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The blood-brain barrier (BBB), a dynamic interface between blood and brain constituted mainly by endothelial cells of brain microvessels, robustly restricts the entry of potentially harmful blood-sourced substances and cells into the brain, however, many therapeutically active agents concurrently cannot gain access into the brain at effective doses in the presence of an intact barrier. On the other hand, breakdown of BBB integrity may involve in the pathogenesis of various neurodegenerative diseases. Besides, certain diseases/disorders such as Alzheimer's disease, hypertension, and epilepsy are associated with varying degrees of BBB disruption. In this review, we aim to highlight the current knowledge on the cellular and molecular composition of the BBB with special emphasis on the major transport pathways across the barrier type endothelial cells. We further provide a discussion on the innovative brain drug delivery strategies in which the obstacle formed by BBB interferes with effective pharmacological treatment of neurodegenerative diseases/disorders. [ABSTRACT FROM AUTHOR]

The blood-brain barrier (BBB) constituted by endothelial cells of brain microvessels is a dynamic interface, which controls and regulates the transport of various substances including peptides, proteins, ions, vitamins, hormones, and immune cells from the circulation into the brain parenchyma. Certain diseases/disorders such as Alzheimer's disease, sepsis, and hypertension can lead to varying degrees of BBB disruption. Moreover, impairment of BBB integrity has been implicated in the pathogenesis of various neurodegenerative diseases like epilepsy. In attempts to explore the wide spectrum of pathophysiologic mechanisms of these diseases/disorders, a variety of experimental insults targeted to the BBB integrity in vitro in cell culture models and in vivo in laboratory animals have been shown to alter BBB permeability causing enhanced transport of certain tracers such as sodium fluorescein, cadaverine-Alexa fluor, horseradish peroxidase, FITC-dextran, albumin-Alexa fluor conjugates, and Evans blue dye across the barrier. The permeability changes in barrier-type endothelial cells can be assessed by intravascular infusion of exogenous tracers and subsequent detection of the extravasated tracer in the brain tissue, which enable functional and structural analysis of BBB integrity. In this chapter, we aimed to highlight the current knowledge on the use of four most commonly performed tracers, namely, Evans blue, sodium fluorescein, albumin-Alexa fluor conjugates, and horseradish peroxidase. The experimental methodologies that we use in our laboratory for the detection of these tracers by macroscopy, spectrophotometry, spectrophotofluorometry, confocal laser scanning microscopy, and electron microscopy are also discussed. Tracing studies at the morphological level are mainly aimed at the identification of the tracers both in the barrier-related cells and brain parenchyma. In addition, BBB permeability to the tracers can be quantified using spectrophotometric and spectrophotofluorometric assays and image analysis by confocal laser scanning microscopy and electron microscopy. The results of our studies conducted under various experimental settings using the mentioned tracers indicate that barrier-type endothelial cells in brain microvessels orchestrate the paracellular and/or transcellular trafficking of substances across BBB. These efforts may not only contribute to designing approaches for the management of diseases/disorders associated with BBB breakdown but may also provide new insights for developing novel brain drug delivery strategies.

Blood-brain barrier (BBB) constituted primarily by the capillary endothelial cells functions to maintain a constant environment for the brain, by preventing or slowing down the passage of a variety of blood-borne substances, such as serum proteins, chemical compounds, ions, and hormones from the circulation into the brain parenchyma. Various diseases such as brain tumors, epilepsy, and sepsis disturb the BBB integrity leading to enhanced permeability of brain microvessels. In animal models, a variety of experimental insults targeted to the BBB integrity have been shown to increase BBB permeability causing enhanced passage of molecules into the brain paranchyma by transcellular and/or paracellular pathways. This alteration can be demonstrated by intravascular infusion of exogenous tracers and subsequent detection of extravasated molecules in the brain tissue. A number of exogenous BBB tracers are available, and they can be used for functional and structural analysis of BBB permeability. In this chapter, we aimed to highlight the basic knowledge on the use of three most commonly performed tracers, namely Evans blue dye, sodium fluorescein, and horseradish peroxidase. The experimental methodologies that we use in our laboratory for the detection of these tracers by macroscopy, spectrophotometry, spectrophotofluorometry, and electron microscopy are also discussed. While tracing studies at the morphological level are mainly aimed at the identification and characterization of the tracers both in the barrier related cells and brain parenchyma, spectrophotometric and spectrophotofluorometric assays enable quantification of BBB permeability. The results of our studies that we performed using the mentioned tracers indicate that barrier type of endothelial cells in brain play an important role in paracellular and/or transcytoplasmic trafficking of macromolecules across BBB under various experimental settings, which may provide new insights in both designing approaches for the management of diseases with BBB breakdown and developing novel trans-BBB drug delivery strategies.

Objective: To evaluate leptomeningeal and subpial inflammatory responses of experimental Staphylococcus aureus bacteriemia following intraperitoneal and intravenous applications and to compare the inflammatory reactions in different regions of central nervous system., Material and Methods: Forty anesthetized rats were divided into four groups equal in number. The rats in group-I were given 1 ml suspension of Staphylococcus aureus intraperitoneally. Group-II was the control group of group I; it was administrated 1 ml 0.9% NaCl in water intraperitoneally. The rats in group-III were given the same amount of bacteria intravenously. Group IV was the control group of the group-III; it was administrated 1 ml 0.9% NaCl solution intravenously. The rats were sacrificed on the 21st day. Inflammatory changes of different regions of the central nervous system were examined under transmission electron microscopy. Statistical analysis was done by using variance analysis, Bonferroni, Tamhane post hoc, Student's t and univariate tests., Results: Thoracic and occipital regions were the most vulnerable zones. Increasing of collagen tissue was the most detected inflammatory change., Conclusion: This experimental model can be used for inducing subpial and leptomeningeal inflammations and it may be developed for investigations of pathogenesis of leptomeningitis during systemic infections.

Distraction osteogenesis is a well established clinical treatment for limb length discrepancy and skeletal deformities. Transforming growth factor beta 1 (TGF-beta1) is a multifunctional peptide which controls proliferation and expression of cells specific to bone like chondrocytes, osteoblasts, osteoclasts including mesenchymal precursor cells. To decrease the external fixation time with increasing the strength of regenerate (newly formed bone after distraction) we tested the effect of locally applied transforming growth factor beta 1 on distraction osteogenesis. A total of 28 mature female white New zealand rabbits weighing 3,5 kg-4,5 kg were studied. 10 animals were belonging to biomechanical testing group (5 for the study and 5 for the control subgroups), and the others were to histology group. In biomechanical group after tibial osteotomy TGF-beta1 was applied subperiosteally for 5 days just proximal to osteotomy site. Control group received only the solvent. Seven days after tibial osteotomy distraction was started at a rate of 0.25 mm/12 hours for 3 weeks with a unilateral fixator. Rabbits were sacrificed at the end of a consolidation period 8 week after tibial osteotomy. We assessed density of the elongation zone of rabbit tibial bones with the computed tomography. Then biomechanical parametres were assessed using the torsional testing using the material testing machine. In histology group rabbits were classified as control and study (rabbits that were given TGF-beta1). Rabbits were sacrificed at the end of first week, second week and fourth week also at the end of consolidation period 8 week after tibial osteotomy. Immunohistochemical and histologic parameters were examined. Biomechanical testing was applied as torsional testing. These values are used in determination of maximal loading, stiffness and energy absorbed during testing (brittleness). The histomorphometric examination looked for the differences between the study and control groups in terms of bone formation pattern, bone quality and quantity. The immunohistochemical studies investigated the mechanism of TGF-beta1, and its presence in different cell types. The results of this study suggest that locally applied TGF-beta1 improves the mineral density of distraction gap and load to failure(energy absorbed during testing). Though there is no significant histomorphometric difference between the study and control groups, there is an increased bone mineral density and an according maximum energy absorbance in the study group. This effect can be explained by the following mechanism: TGF-beta1 exerts its effect on two different receptor types (Type 1 and 2). Type 1 receptors are localized to bone matrix and type 2 receptors are localized to the intracellular space. The specific stains utilized in the current experiment are specific to type 2 receptors. They have been shown to be down-regulated by exogenous TGF-beta1 injections. Most probably, type 1 receptors are up-regulated by this exogenous administration, but unfortunately, there is currently no specific stain on tha market to display type 1 receptors and to prove this explanation.

The aim of this study is to evaluate the prognostic role of thymidine labeling index in patients with breast cancer. Cellular proliferation rates in 155 breast cancer specimens were investigated by 3H-thymidine labeling index (3H-TLI). Median age was 47 years (range: 23-76). At presentation, 11 patients (7.1%) had stage I disease, 76 (49%) had stage II, 64 (41.3%) had stage III disease, and 4 (2.6%) had metastatic involvement. Patients were placed in 2 groups based on their proliferative indices. The cut-off level was assigned as the median TLI value of the whole group. Correlations between proliferative activity of the tumors based on 3H-TLI levels and various previously established prognostic factors, as well as the influence of proliferative activity on survival as a clinical outcome, were analyzed. The mean and median TLI values for the whole group of patients were 4.36 +/- 4.96% and 2.76% (range: 0-23.6), respectively. There was a significant association of nuclear grade with TLI (P = 0.04). Patients who were alive with no sign of disease at the final follow-up examination had a significantly lower median TLI rate than those who were either alive with disease or those who had eventually died with disease progression (3.7% versus 1.9%, respectively; P = 0.04). Patients with locally advanced disease (N2 + N3 involvement) had a significantly higher median TLI rate than those with local nodal involvement (N1) (3.4% versus 1.7%, respectively, P = 0.026). Furthermore, TLI levels showed a significant association with overall survival in patients with node-negative disease (P = 0.02). Based on the results of this study, it can be concluded that TLI plays a significant prognostic role in a subset of patients with node-negative breast cancer. Furthermore, TLI appears to have a predictive value for the clinical outcome of patients with breast cancer. These findings may justify a more aggressive therapeutic approach in patients with high TLI levels. Further large-scale, prospective studies are required before a definite conclusion can be reached.reached.