No abstract

Development of an optimal systemic drug delivery strategy to the brain will require noninvasive or minimally invasive methods to quantify the permeability of the cerebral microvessel wall or blood-brain barrier (BBB) to various therapeutic agents and to measure their transport in the brain tissue. To address this problem, we used laser-scanning multiphoton microscopy to determine BBB permeability to solutes (P) and effective solute diffusion coefficients (Deff) in rat brain tissue 100-250 μm below the pia mater. The cerebral microcirculation was observed through a section of frontoparietal bone thinned with a microgrinder. Sodium fluorescein, fluorescein isothiocyanate (FITC)-dextrans, or Alexa Fluor 488-immunoglobulin G (IgG) in 1% bovine serum albumin (BSA) mammalian Ringer's solution was injected into the cerebral circulation via the ipsilateral carotid artery by a syringe pump at a constant rate of ∼3 ml/min. P and Deff were determined from the rate of tissue solute accumulation and the radial concentration gradient around individual microvessels in the brain tissue. The mean apparent permeability P values for sodium fluorescein (molecular weight (MW) 376 Da), dextran-4k, -20k, -40k, -70k, and IgG (MW ∼160 kDa) were 14.6, 6.2, 1.8, 1.4, 1.3, and 0.54 × 10-7 cm/s, respectively. These P values were not significantly different from those of rat pial microvessels for the same-sized solutes (Yuan et al., 2009, "Non-Invasive Measurement of Solute Permeability in Cerebral Microvessels of the Rat," Microvasc. Res., 77(2), pp. 166-73), except for the small solute sodium fluorescein, suggesting that pial microvessels can be a good model for studying BBB transport of relatively large solutes. The mean Deff values were 33.2, 4.4, 1.3, 0.89, 0.59, and 0.47 × 10-7 cm2/s, respectively, for sodium fluorescein, dextran-4k, -20k, -40k, -70k, and IgG. The corresponding mean ratio of Deff to the free diffusion coefficient Dfree, Deff/Dfree, were 0.46, 0.19, 0.12, 0.12, 0.11, and 0.11 for these solutes. While there is a significant difference in Deff/Dfree between small (e.g., sodium fluorescein) and larger solutes, there is no significant difference in Deff/Dfree between solutes with molecular weights from 20,000 to 160,000 Da, suggesting that the relative resistance of the brain tissue to macromolecular solutes is similar over a wide size range. The quantitative transport parameters measured from this study can be used to develop better strategies for brain drug delivery.

The resonance Raman (RR) spectra of six types of human brain tissues are examined using a confocal micro-Raman system with 532-nm excitation in vitro. Forty-three RR spectra from seven subjects are investigated. The spectral peaks from malignant meningioma, stage III (cancer), benign meningioma (benign), normal meningeal tissues (normal), glioblastoma multiforme grade IV (cancer), acoustic neuroma (benign), and pituitary adenoma (benign) are analyzed. Using a 532-nm excitation, the resonance-enhanced peak at 1548  cm-1 (amide II) is observed in all of the tissue specimens, but is not observed in the spectra collected using the nonresonance Raman system. An increase in the intensity ratio of 1587 to 1605  cm-1 is observed in the RR spectra collected from meningeal cancer tissue as compared with the spectra collected from the benign and normal meningeal tissue. The peak around 1732  cm-1 attributed to fatty acids (lipids) are diminished in the spectra collected from the meningeal cancer tumors as compared with the spectra from normal and benign tissues. The characteristic band of spectral peaks observed between 2800 and 3100  cm-1 are attributed to the vibrations of methyl (─CH3) and methylene (─CH2─) groups. The ratio of the intensities of the spectral peaks of 2935 to 2880  cm-1 from the meningeal cancer tissues is found to be lower in comparison with that of the spectral peaks from normal, and benign tissues, which may be used as a distinct marker for distinguishing cancerous tissues from normal meningeal tissues. The statistical methods of principal component analysis and the support vector machine are used to analyze the RR spectral data collected from meningeal tissues, yielding a diagnostic sensitivity of 90.9% and specificity of 100% when two principal components are used.