Establishment and characterization of a new,spontaneously immortalized,pancreatic ductal cell line from the Syrian golden hamster

Spontaneously immortal pancreatic cell lines are not available. By use of a defined culture medium, such a line (TAKA-1) was established from the Syrian golden hamster. Cytological, cytogenetic, molecular biological, enzymatic and receptor patterns as well as antigenicity were studied and were compared with those of the normal hamster pancreatic ductal cells in vivo. TAKA-1 cells grew exponentially in a monolayer on collagen gel in a defined medium but did not proliferate in soft agar. Ultrastructurally, the cells closely resembled the normal hamster pancreatic ductal cells. Similarities and dissimilarities were found between the normal ductal cells and TAKA-1 cells. Similarities included the presence of cytokeratin, carbonic anhydrase and some tumor-associated antigens. However, unlike the normal ductal cells, TAKA-1 cells expressed blood group A angigen and anti-vimentin, showed affinity to selected lectins, and an abnormality of chromosome 3, which is suggested to be associated with immortality. Moreover, unlike the hamster pancreatic ductal cancer cells but like the normal hamster pancreatic ductal cells, TAKA-1 cells did not have a c-Ki-ras mutation. EGF, TGF- and secretin, but not CCK or GRP, bound to the TAKA-1 cells. TAKA-1 cells produced TGF-, and their growth was stimulated by exogenous EGF in serum-free medium. This cell line presents a suitable model for biologic and pathologic study of the hamster pancreatic ductal cells in vitro.     

RAB2A: a major subacrosomal protein of bovine spermatozoa implicated in acrosomal biogenesis

The perinuclear theca (PT) of mammalian sperm is a unique subcellular structure encapsulating the nucleus. Compositionally, the PT is made up of at least six prominent polypeptides (60, 36, 31, 28, 24, and 15 kDa), of which only two have been sequence identified, as well as many less prominent ones. As an ongoing process in unveiling the protein composition of the PT, we have uncovered the sequence identity of the prominent 24-kDa polypeptide (PT24). Initial N-terminal sequence analysis obtained by Edman degradation suggested that PT24 is a RAB2 protein. This was corroborated by mass spectrometric analyses of trypsin-digested fragments of PT24, identifying RAB2A of the RAB2 subfamily as the best sequence match. Quadrapole/time-of-flight analysis identified 72%% sequence coverage between PT24 and bull, human, mouse, or rabbit RAB2A. Since a genome search only identified two RAB2 subfamily members, RAB2A and RAB2B, the 72%% sequence coverage of PT24 provides assurance that this protein is RAB2A and not a new RAB2 subfamily member. Furthermore, commercial RAB2A antibodies, raised against oligopeptide fragments in the unique C-terminal region of RAB2A, specifically labeled PT24 on Western blot analysis of PT extracts. These anti-RAB2A antibodies, along with immune serum that we raised and affinity purified against isolated PT24, demonstrated at both light and electron microscope levels that RAB2 is associated with the periphery of the growing proacrosomic and acrosomic vesicles in the Golgi and cap phases of spermiogenesis and consequently assembled as part of the PT. This pattern of subacrosomal assembly is reminiscent of the pathway used by SubH2Bv (PT15), another prominent and exclusive subacrosomal protein, indicating a common route for subacrosomal-PT assembly. Traditionally somatic RAB2 proteins are involved in vesicular transport between the endoplasmic reticulum and the cis-side of the Golgi apparatus. Our study suggests an unprecedented direction of RAB2A-mediated vesicular transport in spermatids during acrosomal biogenesis, from the trans-side of the Golgi apparatus to the nuclear envelope.     

Connective tissue activation. XXXIII. Biologically active cleavage products of CTAP-III from human platelets

Evidence for three new isoforms of CTAP-III from human platelets is presented; two NH2-terminal cleavage products were identified, CTAP-III (des 1-13) and CTAP-III (des 1-15). CTAP-III (des 1-13) has a pI of 8.6 and is a relatively stable proteolytic cleavage product that retains the capacity to stimulate [14C]GAG synthesis in human synovial cell cultures. CTAP-III (des 1-15) appears to be an elastase or chymotrypsin cleavage product and identical to NAP-2, an entity thought to have neutrophil activating properties.     

Weight loss effects of quaternary salts of 5-amino-1-(chloromethyl)-1,2-dihydro-3H-benz[e]indoles; structure-activity relationships

Quaternary salt analogues based on the DNA minor groove binder and adenine N3 alkylating agent 5-amino-1-(chloromethyl)-1,2-dihydro-3H-benz[e]indole (aminoCBI) show remarkable effects on the body weight of mice (a long-term failure to gain weight relative to matched controls with no loss of appetite or perceptible deterioration in health) following administration of a single (non-toxic) dose between about 0.5-5 μmol/kg. The nature of the quaternizing group was not important, but a related hydroxyCBI analogue was much less effective. Compounds where the chloro group was replaced by a hydrogen or hydroxy group (thus abrogating DNA alkylating capability) showed no weight control activity. It is speculated, based on other studies, that the marked long-term weight control effect is due to inhibition of bile flow into the intestine and reduced absorption of triglycerides, together with accelerated cell death in spleen and white adipose tissues due to drug accumulation there. This class of compound may serve as interesting tools for further study of these phenomena.     

Neurochemical characteristics of early and late onset types of Alzheimer's disease.

Brains of 49 patients who had died with Alzheimer''s disease and 54 controls were examined. The Alzheimer group exhibited noticeably reduced activity of the cholinergic marker enzyme choline acetyltransferase in the cerebral cortex, but cortical concentrations of noradrenaline, gamma-aminobutyric acid, and somatostatin were also significantly reduced. Analysis of the results according to age at death showed that the older patients, dying in their 9th and 10th decades, had a relatively pure cholinergic deficit confined to temporal lobe and hippocampus, together with a reduced concentration of somatostatin confined to temporal cortex. By contrast, the younger patients, dying in their 7th and 8th decades, had a widespread and severe cholinergic deficit together with the abnormalities of noradrenaline, gamma-aminobutyric acid, and somatostatin, and the younger patients accounted for most of the abnormalities in these systems observed in the overall group. Comparison of the young subjects with Alzheimer''s disease with the older controls did not support the concept of Alzheimer''s disease representing an acceleration of the aging process. These results suggest that Alzheimer''s disease in people aged under 80 may represent a distinct form of presenile dementia which differs in important respects from the dementia of old age.     

Amino Acid Composition, Immunoreactivity, Sequence Analysis, and Function of Bovine Hippocampal Metallothionein Isoforms

Abstract: The high concentration of zinc in the hippocampal mossy fiber axon boutons is localized in the vesicles and is mobilized by exocytosis of the zinc-laden vesicles. Because "free" zinc in excess is a neurotoxic substance inhibiting an extensive number of sulfhydryl-containing enzymes and receptor sites, we hypothesized that low-molecular-weight zinc binding proteins must exist in the hippocampus to regulate the steady-state concentration of zinc. In this communication, we report that the bovine hippocampus synthesizes metallothionein (MT) isoforms that are similar, but not identical, to those of the rat brain MT isoforms and cross-react poorly with antibodies formed against the hepatic MT isoforms, suggesting that the immunologically dominant regions of hippocampal MT (residues 1–29) are not conserved. A comparative sequence analysis of bovine hippocampal MTs and bovine hepatic MT isoforms I and II revealed a 90% sequence identity, being mostly different in residues 1–29. The results of these studies suggest that the hippocampal MT isoforms, which are synthesized on a continuous basis, may play a role in regulating the transport, accumulation, and compartmentation of zinc in the hippocampus.     

Serotonin reciprocally regulates melanocortin neurons to modulate food intake

The neural pathways through which central serotonergic systems regulate food intake and body weight remain to be fully elucidated. We report that serotonin, via action at serotonin1B receptors (5-HT1BRs), modulates the endogenous release of both agonists and antagonists of the melanocortin receptors, which are a core component of the central circuitry controlling body weight homeostasis. We also show that serotonin-induced hypophagia requires downstream activation of melanocortin 4, but not melanocortin 3, receptors. These results identify a primary mechanism underlying the serotonergic regulation of energy balance and provide an example of a centrally derived signal that reciprocally regulates melanocortin receptor agonists and antagonists in a similar manner to peripheral adiposity signals.     

In the eye of the beholder: visual mate choice lateralization in a polymorphic songbird

Birds choose mates on the basis of colour, song and body size, but little is known about the mechanisms underlying these mating decisions. Reports that zebra finches prefer to view mates with the right eye during courtship, and that immediate early gene expression associated with courtship behaviour is lateralized in their left hemisphere suggest that visual mate choice itself may be lateralized. To test this hypothesis, we used the Gouldian finch, a polymorphic species in which individuals exhibit strong, adaptive visual preferences for mates of their own head colour. Black males were tested in a mate-choice apparatus under three eye conditions: left-monocular, right-monocular and binocular. We found that black male preference for black females is so strongly lateralized in the right-eye/left-hemisphere system that if the right eye is unavailable, males are unable to respond preferentially, not only to males and females of the same morph, but also to the strikingly dissimilar female morphs. Courtship singing is consistent with these lateralized mate preferences; more black males sing to black females when using their right eye than when using their left. Beauty, therefore, is in the right eye of the beholder for these songbirds, providing, to our knowledge, the first demonstration of visual mate choice lateralization.