Blood-Brain Barrier Damage Induces Release of
2-Macroglobulin*
Luca Cucullo
,
Nicola Marchi
,
Matteo Marroni
,
Vincent Fazio
,
Shobu Namura
and
Damir Janigro
,
,¶
Department of Neurological Surgery, Cerebrovascular Research Center, Cleveland Clinic Foundation, Cleveland, Ohio 44195
Department of Cell Biology, Cerebrovascular Research Center, Cleveland Clinic Foundation, Cleveland, Ohio 44195
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ABSTRACT
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Blood-brain barrier (BBB) failure occurs in many neurological diseases and is caused in part by activation of proinflammatory factors including matrix metalloproteinases. Counterbalancing, "BBB protective" cascades have recently been described, including NO-mediated interleukin 6 release by glia. Interleukin 6 has been shown to trigger production of matrix metalloproteinase inhibitors such as
2-macroglobulin (
2M). We hypothesized that BBB failure may result in increased
2M release by perivascular astrocytes. This was initially tested in patients undergoing iatrogenic BBB disruption by hyperosmotic mannitol for intra-arterial chemotherapy of brain tumors. Serum samples revealed significantly increased levels of
2M at 4 h after BBB disruption by hyperosmotic mannitol. In parallel in vitro experiments, we observed a similar increase of
2M release by astrocytes under conditions mimicking BBB failure and perivascular edema. For both experiments, protein analysis was initially performed by bidimensional gel electrophoresis and mass spectrometry followed by Western blotting immunodetection. We conclude that, in addition to proinflammatory changes, BBB failure may also trigger protective release of
2M by perivascular astrocytes as well as peripheral source.
The mature extracellular matrix, a heterogeneous substance produced at the BBB1 level by perivascular astrocytes, serves as a tissue skeleton, as a medium of communication between cells, and as a barrier between the cells and the vascular system (1, 2). MMPs are the key components of the system that dynamically control and continuously remodel the mature extracellular matrix in living tissues. The proteolytic activity of MMPs acts as an effector mechanism of tissue remodeling under physiologic and pathologic conditions and as a modulator of inflammation. In the context of neuroinflammatory diseases, MMPs have been implicated in processes such as BBB failure, invasion of neural tissue by blood-derived immune cells, and cellular damage in diseases of the central nervous system (1, 3, 4). In the central nervous system, matrix remodeling by metalloproteinases gelatinase A (MMP-2) and gelatinase B (MMP-9) play a major role in maintaining BBB integrity (57). Activation of proinflammatory factors and altered processes of extracellular proteolysis may contribute to the progression of tissue damage and matrix destruction leading to BBB failure after brain injury (8, 9).
MMPs have also been implicated in the pathogenesis of neurodegenerative diseases (Alzheimers disease) and are believed to be critical events leading to BBB failure with secondary vasogenic edema and hemorrhagic transformation of infarcted brain tissue (1012). However, the mechanism of activation of MMPs remains unclear. MMPs are neutralized by naturally occurring inhibitors such as
2M (13, 14) or tissue inhibitors of metalloproteinases.
2M is a broad spectrum protease inhibitor naturally present in serum and interstitial fluids.
2M production can be stimulated by interleukin 6 (1517). Recent findings suggest that glucocorticoids may stimulate
2M release and inhibit up-regulation of MMP-9 (18, 19). This is consistent with the concept that glucocorticoids may act as repair agents following BBB failure and edema formation (20).
2M is a tetramer of four identical 180-kDa subunits exhibiting a ß-cysteinyl-glutamyl-thiol ester bond and inhibits the target protease by physical entrapment.
2M has a peptide stretch, called the "bait region," that contains specific cleavage sites for different proteinases (21, 22). This protein also binds to a number of different growth factors and cytokines suggesting that
2M may play an additional role in cellular growth regulation (23, 24).
We focused on the role of astrocytes (AS) and endothelial cells (EC) in BBB maintenance by studying in vitro cell culture and serum samples obtained from patients who underwent BBBD procedures during chemotherapy treatment. We hypothesized that BBB impairment may result in increased
2M released by perivascular astrocytes as well as peripheral sources.
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EXPERIMENTAL PROCEDURES
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Cell Isolation and Culture
Specimens were obtained with the consent of patients undergoing brain surgery; the details of this procedure are listed elsewhere (25, 26). Primary AS and primary EC were isolated from human brain tissue and co-cultured in 75-mm-diameter Transwell polycarbonate membrane with pore size of 0.4 µm and total insert growth area of 44 cm2 (Costar 3419). Initially cells were grown to confluence in 150-cm2 flasks, and the growth media was collected every 2 days. Spare media were stored at -20 °C. EC and AS were then detached with trypsin and resuspended in fetal bovine serum.
To study the reciprocal interactions between astrocytes and endothelial cells, both of which can lead to BBB induction, six different culturing conditions were set (Fig. 4). Glial and endothelial cells were seeded at a density of 8 x 104/cm2. Before cell loading, Transwell membranes were precoated with fibronectin on the luminal (i.e. "endothelial") side and with poly-D-lysine on the abluminal (i.e. "astrocytic") side. Protein extracts were obtained from culture media samples of AS, EC, AS + EC, AS exposed to EC-conditioned media, and EC exposed to AS-conditioned media. To mimic an impaired BBB, EC-AS co-cultures were also grown spaced from each other by less than 1 mm of media between the cellular monolayers.
Patients
The six patients whose serum samples were used for the study presented herein all participated in a BBBD trial for the treatment of primary central nervous system lymphoma. The Cleveland Clinic Brain Tumor Institute provides BBBD treatment (2729) for primary central nervous system lymphoma, primitive neuroectodermal tumors, some gliomas, central nervous system germinoma, and some metastatic brain tumors (such as breast, small cell lung, or germ cell). These patients were treated with intra-arterial injection of mannitol causing a temporary disruption of the BBB followed by a selective methotrexate injection into the cerebral circulation directly thereafter. Blood samples were collected before (10 min prior to mannitol injection), during (45 s, 1 min, and 10 min after mannitol injection), and after (up to 6 h) iatrogenic selective intra-arterial osmotic opening of the BBB (30, 31, 33, 51). Efficacy of BBBD was assessed by computed tomography scans and, indirectly, by measuring S100ß level in plasma (30, 31). All patients signed an informed consent form prior to the procedure. Experiments with human subjects were performed in agreement with National Institutes of Health and Cleveland Clinic Foundation guidelines.
Two-dimensional Gel Electrophoresis
The proteins in aliquots of the respective conditioned media (or human serum in the case of BBBD protocol) were precipitated with acetone and reconstituted in an isoelectric focusing buffer consisting of 6 M urea, 2 M thiourea, 2% Chaps, 1% Triton X-100, 1% ampholytes, and 50 mM dithiothreitol. Isoelectric focusing was carried out for 43 kV-h at 20 °C in 11-cm immobilized pH gradient strips (Bio-Rad) covering pI 58, which were loaded with 500 µg of total protein using the active rehydration method. The second dimension was carried out in precast 12% SDS-polyacrylamide gels (Criterion gels, Bio-Rad). Gels were subsequently fixed and stained with Coomassie Blue (GelCode Blue, Pierce).
Mass Spectroscopy
We used a liquid chromatography-mass spectrometry system with a Protana microelectrospray ion source interfaced to a self-packed 10-cm x 75-µm-inner diameter Phenomenex Jupiter C18 reversed-phase capillary chromatography column. Data were analyzed by using all collision-induced dissociation spectra collected in the experiment to search the National Center for Biotechnology Information (NCBI) non-redundant data base with the search program TurboSequest. All matching spectra were verified by manual interpretation. The interpretation process was also aided by additional searches using Mascot and Fasta. S100ß was measured by techniques described elsewhere (32). The enzyme-linked immunosorbent assay kits were kindly provided by Dr. Anne-Charlotte Aronsson (Sangtec Medical, Bromma, Sweden).
Relative Protein Density on Two-dimensional Gel
Two-dimensional protein gels were scanned using a 35-mm camera mounted on a gel-scanning unit interfaced to a PC using GelProTM analysis software. The scanned black and white image was saved in a TIFF format and further analyzed using macros specifically developed for the Adobe Photoshop platform. The histogram function provided information on the number of pixels of the selected area. Tracings were performed in triplicate, and data were averaged and expressed as mean ± S.E. The density of the protein pattern for haptoglobin was used as normalizing value to account for variability between gels.
Western Blotting
Identification of
2M protein was also performed by Western blotting techniques. Serum samples were obtained from the BBBD procedures and media samples from the co-cultures. Protein concentration was estimated according to the Bradford assay method (Bio-Rad). Total proteins (50 µg/lane) were separated on 12% polyacrylamide gels with SDS-PAGE at 80 V and transferred onto a polyvinylidene difluoride membrane (Millipore Corp., Bedford, MA) by electroblotting overnight at 40 mA of constant current. After blocking with Blotto (phosphate-buffered saline, 1% milk powder, and 0.05% Tween 20) for at least 2 h, the membrane was probed overnight at 4 °C with primary
2M goat anti-human antibody (1:1000, Sigma). After a series of washes, the membrane was incubated with secondary horseradish peroxidase-conjugated anti-goat IgG antibody (rabbit) (1:5000, Calbiochem) for 2 h. Specific blots were visualized by enhanced chemiluminescence reagent (ECL Plus, Amersham Biosciences). For media samples, to ensure that the same amount of total protein was electroblotted, polyvinylidene difluoride membranes were incubated for 30 min at 50 °C in a "stripping buffer" (100 mM 2-mercaptoethanol, 2% sodium dodecyl sulfate, 62.5 mM Tris-HCl, pH 6.7). Nonspecific binding blocking was performed as described above; membranes were reprobed with rabbit anti-bovine albumin antibodies (1:1000, Calbiochem; see Fig. 5).
Relative expression of proteins was determined by densitometric analysis using Scion Image Software. Human
2M (Calbiochem) for the BBBD experiment and bovine albumin (Calbiochem) for the cell culture experiment were used as protein standards.
Relative Protein Density on Western Blot
Western blots were scanned on a scanner interfaced with a PC using ScanSoftTM analysis software. The scanned black and white images were saved in a TIFF format and further analyzed using macros specifically developed on a Adobe Photoshop platform. We first calculated a calibration curve by measuring the number of pixels of
2M bovine albumin standards. Density of the protein spots was transformed in nanograms by comparison with the standard.
Radial Immunodiffusion Test
This technique was used to quantitatively determine the concentration of
2M, haptoglobin-1, and S100ß in human serum. Prefabricated immunodiffusion plates were purchased from Kent Laboratories, Inc. (Bellingham, WA) containing antiserum to one of the three proteins in a medium of agarose gel, 0.1 M phosphate-buffered saline, pH 7.0, with 0.1% sodium azide. Standards and samples were pipetted at constant volume (5 µl) to individually prelabeled wells
3 mm in diameter. The plates are manufactured with 24 wells/plate. The proteins were allowed to diffuse to their end point following incubation at room temperature (2024 °C) for 48 h. Following incubation the diameters of the precipitin arcs formed at the periphery of each well were measured with a jewelers loupe (7x magnification) to the nearest 0.1 mm. A linear plot of the radial diffusion (in mm2) was plotted versus concentration of each protein (mg/dl). Samples and standards were run in duplicate to yield slopes consistent with the quality control data provided in the package insert. The R2 value of each standard curve was high, yielding an average result of 0.9954 (
2M), 0.9924 (haptoglobin-1), and 0.9939 (S100ß).
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RESULTS
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Serum samples taken from patients undergoing an osmotic opening of the BBB were analyzed by two-dimensional gel electrophoresis as described under "Experimental Procedures." Samples were also analyzed to determine levels of S100ß, a low molecular weight protein associated with BBB impairment (31). The results of a typical experiment are shown in Fig. 1. Note that distinct protein spots appeared increased after BBBD. Mass spectrometry revealed that these protein spots corresponded to
2M, a 163-kDa protein with pI
6.0 (NCBI accession number 4557225).
The fragments shown in Table I were used for protein identification as described under "Experimental Procedures." The results shown in Fig. 1A refer to blood samples taken from a representative patient before BBBD and 4 h after the procedure. Quantitative analysis was performed by determining the number of pixels in gels from six patients. The mean results of these experiments are shown in the bar graph in Fig. 1B. Only small changes in
2M expression were seen when comparing samples taken minutes after the procedure. However, a significant increase in
2M was observed at 4 h after BBBD. Levels of the spots used as control (haptoglobin-1) remained unchanged during the entire procedure.
Fig. 2 shows quantitative results obtained by radial immunodiffusion technique. An increase in
2M comparable to that observed in the two-dimensional gels was discovered (Fig. 2A). Levels of haptoglobin-1 (Fig. 2C) remained unchanged during the entire procedure as expected (based on the result shown in Fig. 1B). Note that post-BBB disruption samples invariably contained elevated levels of S100ß (Fig. 2B; see also Refs. 30, 31, 33, and 50), indicating effective opening of the blood-brain barrier.
Consistent with the data from radial immunodiffusion tests, increased immunoreactivity for
2M was also detected by Western blotting (Fig. 3B). Quantitative data were obtained by comparing known amounts of
2M standards to experimental results (Fig. 3A, see "Experimental Procedures" for further details). Fig. 3C shows the mean values obtained from these experiments.
A bidimensional tissue culture apparatus was used to grow human brain endothelial cells and astrocytes in a variety of configurations aimed at mimicking various stages of BBB formation or failure. The experimental conditions used are illustrated on the left side of Fig. 4. EC- and AS-conditioned media were obtained from astrocytic and endothelial cultures, respectively. These media were collected regularly every 2 days and stored at -20 °C. Co-culture of EC and AS were designed to mimic an intact BBB (cells in contact on opposite side of Transwell filters) or BBB failure accompanied by enlarged extracapillary space (EC on filter with glia plated on bottom; intercellular distance was
1 mm). Patterns of protein release were monitored by two-dimensional gel electrophoresis. Comparison of gels obtained under different conditions revealed the first appearance of a discrete spot when glia was cultured alone in the presence of EC-conditioned media (Fig. 4). When EC and AS were grown in proximity but not in contact, we noticed an augmentation (in size) of the previous spot and the appearance of two other discrete spots in the immediate proximity. Mass spectroscopy analysis revealed that these changes (see Table II) were due to release of human
2M (NCBI accession number 579592, 163 kDa, pI = 6.04). Western blotting also demonstrated increased
2M immunoreactivity under these conditions (Fig. 5). To avoid possible and unexpected changes in the experimental parameters, all samples were collected and processed simultaneously after 1 week since the cultures and the co-cultures were completely established.
View this table:
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TABLE II Protein fragments were used to identify 2M in media samples from co-culture of AS + EC set to mimic BBB failure (see text for details)
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DISCUSSION
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The main finding from the present work is that the antiproteolytic molecule
2M is increased in serum under conditions of blood-brain barrier failure. This was interpreted as an endogenous strategy aimed at counteracting activity of matrix metalloproteinases presumably released by injured BBB cells. Post-BBB disruption release of
2M was observed under two different experimental conditions. In one set of experiments,
2M serum levels were elevated hours after iatrogenic opening of the BBB. In parallel experiments, we documented
2M release in vitro by astrocytes under conditions mimicking BBB failure. The implications of this novel finding are 2-fold. First, it appears that in addition to tissue inhibitors of metalloproteinases, other endogenous mechanisms exist to limit the extent of matrix remodeling during ischemia or neurodegenerative diseases. Second, enhancing
2M expression may be a useful strategy against MMP-mediated degradation of the extracellular matrix.
Temporal Patterns and Molecular Mechanisms of
2M Release
While most pictorials represent the brain endothelium as a static, tight-junctioned barrier, the BBB has been repeatedly shown to be a dynamic organ that readily responds and adapts to external stimuli (3437). So far, several mediators of BBB maintenance or failure have been described, but the exact molecular and cellular mechanisms involved in endothelium-mediated central nervous system homeostasis are virtually unknown. Recent evidence has shown that MMPs are involved in the enzymatic digestion of the extracellular matrix (8, 10, 39). Interestingly serum levels of MMP-9 correlate with hemorrhagic conversion after thrombolytic therapy for acute stroke (40), suggesting that BBB damage may play a role in tissue plasminogen activator toxicity (4143). The findings presented here demonstrate that, in addition to potentially deleterious molecular events, antiproteolytic molecules such as
2M are also produced. It is not clear whether these are synthesized ex novo in direct response to injury or following activation of metalloproteinases. Alternatively it is possible that the ubiquitous
2M simply accumulates in blood after injury, after being recalled, or after leaking from the central nervous system.
It has been suggested that elevated CSF levels of
2M indicate damage to the blood-to-CSF barrier (44). Thus, it was hypothesized that, during neuroinflammation,
2M extravasates from blood to CSF due to the favorable gradient between blood and CSF and a damaged protein-permeant BBB. Our in vivo experiments did not address this issue directly since CSF samples were for obvious reasons not available. However, comparisons of a time course of
2M level with changes in the putative indicator of BBB damage S100ß suggest that the observed
2M increase was a consequence of BBBD rather than a concomitant event. In fact,
2M levels were significantly increased only after the initial, rapid surge of S100ß (31, 33). Thus the increased levels of
2M after BBB disruption may be a reaction to proinflammatory changes that occur after the insult to the brain endothelium.
Cellular Origin of
2M
Under normal conditions, the plasma level of
2M exceeds the CSF level of
2M by a factor of >1,000 (4547). Thus, if the BBB is permeable to macromolecules (for example after osmotic disruption of endothelial tight junctions)
2M levels can be elevated in the CSF but not in the plasma. However, we observed a marked increase in serum
2M following osmotic opening of the BBB suggesting that the source of
2M may be peripheral. In vitro, in contrast, we observed a detectable increase in
2M presumably released by glia. This latter hypothesis is supported by the fact that
2M was found under the condition where glia was cultured alone in the presence of EC-conditioned media. Under the condition where EC and AS were grown in proximity but not in contact, we noticed an increased release of
2M suggesting the endothelium modulates
2M release by glia. Under this latter condition, the appearance of three spots at the same molecular weight as the
2M band but positioned at slightly different pIs suggests that different isoforms of
2M were also released. By mass spectrometry and Western blot analysis, they were demonstrated to be the same protein. The results obtained from in vivo and in vitro experiments are not necessarily in contradiction since it is possible that
2M is released systemically and by perivascular glia after osmotic challenge. If this was the case, what are the initial triggers and critical cellular substrates of
2M release?
2M is primarily synthesized by liver cells. It has been suggested that
2M can also be synthesized by astrocytes (48), which is consistent with our current findings. It may be argued that the hyperosmotic medium itself could cause
2M release. However, this seems unlikely since hyperosmotic mannitol failed to cause significant release of
2M from human epithelial cells (49). In addition to liver cells and astroglia, blood cells may also produce
2M (50). Given the fact that the observed increases in serum are hard to reconcile exclusively with increased synthesis in brain, it appears that more than one cell type is involved. The critical origin of
2M supply needs to be determined in the future.
Two-dimensional Gel Plasma Electrophoresis: a Proteomic Tool to Study Peripheral Markers of Neurological Disorders
Modern neurodiagnostic techniques use imaging, ultrasound, direct sampling of cerebrospinal fluid, or Doppler techniques to gain insight in brain (dys)function. An array of blood tests would be ideal to examine changes occurring in the brain parenchyma. This is hampered by the presence of the BBB that effectively separates the brain from blood and vice versa (31, 33, 51). However, evidence suggests that the blood-brain barrier is breached in a variety of neurological disorders, allowing communication between the periphery and central nervous system. Thus, diagnosis of diseases may be possible by peripheral blood analysis and routinely used in ischemic heart attack. Creatine phosphokinase and lactate dehydrogenase are reliable markers of myocardiac infarct.
A number of putative markers of Alzheimers dementia consist of proteins or protein fragments that are mutated in the disease condition (52). A mutated form of
2M has been suggested to participate in a neuropathogenic pathway leading to Alzheimers dementia (53). Thus, detection of
2M in serum may afford a routine test for early detection of presenile dementia. We have previously shown the value of the proteomic approach to discovery of markers of BBB damage (31, 33, 51). Limitations of this approach include the relatively poor resolution of high molecular weight signals in serum due to the preponderance of albumin expression, the equally poor resolution of proteins present in low abundance, and the relatively high cost of this technique. However, two-dimensional gel electrophoresis has the immense advantage of allowing rapid identification of proteins by mass spectroscopy. This approach has in the past allowed the evaluation of
2M changes occurring during aging (38), further demonstrating that at least for selected high molecular weight proteins this approach is feasible. Experiments are on-going to explore the possibility of using this approach to measure mutated, polymorphic forms of this protein in Alzheimers dementia and other disorders.
In conclusion, we demonstrated that a rapid increase of
2M in serum occurs in patients after BBBD. Moreover we demonstrated that in vitro BBB failure mimicking this condition up-regulates
2M release from AS. Although MMP activity levels need to be studied in these experimental paradigms, our data suggest that
2M may constitute an endogenous counterbalancing mechanism against BBB failure.
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ACKNOWLEDGMENTS
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The kits used to measure S100ß were a kind gift from Sangtec Medical (Bromma, Sweden).
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FOOTNOTES
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Received, November 26, 2002, and in revised form, April 10, 2003.
Published, MCP Papers in Press, April 24, 2003, DOI 10.1074/mcp.M200077-MCP200
1 The abbreviations used are: BBB, blood-brain barrier; MMP, matrix metalloproteinase; BBBD, BBB disruption by hyperosmotic mannitol;
2M,
2-macroglobulin; AS, astrocytes; EC, endothelial cells; Chaps, 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid; CSF, cerebrospinal fluid. 
* This work was supported by National Institutes of Health Grants HL51614, NS43284, and NS38195 and by the Yamanouchi USA Foundation. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 
¶ To whom correspondence should be addressed: Cerebrovascular Research, Cleveland Clinic Foundation NB20, Neurosurgery, 9500 Euclid Ave./NB20, Cleveland, OH 44195. Tel.: 216-445-0561; Fax: 216-444-1466; E-mail: janigrd{at}ccf.org
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REFERENCES
|
---|
- Rosenberg, G. A., Kornfeld, M., Estrada, E., Kelley, R. O., Liotta, L. A., and Stetler-Stevenson, W. G.
(1992) TIMP-2 reduces proteolytic opening of blood-brain barrier by type IV collagenase.
Brain Res.
576, 203
207[CrossRef][Medline]
- Okada, Y., Copeland, B. R., Fitridge, R., Koziol, J. A., and del Zoppo, G. J.
(1994) Fibrin contributes to microvascular obstructions and parenchymal changes during early focal cerebral ischemia and reperfusion.
Stroke
25, 1847
1853[Abstract]
- Mun-Bryce, S., and Rosenberg, G. A.
(1998) Gelatinase B modulates selective opening of the blood-brain barrier during inflammation.
Am. J. Physiol.
274, R1203
R1211[Medline]
- del Zoppo, G. J., and Hallenbeck, J. M.
(2000) Advances in the vascular pathophysiology of ischemic stroke.
Thromb. Res.
98, 73
81[CrossRef]
- Haas, T. L., and Madri, J. A.
(1999) Extracellular matrix-driven matrix metalloproteinase production in endothelial cells: implications for angiogenesis.
Trends Cardiovasc. Med.
9, 70
77[CrossRef][Medline]
- Sternlicht, M. D., and Werb, Z.
(2001) How matrix metalloproteinases regulate cell behavior.
Annu. Rev. Cell Dev. Biol.
17, 463
516[CrossRef][Medline]
- Nguyen, M., Arkell, J., and Jackson, C. J.
(2001) Human endothelial gelatinases and angiogenesis.
Int. J. Biochem. Cell Biol.
33, 960
970[CrossRef][Medline]
- Lukes, A., Mun-Bryce, S., Lukes, M., and Rosenberg, G. A.
(1999) Extracellular matrix degradation by metalloproteinases and central nervous system diseases.
Mol. Neurobiol.
19, 267
284[Medline]
- Mun-Bryce, S., and Rosenberg, G. A.
(1998) Matrix metalloproteinases in cerebrovascular disease.
J. Cereb. Blood Flow Metab.
18, 1163
1172[Medline]
- Lo, E. H., Wang, X., and Cuzner, M. L.
(2002) Extracellular proteolysis in brain injury and inflammation: role for plasminogen activators and matrix metalloproteinases.
J. Neurosci. Res.
69, 1
9[CrossRef][Medline]
- Rosenberg, G. A., Estrada, E. Y., and Dencoff, J. E.
(1998) Matrix metalloproteinases and TIMPs are associated with blood-brain barrier opening after reperfusion in rat brain.
Stroke
29, 2189
2195[Abstract/Free Full Text]
- Stanimirovic, D., and Satoh, K.
(2000) Inflammatory mediators of cerebral endothelium: a role in ischemic brain inflammation.
Brain Pathol.
10, 113
126[Medline]
- Woessner, J. F., Jr.
(1999) Matrix metalloproteinase inhibition. From the Jurassic to the third millennium.
Ann. N. Y. Acad. Sci.
878, 388
403[Abstract/Free Full Text]
- Beekman, B., Drijfhout, J. W., Ronday, H. K., and TeKoppele, J. M.
(1999) Fluorogenic MMP activity assay for plasma including MMPs complexed to
2-macroglobulin
Ann. N. Y. Acad. Sci.
878, 150
158[Abstract/Free Full Text]
- Aschner, M.
(1998) Immune and inflammatory responses in the CNS: modulation by astrocytes.
Toxicol. Lett.
102103, 283
287[CrossRef]
- Bode, J. G., Fischer, R., Haussinger, D., Graeve, L., Heinrich, P. C., and Schaper, F.
(2001) The inhibitory effect of IL-1ß on IL-6-induced
2-macroglobulin expression is due to activation of NF-
B.
J. Immunol.
167, 1469
1481[Abstract/Free Full Text]
- Gao, F., Bales, K. R., Dodel, R. C., Liu, J., Chen, X., Hample, H., Farlow, M. R., Paul, S. M., and Du, Y.
(2002) NF-
B mediates IL-1ß-induced synthesis/release of
(2)-macroglobulin in a human glial cell line.
Brain Res. Mol. Brain Res.
105, 108
114[Medline]
- Marzolo, M. P., von Bernhardi, R., Bu, G., and Inestrosa, N. C.
(2000) Expression of
(2)-macroglobulin receptor/low density lipoprotein receptor-related protein (LRP) in rat microglial cells.
J. Neurosci. Res.
60, 401
411[CrossRef][Medline]
- Harkness, K. A., Adamson, P., Sussman, J. D., Davies-Jones, G. A., Greenwood, J., and Woodroofe, M. N.
(2000) Dexamethasone regulation of matrix metalloproteinase expression in CNS vascular endothelium.
Brain
123, 698
709[Abstract/Free Full Text]
- Hedley-Whyte, E. T., and Hsu, D. W.
(1986) Effect of dexamethasone on blood-brain barrier in the normal mouse.
Ann. Neurol.
19, 373
377[Medline]
- Bowen, M. E., and Gettins, P. G.
(1998) Bait region involvement in the dimer-dimer interface of human
2-macroglobulin and in mediating gross conformational change. Evidence from cysteine variants that form interdimer disulfides.
J. Biol. Chem.
273, 1825
1831[Abstract/Free Full Text]
- Gettins, P. G., Hahn, K. H., and Crews, B. C.
(1995)
2-Macroglobulin bait region variants. A role for the bait region in tetramer formation.
J. Biol. Chem.
270, 14160
14167[Abstract/Free Full Text]
- Dennis, P. A., Saksela, O., Harpel, P., and Rifkin, D. B.
(1989)
2-Macroglobulin is a binding protein for basic fibroblast growth factor.
J. Biol. Chem.
264, 7210
7216[Abstract/Free Full Text]
- Ronne, H., Anundi, H., Rask, L., and Peterson, P. A.
(1979) Nerve growth factor binds to serum
2-macroglobulin.
Biochem. Biophys. Res. Commun.
87, 330
336[Medline]
- Desai, S., Marroni, M., Cucullo, L., Bengez, L., Hossein, M., Mayberg, M. R., Grant, G. A., and Janigro, D.
(2002) Mechanisms of endothelial survival under shear stress.
Endothelium
9, 89
102[Medline]
- Dombrowski, S., Desai, S., Marroni, M., Cucullo, L., Bingaman, W., Mayberg, M. R., Bengez, L., and Janigro, D.
(2001) Overexpression of multiple drug resistance genes in endothelial cells from patients with refractory epilepsy.
Epilepsia
42, 1504
1507
- Neuwelt, E. A., Goldman, D. L., Dahlborg, S. A., Crossen, J., Ramsey, F., Roman Goldstein, S., Braziel, R., and Dana, B.
(1991) Primary CNS lymphoma treated with osmotic blood-brain barrier disruption: prolonged survival and preservation of cognitive function.
J. Clin. Oncol.
9, 1580
1590[Abstract]
- Kroll, R. A., and Neuwelt, E. A.
(1998) Outwitting the blood-brain barrier for therapeutic purposes: osmotic opening and other means.
Neurosurgery
42, 1083
1099[Medline]
- Roman-Goldstein, S., Mitchell, P., Crossen, J. R., Williams, P. C., Tindall, A., and Neuwelt, E. A.
(1995) MR and cognitive testing of patients undergoing osmotic blood-brain barrier disruption with intraarterial chemotherapy.
Am. J. Neuroradiol.
16, 543
553[Abstract]
- Marchi, N., Fazio, V., Cucullo, L., Kight, K., Masaryk, T. J., Barnett, G., Vogelbaum, M., Kinter, M., Mayberg, M. R., and Janigro, D.
(2002) Serum transthyretin as a possible marker of blood-to-CSF barrier disruption.
J. Neurosci.
23, 1949
1955
- Marchi, N., Rasmussen, P. A., Kapural, M., Fazio, V., Cavaglia, M., and Janigro, D.
(2002) Peripheral markers of brain damage and blood-brain barrier dysfunction.
Restor. Neurol. Neurosci., in press
- Georgiadis, D., Berger, A., Kowatschev, E., Lautenschlager, C., Borner, A., Lindner, A., Schulte-Mattler, W., Zerkowski, H. R., Zierz, S., and Deufel, T.
(2000) Predictive value of S-100ß and neuron-specific enolase serum levels for adverse neurologic outcome after cardiac surgery.
J. Thorac. Cardiovasc. Surg.
119, 138
147[Abstract/Free Full Text]
- Kapural, M., Bengez, L., Barnett, G., Perl, J., II, Masaryk, T. J., Apollo, D., Mayberg, M. R., and Janigro, D.
(2002) S-100ß as a possible serum marker for disruption on the blood-brain barrier.
Brain Res.
940, 102
104[CrossRef][Medline]
- Grant, G. A., Abbott, N. J., and Janigro, D.
(1998) Understanding the physiology of the blood-brain barrier: in vitro models.
News Physiol. Sci.
13, 287
293[Abstract/Free Full Text]
- Revest, P. A., Jones, H. C., and Abbott, N. J.
(1993) The transendothelial DC potential of rat blood-brain barrier vessels in situ.
Cereb. Vasc. Biol.
331, 71
74
- Abbott, N. J., Revest, P. A., and Romero, I. A.
(1992) Astrocyte-endothelial interaction: physiology and pathology.
Neuropathol. Appl. Neurobiol.
18, 424
433[Medline]
- McAllister, M. S., Krizanac-Bengez, L., Macchia, F., Naftalin, R. J., Pedley, K. C., Mayberg, M. R., Marroni, M., Leaman, S., Stanness, K. A., and Janigro, D.
(2001) Mechanisms of glucose transport at the blood-brain barrier: an in vitro study.
Brain Res.
904, 20
30[CrossRef][Medline]
- Kondo, T., Sakaguchi, M., and Namba, M.
(2001) Two-dimensional gel electrophoretic studies on the cellular aging: accumulation of
2-macroglobulin in human fibroblasts with aging.
Exp. Gerontol.
36, 487
495[CrossRef][Medline]
- Rosenberg, G. A.
(2002) Matrix metalloproteinases in neuroinflammation.
Glia
39, 279
291[CrossRef][Medline]
- Silvestre, J. S., Mallat, Z., Tamarat, R., Duriez, M., Tedgui, A., and Levy, B. I.
(2001) Regulation of matrix metalloproteinase activity in ischemic tissue by interleukin-10: role in ischemia-induced angiogenesis.
Circ. Res.
89, 259
264[Abstract/Free Full Text]
- Tran, N. D., Schreiber, S. S., and Fisher, M.
(1998) Astrocyte regulation of endothelial tissue plasminogen activator in a blood-brain barrier model.
J. Cereb. Blood Flow Metab.
18, 1316
1324[Medline]
- Tran, N. D., Correale, J., Schreiber, S. S., and Fisher, M.
(1999) Transforming growth factor-ß mediates astrocyte-specific regulation of brain endothelial anticoagulant factors.
Stroke
30, 1671
1678[Abstract/Free Full Text]
- Wagner, K. R., Xi, G., Ya, H., Zuccarello, M., De Courten-Myers, G. M., Broderick, J. P., and Brott, T. G.
(1999) Ultra-early clot aspiration after lysis with tissue plasminogen activator in a porcine model of intracerebral hemorrhage: edema reduction and blood-brain barrier protection.
J. Neurosurg.
90, 491
498[Medline]
- Kanoh, Y., and Ohtani, H.
(1997) Levels of interleukin-6, CRP and
2 macroglobulin in cerebrospinal fluid (CSF) and serum as indicator of blood-CSF barrier damage.
Biochem. Mol. Biol. Int.
43, 269
278[Medline]
- Schliep, G., Rapic, N., and Felgenhauer, K.
(1974) Quantitation of high-molecular proteins in cerebrospinal fluid.
Z. Klin. Chem. Klin. Biochem.
12, 367
369[Medline]
- Felgenhauer, K.
(1974) Protein size and cerebrospinal fluid composition.
Klin. Wochenschr.
52, 1158
1164[Medline]
- Felgenhauer, K., and Hagedorn, D.
(1980) Two-dimensional separation of human body fluid proteins.
Clin. Chim. Acta
100, 121
132[CrossRef][Medline]
- Bauer, J., Gebicke-Haerter, P. J., Ganter, U., Richter, I., and Gerok, W.
(1988) Astrocytes synthesize and secrete
2-macroglobulin: differences between the regulation of
2-macroglobulin synthesis in rat liver and brain.
Adv. Exp. Med. Biol.199
205
- Koskela, H., Di Sciascio, M. B., Anderson, S. D., Andersson, M., Chan, H. K., Gadalla, S., and Katelaris, C.
(2000) Nasal hyperosmolar challenge with a dry powder of mannitol in patients with allergic rhinitis. Evidence for epithelial cell involvement.
Clin. Exp. Allergy
30, 1627
1636[Medline]
- Lysiak, J. J., Hussaini, I. M., and Gonias, S. L.
(1997)
2-Macroglobulin synthesis by the human monocytic cell line THP-1 is differentiation state-dependent.
J. Cell. Biochem.
67, 492
497[CrossRef][Medline]
- Kanner, A., Marchi, N., Fazio, V., Mayberg, M. R., Koltz, M., Siomin, V., Stevens, G. H. G., Masaryk, T. J., Apollo, D., Vogelbaum, M., Barnett, G., and Janigro, D.
(2003) Serum S100ß: a non-invasive marker of blood-brain barrier function and brain lesions.
Cancer
97, 2806
2813[CrossRef][Medline]
- Teunissen, C. E., de Vente, J., Steinbusch, H. W., and De Bruijn, C.
(2002) Biochemical markers related to Alzheimers dementia in serum and cerebrospinal fluid.
Neurobiol. Aging
23, 485
508[CrossRef][Medline]
- Blacker, D., Wilcox, M. A., Laird, N. M., Rodes, L., Horvath, S. M., Go, R. C., Perry, R., Watson, B., Jr., Bassett, S. S., McInnis, M. G., Albert, M. S., Hyman, B. T., and Tanzi, R. E.
(1998)
2 macroglobulin is genetically associated with Alzheimer disease.
Nat. Genet.
19, 357
360[CrossRef][Medline]