Biochemical, biophysical, and pharmacological characterization of bacterially expressed human agouti-related protein.

The agouti-related protein gene (Agrp) is a novel gene implicated in the control of feeding behavior. The hypothalamic expression of Agrp is regulated by leptin, and overexpression of Agrp in transgenic animals results in obesity and diabetes. By analogy with the known actions of agouti, these data suggest a role for the Agrp gene product in the regulation of melanocortin receptors expressed in the central nervous system. The availability of recombinant, highly purified protein is required to fully address this potential interaction. A nearly full-length form of AGRP (MKd5-AGRP) was expressed in the cytosolic or soluble fraction of Escherichia coli and appeared as large intermolecular disulfide-bonded aggregates. Following oxidation, refolding, and purification, this protein was soluble, and eluted as a single symmetric peak on RP-HPLC. Circular dichroism studies indicated that the purified protein contains primarily random coil and beta-sheet secondary structure. Sedimentation velocity studies at neutral pH demonstrated that MKd5-AGRP is monomeric at low micromolar concentrations. Mobility shifts observed using SDS-PAGE under reducing and nonreducing conditions for bacterially expressed and mammalian expressed AGRP were identical, an indication of a similar disulfide structure. The purification to homogeneity of a second, truncated form of AGRP (Md65-AGRP) which was expressed in the insoluble or inclusion body fraction is also described. Both forms act as competitive antagonists of alpha-melanocyte stimulating hormone (alpha-MSH) at melanocortin-3 (MC-3) and melanocortin-4 receptors (MC-4). The demonstration that AGRP is an endogenous antagonist with respect to these receptors is a unique mechanism within the central nervous system, and has important implications in the control of feeding.

Cloning and functional expression of cDNAs encoding human and rat pancreatic polypeptide receptors.

PCR was used to isolate nucleotide sequences that may encode novel members of the neuropeptide Y receptor family. By use of a PCR product as a hybridization probe, a full-length human cDNA was isolated that encodes a 375-aa protein with a predicted membrane topology identifying it as a member of the G-protein-coupled receptor superfamily. After stable transfection of the cDNA into human embryonic kidney 293 cells, the receptor exhibited high affinity (Kd = 2.8 nM) for 125I-labeled human pancreatic polypeptide (PP). Competition binding studies in whole cells indicated the following rank order of potency: human PP = bovine PP > or = human [Pro34]peptide YY > rat PP > human peptide YY = human neuropeptide Y. Northern blot analysis revealed that human PP receptor mRNA is most abundantly expressed in skeletal muscle and, to a lesser extent, in lung and brain tissue. A rat cDNA clone encoding a high-affinity PP receptor that is 74% identical to the human PP receptor at the amino acid level was also isolated. These receptor clones will be useful in elucidating the functional role of PP and designing selective PP receptor agonists and antagonists.

CT detection of intracranial aneurysms in subarachnoid hemorrhage.

Contrast enhanced computed tomography (CT) scans of 59 patients with subarachnoid hemorrhage were evaluated for intracranial aneurysms. After prospective study, the CT results were compared with angiography. Depending on the location, the CT detection rate varied from a low of 36% for the internal carotid artery complex to a high of 76% for anterior and middle cerebral artery aneurysms. It was concluded that angiography may be deferred if the causative aneurysm is demonstrated by CT.

Basal ganglia calcification in Down's syndrome.

Basal ganglia calcification has been observed in autopsy material of 7% of children suffering from Down's Syndrome. The incidence of this finding in the general neurological population is approx. 0.3%. The present report is the first case of Down's Syndrome with basal ganglia calcification diagnosed in life by computed axial tomography. This case illustrates that dense basal ganglia calcification can exist with no signs or symptoms of a movement disorder.

Essential facets of radiological diagnosis of extremity trauma.

Trauma to the extremities is sometimes dismissed as relatively unimportant in certain radiologic circles. In many instances, radiologic responsibility is denied and relegated to the orthopedist. Although the radiologic manifestations of much trauma to the extremities is clear cut and does not require sophisticated techniques or interpretation, there are many diagnoses which require innovative techniques and a knowledge of mechanisms of injury as well as the subtle oseous and soft tissue manifestations which may be confronted radiologically. The above factors will be stressed along with the need for the fundamental knowledge of radiologic anatomy which is required in order to appreciate some of the more subtle changes of extremity trauma.

Computed tomography of the spinal canal and cord.

Selective cases have been presented to demonstrate the usefulness of computed tomography in disorders of the spine and spinal cord. The osseous structures are more readily demonstrated than the soft tissues. The cord is difficult and inconsistently identified. However, cystic lesions (syringomyelia) may, at times, be demonstrated. The transverse projection of the CT scan is an added benefit in demonstrating bony spinal lesions. The use of metrizamide with faster scanners should improve the usefulness of this technique in evaluating intraspinal lesions.