Polarization of porous hydroxyapatite scaffolds: influence on osteoblast cell proliferation and extracellular matrix production.

Improvements to clinically used biomaterials such as hydroxyapatite (HA) are of potential benefit to the patient. One modification, the addition of surface charges, has been shown to have an important role influencing cell response. In this study, porous HA scaffolds with both positive and negative surface charges were manufactured. The samples were sintered in air to produce porous HA ceramic scaffolds in the form of cylinders 12 mm in height × 7 mm in diameter. These were polarized with a dc voltage of 3 kV/cm. MC3T3E1 cells were placed on either negative or positive ends of the charged (or unpoled control) HA scaffolds. At 7 days, picogreen analysis was performed to analyze the cell number at the negative (4 mm), central (4 mm), and positive (4 mm) portions of the 12 mm cylindrical scaffold. At 4 weeks, micro-CT analysis was performed to quantify the regional volume of mineralized matrix deposition on the 3D scaffold. At 7 days, there were significantly more cells present at the negative end of the scaffold when seeded from the negative end in comparison to the other samples tested. Micro-CT data at 4 weeks correlated with this finding, demonstrating an increase in mineralized matrix at the negatively charged end of the scaffold seeded from the negative end in comparison to the positively charged and unpoled control scaffolds. The results indicate that the charge on HA influences cell activity and that this phenomenon can be translated to a clinically relevant porous scaffold structure.

Transgenerational marking of freshwater fishes with enriched stable isotopes: a tool for fisheries management and research.

A promising new method of marking larval freshwater fishes with enriched stable isotopes by means of injecting the maternal parent with the marking agent was investigated. The (138)Ba:(137)Ba ratios in the otoliths of larval golden perch Macquaria ambigua were compared to determine the effect of injecting female broodstock with different dosages of enriched (137)Ba at various times before spawning. There was 100% mark success in the progeny of fish injected with 20 microg g(-1) of enriched (137)Ba 24 h before inducing spawning with hormones and 40 microg g(-1) administered at the same time as inducement of spawning. Injection of 40 microg g(-1) enriched (137)Ba 21 days before spawning resulted in only 81% mark success and suggests rapid elimination of barium in M. ambigua. Injection with enriched (137)Ba did not significantly affect the fertilization rate, number of fertilized eggs or hatching rate compared with long-term hatchery records. These results suggest that transgenerational marking is an effective and affordable means of mass-marking larval fishes. Thousands of larval fishes can be permanently marked with a unique artificial isotopic mark via a single injection into the maternal parent, thus avoiding the handling of individual fishes or having to deal with chemical baths. Because no single mark or tagging method is suitable for all situations, transgenerational marking with enriched stable isotopes provides another method for researchers and managers to discriminate both hatchery-reared and wild fishes.

CD19 regulation of human B cell responses. B cell proliferation and antibody secretion are inhibited or enhanced by ligation of the CD19 surface glycoprotein depending on the stimulating signal used.

The regulation of human B cell proliferation and differentiation by the CD19 surface glycoprotein was investigated. As expected, proliferation induced by costimulation with anti-IgM plus IL-4 or IL-2, or with G28.8 antibody plus IL-4 was inhibited by antibody ligation of CD19. In contrast, proliferation of tonsillar B cells to mitogenic doses of PMA (5 ng/ml) or to EBV were enhanced, and proliferation of B cell lines to BCGF(low) was unaffected. Similarly, specific antibody responses by tonsillar B cells to influenza virus, and Ig secretion by the CESS lymphoblastoid cell line in response to IL-6 were inhibited, whereas polyclonal Ig production in response to EBV was enhanced. These results show that human B cell responses may be inhibited or enhanced by CD19 depending on the stimulating signal used. The difference in response to CD19 ligation did not depend on whether proliferation or differentiation was being measured, or whether stimulation was by surface Ig. In experiments using PMA as a T cell independent mitogen, it was found that ligation of CD19 inhibited proliferation of B cells costimulated with low doses of PMA plus G28.5 (CD40) antibody, but enhanced the response to higher (mitogenic) doses with or without costimulation with G28.5. The change from inhibition to enhancement occurred over a very small increase in PMA dose (0.5-1.0 ng/ml) that corresponded exactly to the lowest dose required for mitogenic activity. Finally, we showed that CD19 ligation inhibited the increase in surface expression of CD23, but not IgM, induced by IL-4, showing that CD19 ligation can have opposed effects on different responses to the same signal. Together our results suggest that CD19 activation of human B cells interacts with other signaling events to enhance or inhibit the subsequent response.

Independent regulation of interleukin 4 (IL-4)-induced expression of human B cell surface CD23 and IgM: functional evidence for two IL-4 receptors.

Activation of human B cells with interleukin 4 (IL-4) is known to result in increased expression of CD23 (the low-affinity receptor for IgE) and sIgM. However, whereas CD23 expression is increased by several B cell mitogens, including phorbol 12-myristate 13-acetate, Epstein-Barr virus, anti-immunoglobulin (Ig), and IL-4, surface IgM (sIgM) expression is increased only with IL-4, suggesting that expression of each surface antigen is regulated independently. This was confirmed in three different ways. First, in dose-response experiments, it was shown that 10 times the concentration of IL-4 was required for CD23 than for sIgM expression. Similar or even higher concentrations of IL-4 were required for proliferation. In fact, optimal sIgM expression was obtained in some experiments with concentrations of IL-4 (1-5 units/ml) which had little or no effect on either CD23 expression or B cell proliferation. Secondly, IL-4 is known to activate the phosphatidyl inositol pathway in human B cells followed 8-10 min later by an increase in cAMP. Pharmacologically mimicking this pathway by brief exposure of resting B cells to phorbol dibutyrate plus ionomycin followed 10 min later with dibutyryl cAMP resulted in an increase in expression of CD23 but not sIgM. Thirdly, CD19 monoclonal antibody, which inhibits B cell proliferation in response to IL-4 plus anti-Ig, was found to inhibit IL-4-induced CD23 but not sIgM expression. These results show that CD23 and sIgM expression are regulated independently and are consistent with the existence of two separate signal transduction pathways stimulated by IL-4, which may be coupled to distinct IL-4 receptors.