Nkx2.5‐negative myocardium of the posterior heart field and its correlation with podoplanin expression in cells from the developing cardiac pacemaking and conduction system

Recent advances in the study of cardiac development have shown the relevance of addition of myocardium to the primary myocardial heart tube. In wild‐type mouse embryos (E9.5–15.5), we have studied the myocardium at the venous pole of the heart using immunohistochemistry and 3D reconstructions of expression patterns of MLC‐2a, Nkx2.5, and podoplanin, a novel coelomic and myocardial marker. Podoplanin‐positive coelomic epithelium was continuous with adjacent podoplanin‐ and MLC‐2a‐positive myocardium that formed a conspicuous band along the left cardinal vein extending through the base of the atrial septum to the posterior myocardium of the atrioventricular canal, the atrioventricular nodal region, and the His‐Purkinje system. Later on, podoplanin expression was also found in the myocardium surrounding the pulmonary vein. On the right side, podoplanin‐positive cells were seen along the right cardinal vein, which during development persisted in the sinoatrial node and part of the venous valves. In the MLC‐2a‐ and podoplanin‐positive myocardium, Nkx2.5 expression was absent in the sinoatrial node and the wall of the cardinal veins. There was a mosaic positivity in the wall of the common pulmonary vein and the atrioventricular conduction system as opposed to the overall Nkx2.5 expression seen in the chamber myocardium. We conclude that we have found podoplanin as a marker that links a novel Nkx2.5‐negative sinus venosus myocardial area, which we refer to as the posterior heart field, with the cardiac conduction system. Anat Rec, 290:115–122, 2007. © 2006 Wiley‐Liss, Inc.     

Macrophage-derived chemokine production by activated human T cells in vitro and in vivo: preferential association with the production of type 2 cytokines

Macrophage-derived chemokine (MDC), a potent chemoattractant for chronically activated Th2 lymphocytes, is constitutively expressed by dendritic cells, B cells, macrophages, and thymic medullary epithelial cells, whereas monocytes, NK cells, and T lymphocytes produce MDC only upon appropriate stimulation. In this study, we show in vitro MDC production also by activated T cells, which preferentially associate with the production of Th2 cytokines, IL-4, IL-5, and IL-6, and inversely correlate with the production of the Th1 cytokine, IFN-gamma. Moreover, high levels of MDC were detected in the sera of the great majority of subjects suffering from mycosis fungoides/Sézary syndrome or atopic dermatitis, which are considered as disorders characterized by the predominant expansion and activation of Th2 cells, respectively. By contrast, serum MDC levels in subjects with multiple sclerosis or Crohn's disease, which are characterized by a Th1 predominance, did not differ significantly from those of healthy controls. Finally, MDC expression was detected in the skin biopsy specimens of subjects with atopic dermatitis, where it was expressed by both dendritic cells and T lymphocytes. Taken together, these findings suggest that MDC production by activated T cells may occur both in vitro and in vivo, particularly in association with Th2 cytokines, thus providing an important amplification circuit for Th2-mediated responses.     

The pulmonary biology of isoprostanes

Isoprostanes were first recognized as convenient markers of oxidative stress, but their powerful effects on a variety of cell functions are now also being increasingly appreciated. This is particularly true of the lung, which is comprised of a wide variety of different cell types (smooth muscle, innervation, epithelium, lymphatics, etc.), all of which have been shown to respond to exogenously applied isoprostanes. In this review, we summarize these biological responses in the lung, and also consider the roles that isoprostanes might play in a range of pulmonary clinical disorders.     

Trisomy 8 in myelodysplasia and acute leukemia is constitutional in 15-20% of cases.

The trisomy 8 found in malignancies may derive from a constitutional trisomy 8 mosaicism (CT8M), and in these cases the trisomy itself may be regarded as the first mutation in a multistep carcinogenetic process. To assess the frequency of CT8M in hematological dysplastic and neoplastic disorders with trisomy 8, an informative sample of 14 patients was collected. The data ascertained included chromosome analyses of fibroblast cultures and of PHA-stimulated blood cultures in patients with normal blood differential count, as well as possible CT8M clinical signs. One patient showed trisomy 8 in all cell types analyzed and undoubtedly has a CT8M; a second patient consistently showed trisomy 8 in PHA-stimulated blood cultures when no immature myeloid cells were present in blood and should be considered as having CT8M; a third patient, with Philadelphia-positive chronic myelocytic leukemia, was more difficult to interpret, but the possibility that she had CT8M is likely. A few clinical signs of CT8M were also present in these three patients. Our data indicate that the frequency of CT8M in hematological dysplastic and neoplastic disorders with trisomy 8 is approximately 15-20%.     

Effects of selective NMDA and non-NMDA blockade in the nucleus accumbens on the plus-maze test

Effect of blocking N-methyl-D-aspartic acid (NMDA) and non-NMDA-glutamatergic receptors on performance in the plus-maze was studied in male rats bilaterally cannulated into the nucleus accumbens (Acc). Rats were divided into seven groups that received either 1 microl injections of saline, (+/-)2-amino-7-phosphonoheptanoic acid (AP-7, 0.2, 0.5, or 1 microg) or 2,3 dioxo-6-nitro-1,2,3,4,tetrahydrobenzo-(f)quinoxaline-7-sulphonamide disodium (NBQX, 0.2, 0.5, or 1 microg) 15 min before testing. Time spent in open arm, time per entry, end arrivals, open, closed, and total arm entries, relationship between open-, closed-, and total arm entries, rearing, face-, head-, and body grooming, and number of fecal boli were recorded. Time spent in the open arm increased under AP-7 (0.5 and 1 microg; P<.01) and NBQX (1 microg; P<.05) treatment, whereas time per entry was increased only with AP-7 (1 microg; P<.05). Open arm entries were increased by the intermediate doses of AP-7 (0.5 microg; P<.01) and NBQX (0.5 microg; P<.05); end arrivals were increased by the intermediate dose of AP-7 (0.5 microg/1 microl, P<.05). The frequency of rearing, grooming, and closed arm entries was not affected by the treatment. We conclude that NMDA and non-NMDA-glutamatergic blockade in the Acc lead to a behavioral disinhibition of cortical influences with the median doses, but that at higher doses the blockers have an anxiolytic-like effect.     

Identification, characterization and biological activity of oxytocin receptor in the developing human penis

Although abnormalities of the male external genitalia (MEG) are a relatively common problem, little is known concerning the molecular mechanisms that finely regulate penile development. We report here the expression of the oxytocin receptor (OTR) gene by real-time RT-PCR in human fetal tissues (11th-12th week of gestation), including the MEG. The developing penis expressed a very high level of OTR mRNA, only a half log(10) unit lower than fetal central nervous system, used as a positive control. The OTR protein is also highly expressed (western, immunohistochemistry and binding studies) and immunolocalized both in the mesenchymal body and in the surrounding blood capillaries, which will later constitute penile trabeculae and sinusoids. Binding studies using [125I]oxytocin antagonist ([125I]OTA) in cultured human fetal penile smooth muscle cells (hfPSMC) revealed the presence of specific OTR with a high capacity and affinity for oxytocin (OT) and for OTA. Increasing concentrations of OT dose-dependently induced intracellular Ca2+ mobilization. Furthermore, OTR mediated an increase in the proliferation and the migration of hfPSMC. In conclusion, we demonstrate that in the developing human MEG, OTR is highly expressed and might be involved in coordinating timely and appropriate proliferation and migration of the penile cells. Thus, OTR might represent an additional target for investigating human fetal MEG organogenesis.     

Receptors for IgM: a feature of subpopulations of both T and B human lymphocytes.

Receptors for IgM were detected on peripheral blood and tonsil human lymphocytes by a rosette technique with ox red blood cells (ORBC) coated with anti-ORBC rabbit IgM. It was found that the receptors are very sensitive to handling procedures of cells and to low temperatures. An overnight incubation period at 37 degrees C was the optimal condition for the maximum expression of receptors for IgM, but the use of IgM-free media in these cultures was neither an essential nor favourable factor for an optimal rosette formation, when ORBC heavily coated with rabbit IgM were used. The great majority of chronic lymphocytic leukaemia (CLL) patients presented a high number of EA(IgM)-RFC, either on freshly drawn or cultured lymphocytes. By fractionation procedures of normal peripheral blood and tonsil lymphocytes, it was found that a subpopulation of B cells, like T cells, also posses a lower avidity for IgM than those present on T lymphocytes.     

Evaluation of R-Mix shell vials for the diagnosis of viral respiratory tract infections.

Respiratory viruses cause significant morbidity and mortality. The management of these infections can be improved by a rapid diagnosis and administration of available virus-specific therapy. The goal of this study was to compare R-Mix, an engineered tissue monolayer for rapid shell vial (SV) diagnosis of viral respiratory infections, with conventional tissue culture (TC) and conventional respiratory SV (primary rhesus monkey kidney (RhMK) and Hep2 monolayers). The primary outcome measure was sensitivity for detection of influenza A and B, respiratory syncytial virus, parainfluenza 1-3, and adenovirus. The study was performed in two phases: (1) the three methods were compared using 250 nasal washes from children with lower respiratory tract infections; (2) a modified R-Mix SV harvesting schedule (SV were harvested at 24 and 120 h) was compared with TC and conventional RhMK/Hep2 SV using 311 respiratory specimens. A total of 110 viruses were identified in the first and 55 in the second phase. Diagnostic accuracies of R-Mix harvested at 24, 48, and 120 h were 98%, whereas for TC varied between 99 and 100%, and for RhMK/Hep2 SV between 98 and 99%. Sensitivities of R-Mix harvested at 24, 48, and 120 h were 26, 75, and 47%, respectively, whereas for TC varied between 60 and 94%, and for RhMK/Hep2 SV between 62 and 85%. R-Mix harvested at 48 h represent a valuable substitute for RhMK/Hep2 SV because they have comparable sensitivities and diagnostic accuracies, but R-Mix offers several technical advantages. In contrast, R-Mix harvested at 24h did not seem a very useful diagnostic tool. The utility of R-Mix harvested at 120 h, which accelerated the diagnosis of 16% of positive specimens in study phase 2, needs further investigation.     

Similarities and differences in the action of resiniferatoxin and capsaicin on central and peripheral endings of primary sensory neurons.

We have compared the ability of capsaicin and resiniferatoxin, a natural diterpene present in the latex of plants of the Euphorbia family to excite and desensitize capsaicin-sensitive primary afferents in a variety of models. Both capsaicin and resiniferatoxin inhibited the twitch contractions of the rat isolated vas deferens and prevented, in a concentration-related manner, the effect of a subsequent challenge with 1 microM capsaicin (desensitization). Resiniferatoxin was 1000-10,000 times more potent than capsaicin in both cases. The time course of action of resiniferatoxin was much slower than that of capsaicin, suggesting a slower penetration rate in the tissue. The action of resiniferatoxin was blocked by Ruthenium Red, a proposed antagonist at the cation channel coupled to the capsaicin receptor. Both capsaicin and resiniferatoxin produced a contraction of the rat isolated urinary bladder. Resiniferatoxin was about as potent as capsaicin in this assay although it was 500-1000 times more potent than capsaicin in desensitizing the primary afferents to a subsequent challenge with capsaicin itself. Resiniferatoxin did not affect motility in the isolated vasa deferentia or urinary bladder from capsaicin-pretreated rats. After topical application onto the rat urinary bladder both resiniferatoxin (10 nM) and capsaicin (10 microM) increased bladder capacity as assessed in a volume-evoked micturition reflex model in rats without affecting micturition contraction. Intrarterial injection of resiniferatoxin or capsaicin in the ear of anesthetized rabbits evoked a systemic depressor reflex due to activation of paravascular nociceptors, resiniferatoxin being about three times more potent than capsaicin.(ABSTRACT TRUNCATED AT 250 WORDS)