Evaluation of BBG2Na in infant macaques: specific immune responses after vaccination and RSV challenge

We have addressed the safety of alum-adsorbed BBG2Na, a recombinant respiratory syncytial virus (RSV) subunit vaccine, in infant macaques. Animals received two vaccinations, and were challenged 4 months later with RSV. In two of four BBG2Na-vaccinated animals, specific IL-13 producing T cells were detected. Upon challenge, low level pulmonary eosinophilia was observed in the same two animals. Although the levels of these responses were substantially lower than those observed in the FI-RSV controls, these data suggest that more extensive studies focusing on immunopathological safety of alum-adsorbed BBG2Na in non-human primates would be required before proceeding to clinical trials in seronegative infants.     

Concordance between women's physiological and subjective sexual arousal is associated with consistency of orgasm during intercourse but not other sexual behavior

Many studies have found a discordance between women's genital (vaginal pulse amplitude) and subjective sexual arousal responses to erotica. We hypothesized that the association between the physiological and subjective domains would be greater for women with greater orgasmic consistency during penile-vaginal intercourse but not for orgasm consistency during other sexual behaviors. We confirmed this specific hypothesis in a sample (N = 38) of postmenopausal women. In addition, we discovered that the correlation between the domains was unrelated to social desirability responding, that orgasm consistency was not less for intercourse than for other sexual activity, and that orgasm consistency during intercourse was uncorrelated with orgasm consistency during masturbation. We discuss the results in terms of the unique nature of penile-vaginal intercourse, our study's implications for sex therapy, and orgasm consistency during intercourse being an operational measure of functional vaginal sensitivity and sexual pleasure integration and organization.     

Influence of a Microemulsion Vehicle on Cutaneous Bioequivalence of a Lipophilic Model Drug Assessed by Microdialysis and Pharmacodynamics

Purpose. The aim of the study was to investigate the cutaneous bioequivalence of a lipophilic model drug (lidocaine) applied in a novel topical microemulsion vehicle, compared to a conventional oil–in–water (O/W) emulsion, assessed by a pharmacokinetics microdialysis model and a pharmacodynamic method. Methods. Dermal delivery of lidocaine was estimated by microdialysis in 8 volunteers. Absorption coefficients and lag times were determined by pharmacokinetic modelling of the microdialysis data. Subsequently, the anaesthetic effect of the treatments was assessed by mechanical stimuli using von Frey hairs in 12 volunteers. Results. The microemulsion formulation increased the cutaneous absorption coefficient of lidocaine 2.9 times (95% confidence interval: 1.9/4.6) compared with the O/W emulsion–based cream. Also, lag time decreased from 110 ± 43 min to 87 ± 32 min (P = 0.02). The compartmental pharmacokinetic model provided an excellent fit of the concentration–time curves with reliable estimation of absorption coefficient and lag time. A significant anaesthetic effect was found for both active treatments compared to placebo (P < 0.02), but the effect did not diverge significantly between the two formulations. Conclusions. The microemulsion vehicle can be applied to increase dermal drug delivery of lipophilic drugs in humans. The microdialysis technique combined with an appropriate pharmacokinetic model provides a high sensitivity in bioequivalence studies of topically applied substances.     

Transformation of an established mouse mammary epithelial cell line following transfection with a human transforming growth factor alpha cDNA

To determine whether the enhanced expression of transforming growth factor alpha (TGF alpha) is sufficient to induce the neoplastic transformation of an immortalized population of mammary epithelial cells, we cotransfected NOG-8 cells, a cloned mouse mammary epithelial cell line, with a simian virus 40-human TGF alpha cDNA expression vector plasmid and a pSV2neo plasmid. After cotransfection, nine G418-resistant NOG-8 colonies were cloned and expanded. All clones were subsequently analyzed for TGF alpha mRNA expression by northern blot analysis, TGF alpha secretion, anchorage-dependent growth in serum-free medium, anchorage-independent growth in soft agar, and tumorigenicity in nude mice. Three TGF alpha-transfected NOG-8 clones expressed high levels of a specific TGF alpha mRNA, secreted elevated levels of TGF alpha into the culture medium (177-595 ng/10(8) cells/48 h), exhibited an enhanced growth rate, grew aggressively as colonies in soft agar, and formed undifferentiated, invasive carcinomas in nude mice. A neutralizing mouse monoclonal antibody generated against the low molecular weight human TGF alpha peptide was able to inhibit colony formation in soft agar by TGF alpha-transfected NOG-8 clones that produced high levels by TGF alpha. This inhibition suggested that TGF alpha acted through an external autocrine loop. NOG-8 cells and NOG-8 cells transfected with a pSV2neo plasmid alone secreted very low levels of TGF alpha, failed to grow as colonies in soft agar and did not form tumors in nude mice. These results demonstrate that overexpression of a human TGF alpha cDNA in immortalized, nontransformed mouse mammary epithelial cells can induce a transformed phenotype in vitro and can facilitate tumor formation in vivo.     

Engineered CaM2 modulates nuclear calcium oscillation and enhances legume root nodule symbiosis

The key physiological event essential to the establishment of nitrogen-fixing bacteria and phosphate-delivering arbuscular mycorrhizal symbioses is the induction of nuclear calcium oscillations that are required for endosymbioses. These regular fluctuations in nucleoplasmic calcium concentrations are generated by ion channels and a pump located at the nuclear envelope, including the CYCLIC NUCLEOTIDE GATED CHANNEL 15 (CNGC15). However, how the CNGC15s are regulated in planta to sustain a calcium oscillatory mechanism remains unknown. Here, we demonstrate that the CNGC15s are regulated by the calcium-bound form of the calmodulin 2 (holo-CaM2), which, upon release of calcium, provides negative feedback to close the CNGC15s. Combining structural and evolutionary analyses of CaM residues with bioinformatic analysis, we engineered a holo-CaM2 with an increased affinity for CNGC15s. In planta, the expression of the engineered holo-CaM2 accelerates the calcium oscillation frequency, early endosymbioses signaling and is sufficient to sustain over time an enhanced root nodule symbiosis but not an increased arbuscular mycorrhization. Together, these results reveal that holo-CaM2 is a component of endosymbiosis signaling required to modulate CNGC15s activity and the downstream root nodule symbiosis pathway.

Nutrient acquisition is fundamental to life. Plants have evolved strategies to overcome soil phosphate limitation and gain access to atmospheric dinitrogen by developing beneficial associations with arbuscular mycorrhizal (AM) fungi and nitrogen-fixing bacteria, respectively. Unlike other crops, the vast majority of legumes have mastered associations with both endosymbionts, positioning them as key crops to develop sustainable agricultural practices in both developed and developing countries (1).The entry of nitrogen-fixing bacteria, known as rhizobia, and AM fungi into legume roots is initiated by the recognition of the endosymbiont. Host plants have plasma-membrane receptor-like kinases (2–6) that recognize rhizobial elicitors, lipochitooligosaccharides (LCOs), also known as Nod factors (7), and mycorrhizal factors composed of derivatives of LCOs and shorter chain chitooligosaccharides (8, 9). Although rhizobial and AM elicitors are recognized by different complexes of receptor-like kinases (10, 11), both symbionts require the activation of calcium oscillations in root epidermal nuclei (9, 12, 13) to set off the endosymbiosis program. In the model legume Medicago truncatula, two types of nuclear envelope localized ion channels are required to generate the calcium oscillation; the DOESN’T MAKE INFECTIONS1 (DMI1) channel and paralogs of CYCLIC NUCLEOTIDE GATED CHANNEL 15 (CNGC15) (14), and the calcium pump, MCA8 (15). Similar to the animal CNGCs, plant CNGCs are tetrameric ion channels that can include different CNGC units (16, 17). In M. truncatula, CNGC15a, CNGC15b, and CNGC15c are all involved in nuclear calcium oscillation in the root epidermis, nodulation, and arbuscular mycorrhization, suggesting that the three units could assemble into a heterocomplex at the nuclear envelope (14). However, how CNGC15s are regulated in planta to sustain a calcium oscillatory mechanism remains unknown.In this study, we demonstrate that CNGC15s are regulated by the calcium-bound form of the calmodulin 2 (holo-CaM2) in planta, which shapes the oscillatory pattern of nucleoplasmic calcium concentration by providing negative feedback on CNGC15s to cause its closure. By engineering CaM2 to generate CaM2^R91A, which specifically increased holo-CaM2 binding affinity to each CNGC15 unit, we accelerated closure of CNGC15s and increased the calcium oscillation frequency. We further show that accelerating the calcium oscillation frequency was sufficient to accelerate the early endosymbiosis signaling and that the expression of CaM2^R91A resulted in an enhanced root nodule symbiosis but not enhanced AM colonization. Our data reveal differential regulation of rhizobia and AM endosymbioses by CaM2^R91A and suggest that modulating calcium signaling can be used as a strategy to positively impact symbiosis with nitrogen-fixing bacteria.     

Actions of growth factors on plasma calcium. Epidermal growth factor and human transforming growth factor-alpha cause elevation of plasma calcium in mice.

Specific humoral substances produced and secreted by human tumors that cause hypercalcemia have not been identified. Certain growth factors (such as epidermal growth factor, platelet-derived growth factor, and transforming growth factors-alpha and -beta) have been shown to stimulate the resorption of bone in organ culture by both prostaglandin-dependent and prostaglandin-independent pathways. In this report we demonstrate that epidermal growth factor and recombinant human transforming growth factor-alpha induce a significant rise in plasma calcium concentration when administered repeatedly to intact mice for periods ranging from 24 h to 16 d. The elevation of plasma calcium is not dependent on dietary calcium and is not invariably accompanied by an increase in systemic levels of the prostaglandin E2 metabolite 13,14-dihydro-15-keto-prostaglandin E2. The in vivo calcium-mobilizing activity of epidermal growth factor and transforming growth factor-alpha indicate that these or related growth factors need be considered as potential mediators of tumor-induced hypercalcemia.     

Subdivision of the sural nerve: step towards individual facial reanimation

Long term facial paralysis is a serious affliction and upsetting for the patient. Dynamic facial reanimation has become the treatment of choice. Various techniques that use different donor muscles have been developed since the first functional muscle transplant for facial paralysis more than 30 years ago. The concept of using a single muscle was refined into the use of dividable muscle slips such as serratus muscle or separate muscular subunits to avoid the resulting mass movements. Because the results are still not satisfactory, efforts were put into also dividing the donor nerve transplant into corresponding subunits to create a continuous line of individual action. Twenty human cadaveric sural nerves were successfully dissected into three completely separate subunits, transecting the interfascicular bridges. This anatomical study gives the potential to allow an independent triple innervation of three separate serratus anterior muscle slips, so decreasing further the mass movement after facial reanimation.     

Effects of a nonpeptide motilin receptor antagonist on proximal gastric motor function

AIM: To assess the effects of the motilin receptor antagonist RWJ-68023 on basal and motilin-stimulated proximal gastric volume. METHODS: Eighteen healthy male volunteers received RWJ-68023 in two different doses or placebo for 135 min. After 45 min, subjects received a motilin infusion for 90 min. Proximal gastric volume was measured with a barostat at constant pressure and during isobaric distensions. Abdominal symptoms were scored using visual analogue scales. Motilin and RWJ-68023 concentrations were assessed by radioimmunoassay and liquid chromatography-mass spectrometry, respectively. RESULTS: Both dosages of RWJ-68023 were safe and well tolerated. The most common adverse events were of gastrointestinal origin. RWJ-68023 did not affect basal proximal gastric volume, but the high-dose RWJ-68023 reduced the contractile effect of motilin on the stomach. This antagonizing effect of RWJ-68023 was only significant (P = 0.014) during the distension procedure. CONCLUSIONS: The RWJ-68023 doses used in this study were selected to accomplish plasma concentrations that would block the motilin effect entirely. However, the antagonizing effect of RWJ-68023 was partial and only present when the tonic condition of the stomach was modulated by motilin.