Parathyroid hormone (PTH) rapidly enhances CFTR-mediated HCO₃⁻ secretion in intestinal epithelium-like Caco-2 monolayer: a novel ion regulatory action of PTH

Besides being a Ca2-regulating hormone, parathyroid hormone (PTH) has also been shown to regulate epithelial transport of certain ions, such as Cl, HCO?, and Na, particularly in the kidney. Although the intestinal epithelium also expressed PTH receptors, little was known regarding its mechanism in the regulation of intestinal ion transport. We investigated the ion regulatory role of PTH in intestinal epithelium-like Caco-2 monolayer by Ussing chamber technique and alternating current impedance spectroscopy. It was found that Caco-2 cells rapidly responded to PTH within 1 min by increasing apical HCO?- secretion. CFTR served as the principal route for PTH-stimulated apical HCOV efflux, which was abolished by various CFTR inhibitors, namely, NPPB, glycine hydrazide-101 (GlyH-101), and CFTRinh-172, as well as by small interfering RNA against CFTR. Concurrently, the plasma membrane resistance was decreased with no changes in the plasma membrane capacitance or paracellular permeability. HCOV was probably supplied by basolateral uptake via the electrogenic Na?-HCO?? cotransporter and by methazolamide-sensitive carbonic anhydrase, while the resulting intracellular H? might be extruded by both apical and basolateral Na/H exchangers. Furthermore, the PTH-stimulated HCO?-secretion was markedly reduced by protein kinase A (PKA) inhibitor (PKI 14-22 amide) and phosphoinositide 3-kinase (PI3K) inhibitors (wortmannin and LY-294002), but not by intracellular Ca2? chelator (BAPTA-AM) or protein kinase C inhibitor (GF-109203X). In conclusion, the present study provided evidence that PTH directly and rapidly stimulated apical HCO?- secretion through CFTR in PKA- and PI3K-dependent manner, which was a novel noncalciotropic, ion regulatory action of PTH in the intestinal epithelium.     

Multi-institutional evaluation using the end-to-end test for implementation of dynamic techniques of radiation therapy in Thailand

AimIn this study, an accuracy survey of intensity-modulated radiation therapy (IMRT) and volumetric arc radiation therapy (VMAT) implementation in radiotherapy centers in Thailand was conducted.BackgroundIt is well recognized that there is a need for radiotherapy centers to evaluate the accuracy levels of their current practices, and use the related information to identify opportunities for future development.Materials and methodsAn end-to-end test using a CIRS thorax phantom was carried out at 8 participating centers. Based on each center's protocol for simulation and planning, linac-based IMRT or VMAT plans were generated following the IAEA (CRP E24017) guidelines. Point doses in the region of PTVs and OARs were obtained from 5 ionization chamber readings and the dose distribution from the radiochromic films. The global gamma indices of the measurement doses and the treatment planning system calculation doses were compared.ResultsThe large majority of the RT centers (6/8) fulfilled the dosimetric goals, with the measured and calculated doses at the specification points agreeing within ±3% for PTV and ±5% for OARS. At 2 centers, TPS underestimated the lung doses by about 6% and spinal cord doses by 8%. The mean percentage gamma pass rates for the 8 centers were 98.29 ± 0.67% (for the 3%/3 mm criterion) and 96.72 ± 0.84% (for the 2%/2 mm criterion).ConclusionsThe 8 participating RT centers achieved a satisfactory quality level of IMRT/VMAT clinical implementation.     

Control over the morphology and segregation of Zebrafish germ cell granules during embryonic development

Background

The founding member of the EMAP-like protein family is the Echinoderm Microtubule-Associated Protein (EMAP), so-named for its abundance in sea urchin, starfish, and sand dollar eggs. The EMAP-like protein family has five members in mammals (EML1 through EML5) and only one in both Drosophila (ELP-1) and C. elegans (ELP-1). Biochemical studies of sea urchin EMAP and vertebrate EMLs implicate these proteins in the regulation of microtubule stability. So far, however, the physiological function of this protein family remains unknown.

Results

We examined the expression pattern of C. elegans ELP-1 by means of transgenic gene expression in living embryos and adults, and by immunolocalization with an ELP-1-specific antibody in fixed tissues. In embryos, ELP-1 is expressed in the hypodermis. In larvae and adults, ELP-1 is expressed in the body wall, spermatheca and vulval muscles, intestine, and hypodermal seam cells. In muscle, ELP-1 is associated with adhesion complexes near the cell surface and is bound to a criss-crossing network of microtubules in the cytoplasm. ELP-1 is also expressed in a subset of mechanoreceptor neurons, including the ray neurons in the male tail, microtubule-rich touch receptor neurons, and the six ciliated IL1 neurons. This restricted localization in the nervous system implies that ELP-1 plays a role in mechanotransmission. Consistent with this idea, decreasing ELP-1 expression decreases sensitivity to gentle touch applied to the body wall.

Conclusion

These data imply that ELP-1 may play an important role during the transmission of forces and signals between the body surface and both muscle cells and touch receptor neurons.     

Prolactin stimulates transepithelial calcium transport and modulates paracellular permselectivity in Caco-2 monolayer: mediation by PKC and ROCK pathways

Prolactin (PRL) was previously demonstrated to rapidly enhance calcium absorption in rat duodenum and the intestine-like Caco-2 monolayer. However, its mechanism was not completely understood. Here, we investigated nongenomic effects of PRL on the transepithelial calcium transport and paracellular permselectivity in the Caco-2 monolayer by Ussing chamber technique. PRL increased the transcellular and paracellular calcium fluxes and paracellular calcium permeability within 60 min after exposure but decreased the transepithelial resistance of the monolayer. The effects of PRL could not be inhibited by RNA polymerase II inhibitor (5,6-dichloro-1-beta-D-ribobenzimidazole), confirming that PRL actions were nongenomic. Exposure to protein kinase C (PKC) or RhoA-associated coiled-coil forming kinase (ROCK) inhibitors (GF-109203X and Y-27632, respectively) abolished the stimulatory effect of PRL on transcellular calcium transport, whereas ROCK inhibitor, but not PKC inhibitor, diminished the PRL effect on paracellular calcium transport. Knockdown of the long isoform of PRL receptor (PRLR-L) also prevented the enhancement of calcium transport by PRL. In addition, PRL markedly increased paracellular sodium permeability and the permeability ratio of sodium to chloride, which are indicators of the paracellular charge-selective property and are known to be associated with the enhanced paracellular calcium transport. The permeability of other cations in the alkali metal series was also increased by PRL, and such increases were abolished by ROCK inhibitor. It could be concluded that PRL stimulated transepithelial calcium transport through PRLR-L and increased paracellular permeability to cations in the Caco-2 monolayer. These nongenomic actions of PRL were mediated by the PKC and ROCK signaling pathways.     

Electrogenic Na⁺/HCO₃⁻ co-transporter-1 is essential for the parathyroid hormone-stimulated intestinal HCO₃⁻ secretion

Acidosis, associated with metabolic disorders, leads to the pathological changes of cognition and behavior in the clinical practices of neurology and psychology. The cellular mechanisms underlying these cerebral dysfunctions remain unclear. By using electrophysiological approach and changing extracellular pH, we have investigated the effects of acidic environment on cortical GABAergic neurons in terms of their abilities of firing spikes and responses to synaptic inputs. Artificial cerebral spinal fluid in low pH impairs the responses to excitatory synaptic inputs and the abilities of encoding sequential spikes at these GABAergic neurons. The impairments of neuronal spiking are associated with the increases of refractory periods and threshold potentials. Our studies reveal that acidosis may impair cortical GABAergic neurons and in turn deteriorate brain functions, in which their final targets are voltage-gated sodium channels and glutamate receptor-channels.