The effect of miR-224 down-regulation on SW80 cell proliferation and apoptosis and weakening of ADM drug resistance.

Since this article has been suspected of research misconduct and the corresponding authors did not respond to our request to prove originality of data and figures, "The effect of miR-224 down-regulation on SW80 cell proliferation and apoptosis and weakening of ADM drug resistance, by C.-Q. Liang, Y.-M. Fu, Z.-Y. Liu, B.-R. Xing, Y. Jin, J.-L. Huang, published in Eur Rev Med Pharmacol Sci 2017; 21 (21): 5008-5016-PMID: 29164556" has been withdrawn. The Publisher apologizes for any inconvenience this may cause. https://www.europeanreview.org/article/13747.

The effect of miR-224 down-regulation on SW80 cell proliferation and apoptosis and weakening of ADM drug resistance.

OBJECTIVE: Glycogen synthase kinase-3ß (GSK-3ß) can phosphorylate and degrade ß-catenin, and negatively regulates Wnt/ß-catenin signal pathway. MiR-224 up-regulation is associated with colorectal cancer (CRC). Bioinformatics analysis showed complementary binding sites between miR-224 and GSK-3ß. This study investigated if miR-224 plays a role in mediating GSK-3ß expression, Wnt/ß-catenin pathway activity, CRC cell proliferation, apoptosis as well as drug sensitivity of Adriamycin (ADM). MATERIALS AND METHODS: Dual luciferase gene reporter assay demonstrated the regulatory relationship between miR-224 and GSK-3ß. Expression of miR-224, GSK-3ß, ß-catenin, and Survivin was measured in normal colon epithelium NCM460, CRC cell line SW480, and drug-resistant SW480/ADM cell line. Flow cytometry measured apoptosis under ADM with an IC50 concentration of SW480 cells, followed by CCK-8 analysis of cell proliferation. SW480/ADM cells were treated with miR-224 inhibitor and/or pSicoR-GSK-3ß, followed by analysis of the expressions of GSK-3ß, ß-catenin and Survivin, cell apoptosis, and cell proliferation by EdU staining. RESULTS: MiR-224 targeted and inhibited GSK-3ß expression. In SW480/ADM cells, GSK-3ß expression and cell apoptosis rate were lower than those in SW480 cells, whilst miR-224, ß-catenin, and Survivin expression or proliferation were higher than those in SW480 cells. Transfection of miR-224 inhibitor and/or pSicoR-GSK-3ß significantly increased GSK-3ß expression in SW480/ADM cells, and decreased ß-catenin and Survivin expression, leading to reduced proliferation potency, enhanced cell apoptosis and suppressed ADM resistance. CONCLUSIONS: MiR-224 up-regulation is associated with ADM resistance of CRC cells. Suppression of miR-224 expression up-regulated GSK-3ß expression, inhibited Wnt/ß-catenin signal pathway activity and Survivin expression, as well as reduced ADM resistance of CRC SW480 cells.

Effect of calcium on the uptake of glutamate by synaptosomes: possible involvement of two different mechanisms.

The effects of Ca2+ on glutamate uptake by synaptosomes were investigated. Glutamate uptake was decreased in the absence of Ca2+ with 2 mM EGTA added to the medium. After a 8-min preincubation, the uptake reduced to the greatest extent (about 64% of the uptake measured in Ca2+ medium). According to the kinetic analysis, lack of Ca2+ resulted in a reduction of the Vmax for glutamate uptake. These results suggested external Ca2+ was necessary for the optimal uptake of glutamate. TTX, an inhibitor of the voltage-dependent Na+ channel, almost reversed the reduction of glutamate uptake, indicating that the uptake might be impaired via the partial activation of the voltage-dependent tetrodotoxin-sensitive Na+ channel. On the other hand, glutamate uptake was decreased by trifluoperazine, a calmodulin antagonist, and KN-62, an inhibitor of Ca2+/calmodulin-dependent protein kinase II, implying that intrasynaptosomal Ca2+ might also have a role in the uptake of glutamate, and Ca2+/calmodulin-dependent processes, such as Ca2+/calmodulin-dependent protein kinase II, might be involved in the modulation of glutamate transporter activity.

[Rapid effects of hydrocortisone on the cholinergic synaptic transmission of B neurons in bullfrog sympathetic ganglia].

Rapid effects of hydrocortisone 21-hemisuccinate (F-suc) on cholinergic synaptic transmission of B neurons in bullfrog sympathetic ganglia (BFSG) were studied with intracellular recording technique in vitro. The main results are as follows: (1) F-suc blocked cholinergic synaptic transmission of 52 B neurons within 0.5-3 min, which was concentration-dependent and partially suppressed by the cytoplasm receptor antagonist of steroid hormones, RU38486; (2) after protein synthesis was suppressed by actidione, the above effect persisted; and (3) the effect of F-suc could be potentiated by atropine. All these results suggest that the F-suc induced rapid effects of cholinergic synaptic transmission of B neurons in BFSG may be mediated by non-genomic mechanisms.

[Rapid effect of glucocorticoid on excitatory postsynaptic potential of B cells in bullfrog paravertebral ganglia].

In the present study, the rapid effect of hydrocortisone 21-hemisuccinate (F-suc) on the amplitude of f-EPSP of B cells in isolated bullfrog sympathetic ganglia was examined with the use of intracellular recording technique. It was found that the amplitude of f-EPSP of B cells was decreased by F-suc, with a latency less than 3 min after the beginning of perfusion of F-suc. The inhibition of F-suc was concentration-dependent and could be blocked by the antagonist of intracellular glucocorticoid receptor RU38486, but not by the protein synthesis inhibitor actidione. These findings suggest that the inhibition of f-EPSP of B cells by F-suc is probably mediated through a nongenomic mechanism.

[Ionic mechanism of hyperpolarization induced by glucocorticoid in mammalian neurons].

The rapid membrane actions of glucocorticoid were investigated by intracellular electrical recording from 383 coeliac ganglion neurons of guinea-pig in vitro. Thirty-eight neurons were hyperpolarizaed by 2-12 mV when perfused with 1 mumol/L hydrocortisone 21-hemisuccinate (F-suc), associated with a decrease in input membrane resistance. The hyperpolarization was dose-dependent. Nine neurons were depolarized, and the other 336 neurons were unresponsive. The membrane current was also observed with discontinuous single-electrode voltage clamp technique under perfusion of F-suc in another 43 neurons. In five neurons the current was found outward, but it was inward in one neuron. The hyperpolarization persisted after the elimination of synaptic input by low Ca2+ high Mg2+ perfusion and the suppression of protein synthesis by antinomycin D. The reversal potential of F-suc hyperpolarization is -79 +/- 4.3 mV (n = 5). F-suc induced hyperpolarization and GABA induced depolarization could occur in same neuron. The later action could be blocked by picrotoxin. F-suc induced hyperpolarization could be inhibited by TEA and 4-AP, but not picrotoxin. It is suggested that the F-suc's hyperpoalrization is mediated by potassium channel rather than Cl- channel in the sympathetic ganglion neurons.

[Fast modulation of glutamate and GABA receptor--mediate electrophysiological responses by glucocorticoid].

Fast modulation by glucocorticoid on gluatmate and GABA receptor-mediated electrophysiological responses was investigated by extracellular iontophoresis and intracellular techniques in rat hypothalamic brain slices and guinea pig coeliac ganglion preparations respectively. The results were as follows: Glucocorticoid inhibited the glutamate receptor-mediated excitatory response and potentiated the GABA receptor-mediated inhibitory response within 5 min after perfusion. All these responses are reversible after removal of the glucocorticoid. Low Ca2+ high Mg2+ medium was ineffective to block the fast modulatory effects of glucocorticoid on the gluatmate and GABA receptor-mediated responses. It appears that the latter responses are mediated through some non-genomic mechanism without an involvement of synaptic circuitry.

[The pharmacology of the GABA-induced biphasic response in guinea-pig celiac ganglion neurons in vitro].

In guinea-pig celiac ganglion neurons, GABA induced a biphasic response, i.e. a depolarizing response associated with a resistance decrease followed by a hyperpolarizing response associated with a resistance increase. Such response persisted in low Ca2+ high Mg2+ medium. The GABA-induced biphasic response was mimicked by muscimol and inhibited by bicuculline methiodide (100 mumol/L) and picrotoxin (100-300 mumol/L). At doses producing almost equal depolarizing responses, muscimol was weaker in producing hyperpolarizing responses than GABA. Baclofen had no effect on membrane potentials of the ganglion cells. These results suggest that the depolarizing phase of the GABA-induced biphasic response is mediated by GABAA/Cl- ionophore receptor complex and the hyperpolarizing phase may be mediated by a novel type of GABA receptor possessing similar pharmacological properties as the classical GABAA receptor.

An electrophysiological study on the membrane receptor-mediated action of glucocorticoids in mammalian neurons.

The action of glucocorticoids (GC) on neuronal cell membrane was studied in isolated and superfused guinea pig coeliac ganglia by the intracellular recording technique. Cortisol succinate (F) hyperpolarized the membrane potential of 47 of 179 cells and changed the cell's input resistance with a latency of less than 2 min in vitro. The effect persisted under low Ca2+/high Mg2+ superfusing condition and could be blocked by RU 38486, a competitive antagonist of GC cytosolic receptor. Cortisol-21-bovine albumin conjugant exhibited the same effect. Corticosterone (B) elicited hyperpolarization in another 15 of 83 cells, but dexamethasone (Dex) did not. Dex, however, depolarized 2 of 18 cells. Aldosterone, cholesterol and vehicle (ethyl alcohol) caused no detectable change in membrane potential. In vivo studies by iontophoretic application of steroids to hypothalamic paraventricular (PVN) neurons showed that F inhibited the unit discharges in 68 of 97 PVN neurons, and the effect could be antagonized by RU 38486. Dex excited 30 of 100 neurons. Estradiol (E) also inhibited the discharges, but this inhibition was not antagonized by RU 38486. The effect of GC on PVN neurons was also examined in hypothalamic slices including the paraventricular nucleus. B inhibited 28 of 104 units and excited 7 of 104 cells, and both effects could be antagonized by RU 38486. The threshold of inhibitory response was about 10(-7) M, which is close to the physiological level of the hormone in plasma. The results suggest that GC can act non-genomically and specifically through its membrane receptor on the neuronal surface, and that there might be a chemical similarity between the membrane receptor and the traditional cytosolic GC receptor.(ABSTRACT TRUNCATED AT 250 WORDS)

[Effects of corticosterone on spontaneous discharges of rat hypothalamic paraventricular neurons in vitro].

Experiments were performed to investigate the effects of corticosterone on electrical activities of paraventricular neurons in rat hypothalamic slices. In unstimulated conditions, most of the neurons had either a slow irregular (48.1%) or fast continuous pattern (42.5%) of discharge. Only a small portion (9.4%) of the cells shows phasic activity. During perfusion of the slices with corticosterone (10(-7), 10(-6) mol/L), 25 out of 104 paraventricular neurons were profoundly inhibited with a short latency, 8 neurons were excited and the remaining cells showed no response. The response, the magnitude of the cells which depended upon the concentration of corticosterone, could be reversed by concomitant application of RU38486 (10(-7), 10(-6) mol/L), a competitive antagonist of glucocorticoid's cytosolic receptor. These results provide further evidence that corticosterone has rapid effects on rat hypothalamic paraventricular neurons, and suggests the existence of membrane receptors for glucocorticoid in paraventricular neurons.

[Effects of noradrenaline and acetylcholine on electric activity of paraventricular neurons of rat hypothalamic slices].

The effects of noradrenaline (NA) and acetylcholine (ACh) on 90 paraventricular neurons from 31 hypothalamic slices prepared from rat were observed. After perfusing the brain slices with NA (10(-6) mol/L, 3 min), discharge rate of 14/73 (17%) nonphasic and 7/12 (58.3%) phasic neurons was significantly increased, while that of 10/73 (12%) nonphasic and 2/12 (16.6%) phasic neurons was significantly decreased, even ceased. 50/73 (64%) of the nonphasic and 3/12 (25%) of the phasic neurons were non-responsive. The effects of NA on the nonphasic neurons were completely blocked by alpha receptor blocker phentolamine. The effects of NA on phasic neurons were partly blocked by phentolamine or propranolol (beta receptor blocker). After perfusing hypothalamic slices with ACh (10(-7) mol/L, 3 min), the firing rate of 15/73 (19%) nonphasic and 6/12 (50%) phasic neurons was increased, while that of 9/73 (11%) nonphasic and 2/12 (16.6%) phasic neurons was decreased. 54/73 (69%) nonphasic and 4/12 (33.3%) phasic neurons were non-responsive. The effect of ACh on the neurons could be reversed by atropine. The effect of scopolamine (M blocker) is similar to that of atropine. The experimental results suggest that the excitatory or inhibitory effect of NA or ACh on paraventricular neurons has mediated respectively by alpha, beta or M receptors.

[Effects of serotonin on the electric activity of paraventricular neurons of in vitro rat hypothalamic slices].

Hypothalamic slices were prepared with a vibratome. The spontaneous discharge of 46 paraventricular neurons was recorded by glass microelectrode from 20 such slices, and effects of serotonin on spontaneous discharge were observed. After perfusing the brain slices with artificial cerebro-spinal fluid containing serotonin (10(-6) mol/L) for 3 min, discharge rate of 16 units is significantly increased with latency 1.21 +/- 1.21 (mean + SD) min, this response is blocked by serotonin antagonist, cyproheptadine. The discharges of 3 units were decreased, but 25 units non-responsive. The experimental results suggest that about 1/3 of paraventricular neurons can be activated by topical application of serotonin via the activation of 5-HT receptors.

[A simple vibratome for brain slice].

A simple vibratome was fabricated using double-function electric shaver and microscopic platform. Spontaneous discharge of neurons in hippocampal and hypothalamic brain slices (in 300-400 microns thick) prepared by the vibratome could kept above 12 hours in artificial cerebro-spinal fluid.