Prolonged cultivation of separated rat lung mast cells (LMC) in vitro is necessary to better investigate a possible role of LMC in different stages of tissue remodeling induced by hypoxia. Rat lung mast cells (LMC) were separated using a protocol including an improved proteolytic extraction and two subsequent density gradient separations on Ficoll-Paque PLUS and a new generation of Percoll, i.e. Percoll PLUS. Instead of usual isotonic stock Percoll solution, an alternative "asymptotically isotonic" stock solution was more successful in our density separation of LMC on Percoll PLUS. Separated cells were cultivated for six days in media including stem cell factor, interleukins IL-3 and IL-6, and one of two alternative mixtures of antibiotics. These cultivations were performed without any contamination and with only rare changes in cell size and morphology. Model co-cultivation of two allogenic fractions of LMC often caused considerable rapid changes in cell morphology and size. In contrast to these observations no or rare morphological changes were found after cultivation under hypoxic conditions. In conclusions, we modified separation on Percoll PLUS to be widely used, altered LMC separation with respect to purposes of long-lasting cultivation and observed some model morphological changes of LMC.
- MeSH
- centrifugace - gradient hustoty MeSH
- isotonické roztoky chemie MeSH
- krysa rodu Rattus MeSH
- mastocyty cytologie MeSH
- oxid křemičitý chemie MeSH
- plíce cytologie metabolismus MeSH
- povidon chemie MeSH
- separace buněk metody MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- isotonické roztoky MeSH
- oxid křemičitý MeSH
- Percoll MeSH Prohlížeč
- povidon MeSH
Chronic hypoxia results in hypoxic pulmonary hypertension characterized by fibrotization and muscularization of the walls of peripheral pulmonary arteries. This vessel remodeling is accompanied by an increase in the amount of lung mast cells (LMC) and the presence of small collagen cleavage products in the vessel walls. We hypothesize that hypoxia activates LMC, which release matrix metalloproteinases (MMPs) cleaving collagen and starting increased turnover of connective tissue proteins. This study was designed to determine whether in vitro hypoxia stimulates production of MMPs in rat LMC and increases their collagenolytic activity. The LMC were separated on the Percoll gradient and then were divided into two groups and cultivated for 24 h in 21 % O(2) + 5 % CO(2) or in 10 % O(2) + 5 % CO(2). Presence of the rat interstitial tissue collagenase (MMP-13) in LMC was visualized by immunohistological staining and confirmed by Western blot analysis. Total MMPs activity and tryptase activity were measured in both cultivation media and cellular extracts. Exposure to hypoxia in vitro increased the amount of cells positively labeled by anti-MMP-13 antibody as well as activities of all measured enzymes. The results therefore support the concept that LMC are an important source of increased collagenolytic activity in chronic hypoxia.
- MeSH
- hypoxie buňky MeSH
- imunohistochemie MeSH
- kolagen metabolismus MeSH
- krysa rodu Rattus MeSH
- kultivační média metabolismus MeSH
- kultivované buňky MeSH
- mastocyty enzymologie MeSH
- matrixová metaloproteinasa 13 metabolismus MeSH
- matrixová metaloproteinasa 2 metabolismus MeSH
- matrixová metaloproteinasa 9 metabolismus MeSH
- matrixové metaloproteinasy metabolismus MeSH
- plíce cytologie enzymologie MeSH
- potkani Wistar MeSH
- separace buněk MeSH
- tryptasy metabolismus MeSH
- upregulace MeSH
- western blotting MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- kolagen MeSH
- kultivační média MeSH
- matrixová metaloproteinasa 13 MeSH
- matrixová metaloproteinasa 2 MeSH
- matrixová metaloproteinasa 9 MeSH
- matrixové metaloproteinasy MeSH
- Mmp13 protein, rat MeSH Prohlížeč
- Mmp2 protein, rat MeSH Prohlížeč
- Mmp9 protein, mouse MeSH Prohlížeč
- tryptasy MeSH
To assess the light-induced electrical activity of rodent retinal ganglion cells (RGCs) regenerating into a peripheral nerve (PN) graft we used non-invasive recording of electroretinographic responses to the contrast-reversal of sinusoidal gratings (p-ERG). On comparing the retinas that received a PN graft and retinas with only optic nerve (ON) transection, p-ERG responses were present in grafted retinas as late as 20 months after the surgery while they completely disappeared in non-transplanted controls within 4 months of ON transection. Next, the ability of regenerating RGCs to form functional connections with their targets in the superior colliculus (SC) was tested by a light-escape task. While the bilaterally blinded animals did not improve during the test, unilaterally grafted animals (with the contralateral eye blinded) reached 26% success in the last quartile of the light-escape task. This performance was significantly better than that of blind animals (ANOVA and Student-Newman-Keuls test; p<0.05), but did not reach the level of intact rats (87%). The transplanted rats, therefore, were capable of light perception, but at a sub-normal ability. In addition, we were also able to correlate the amplitude of the p-ERG response with the visual behavioral performance for each transplanted animal. This finding indicates that there is a direct link between the RGC electrophysiological activity and the functional capacity of the regenerated visual pathway. In conclusion, the above results indicate that (a) PN grafts help to preserve the normal electroretinographic activity of injured and regenerating RGCs (b) the regenerated visual pathway is functional and capable of mediating simple visual behavior and that (c) there is a correlation between the light-evoked RGC electrical activity and visual behavior and, finally, that (d) the effect of PN graft on the electrophysiological and functional restoration of the visual pathway is long-lasting or even permanent.
- MeSH
- elektroretinografie MeSH
- krysa rodu Rattus MeSH
- periferní nervy transplantace MeSH
- potkani Long-Evans MeSH
- regenerace nervu * MeSH
- retinální gangliové buňky fyziologie transplantace MeSH
- světlo MeSH
- učení vyhýbat se fyziologie MeSH
- zraková percepce fyziologie MeSH
- zrakové dráhy fyziologie MeSH
- zrakové evokované potenciály fyziologie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The effects of 14-day treatments with nerve growth factor (NGF), basic fibroblast growth factor (b-FGF), or the peptidergic drug Cerebrolysin on postlesion acquisition of a water maze task and on motor activity were evaluated. Rats were tested in the Morris water maze 14 days (early test) and 7 to 8 months (delayed test) after a bilateral lesion of the frontoparietal (sensorimotor) cortex. Only the rats treated with Cerebrolysin performed the water maze task at the level of the nonlesioned controls in the early test. No short-term effect of NGF (6.5 ng/14 days; 38 ng/ml) or b-FGF (17 ng/14 days; 100 ng/ml) treatment was found. The delayed test revealed that water maze performance was restored in rats treated with b-FGF in comparison with intact controls. The data showed that b-FGF can support or initiate processes in the CNS that lead to a delayed functional amelioration and/or compensation for a water maze performance deficit. NGF did not influence the acquisition impairment caused by a sensorimotor cortical lesion. Two-week administration of Cerebrolysin had a time-dependent influence: it attenuated the acquisition deficit and increased the motor activity of rats, both effects declined to the level of lesioned controls within 8 months.
- MeSH
- aminokyseliny farmakologie MeSH
- bludiště - učení účinky léků MeSH
- časové faktory MeSH
- čelní lalok chirurgie MeSH
- fibroblastový růstový faktor 2 farmakologie MeSH
- krysa rodu Rattus MeSH
- neurotrofní faktory farmakologie MeSH
- psychomotorický výkon účinky léků MeSH
- stárnutí účinky léků MeSH
- temenní lalok chirurgie MeSH
- voda MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- aminokyseliny MeSH
- cerebrolysin MeSH Prohlížeč
- fibroblastový růstový faktor 2 MeSH
- neurotrofní faktory MeSH
- voda MeSH
Our previous studies showed that the nootropic drug Cerebrolysin, applied immediately after the traumatic or excitotoxic brain lesion influenced spatial learning and memory. Long-lasting ameliorative effect of Cerebrolysin was found after its 4-week-administration, while two-week-treatment had only temporal effect. With the aim to verify the capability of Cerebrolysin to restore chronically deteriorated learning and memory. The drug was applied 4 months after lesioning the rat's CNS. The present study shows that Cerebrolysin restored learning capability of the lesioned rats. Although their spatial memory was improved in comparison to lesion untreated controls, it did not reach the level of intact controls. The effect was more pronounced after the application of 1.25 ml/kg b. w. of Cerebrolysin than after the application of 2.5 ml/kg b. w.
- MeSH
- aminokyseliny terapeutické užití MeSH
- chování zvířat účinky léků MeSH
- krysa rodu Rattus MeSH
- nootropní látky terapeutické užití MeSH
- poranění mozku komplikace patologie psychologie MeSH
- poruchy paměti farmakoterapie etiologie MeSH
- poruchy učení farmakoterapie etiologie MeSH
- potkani Long-Evans MeSH
- vnímání prostoru účinky léků MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- aminokyseliny MeSH
- cerebrolysin MeSH Prohlížeč
- nootropní látky MeSH
- MeSH
- aminokyseliny farmakologie MeSH
- časové faktory MeSH
- fibroblastové růstové faktory farmakologie MeSH
- hipokampus metabolismus MeSH
- krysa rodu Rattus MeSH
- neurotrofní faktory farmakologie MeSH
- paměť účinky léků MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- aminokyseliny MeSH
- cerebrolysin MeSH Prohlížeč
- fibroblastové růstové faktory MeSH
- neurotrofní faktory MeSH
In an attempt to compare effects of different neurotrophic factors on impaired memory function, young adult naive rats were trained to find the hidden platform in the Morris water maze (3 consecutive days, eight trials/day). The fimbria-fornix was unilaterally removed by aspiration and nerve growth factor (NGF) (11 micrograms/ml and 0.5 microgram/ml; groups NGF and ngf, respectively) or basic fibroblast growth factor (bFGF) (0.2 microgram/ml, group FGF) were applied via intra-cerebroventricular infusion by the osmotic minipump (flow rate 0.5 microliter/h, 14 days). Nootropic drug Cerebrolysin (EBEWE Arzneitmittel; 2.5 ml/kg/day, group CER) was applied via intraperitoneal injection (14 days). One group was formed by the rats treated with NGF (11 micrograms/ml) and Cerebrolysin (group NGFCER). Non-lesioned and lesioned only rats served as controls (groups INT and LES). After a 14-day treatment, rats were tested using the retention test (1 day, four trials). On the next day, the rats were tested using transfer test (3 days, eight trials/day). Escape latency and length of trajectory was recorded. Groups NGF, ngf, FGF and LES were similarly impaired in their ability to retrieve the old position of the platform (retention test), as well as in their ability to navigate to the new position of the platform (transfer test). In the latter, NGF group significantly differed from lesioned animals. Groups CER and NGFCER were comparable to group INT in the retention or transfer test. It is concluded that anterograde amnesia elicited by fimbria-fornix lesion can be abbreviated by NGF and/or CER, while retrograde amnesia is absent only in rats treated by CER. No short-term influence of bFGF was found. It is suggested that biochemical systems other than the cholinergic one are involved.
- MeSH
- aminokyseliny farmakologie MeSH
- bludiště - učení účinky léků MeSH
- fibroblastový růstový faktor 2 farmakologie MeSH
- hipokampus cytologie fyziologie MeSH
- krysa rodu Rattus MeSH
- neurotrofní faktory farmakologie MeSH
- nootropní látky farmakologie MeSH
- parasympatický nervový systém cytologie účinky léků MeSH
- poruchy paměti farmakoterapie psychologie MeSH
- viabilita buněk účinky léků MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- aminokyseliny MeSH
- cerebrolysin MeSH Prohlížeč
- fibroblastový růstový faktor 2 MeSH
- neurotrofní faktory MeSH
- nootropní látky MeSH
Fetal neocortical tissue (ED 14) was grafted unilaterally into a cortical cavity made bilaterally in the sensorimotor cortex of adult rats. Transplantation was done immediately after the lesion (group TR0, n = 8) or with 14-day delay (group TR14, n = 8). Six rats served as lesion only controls (group LES). After long-term survival (up to 15 months) the brains were photographed and surface areas of transplant and contralateral cavities were measured by means of a graphic tablet. The results show that (a) the presence of a transplant in one lesion cavity in the cortex decrease the size of a similar cavity in the contralateral cortex and that (b) the better host transplant integration there is, the greater the effect on the contralateral lesion. No correlation between the size of the transplant and the size of the symmetric traumatic lesion was found. The ameliorating effect of the transplant on the contralateral cortical lesion size is most likely related the long-term influence of growth of trophic factors released by transplanted cells which lead to the healing of the symmetric lesion.
- MeSH
- funkční lateralita fyziologie MeSH
- krysa rodu Rattus MeSH
- mozková kůra zranění fyziologie patofyziologie MeSH
- poranění mozku patofyziologie MeSH
- transplantace fetální tkáně fyziologie MeSH
- transplantace mozkové tkáně fyziologie MeSH
- viabilita buněk účinky léků MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The authors studied the metabolic activity of rat embryonic cerebral cortex grafts (ED 15-16) implanted into rat brains immediately (TR0) and 14 days (TR14) after cavity formation. Over a period of two months, the ATP, lactate and glucose concentration in TR0 transplants remained at the same level as observed in the intact cortex, whereas in TR14 transplants the ATP and glucose concentration fell significantly and the lactate concentration rose. The DNA concentration rose in both types of transplants, but the increase was more pronounced in TR0 grafts. Choline acetyltransferase activity (a neuron marker) fell significantly in both cases, but the decrease was greater in TR14 transplants. The results indicate that grafts implanted into the brain immediately after cavities had been formed have better metabolic activity and are capable of longer survival than grafts implanted 14 days after cavitation.
- MeSH
- acetylcholin metabolismus MeSH
- adenosintrifosfát metabolismus MeSH
- aminokyseliny metabolismus MeSH
- DNA metabolismus MeSH
- fosfokreatin metabolismus MeSH
- glukosa metabolismus MeSH
- krysa rodu Rattus MeSH
- laktáty metabolismus MeSH
- mozková kůra embryologie zranění metabolismus transplantace MeSH
- RNA metabolismus MeSH
- zvířata MeSH
- Check Tag
- krysa rodu Rattus MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- acetylcholin MeSH
- adenosintrifosfát MeSH
- aminokyseliny MeSH
- DNA MeSH
- fosfokreatin MeSH
- glukosa MeSH
- laktáty MeSH
- RNA MeSH
The authors studied the morphological picture of implanted foetal brain tissue. Macroscopically, they found that the grafts used for morphological study had taken in every case and that the size of the graft had doubled or trebled. The surface of the implant grew above the level of the recipient's cortex and numerous branching blood vessels were observed in the graft-cortex contact area. In the light microscope, organization of the cells in circular clusters or strips was found in the implant. Large numbers of blood vessels of varying calibre were present in the vicinity of large cell concentrations. Distinct differences between the cytoarchitectonics of the normal tissue of the recipient and the graft tissue were seen. Cells with a normal appearance or an apolar form, whose nucleus contained irregularly scattered chromatin and mosaic-like nucleoli, were observed at ultrastructural level. The nuclear membrane was thrown into multiple folds which invaginated deep into the nuclear matter. The Golgi complex covered a large area. Axodendritic synapses indistinguishable from the control were found in the neuropil. No changes were observed in the walls of the capillaries. The pericapillary zone was characterized by the presence of light astrocyte processes.
