Xuhui Tong

Cx43 reverses the resistance of A549 lung adenocarcinoma cells to cisplatin by inhibiting EMT.

Cisplatin (CDDP) is one of the standard first-line chemotherapeutic agents for advanced non-small cell lung cancer (NSCLC). Unfortunately, prolonged exposure to CDDP results in acquired resistance which prevents the successful treatment of lung cancer patients. Thus, it is necessary to explore the mechanism underlying the resistance of NSCLC to CDDP. In the present study, a CDDP-resistant human lung cancer cell line A549/CDDP was established from the parental cell line A549. The results demonstrated that A549/CDDP cells acquired an epithelial-mesenchymal transition (EMT) phenotype, with morphological changes including acquisition of a spindle-like fibroblastic phenotype, downregulation of E-cadherin, upregulation of mesenchymal markers (vimentin, Snail and Slug), and increased capability of invasion and migration. Compared with A549 cells, the A549/CDDP cells showed decreased connexin43 (Cx43) expression. Overexpression of Cx43 reversed EMT and CDDP resistance in the A549/CDDP cells. Conversely, knockdown of Cx43 expression by siRNA-Cx43 initiated EMT and induced CDDP insensitivity in A549 cells. In summary, Cx43 reverses CDDP resistance in A549 CDDP-resistant cells by preventing EMT, making Cx43 a possible therapeutic target for lung cancer.

Role of gap junction intercellular communication in testicular leydig cell apoptosis induced by oxaliplatin via the mitochondrial pathway

Platinum agents are widely used in the chemotherapy of testicular cancer. However, adverse reactions and resistance to such agents have limited their application in antineoplastic treatment. The aim of the present study was to determine the role of gap junction intercellular communication (GJIC) composed of Cx43 on oxaliplatin‑induced survival/apoptosis in mouse leydig normal and cancer cells using MTT, Annexin V/PI double staining assays and western blot analysis. The results showed that GJIC exerted opposite effects on the mouse leydig cancer (I-10) and normal (TM3) cell apoptosis induced by oxaliplatin. In leydig cancer cells, survival of cells exposed to oxaliplatin was substantially reduced when gap junctions formed as compared to no gap junctions. Pharmacological inhibition of gap junctions by oleamide and 18-α-glycyrrhetinic acid resulted in enhanced survival/decreased apoptosis while enhancement of gap junctions by retinoic acid led to decreased survival/increased apoptosis. These effects occurred only in high‑density cultures (gap junction formed), while the pharmacological modulations had no effects when there was no opportunity for gap junction formation. Notably, GJIC played an opposite (protective) role in normal leydig cells survival/apoptosis following exposure to oxaliplatin. Furthermore, this converse oxaliplatin‑inducing apoptosis exerted through the functional gap junction was correlated with the mitochondrial pathway‑related protein Bcl-2/Bax and caspase‑3/9. These results suggested that in testicular leydig normal/cancer cells, GJIC plays an opposite role in oxaliplatin‑induced apoptosis via the mitochondrial pathway.

Components of Panax notoginseng saponins enhance the cytotoxicity of cisplatin via their effects on gap junctions

Previously, Panax notoginseng saponin (PNS)-induced enhancement of gap junction (GJ) formation or function was observed to be responsible for the increased cytotoxic action of cisplatin. PNS has three constituents, ginsenoside Rg1 and Rb1, and notoginsenoside R1. The active compounds in PNS responsible for enhancing the cytotoxicity of cisplatin remain unknown. Thus, the effects of the main components of PNS on the cytotoxicity of cisplatin were investigated, as well as the correlation with the modulation of GJ function in transfected HeLa cells. The cytotoxicity of cisplatin (0.25-1 µg/ml) was increased in the presence of GJs. By contrast, the cytotoxicity of cisplatin was decreased when GJs were inhibited by a GJ blocker or by the inhibition of connexin expression. Ginsenoside Rg1 (100 µM) and notoginsenoside R1 (100 µM) were observed to significantly enhance cisplatin cytotoxicity in cells with functional GJs. Ginsenoside Rb1 had no effect on the cytotoxicity of cisplatin in the presence or absence of functional GJs. Cell exposure to ginsenoside Rg1 and notoginsenoside R1 for 4 h led to significant enhancement of a dye-coupled GJ in a dose-dependent manner; however, no effect was observed in cells exposed to ginsenoside Rb1. The present results indicate that ginsenoside Rg1 and notoginsenoside R1 are the active compounds responsible for enhancing the cytotoxic action of cisplatin induced by PNS in the presence of functional GJs.

In vitro inhibited effect of gap junction composed of Cx43 in the invasion and metastasis of testicular cancer resistanced to cisplatin

The effect of gap junction intercellular communication composed of connexin on cancer invasion/metastasis has been thoroughly explored; however, its effect on testicular cancer resistanced to chemotherapy is still unclear. In this study, we found that the capability of invasion and migration of I-10/DDP (cisplatin (DDP)-resistance) cells were elevated. Furthermore, the expression of Cx43 and the function of gap junction (GJ) in I-10/DDP cells were decreased compared with parental I-10 cells. Pharmacological inhibition of GJs by oleamide (Olea) enhanced invasion and migration. However, enhancement of GJs by retinoic acid (RA) decreased invasion and migration of I-10/DDP cells. To further clarify the invasion/migration inhibited effect of GJ in the testicular cancer resistanced to DDP, GJ function was modulated by overexpression and knockdown of Cx43 expression. Overexpression of Cx43 reduced invasion and migration of I-10/DDP cells. Conversely, knockdown of Cx43 expression increased invasion and migration of I-10/DDP cells. In summary, GJ composed of Cx43 inhibits I-10/DDP cells invasion and migration, and it may become the potential therapeutic target for testicular cancer chemotherapy.

Influence of gap junction intercellular communication composed of connexin 43 on the antineoplastic effect of adriamycin in breast cancer cells

Gap junctions (GJs) serve the principal role in the antineoplastic (cytotoxicity and induced apoptosis) effect of chemical drugs. The aim of the present study was to determine the effect of GJ intercellular communication (GJIC) composed of connexin 43 (Cx43) on adriamycin cytotoxicity in breast cancer cells. Four cell lines (Hs578T, MCF-7, MDA-MB-231 and SK-BR-3) with different degree of malignancy were used in the study. The results of western blotting and immunofluorescence revealed that, in Hs578T and MCF-7 cells, which have a low degree of malignancy, the expression levels of Cx43 and GJIC were higher than those in MDA-MB-231 and SK-BR-3 cells (which have a high degree of malignancy). In Hs578T and MCF-7 cells, where GJ could be formed, the function of GJ was modulated by a pharmacological potentiators [retinoid acid (RA)]/inhibitors [oleamide and 18-α-glycyrrhetinic acid (18-α-GA)] and small interfering RNA (siRNA). In high-density cells (where GJ was formed), enhancement of GJ function by RA increased the cytotoxicity of adriamycin, while inhibition of GJ function by oleamide/18-α-GA and siRNA decreased the cytotoxicity caused by adriamycin. Notably, the modulation of GJ did not affect the survival of cells treated with adriamycin when cells were in low density (no GJ was formed). The present study illustrated the association between GJIC and the antitumor effect of adriamycin in breast cancer cells. The cytotoxicity of adriamycin on breast cancer cells was increased when the function of gap junctions was enhanced.

Role of Pannexin1 channels in the resistance of I-10 testicular cancer cells to cisplatin mediated by ATP/IP3 pathway

Cisplatin (DDP) is the most commonly used drug in testicular cancer. However, drug resistance severely limits its clinical use and the underlying mechanisms need to be further clarified. The aim of present study was to investigate the role of ATP/IP3 pathway mediated by pannexin1 (Panx-1) channels on DDP-induced apoptosis and to reveal the potential mechanisms of DDP-resistance in testicular cancer. We found that the expression of Panx-1 in I-10/DDP cells (DDP-resistance) was decreased compared with parental I-10 cells determined by western blotting and immunofluorescence assay. To further clarify the role of Panx-1 in DDP resistance, Panx-1 function was modulated by overexpression and knockdown of Panx-1 expression. Panx-1 overexpression increased DDP-induced apoptosis, ATP release and IP3 levels. On the contrary, Panx-1 silencing decreased DDP-induced apoptosis, ATP release and IP3 levels. Apyrase (hydrolyzing extracellular ATP) or xestospongin C (antagonizing IP3 receptor) also decreased DDP-induced apoptosis. Our findings demonstrate that Panx-1 is involved in DDP-resistance and ATP/IP3 pathway mediated by Panx-1 channels participates in DDP-induced apoptosis in testicular cancer. Panx-1 modulation may be interesting to amplify the clinical effect of DDP and reverse the resistance of testicular cancer cells to DDP.

Influence of Berberine on Cisplatin Antineoplastic Effect in A549 Cells

BACKGROUND AND OBJECTIVE Cisplatin is a standard first-line chemotherapeutic agents for treating advanced non-small cell lung cancer. Unfortunately, the clinical application cisplatin is restricted because it induces serious adverse reaction. The aim of this study is to investigate the influence and probable mechanism of berberine on cisplatin antineoplastic effect on lung cancer A549 cells. METHODS The total Cx43 protein amount, localization of Cx43 on cell membrane, and gap junction function were observed after the A549 cells were treated with berberine. The influence of berberine on the antitumor action of cisplatin was detected by standard colony-forming assay. Protein kinase C (PKC) protein, which regulates the gap junction, was subsequently determined. RESULTS Berberine did not affect cell survival at concentrations of 0 μM to 10 μM in the A549 cells. The gap junction function between the cells was enhanced through increased Cx43 protein expression and localization of Cx43 on the membrane after berberine treatment. The intercellular dye coupling through gap junction increased when the cells exposed to 0.1 μM, 1 μM, 10 μM berberine [33.3% (P=0.002,3), 67.0% (P<0.001), 160.0% (P<0.001)] compared withcontrols. This effect was associated with the PKC activity. The cisplatin-induced inhibition of colony growth was enhanced when berberine was combined with cisplatin. CONCLUSIONS Berberine can obviously increase the antitumor effect of cisplatin by enhancing the function of the gap junction possibly in A549 cells.
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JAK2/STAT3 pathway is involved in the protective effects of epidermal growth factor receptor activation against cerebral ischemia/reperfusion injury in rats

Cerebral ischemia and reperfusion is a common pathophysiologic process, which is involved in stroke and brain trauma. Recent studies revealed that activating epidermal growth factor receptor (EGFR) ameliorates cerebral ischemia/reperfusion (I/R) injury, however, the precise mechanisms remain to be illuminated. In this study, the neurological behavior was evaluated by Longa score. The infarct volume was performed by 2, 3, 5-triphenyltetrazolium chloride (TTC) staining and the expression of p-EGFR, p-STAT3, connexin (Cx43), Bax and Bcl-2 were detected by Western blot. The neurological behavior and infarct volume were increased in rats with cerebral I/R injury. Epidermal growth factor (EGF) pretreatment significantly decreased neurological deficit and infarct volume. However, the antagonist of EGFR, AG1478 attenuated the EGF-induced reduction of neurological deficit and infarct volume. Moreover, the inhibitor of JAK2/STAT3, AG490 undermined the protective effects stimulated by activating EGFR in rats with I/R injury. In addition, EGF pretreatment increased the expression of Bcl-2 and reduced the expression of Bax and Cx43, and the effects were abolished after using AG1478 and AG490. These findings implicate that JAK2/STAT3 pathway plays the vital role in I/R injury protection from activating EGFR. And the neuroprotective effects may associate with inhibiting the Cx43 expression and the inhibition of apoptosis.

Connexin 32 affects doxorubicin resistance in hepatocellular carcinoma cells mediated by Src/FAK signaling pathway

Doxorubicin (DOX) is first-line chemotherapy for hepatocellular carcinoma (HCC), but the effect is not satisfactory. The resistance of HCC cells to DOX is the main reason leading to treatment failure. Therefore, it is necessary to study the mechanism of DOX resistance in HCC. In this study, expression of connexin (Cx)32 was significantly decreased in HCC tissues compared with corresponding paracancerous tissues, and activity of the Src/focal adhesion kinase (FAK) signaling pathway was significantly enhanced. Expression of Cx32 was closely associated with activity of the Src/FAK signaling pathway, Cx32, and the Src/FAK signaling pathway was also correlated with degree of HCC differentiation. In DOX-resistant HepG2 cells, compared with DOX-sensitive HepG2 cells, expression of Cx32 was significantly reduced and activity of the Src/FAK pathway increased. After silencing Cx32 in HepG2 cells, activity of the Src/FAK pathway increased and sensitivity to DOX decreased. In contrast, overexpression of Cx32 in HepG2/DOX cells decreased activity of the Src/FAK pathway and increased sensitivity to DOX. Dasatinib and KX2-391, inhibitors of the Src/FAK pathway, significantly increased the sensitivity of HepG2/DOX cells to DOX. The results suggest that Src/FAK is a downstream regulator of Cx32 and Cx32 regulates the sensitivity of HCC cells to DOX via the Src/FAK signaling pathway. Our study demonstrates a potential mechanism of DOX resistance in HCC cells and supports that Cx32-Src/FAK is an important target for reversing drug resistance of HCC.