Andrenocorticotropin and β-lipotropin in the hypothalamus

Adrenocorticotropin and β-lipotropin (β-LPH) have been localized by immunoperoxidase methods in nerve cells and fibers of the hypothalamus and brain stem of the ewe. 6-μm sections were immunostained first for either ACTH or β-LPH. The reaction products and the antibody complexes were then eluted completely from the tissue, and the same section was immunostained for the second peptide. Absorption of the primary antisera with a variety of peptide fragments of ACTH and β-LPH demonstrated, immunocytochemically as well as by radioimmunoassay, that the ACTH and β-LPH antisera were directed to the COOH- and NH(2)-termini of the peptides, respectively. Neither antiserum recognized any portion of the heterologous peptide. In the sequential staining procedure on the same tissue section, preincubation of the antisera with the homologous peptide abolished the staining, whereas preincubation with the heterologous peptide did not affect it, regardless of the order followed. Every nerve cell in the arcuate nucleus that contained ACTH also contained β-LPH, but β-LPH cells appeared, probably falsely, to be twice as numerous as ACTH cells. β-LPH-positive fibers in and beyond the hypothalamus were also more numerous and stained more intensively than ACTH fibers. The salient exception was fibers in the infundibular zona externa, where the opposite was true. Our observations establish that ACTH and β-LPH are contained in the same nerve cells They stongly favor biosynthesis in brain, probably from a common precursor molecule, as has been demonstrated in the pituitary gland. The complexity of the cytologic distribution pattern described suggests that the two peptides are not processed in the same manner by the nerve cell.     

Bactericidal Immunity to Salmonella in Africans and Mechanisms Causing Its Failure in HIV Infection

Background

Nontyphoidal strains of Salmonella are a leading cause of death among HIV-infected Africans. Antibody-induced complement-mediated killing protects healthy Africans against Salmonella, but increased levels of anti-lipopolysaccharide (LPS) antibodies in some HIV-infected African adults block this killing. The objective was to understand how these high levels of anti-LPS antibodies interfere with the killing of Salmonella.

Methodology/Principal Findings

Sera and affinity-purified antibodies from African HIV-infected adults that failed to kill invasive S. Typhimurium {"type":"entrez-nucleotide","attrs":{"text":"D23580","term_id":"427513","term_text":"D23580"}}D23580 were compared to sera from HIV-uninfected and HIV-infected subjects with bactericidal activity. The failure of sera from certain HIV-infected subjects to kill Salmonella was found to be due to an inherent inhibitory effect of anti-LPS antibodies. This inhibition was concentration-dependent and strongly associated with IgA and IgG2 anti-LPS antibodies (p<0.0001 for both). IgG anti-LPS antibodies, from sera of HIV-infected individuals that inhibit killing at high concentration, induced killing when diluted. Conversely, IgG, from sera of HIV-uninfected adults that induce killing, inhibited killing when concentrated. IgM anti-LPS antibodies from all subjects also induced Salmonella killing. Finally, the inhibitory effect of high concentrations of anti-LPS antibodies is seen with IgM as well as IgG and IgA. No correlation was found between affinity or avidity, or complement deposition or consumption, and inhibition of killing.

Conclusion/Significance

IgG and IgM classes of anti-S. Typhimurium LPS antibodies from HIV-infected and HIV-uninfected individuals are bactericidal, while at very high concentrations, anti-LPS antibodies of all classes inhibit in vitro killing of Salmonella. This could be due to a variety of mechanisms relating to the poor ability of IgA and IgG2 to activate complement, and deposition of complement at sites where it cannot insert in the bacterial membrane. Vaccine trials are required to understand the significance of lack of in vitro killing by anti-LPS antibodies from a minority of HIV-infected individuals with impaired immune homeostasis.     

Type 1 plasminogen activator inhibitor binds to fibrin via vitronectin

Type 1 plasminogen activator inhibitor (PAI-1), the primary inhibitor of tissue-type plasminogen activator (t-PA), circulates as a complex with the abundant plasma glycoprotein, vitronectin. This interaction stabilizes the inhibitor in its active conformation In this report, the effects of vitronectin on the interactions of PAI-1 with fibrin clots were studied. Confocal microscopic imaging of platelet-poor plasma clots reveals that essentially all fibrin-associated PAI-1 colocalizes with fibrin-bound vitronectin. Moreover, formation of platelet-poor plasma clots in the presence of polyclonal antibodies specific for vitronectin attenuated the inhibitory effects of PAI-1 on t-PA-mediated fibrinolysis. Addition of vitronectin during clot formation markedly potentiates PAI-1-mediated inhibition of lysis of (125)I-labeled fibrin clots by t-PA. This effect is dependent on direct binding interactions of vitronectin with fibrin. There is no significant effect of fibrin-associated vitronectin on fibrinolysis in the absence of PAI-1. The binding of PAI-1 to fibrin clots formed in the absence of vitronectin was characterized by a low affinity (K(d) approximately 3.5 micrometer) and rapid loss of PAI-1 inhibitory activity over time. In contrast, a high affinity and stabilization of PAI-1 activity characterized the cooperative binding of PAI-1 to fibrin formed in the presence of vitronectin. These findings indicate that plasma PAI-1.vitronectin complexes can be localized to the surface of fibrin clots; by this localization, they may modulate fibrinolysis and clot reorganization.     

Mutation spectra in Salmonella TA98, TA100, and TA104 of two phenylbenzotriazole mutagens (PBTA-1 and PBTA-2) detected in the Nishitakase River in Kyoto, Japan.

Previous studies have identified two potent aromatic amine mutagens in the Nishitakase River, a tributary of the Yodo River, which serves as the main drinking water supply for the Osaka area in Japan. The two potent mutagens are 2-[2-(acetylamino)-4-[bis(2-methoxyethyl)amino]-5-methoxyphenyl]-5-am ino-7-bromo-4-chloro-2H-benzotriazole (PBTA-1) and 2-[2-(acetylamino)-4-[N-(2-cyanoethyl)ethylamino]-5-methoxyphenyl]-5- amino-7-bromo-4-chloro-2H-benzotriazole (PBTA-2). PBTA-1 and PBTA-2 are presumed to be formed from azo dyes discharged in a reduced form from dye factories to sewage treatment plants where they become chlorinated and are then discharged into the river. PBTA-1 and PBTA-2 account for 21% and 17% of the mutagenic activity of the Nishitakase River, respectively. Here we determined the mutation spectra induced by these two mutagens in TA98, TA100, and TA104 at 30-35, 8-10, and 2x, respectively, above the background. In TA98, the PBTA compounds produced identical mutation spectra, with 100% of the revertants containing the hotspot 2-base deletion of CG within the (CG)(4) sequence. In TA100, 73% of the revertants were GC-->TA transversions, with most of the remaining being GC-->AT transitions; the spectra produced by the two compounds in TA100 were not significantly different (p=0.8). In TA104, as in TA100, the majority (83%-87%) of the revertants were GC-->TA transversions, with most of the remaining revertants (11%-13%) being AT-->TA transversions. Thus, 83%-87% of the mutations induced by the PBTA compounds in TA104 were at G/C sites. The mutation spectra produced by the two compounds in TA104 were not significantly different (p0.08). PBTA-1 and PBTA-2 are structurally similar and have similar mutagenic potencies and mutation spectra in the respective strains. The mutation spectra produced by the PBTA compounds (100% hotspot deletion in TA98 and primarily GC-->TA transversions in TA100 and TA104) are similar to those produced by other potent aromatic amines, which is the class of compounds from which the PBTA mutagens derive.     

Cranial ontogeny of otariid seals

Over 2000 otariid skulls were measured for a morphometric study of cranial ontogeny in fur seals and sea lions. Few interspecific differences in cranial ontogeny were observed in the Otariidae, with only minor differences in rates of growth. Sexual dimorphism was significant in all otariids but was more apparent in the larger species. Female otariids of each species showed monophasic development in all characteristics, whereas males expressed monophasic growth for some characters and biphasic growth for others. Biphasic development in skulls of male otariids occurred well after physical maturity had been reached, usually at a suture index of 27. The rate of development varied between skull characters; components relating to the nervous system completed growth well before the rest of the skull, whereas those related to feeding, respiration and vocalisation developed in synchrony with the overall growth of total skull length. Sutures of the calvaria, or braincase, were the first to show partial closure and those uniting the facial bones were usually the last to fuse. As with the calvaria, the orbits and otic capsules developed quickly, suggesting a need for good hearing and vision early in independent postnatal life. Development of the rostral and palatal regions required significantly longer to complete growth.     

Consequences of daily administered parathyroid hormone on myeloma growth, bone disease, and molecular profiling of whole myelomatous bone

Background

Induction of osteolytic bone lesions in multiple myeloma is caused by an uncoupling of osteoclastic bone resorption and osteoblastic bone formation. Current management of myeloma bone disease is limited to the use of antiresorptive agents such as bisphosphonates.

Methodology/Principal Findings

We tested the effects of daily administered parathyroid hormone (PTH) on bone disease and myeloma growth, and we investigated molecular mechanisms by analyzing gene expression profiles of unique myeloma cell lines and primary myeloma cells engrafted in SCID-rab and SCID-hu mouse models. PTH resulted in increased bone mineral density of myelomatous bones and reduced tumor burden, which reflected the dependence of primary myeloma cells on the bone marrow microenvironment. Treatment with PTH also increased bone mineral density of uninvolved murine bones in myelomatous hosts and bone mineral density of implanted human bones in nonmyelomatous hosts. In myelomatous bone, PTH markedly increased the number of osteoblasts and bone-formation parameters, and the number of osteoclasts was unaffected or moderately reduced. Pretreatment with PTH before injecting myeloma cells increased bone mineral density of the implanted bone and delayed tumor progression. Human global gene expression profiling of myelomatous bones from SCID-hu mice treated with PTH or saline revealed activation of multiple distinct pathways involved in bone formation and coupling; involvement of Wnt signaling was prominent. Treatment with PTH also downregulated markers typically expressed by osteoclasts and myeloma cells, and altered expression of genes that control oxidative stress and inflammation. PTH receptors were not expressed by myeloma cells, and PTH had no effect on myeloma cell growth in vitro.

Conclusions/Significance

We conclude that PTH-induced bone formation in myelomatous bones is mediated by activation of multiple signaling pathways involved in osteoblastogenesis and attenuated bone resorption and myeloma growth; mechanisms involve increased osteoblast production of anti-myeloma factors and minimized myeloma induction of inflammatory conditions.