Investigation of IL-8Rbeta mRNA expression profile in single human neutrophil / 中国应用生理学杂志

<p><b>AIM</b>To validate the abundance of Interleukin 8 receptor beta (IL-8Rbeta) mRNA in single human neutrophil.</p><p><b>METHODS</b>Human neutrophils were isolated and purified from volunteers, total RNA was extracted and a regular RT-PCR aiming at IL-8Rbeta mRNA was performed to ascertain its expression profile in human neutrophils and optimize the reaction conditions for the following single-cell RT-PCR procedures. Subsequently, single neutrophil or the cellular content was harvested to conduct reverse transcription and two-round PCR with the same primer pairs used before. Serial dilution of single neutrophil cDNA pool was carried out at the same time with the exact two-round PCR followed. The specificity of this single-cell RT-PCR procedure was verified by the BamHI restriction endonuclease digestion on the final cDNA products.</p><p><b>RESULTS</b>Regular RT-PCR indicated IL-8Rbeta mRNA expression in human neutrophils. While single-cell RT-PCR was sensitive enough to detect trace IL-8Rbeta mRNA as predicted cDNA product could be amplified from a 10 000 times diluted intracellular specimen from single neutrophil, which indicated an abundant expression of this mRNA in human neutrophil. Moreover, BamHI digestion on the final cDNA product clarified the specificity of this single-cell RT-PCR procedure.</p><p><b>CONCLUSION</b>This simplified semi-quantitative single-cell RT-PCR procedure specifically confirmed that IL-8Rbeta mRNA was highly expressed in human neutrophil, which also provided the possibility of comparing mRNA abundance at single cell level.</p>

Electrophysiological study on rat conduit pulmonary artery smooth muscle cells under normoxia and acute hypoxia / 生理学报

The present study was designed to investigate the electrophysiological characteristics of rat conduit pulmonary artery smooth muscle cells (PASMCs) and the response to acute hypoxia. PASMCs of the 1st to 2nd order branches in the conduit pulmonary arteries were obtained by enzymatic isolation. The PASMCs were divided into acute hypoxia preconditioned group and normoxia group. Hypoxia solutions were achieved by bubbling with 5% CO2 plus 95% N2 for at least 30 min before cell perfusion. Potassium currents were compared between these two groups using whole-cell patch clamp technique. The total outward current of PASMCs was measured under normoxia condition when iBTX [specific blocking agent of large conductance Ca-activated K(+) (BK(Ca)) channel] and 4-AP [specific blocking agent of delayed rectifier K(+) (K(DR)) channel] were added consequently into bath solution. PASMCs were classified into three types according to their size, shape and electrophysiological characteristics. Type I cells are the smallest with spindle shape, smooth surface and discrete perinuclear bulge. Type II cells show the biggest size with banana-like appearance. Type III cells have the similar size with type I, and present intermediary shape between type I and type II. iBTX had little effect on the total outward current in type I cells, while 4-AP almost completely blocked it. Most of the total outward current in type II cells was inhibited by iBTX, and the remaining was sensitive to 4-AP. In type III cells, the total outward current was sensitive to both iBTX and 4-AP. Acute hypoxia reduced the current in all three types of cells: (1614.8+/-62.5) pA to (892.4+/-33.6) pA for type I cells (P<0.01); (438.3+/-42.8) pA to (277.5+/-44.7) pA for type II cells (P<0.01); (1 042.0+/-37.2) pA to (613.6+/-23.8) pA for type III (P<0.01), and raised the resting membrane potentials (E(m)) in all these three types of cells: (-41.6+/-1.6) mV to (-18.6+/-1.5) mV (P<0.01), (-42.3+/-3.8) mV to (-30.6+/-3.0) mV (P<0.01), (-43.3+/-1.6) mV to (-28.4+/-1.4) mV (P<0.01), for type I, II, III cells, respectively. These results suggest that acute hypoxia suppresses the potassium current and improves the E(m) in PASMCs. These effects may be involved in the modulation of constriction/relaxation of conduit artery under acute hypoxia. Different distribution of K(DR) and BK(Ca) channels in these three types of PASMCs might account for their different constriction/relaxation response to acute hypoxia.

Effects of Yizhi Capsule on learning and memory disorder and beta-amyloid peptide induced neurotoxicity in rats / 中国结合医学杂志

<p><b>OBJECTIVE</b>To explore the effects of Yizhi Capsule (YZC) on learning and memory disorder and beta-amyloid peptide induced neurotoxicity in rats.</p><p><b>METHODS</b>Various doses of YZC were administered to Sprague-Dawley (SD) rats for 8 consecutive days, twice a day. On the 8th day of the experiment, scopolamine hydrobromide was intraperitoneally injected to every rat and Morris water maze test and shuttle dark avoidance test were carried out respectively to explore the changes of learning and memory capacities in the rats. Besides, after the cerebral cortical neurons of newborn SD rats aged within 3 days were cultured in vitro for 7 days, drug serum containing YZC was added to the cultured neurons before or after beta amyloid peptide(25 - 35) (Abeta(25 - 35)) intoxication to observe the protective effect of YZC on neurotoxicity by MTT assay and to determine the LDH content in the supernatant.</p><p><b>RESULTS</b>Compared with those untreated with YZC, the rats having received YZC treatment got superiority in shorter time of platform seeking in Morris water maze test, as well as elongated latent period and less times of error in shuttle dark avoidance test. On the cultured neurons, YZC drug serum could effectively increase the survival rate of Abeta(25 - 35) intoxicated neurons and reduce the LDH contents in cultured supernatant.</p><p><b>CONCLUSION</b>YZC has an action of improving learning and memory disorder, and good protective effect on Abeta(25 - 35) induced neurotoxicity in SD rats.</p>