishvili et al., 2003; Rukhadze et al., 2005). We had also determined the molecular weights of TSPC components. In particular, electrophoresis in polyacrylamide gel showed the presence of a low molecular weight subfraction of 12–17
Focusing our attention here on the hydrophilic component of TSPC, the problem is whether this hydrophilic component and the low molecular weight fraction described earlier (Giorgobiani et al., 2002) are the same fraction. Chromatography of the low molecular weight TSPC component (Fig. 3) shows that the 12–17
Chromatogram of low molecular weight TSPC component.
Spectrum of hydrophilic component in the UV region (190–360
Chromatograms in Fig. 5 show hydrophilic and hydrophobic components of TSPC extracted from different organs (heart, liver, kidney, brain) of the same animal (adult white rat). According to the results, hydrophilic fractions of TSPC of all investigated organs seemed identical. The presence of the hydrophilic fraction in the myocardium of different animals indicates that a phylogenetically conservative group of thermostable proteins inhibiting proliferation processes has been found.
Chromatograms of hydrophilic and hydrophobic components of TSPC extracted from different organs (heart, liver, kidney, brain) of rat.
In the light of these results, we suggest two approaches for the use of hydrophilic (12–17
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