[Changing of TSG101 expression on the invasion and metastasis of gastric cancer cells].

OBJECTIVE: To explore the functions of tumor susceptibility gene 101 (TSG101) in the invasion and metastasis of gastric cancer cells by cell culture. METHODS: The TSG101 eukaryotic expression and empty plasmids were transfected into gastric cancer cell line SGC7901. After screening with G418, single cell clone was selected and cultured. The expression of TSG101 was detected by reverse transcription (RT)-PCR and Western blotting. Cells were divided into TSG101 eukaryotic expression plasmid and blank control groups. Then the relationship was examined between TSG101 expression and tumor invasion and metastasis through the invasion, mobile, adhesion and damage scar experiment. RESULTS: The expression levels of TSG101 in mRNA and protein in the TSG101 eukaryotic expression group were significantly higher than those of the plasmid and blank control groups (0.85 ± 0.09 vs 0.55 ± 0.07, 0.45 ± 0.07 and 29.4 ± 1.2 vs 17.0 ± 0.4, 15.9 ± 0.4, all P < 0.05). The cell number of TSG101 eukaryotic expression group through Matrigel, laminin, type IV collagen protein (84 ± 14, 128 ± 10, 62 ± 7) were significantly higher than those of the plasmid group (55 ± 9, 77 ± 10, 31 ± 6) and blank control group (48 ± 8, 76 ± 9, 24 ± 5, all P < 0.01). The number of cells adherent to Matrigel, laminin, type IV collagen protein of the TSG101 eukaryotic expression group (0.97 ± 0.04, 1.34 ± 0.04, 0.90 ± 0.01) were obviously higher than those of the plasmid group (0.53 ± 0.03, 0.75 ± 0.05, 0.42 ± 0.02) and blank control group (0.60 ± 0.03, 0.72 ± 0.03, 0.40 ± 0.01, all P < 0.01). The number of TSG101 eukaryotic expression group cell migrating to membrane lower surface was obviously higher than that of the plasmid group and blank control group (87 ± 13 vs 54 ± 8, 48 ± 7, all P < 0.01). The fusion speed of the TSG101 eukaryotic expression group was faster than that of plasmid and blank control groups after cultivating for 24 and 48 h. CONCLUSIONS: TSG101 expression increases significantly in SGC-7901 cells after a stable transfection of TSG101 eukaryotic expression plasmids. Also the capacities of invasion and metastasis become markedly enhanced.

[Effect of cisplatin on expression of matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1 and their correlations in Lewis lung cancer in mice].

BACKGROUND AND OBJECTIVE: Metastasis of lung cancer is the leading cause of disease progression and treatment failure. Matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1) are related to the metastasis of lung cancer via regulating the degradation of extracellular matrix. This study was to observe the impacts of cisplatin (DDP) on the expression of MMP-9 and TIMP-1 in Lewis lung cancer, and explore their correlations and roles in metastasis. METHODS: Lewis lung cancer model was established in C57BL/6 mice. DDP group was given intraperitoneal DDP injection, and compared with normal control and tumor-bearing groups. The expression of MMP-9 and TIMP-1 were determined by ELISA in serum and detected by immunohistochemistry in tumor tissues. RESULTS: The inhibition rates of tumor growth and metastasis were 41.2% and 39.0% in DDP group, respectively. The positive rates of MMP-9 and TIMP-1 were 100% in tumor-bearing group, and their serum concentrations were significantly higher in tumor-bearing group than in normal control group (P<0.05). Serum concentrations of MMP-9 and TIMP-1, and positive rate of MMP-9 were all significantly lower in DDP group than in tumor-bearing group (P<0.05). Serum concentration of MMP-9 and positive rates of MMP-9 and TIMP-1 were positively correlated to tumor weight (r=0.665, 0.749 and 0.615, all P<0.05) and lung metastasis (r=0.668, 0.545 and 0.664, all P<0.05). MMP-9 expression was positively correlated to TIMP-1 expression both in serum and tumor (r=0.617 and 0.695, all P<0.05). The ratio of sMMP-9/TIMP-1 became a constant in normal distribution, with a mean of 1.72. CONCLUSIONS: Both MMP-9 and TIMP-1 are highly expressed in Lewis lung cancer, correlated to tumor invasion and metastasis. DDP may suppress tumor metastasis via down-regulating the expression of MMP-9 and TIMP-1 in serum and tumor.

Neurosteroid dehydroepiandrosterone sulfate enhances spontaneous glutamate release in rat prelimbic cortex through activation of dopamine D1 and sigma-1 receptor.

This paper studied the effect of neurosteroid dehydroepiandrosterone sulfate on spontaneous glutamate release in the prelimbic cortex by using electrophysiological and biochemical methods combined with a pharmacological approach and made some comparisons with those in the hippocampus. The results showed that dehydroepiandrosterone sulfate increased the frequency of miniature excitatory postsynaptic currents in the prelimbic cortex and hippocampus; sigma-1 receptor antagonist partially blocked the effect of dehydroepiandrosterone sulfate in the prelimbic cortex, but completely blocked it in the hippocampus; D1 receptor antagonist, adenylyl cyclase inhibitor and protein kinase A inhibitor completely blocked the effect of dehydroepiandrosterone sulfate in the prelimbic cortex; dehydroepiandrosterone sulfate increased the activity of protein kinase A in the prelimbic cortex and hippocampus; the effect of dehydroepiandrosterone sulfate on protein kinase A was completely blocked by sigma-1 receptor antagonist in the hippocampus, but was partially blocked in the prelimbic cortex; interestingly, here again, the effect of dehydroepiandrosterone sulfate on protein kinase A was completely blocked by D1 receptor antagonist in the prelimbic cortex. These results suggest that dehydroepiandrosterone sulfate promotes presynaptic glutamate release in the prelimbic cortex via activation of D1 and sigma-1 receptors.

Neurosteroid pregnenolone sulfate inhibits stimulus-evoked EPSC via presynaptic inhibition of protein kinase A in rat prelimbic cortical neurons.

Pregnenolone sulfate is one of the most abundantly produced neurosteroids in the brain. The present paper studies the effect of pregnenolone sulfate on excitatory synaptic transmission in the pyramidal cells of the layer V-VI of the prelimbic cortex using whole-cell patch-clamp in slices. We found that pregnenolone sulfate inhibited stimulus-evoked excitatory postsynaptic currents (EPSC); the effect of pregnenolone sulfate was significant at concentration of 1 microM and increased with an increase in concentrations; pregnenolone sulfate had no effect on the amplitude and frequency of miniature excitatory spontaneous postsynaptic currents; pregnenolone sulfate significantly enhanced the paired-pulse facilitation (PPF) and inhibited dopamine and 5-HT-evoked increase in the frequency of spontaneous excitatory postsynaptic currents; the protein kinase A inhibitor H89 and Rp-cAMPS enhanced PPF and canceled the effect of pregnenolone sulfate on PPF; pregnenolone sulfate inhibited the protein kinase A agonist forskolin-evoked increase in the frequency of spontaneous excitatory postsynaptic currents; the G(i) protein inhibitor N-ethylmaleimide canceled the effect of pregnenolone sulfate on PPF. These results suggest that pregnenolone sulfate inhibits stimulus-evoked EPSC via presynaptic inhibition of protein kinase A in rat prelimbic cortical neurons.

Progesterone inhibition of dopamine-induced increase in frequency of spontaneous excitatory postsynaptic currents in rat prelimbic cortical neurons.

We examined the effects of progesterone on frequency of miniature excitatory postsynaptic currents (mEPSCs) and spontaneous excitatory postsynaptic currents (sEPSCs), and dopamine-induced increase in the frequency of sEPSCs in pyramidal cells of layers V-VI of the rat prelimbic cortex using whole-cell patch-clamp techniques in slices. The results showed that progesterone 100 microM had no effects on the frequency of mEPSCs and sEPSCs, but significantly inhibited dopamine-induced increase in frequency of sEPSCs. This was in contrast to the effect of progesterone on the effect of 5-HT, which showed no changes after progesterone. When studying the mechanism of the progesterone effect, we observed that GABA(A) receptor antagonist and progesterone receptor antagonist did not influence the effect of progesterone; progesterone had no effects on D1 receptor agonist, protein kinase A and protein kinase C activator-induced increase in the frequency of sEPSCs. Interestingly, sigma(1) receptor antagonist could inhibit the effect of dopamine and sigma(1) receptor agonist had a synergistic effect on the effect of D1 receptor agonist. These results suggest that progesterone may inhibit dopamine-induced increase in frequency of sEPSCs in rat prelimbic cortical neurons via inhibition of sigma(1)/D1 receptor synergism because progesterone has been known to be an antagonist of sigma(1) receptor.