Key words: Blood-brain barrier; Astrocytes; Endothelial cells; Neuroregeneration; Cytokines; Trafficking; Spinal cord injury; EAE; Neuroinflammation; Obesity

The BBB Group is composed of two laboratories that share resources and provide mutual support.

Blood-Brain Barrier Laboratory - I (Dr. Weihong Pan, Dr. Kirsten Prufer, Dr. Xiaojun Wu, Mr. Michael Cash)

General Theme - To determine the role of the blood-brain barrier (BBB) in neuroinflammation and neuroregeneration.

Current Projects

• Regulation of the interleukin-15 (IL15) system in the CNS: IL15 is a unique T helper cell cytokine that plays important roles in CNS development and physiological functions, response to inflammation, and development of autoimmune disease. Tumor necrosis factor alpha (TNF), lipopolysaccharide, and experimental autoimmune encephalomyelitis are among the strongest inducers that activate the IL15 system in the CNS. Chuanhui (Jasmine) and Xiaojun employ different molecular, cellular, and animal models to determine the mechanisms of regulation of IL15 and its receptors. This involves intracellular trafficking studies by use of confocal microscopy, generation and expression of different mutant constructs, analyses of transcriptional regulation by genetic studies on promoters and untranslated RNAs, studies of protein turnover, transport assays, and a series of neurobehavioral analyses. We hope to identify some novel answers about how transcytosis occurs and what mechanisms drive the upregulation of the transport.
• Functional implications of upregulated transport: Cytokines are dynamically involved in neuroinflammation, neuroendocrine changes, and regenerative processes. The specific changes of cytokine transport systems might just reflect an adaptive change of the BBB to CNS insults, or they may serve beneficial roles in promoting functional recovery. Thus, we address the specific consequences of cytokine transport by use of overexpression and knockdown approaches in combination with histological and behavioral parameters.
• Signal modification at the BBB: We have shown at the BBB level that one proinflammatory cytokine can affect the signal transduction and transport of another. This indicates that the BBB plays a crucial role in integrating peripheral stimuli and in relaying messages to the CNS after its “interpretation”. We know only very little from studies with LIF receptors, which are subject to modulation by TNF and lipopolysaccharide. Our ongoing studies mainly focus on the interleukin-15 system and the P-glycoprotein efflux transporter.

Figure from Pan et al., J Neuroimmunol 07: TNF transport across the mouse BBB after iv injection. The EM autoradiography shows the TNF molecule (black silvergrain) transversing the lumen of microvessels to reach CNS parenhyma (in collaboration with the von Bartheld lab at Univ of Nevada).

Blood-Brain Barrier Laboratory - II (Dr. Abba Kastin, Mr. Hung Hsuchou, Dr. Reas Khan Sulaimankutty, Ms. Emily Markadakis)

Focus - To determine the role of the blood-brain barrier (BBB) in neuroendocrine control, particularly related to peptides/polypeptides involved in feeding behavior.

Current Projects

We pioneered the concept that peptides in the periphery have CNS effects several decades ago, and are now showing the mechanisms of such actions. Concurrent with the publication of the 216-chapter book of Biologically Active Peptides, Dr. Kastin recently received another honorary doctorate, this time from Uppsala University in Sweden. We have two exciting areas of focus this year: the astrocytic leptin receptor (ObR) and the naturally occuring brain tripeptide Pro-Leu-Gly-amide (MIF-1).

The 16 kD leptin is an adipokine (a polypeptide produced by fat cells, or adipocytes) and a neuroendocrine hormone. In the course of studying the roles of ObR subtypes in leptin transport across the cerebral endothelia composing the BBB, we incidentally found that obesity induces robust upregulation of ObR in astrocytes. It took a long while for the research community to accept this fact, but we succeed by a series of immunofluorescent histochemistry, double-labeling fluorescent in-situ hybridization, quantitative PCR, calcium imaging, and cellular signaling assays. Now it is time to show how this happens and what the consequence is in the regulatory changes in subjects with obesity and metabolic syndrome.

Some of the questions we have addressed so far include the following (if you do not know the answers, check out our recent papers from Hong and Hung in the Publications webpage): How does the transport of feeding-related peptides change in altered feeding and nutritional states? How do different peptides and polypeptides, such as urocortin and leptin, interact at the BBB level? What isoforms of the leptin receptor are involved in transcytosis and also exocytosis? Are there differences in expression of leptin receptors and leptin transport between neonatal and adult mice? Do Avy mice, which develop obesity later in life, transport ingestive peptides differently than their controls? And then, how can we use this information to assist in the treatment of pathological obesity?

The tripeptide MIF-1 (Pro-Leu-Gly-amide) was borne as Kastin's brain baby three decades ago. Isolated from brain tissue, MIF-1 has been active in several animal models of Parkinson's disease and mental depression, as well as clinical studies in these disorders where it has a surprisingly rapid onset. In collaboration with our Proteomics Core (Kheterpal) and Applied Biosystems, we detected MIF-1 in different regions of mouse brain by ultrasensitive MRM mass spectrometry techniques. Our focus now is on its mechanism of action, probably different from other agents used in these diseases.

It seems that our postdoctoral fellow Reas indeed links the projects (at least leptin and MIF-1) with glutamate. Reas also established a mouse model of epilepsy and EEG sleep recording. The next thing is to recruit one or two more postdocs to devote the same enthusiasm and technical expertise.

Figure from Pan et al., Endocrinology 08: Astrocytes (green) express leptin receptor (red). The co-localization (orange) is shown by confocal microscopy.