Collagen/hyaluronan membrane as a scaffold for chondrocytes cultivation

Rabbit chondrocytes were cultivated in vitro using the collagen/hyaluronan membrane. The membrane did not show any adverse effects on chondrocyte viability during in vitro cultivation. The inoculated cells grew without any negative changes. According to the histochemical analyses: (i) hematoxylin and eosin; (ii) safranin O; and (iii) rabbit anti-human collagen type II staining, the rabbit chondrocytes maintained their morphology and phenotype during in vitro cultivation. The collagen/hyaluronan membrane became more stable and stiffer after long time cultivation. The proliferation of the chondrocytes stabilised the structure of the membrane. The collagen/hyaluronan membrane is suitable material for the chondrocyte growth and could provide functional tissue-engineered scaffold for cartilage repair.     

Lipid peroxidation and biochemical parameters in umbilical cord blood of well-adapted term newborns

Lipid peroxidation (LPX) can play an important role in the development of pathological changes of foetal and neonatal tissues. We investigated LPX and biochemical parameters in plasma from mixed umbilical cord (m.u.c.) blood and acid-base balance (ABB) parameters in m.u.c. blood of well-adapted full-term newborns. LPX products were estimated as thiobarbituric acid reacting substances (TBARS) and were expressed by using of malondialdehyde (MDA) as a standard solution. Intensity of LPX was estimated in vitro in m.u.c. blood plasma without and with added LPX activator (125 μM L-ascorbate plus 5 μM FeSO₄) and in the incubated plasma (30 min, 37°C) under both conditions. Actual TBARS (3.51 ± 0.49 nmol/mL) were determined in the non-incubated plasma without the added LPX activator. Approximately twice higher TBARS were found in the incubated plasma without the LPX activator (7.29 ± 2.17 nmol/mL) or with it (8.57 ± 2.20 nmol/mL), as well as in the non-incubated plasma after its addition (7.38 ± 1.98 nmol/mL). All analysed biochemical parameters (Fe, total iron-binding capacity, uric acid, proteins, Mg, Ca, phosphate, glucose, K, Na, Cl, ALT, AST, GMT, CK, LD, HBD, AMS, ALP, ACP) and ABB parameters were within their reference ranges. The actual TBARS levels were found being positively correlated with α-hydroxybutyrate dehydrogenase (HBD) activity and negatively with pO₂. These results suggest that LPX in m.u.c. blood plasma might be activated. This activation could probably depend on extent of hypoxia. TBARS and their formation in vitro could be suitable parameters of LPX in m.u.c. blood.     

Effect of PRP and PPP on proliferation and migration of human chondrocytes and synoviocytes in vitro

Cartilage tissue engineering can provide substantial relief to people suffering from degenerative cartilage disease, such as osteoarthritis. The autologous platelet-rich plasma (PRP) application appears to improve cartilage healing due to its ability to positively influence cellular mechanisms, mainly in cells from synovium and cartilage. Primary cultures of human synovial fluid stem cells (synoviocytes, SCs) and chondrocytes (CCs) were exposed to various concentrations of non-activated PRP and plateletpoor plasma (PPP) prepared by apheresis. Cell proliferation and migration were evaluated in real-time with the non-invasive xCELLigence System. It was found that PRP had a similar effect on the growth of cells as fetal bovine serum (FBS). Surprisingly, our proliferation assay results indicated that 50% PPP had the largest effect on both cell types, with a statistically significant increase in cell number (P<0.001) compared to the (0% FBS) in vitro control. The migratory ability of SCs was significantly enhanced with 10% PRP and 0.8% hyaluronic acid (HA). HA also augmented migration of CCs. In summary, these results demonstrate that directed cell proliferation and migration are inducible in human articular CCs and SCs, and that both platelet-derived fractions may exert a positive effect and modulate several cell responses that are potentially involved in tissue integration during cartilage repair.     

Proteomic analysis of iron overload in human hepatoma cells

Iron-mediated organ damage is common in patients with iron overload diseases, namely, hereditary hemochromatosis. Massive iron deposition in parenchymal organs, particularly in the liver, causes organ dysfunction, fibrosis, cirrhosis, and also hepatocellular carcinoma. To obtain deeper insight into the poorly understood and complex cellular response to iron overload and consequent oxidative stress, we studied iron overload in liver-derived HepG2 cells. Human hepatoma HepG2 cells were exposed to a high concentration of iron for 3 days, and protein expression changes initiated by the iron overload were studied by two-dimensional electrophoresis and mass spectrometry. From a total of 1,060 spots observed, 21 spots were differentially expressed by iron overload. We identified 19 of them; 11 identified proteins were upregulated, whereas 8 identified proteins showed a decline in response to iron overload. The differentially expressed proteins are involved in iron storage, stress response and protection against oxidative stress, protein folding, energy metabolism, gene expression, cell cycle regulation, and other processes. Many of these molecules have not been previously suggested to be involved in the response to iron overload and the consequent oxidative stress.     

ApoE deficiency leads to a progressive age-dependent blood-brain barrier leakage

Previously, we reported a defect in the blood-brain barrier (BBB) of apolipoprotein E-deficient (apoE–/–) mice (24). Here, we investigate BBB permeability in wild-type (WT) and apoE–/– mice as a function of age. Both WT and apoE–/– mice showed significantly increased cortical BBB leakage with age. However, in apoE–/– mice, the leakage increased at a 3.7x higher rate compared with WT mice. Surprisingly, the cerebellum showed significantly more leakage than other brain regions across age, while there was no difference between the two hemispheres. To determine the contribution of tissue- vs. blood-borne apoE to vascular permeability, we generated chimeric mice by bone marrow transplantation and measured their BBB leakage. These experiments suggest that both blood- and tissue-derived apoE are equally important for BBB function. In sum, we find an age-dependent defect in the BBB that is exacerbated in apoE–/– mice. Since vascular defects are found in patients with age-related neurodegenerative diseases, such as Alzheimer's, age-related BBB leakage could underlie these defects and may thus be an important contributor to the cumulative neuronal damage of these diseases. apolipoprotein E; aging; age-related neurodegeneration; inflammation     

Combined pharmacological therapy of the acute radiation disease using a cyclooxygenase-2 inhibitor and an adenosine A3 receptor agonist

Combined approaches to the treatment of acute radiation disease are preferred to single-agent therapies due to proven or anticipated better outcomes comprising increased therapeutic efficacy and decreased incidence of undesirable side effects. Our studies on post-exposure treatment of mice irradiated by sublethal or lethal doses of ionizing radiation included testing the effectiveness of meloxicam, a cyclooxygenase-2 inhibitor, and IB-MECA, an adenosine A3 receptor agonist. The efficacy of meloxicam and IB-MECA to positively influence the progress of the acute radiation disease has been tested in situations of their combined administration with granulocyte colony-stimulating factor (G-CSF) or with each other. The results of our studies revealed a significantly improved regeneration of hematopoietic cell populations ranging from the bone marrow progenitor cells to mature blood cells following combined treatments. Also, survival of mice exposed to lethal radiation doses was highest in the animals treated with a combination of the two drugs. It can be inferred from the results that if the drug combinations employed were used in humans, e.g. in the treatment of victims of radiation accidents, a better therapeutic outcome could be expected. Therefore, further studies directed at clinical applications of meloxicam and IB-MECA in radiation victims is recommended.