Biomedicine

A research conducted by L. Wei’s group showed that peripheral stimulation and physiotherapy could promote neurovascular plasticity and functional repair after central nervous system disorders, such as ischemic stroke. Ischemia is a local anaemia often caused by the vascular factor (arterial luminal narrowing or obturation) and leading to temporary dysfunction or stable injury of the tissue or organ. Organs of the central nervous system and myocard are the most ischemia-sensitive organs. Researchers from the group headed by L. Wei provoked ischemic stroke in the right cortical sensorimotor area of the adult mice’s brain cortex, or they simulated that injury. Three days after that manipulation all mice obtained BrdU (5-bromo-2'-deoxyuridine) injection and every second mouse got whisker stimulation to enhance afferent signals to the cortex of the injured area. In 14 days after stroke, whisker stimulation considerably increased the level of the vascular endothelial growth factor (VEGF) and the stromal cell-derived factor-1 (SDF-1); also, local cerebral blood circulation improved in those animals. Neurogenesis, detected by the level of the nuclear protein (NeuN) and by BrdU staining, improved as well. Thus, it was found that after peripheral stimulation, neurogenesis and cell migration improved in mice undergone focal ischemia.

Adding four factors only: Oct3/4, Sox2, c-Myc and Klf4, — to cultured fibroblasts leads to cell reprogramming. This method is effective for embryonal as well as for mature fibroblasts. The cells obtained are called induced pluripotent stem cell (iPS). They had morphology and proliferative ability peculiar to embryonal stem cells. Also, they expressed markers of embryonal stem cells. Subcutaneous introduction of iPS cells into mice provoked formation of neoplasms which contained spots of tissues derived from all three germ layers; iPS injected into mouse’s blastocytes took part in the formation of an embryo. Human iPS show numerous properties of embryonal stem cells: morphology, proliferative ability, surface antigens, epigenetic state of genes that are specific for the pluripotent state, telomerase activity. It was shown during in vitro and teratoma experiments that iPS could differentiate into cells of all three germ layers. In the in vitro experiments, factors necessary for reprogramming were delivered by retroviral vectors. Those vectors had the ability to integrate into genome that was fraught with carcinogenesis. In 2008, a mode was developed allowing to administer Oct3/4, Sox2, c-Myc and Klf4 into cells by means of cDNA. So scientists developed iPS which did not provoke formation of teratomas after they had been injected. The second factor increasing potential risk of neoplasm formation is с-Myc. To eliminate this problem, the scientists developed a method allowing to obtain iPS without using c-Myc. That method is less effective, but safer. Not only fibroblasts, but also liver and stomach cells may be used as the basis for iPS obtaining. Success of the experiments on mouse fibroblasts was lately reproduced in the experiments on human fibroblasts. Addition of the same four factors: Oct3/4, Sox2, c-Myc и Klf4, — led to iPS formation. Thus, it was the first time in 2007 when embryonal stem cells were obtained from somatic cells without using embryonal material.

Prof. J. Holloszy studied 18 member of the Caloric restriction society who practised low-caloric diet (1100–1950 calories a day) during six years. 18 members of the control group consumed 1975–3550 calories a day that is more typical for Americans. The aim of the research was to determine how caloric restriction affected health and aging processes in the human organism. It was found that blood pressure and cholesterol level in the middle-aged Americans who were on a low-caloric diet corresponded with those typical for far younger people. The level of C-reactive protein (a marker of inflammation) was 12,5 times lower in the subjects of the experiment than in middle-aged persons. Moreover, J. Holloszy’s group studied long-term effects of caloric or protein restriction on IGF-1 and IGFBP-3 concentration in the blood serum in human. Experiments showed that, in contrast to rats, long-term serious caloric restriction did not lower neither IGF-1 concentration in the blood serum, nor IGF-1: IGFBP-3 ratio in human. It was also found out that consumption of proteins was the key factor determining IGF-1 regulation in human. The investigations showed that caloric restriction reduced the risk of atherosclerosis and diabetes, as well as inflammation level. Thus, restriction of protein consumption may become important component of anti-tumor and anti-aging diets.

Poral tissue-engineered constructions can be used to transport genetic material (a gene for the blood-vessel growth factor) and stimulate new tissue formation. In L. Shea’s laboratory, poral tissue-engineered nanoconstructions were created using polylactide-co-glycoside with encapsulated plasmids which carry gene insertions for transfection. It was found that porization increased the level of transgene expression in vivo. Transfected cells were at first found at the periphery of the construction, but in 4 months, transgene expression was observed throughout tissue construction. Stable expression of VEGE, born on the plasmid, led to increased density of blood vessels in transgene tissue compared to the control. Combining new nanoconstructions with gene delivery technology by the use of nonviral vectors will facilitate manipulations with different tissues in the future, favouring the development of the regenerative medicine.

The area of scientific interest includes a research on the role of poly-ADP-ribosylation in DNA reparation and aging, and on prion protein neurotoxicity. In 1992, A. Bürkle and K. Grube characterized positive correlation between poly-ADP-ribosylation activity (PARP1) in peripheral monocytes, induced by single-strand breaks, and life span of mammals. PARP1 activity in human peripheral monocytes was 5 times higher than in rat’s monocytes due to self-activation of the enzyme by means of ribosylation. The level of that enzyme was 2 times higher that content of ribosylated PARP-1 in the rat’s cells. As an organism grows older, PARP activity lowers both in human and in rats. On the other hand, PARP activity in immortalized lymphocytes taken from French long-lived persons, who were 100- and more years old, was higher than that in the cells belonging to the representatives of the control group (20- to 70 years old). Thus, high and carefully controlled PARP activity may promote life extension via supression of genome instability and oncogenesis.

Workers of prof. A. Badley’s laboratory study how viral proteins can modify the cell immune response, and how we can operate cell apoptosis in order to treat human diseases. A. Badley’s laboratory studied the process of neutrophil apoptosis (neutrophils are the phagocytes produced in the bone marrow). After neutrophils get into the excitation focus, they quickly become senescent. Then neutrophils come back into the marrow bone and undergo apoptosis. Return of the senescent neutrophils into the marrow bone is controlled by the stromal cell-derived factor 1 (SDF-1) which interacts with CXCR4 — the molecule senescent neutrophils express. If senescent neutrophils do not undergo apoptosis by any reason, they can cause various autoimmune diseases. It is revealed that recombinant TRAIL induced apoptosis of the senescent neutrophils. Scientists conclude that recombinant TRAIL can theoretically be used for selective elimination of the senescent neutrophils.

One of Dr. R. J. Shmookler Reis’s research areas is the search of genes regulating life span of nematode Caenorhabditis elegans. Shmookler Reis’s laboratory has created mutant nematodes that have age-1 gene with truncated phosphatidylinositol 3-kinase catalytic subunit (PI3KCS). Such PI3KCS mutants have had remarkable longevity. They also have had prolonged period of development and normal mobility. On the other hand, C. elegans PI3KCS mutants have sacrificed their body size and metabolic rate. Such worms have also had exceptional resistance to oxidative and electrophilic stress as compared to worms with normal alleles or alleles associated with lesser life spans. Mutant worms have lived 145–190 days at 20 degrees centigrade showing ten times extension of average and maximum life span.

When a patient has serious and irreversible disorders of the gastrointestinal tract, intestinal tissue engineering can be used as a promising approach. This approach proposes to replace dysfunctional or missing intestinal tissues with an implant that imitate physiological functions of the intestine. Different research groups have developed methods allowing to harvest intestinal stem cells from animal embryos or newborns. Such cells can be introduced into natural or synthetic carrier materials. At present, this methodology is rather standardized. That allows to create new intestinal tissue that resembles natural intestine in many morphological features and functional characteristics. Intestines created in that way have already allowed to improve or even cure of clinical syndromes of malabsorption in researches on rodents. Later, new intestinal tissues of large animals have been created. Prof. M. Stelzner’s researches are concentrated on the stem cell therapy in application to intestinal mucosa. They cover the use of bioengineered scaffold supporting the growth of cells transplanted into the patients who have some parts of their intestinal tract removed surgically. Intestinal stem cell organoid transplantation generates functional intestinal neomucosa and is used therapeutically to improve nutrient absorption and cure bile acid malabsorption in rats. The researchers from M. Stelzner’s team hypothesized that intestinal organoids can be harvested and transplanted to generate intestinal neomucosa in a large animal model — a dog. As the result of the experiments, in the the group where intestinal organoids were harvested from fetuses and allotransplanted into 10-month old mother animals, neomucosal growth was identified in 11 of 12 polyglycolic acid scaffolds after allotransplantation of fetal organoids. The neomucosa resembled normal canine mucosa in structure and composition. Intestinal stem cell organoid transplantation can be used to generate neomucosa in dogs. This is a great success and an important step towards clinical use of the intestinal tissue engineering in people. This is the first report of successful generation of intestinal neomucosa using intestinal stem cell organoid transplantation in a large animal model.