Activation of the Transcription Factor FosB/Activating Protein-1 (AP-1) Is a Prominent Downstream Signal of the Extracellular Nucleotide Receptor P2RX7 in Monocytic and Osteoblastic Cells

Activation of the ionotropic P2RX7 nucleotide receptor by extracellular ATP has been implicated in modulating inflammatory disease progression. Continuous exposure of P2RX7 to ligand can result in apoptosis in many cell types, including monocytic cells, whereas transient activation of P2RX7 is linked to inflammatory mediator production and the promotion of cell growth. Given the rapid hydrolysis of ATP in the circulation and interstitial space, transient activation of P2RX7 appears critically important for its action, yet its effects on gene expression are unclear. The present study demonstrates that short-term stimulation of human and mouse monocytic cells as well as mouse osteoblasts with P2RX7 agonists substantially induces the expression of several activating protein-1 (AP-1) members, particularly FosB. The potent activation of FosB after P2RX7 stimulation is especially noteworthy considering that little is known concerning the role of FosB in immunological regulation. Interestingly, the magnitude of FosB activation induced by P2RX7 stimulation appears greater than that observed with other known inducers of FosB expression. In addition, we have identified a previously unrecognized role for FosB in osteoblasts with respect to nucleotide-induced expression of cyclooxygenase-2 (COX-2), which is the rate-limiting enzyme in prostaglandin biosynthesis from arachidonic acid and is critical for osteoblastic differentiation and immune behavior. The present studies are the first to link P2RX7 action to FosB/AP-1 regulation in multiple cell types, including a role in nucleotide-induced COX-2 expression, and support a role for FosB in the control of immune and osteogenic function by P2RX7.     

The Biology of Rose Rosette Disease: A Mite-associated Disease of Uncertain Aetiology

Spread of rose rosette disease was best described by a logistic population growth model. In Iowa, peak susceptibility to graft transmission of the disease in the field occurred from May 27 thrrough July 10. Populations of the eriophyid mite vector were low until mid-July and reached peak levels at the end of August. Host range was limited to multiflora rose and hybrid ornamental roses. No evidence was obtained for seed-or soil-transmission of the aetiologic agent and no symptom remission was obtained with either heat therapy or tetracycline treatment. A significant decrease in stored starch corresponded with a major decrease in sucrose and increases in fructose and glucose in infected tissue. Because of its limited host range, the agent may have potential for biocontrol of multiflora rose.     

SOME CHARACTERISTICS OF THE SPERM ATTRACTANT FROM OEDOGONIUM CARDIACUM1

A bioassay for the sperm attractant from Oedogonium cardiacum was developed. The attractant was confirmed to be highly water soluble, destroyed by heating to 100 C and by dilute acid or alkali. It was fractionated by gel filtration with the attractant accompanied by yellow pigments. It has it molecular weight between 500 and about 1500, based on gel filtration, ultrafiltration, and dialysis. Freezing changes this to both larger and smaller molecules. It appears to have cationic groups. A paper chromatographic separation was found that divided the pigment into at least 3 major bands, one of which was closely associated with the attractant. An absorption spectrum of this pigment showed 1 peak in the visible spectrum at 417 mμ and 3 well-defined peaks in the ultraviolet region. The lack of a peak at 280 mμ indicates the attractant is not a polypeptide.     

Bone alkaline phosphatase in Paget's disease.

The measurement of bone proteins and peptides and their derived products has been very useful in the diagnosis and management of patients with skeletal disease. This group of assays includes alkaline phosphatase (AP) and bone Gla protein (BGP). We here describe the comparison of a new immunoassay that is specific for bone alkaline phosphatase (BAP) to measurements of total alkaline phosphatase (TAP) and BGP in Paget's disease. In our studies, we demonstrated that BAP was increased in the serum of patients with Paget's disease. Comparisons with the other measurements revealed that BAP correlated better with total AP (r = 0.92) than with BGP (r = 0.51); the lowest correlation occurred between BGP and total AP (r = 0.26). In patients with liver disease, the BAP was indistinguishable from normal whereas the TAP was elevated. These studies indicate that BAP assesses different aspects of bone cell function than BGP in Paget's disease. This discordancy also exists between BGP and total serum AP activity. BAP measurements by immunoassay offer a novel method of assessing skeletal status. Thus, the information that measurement of different bone-specific proteins provides should be separately useful in assessing the skeleton for a variety of metabolic bone diseases.     

Aromatic Amino Acid Activation of Signaling Pathways in Bone Marrow Mesenchymal Stem Cells Depends on Oxygen Tension

The physiologic oxygen pressures inside the bone marrow environment are much lower than what is present in the peripheral circulation, ranging from 1–7%, compared to values as high as 10–13% in the arteries, lungs and liver. Thus, experiments done with bone marrow mesenchymal stem cells (BMMSCs) using standard culture conditions may not accurately reflect the true hypoxic bone marrow microenvironment. However, since aging is associated with an increased generation of reactive oxygen species, experiments done under 21%O₂ conditions may actually more closely resemble that of the aging bone marrow environment. Aromatic amino acids are known to be natural anti-oxidants. We have previously reported that aromatic amino acids are potent agonists for stimulating increases in intracellular calcium and phospho-c-Raf and in promoting BMMSC differentiation down the osteogenic pathway. Our previous experiments were performed under normoxic conditions. Thus, we next decided to compare a normoxic (21% O₂) vs. a hypoxic environment (3% O₂) alone or after treatment with aromatic amino acids. Reverse-phase protein arrays showed that 3% O₂ itself up-regulated proliferative pathways. Aromatic amino acids had no additional effect on signaling pathways under these conditions. However, under 21%O₂ conditions, aromatic amino acids could now significantly increase these proliferative pathways over this “normoxic” baseline. Pharmacologic studies are consistent with the aromatic amino acids activating the extracellular calcium-sensing receptor. The effects of aromatic amino acids on BMMSC function in the 21% O2 environment is consistent with a potential role for these amino acids in an aging environment as functional anti oxidants.