Gap junctions mediate direct communication between cells; however, toxicological cascade triggered by nonessential metals can abrogate cellular signaling mediated by gap junctions. Although cadmium (Cd) is known to induce apoptosis in organs and tissues, the mechanisms that underlie gap junction activity in Cd-induced apoptosis in BRL 3A rat liver cells has yet to be established. In this study, we showed that Cd treatment decreased the cell index (a measure of cellular electrical impedance) in BRL 3A cells. Mechanistically, we found that Cd exposure decreased expression of connexin 43 (Cx43), increased expression of p-Cx43 and elevated intracellular free Ca2+ concentration, corresponding to a decrease in gap junctional intercellular communication. Gap junction blockage pretreatment with 18β-glycyrrhizic acid (GA) promoted Cd-induced apoptosis, involving changes in expression of Bax, Bcl-2, caspase-3 and the mitochondrial transmembrane electrical potential (Δψm). Additionally, GA was found to enhance ERK and p38 activation during Cd-induced activation of mitogen-activated protein kinases, but had no significant effect on JNK activation. Our results indicated the apoptosis-related proteins and the ERK and p38 signaling pathways may participate in gap junction blockage promoting Cd-induced apoptosis in BRL 3A cells.
Animals ; Apoptosis/*drug effects ; Cadmium/*toxicity ; Calcium/metabolism ; Cell Communication/drug effects ; Connexin 43/genetics ; Enzyme Activation/drug effects ; Gap Junctions/*drug effects ; Gene Expression Regulation/drug effects ; Hepatocytes/cytology/*drug effects ; Rats ; Signal Transduction/drug effects

OBJECTIVETo investigate the effects of the quinoline derivative PQ1 combined with cisplatin on the proliferation and gap junction communication of prostate cancer PC3 cells.

METHODSWe cultured in vitro prostate cancer PC3 cells and divided them into DMSO blank control, cisplatin control, and cisplatin (10 mg/ml) plus PQ1 (1, 2, 5, 10, and 15 μmol/L) groups. We measured the proliferation of the prostate cancer PC3 cells, determined the expressions of the connexin 43 (Cx43) mRNA and protein by RT-PCR and Western blot, and compared the indexes among different groups.

RESULTSCisplatin combined with PQl at 1 - 10 μmol/L significantly inhibited the proliferation of the PC3 cells and the inhibition rate rose in a concentration- and time-dependent manner, from (48.72 ± 0.98)% vs (50.33 ± 0.62)% at 0 μmol/L to (77.38 ± 1.12)% vs (83.50 ± 1.05)% at 15 μmol/L at 24 and 48 hours (P < 0.05). Compared with the cisplatin control, cisplatin combined with PQ1 at 1, 2, 5, 10, and 15 μmol/L increased the expression of Cx43 mRNA from 0.379 ± 0.113 to 0.669 ± 0.031, 0.831 ± 0. 127, 0.769 ± 0.100, 0.532 ± 0.086, and 0.475 ± 0.134, respectively (P < 0.05), and cisplatin combined with PQ1 at 1, 2, 5, and 10 μmol/L elevated that of Cx43 protein from 0.138 ± 0.146 to 0.263 ± 0.111, 0.306 ± 0.152, 0.415 ± 0.280, and 0.643 ± 0.310, respectively (P < 0.05).

CONCLUSIONThe quinoline derivative PQ1 can promote the gap junction communication of prostate cancer PC3 cells and enhance the killing effect of cisplatin on PC3 cells by upregulating the expressions of Cx43 mRNA and protein.

Aminoquinolines ; pharmacology ; Antineoplastic Combined Chemotherapy Protocols ; pharmacology ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Cisplatin ; pharmacology ; Connexin 43 ; genetics ; metabolism ; Dose-Response Relationship, Drug ; Gap Junctions ; drug effects ; physiology ; Humans ; Male ; Prostatic Neoplasms ; metabolism ; pathology ; physiopathology ; RNA, Messenger ; metabolism ; Time Factors

OBJECTIVETo investigate the effect of baicalein on the gap junction intercellular communication (GJIC) in the TM4 Sertoli cells of the mouse testis and its related mechanism.

METHODSWe measured the cytotoxicity of different concentrations of baicalein on the TM4 Sertoli cells in the mouse testis by MTT, detected the fluorescence transfer of the TM4 Sertoli cells by parachute assay, and determined the expression of the protein connexin 43 ( Cx43) in the baicalein-treated cells by Western blot and immunofluorescence assay.

RESULTSBaicalein produced no obvious cytotoxicity on the TM4 Sertoli cells at the concentration below 60 µmol/L but significantly increased their GJIC at 0-20 µmol/L (P < 0.01). Western blot and immunofluorescence assay showed that 0-20 µmol/L baicalein remarkably elevated the expression of Cx43 in the TM4 cells (P < 0.01) and on the membrane of the TM4 cells.

CONCLUSIONBaicalein at the concentration of 0-20 µmol/L can significantly enhance GJIC in mouse TM4 Sertoli cells by increasing the expression of the Cx43 protein.

Animals ; Cell Communication ; drug effects ; Connexin 43 ; metabolism ; Flavanones ; administration & dosage ; pharmacology ; Gap Junctions ; drug effects ; Male ; Mice ; Sertoli Cells ; drug effects ; metabolism ; ultrastructure

OBJECTIVETo investigate the effect of sodium valproate, a histone deacetylase inhibitor, on the cytotoxicity of doxorubicin in breast cancer cells.

METHODSWestern blotting was used to assess Cx43 protein expression in breast cancer Hs578T cells exposed to doxorubicin and sodium valproate. MTT assay was used to determine the cytotoxicity of doxorubicin; annexin V/PI double staining and Hochest 33258 fluorescence staining were employed to detect doxorubicin-induced early and late apoptosis, respectively.

RESULTSWestern blotting showed that sodium valproate significantly increased Cx43 protein expression in Hs578T cells (P/0.01). The cells exposed to both sodium valproate and doxorubicin showed significantly lowered cell viability compared with the cells exposed to doxorubicin alone (P/0.01). Exposure to both sodium valproate and doxorubicin resulted in significantly increased early and late cell apoptosis rate compared with doxorubicin treatment alone (P/0.01).

CONCLUSIONsodium valproate can significantly enhance the cytotoxicity of doxorubicin and increase doxorubicin-induced apoptosis in breast cancer cells in vitro possibly by enhancing the gap junction function.

Apoptosis ; drug effects ; Breast Neoplasms ; pathology ; Cell Line, Tumor ; drug effects ; Cell Survival ; drug effects ; Connexin 43 ; metabolism ; Doxorubicin ; pharmacology ; Drug Synergism ; Gap Junctions ; Histone Deacetylase Inhibitors ; pharmacology ; Humans ; Valproic Acid ; pharmacology

The aim of the present study is to investigate the effect of 18β-glycyrrhetinic acid (18β-GA) on KCl- and PE-induced constriction of rat renal interlobar artery (RIA). Pressure myograph system was used to observe the constriction induced by KCl and PE (endothelial independent vasoconstrictor) in acutely separated RIA of Wistar rats with or without 18β-GA pretreatment. Whole-cell patch clamp recordings were used to observe the effect of 18β-GA on membrane input capacitance (C(input)), membrane input conductance (G(input)) or membrane input resistance (R(input)) of smooth muscle cells embedded in arteriole segment. The results showed that both KCl (30-100 mmol/L) and PE (0.1-30 μmol/L) induced contraction of RIA in a concentration-dependent way. After pretreatment with 18β-GA (100 μmol/L), KCl- or PE-induced constriction of RIA was significantly decreased. After application of 18β-GA (100 μmol/L), the C(input), G(input) and R(input) of the in situ smooth muscle cells were very close to those of dispersed single smooth muscle cells. These results suggest 18β-GA inhibits the contraction induced by KCl and PE, and the underlying mechanism may involve the inhibitory effect of 18β-GA on gap junction.
Animals ; Arteries ; drug effects ; physiopathology ; Constriction ; Gap Junctions ; Glycyrrhetinic Acid ; analogs & derivatives ; pharmacology ; In Vitro Techniques ; Myocytes, Smooth Muscle ; cytology ; Patch-Clamp Techniques ; Rats ; Rats, Wistar

OBJECTIVETo investigate the effects of total flavonoids of Litsea Coreana (TFLC) on the gap junction (GJ) intercellular communication in TM3 testicular Leydig cells and whether TFLC can reduce the cytotoxicity of oxaliplatin (OHP) in vitro.

METHODSWe detected the effect of TFLC on the dye spread of the in vitro cultured TM3 cells by parachute assay, observed changes in the expression of connexin 43 (Cx43) total protein in the TFLC-treated TM3 cells by Western blot, and determined the effects of TFLC on the expression of Cx43 on the membrane of the TM3 cells by immunofluorescence assay and on the cytotoxicity of OHP by MTT assay.

RESULTSTFLC obviously enhanced the GJ function with the increasing of the TFLC concentration in the TM3 cells. Western blot and immunofluorescence assay confirmed that TFLC significantly enhanced the expression of Cx43 total protein and Cx43 expression on the membrane of the TM3 cells. MTT assay showed that at a high cell density (confluent with GJ formation), 20 microg/ml TFLC enhanced the GJ function of the TM3 cells and reduced the cytotoxicity of OHP (P < 0.05), while at a low density (preconfluent with no GJ formation), TFLC exhibited no effect on the cytotoxicity of OHP (P > 0.05).

CONCLUSIONTFLC increases the Cx43 expression and GJ function in normal TM3 Leydig cells, and the enhancement of GJ function reduces the cytotoxicity of OHP.

Antineoplastic Agents ; toxicity ; Cell Communication ; drug effects ; physiology ; Cell Count ; Connexin 43 ; metabolism ; Flavonoids ; pharmacology ; Gap Junctions ; drug effects ; Humans ; In Vitro Techniques ; Leydig Cells ; drug effects ; ultrastructure ; Litsea ; chemistry ; Male ; Organoplatinum Compounds ; antagonists & inhibitors ; toxicity ; Proteins ; metabolism

OBJECTIVETo explore the repair of Xiangsha Liujunzi Decoction (XSLJZD) on interstitial cells of Cajal (ICC) and gap junction (GJ) in the gastric muscular layer of rats of Pi-qi deficiency syn- drome (PQDS).

METHODSPQDS was established using purgative method with bitter and cold drugs in 30 healthy Wistar rats. After successful modeling they were randomly divided into the treatment group and the model group, 15 in each group. Another 15 healthy Wistar rats were recruited as the healthy control group. Rats in the treatment group were gastric administered with XSLJZD at 2 mL/100 g body weight, once daily for 14 successive days. Equal volume of normal saline was gastrically administered to those in the healthy control group and the model group. The gastric muscle tissues were taken out before modeling, before intervention, and after intervention, respectively. Ultrastructural changes of ICC and GJ were observed using transmission electron microscope (TEM). The number and distribution of Connexin43 (Cx43) were detected using immunohistochemistry.

RESULTSResults of TEM indicated that compared with the healthy control group, both ICC and GJ in the model group showed obvious injury. ICC and GJ were apparently repaired after intervention in the treatment group. Compared with the same group before modeling, the integrated optical density (IOD) of the Cx43 expression significantly decreased in the model group before and after intervention (P <0.05). Compared with before intervention, the IOD of the Cx43 expression significantly increased in the treatment group (P <0.05). Compared with the healthy control group, the IOD of the Cx43 expression significantly decreased in the model group before and after intervention (P <0.05). Compared with the model group, the IOD of the Cx43 expression significantly increased in the treatment group (P <0.05).

CONCLUSIONSUltrastructures of ICC and GJ in the gastric muscular layer of rats of PQDS were obviously damaged. XSLJZD could repair the structural damage of ICC and GJ in the gastric muscle tissues of rats of PQDS.

Animals ; Connexin 43 ; Drugs, Chinese Herbal ; pharmacology ; Gap Junctions ; Interstitial Cells of Cajal ; drug effects ; Leydig Cells ; Male ; Muscle, Smooth ; Qi ; Rats, Wistar ; Syndrome

OBJECTIVEThis study compared Wistar rat with spontaneously hypertensive rat (SHR) on the electrophysiology and coupling force of the smooth muscle cells in the cerebral arteriolar segments and observe the influence of 18beta-glycyrrhetinic acid(18beta-GA) on the gap junctions between the arterial smooth muscle cells.

METHODSThe outer layer's connective tissue of the cerebral arteriolar segments was removed. Whole-cell patch clamp recordings were used to observe the 18beta-GA's impaction on the arteriolar segment membrane's input capacitance (C(input)), input conductance (G(input)) and input resistance (R(input)) of the smooth muscle cells.

RESULTS(1) The C(input) and G(input) of the SHR arteriolar segment smooth muscle cells was much higher than the Wistar rats, there was significant difference (P < 0.05). (2) 18beta-GA concentration-dependently reduced C(input) and G(input) (or increase R(input)) on smooth muscle cells in arteriolar segment. IC50 of 18beta-GA suppression's G(input) of the Wistar rat and SHR were 1.7 and 2.0 micromol/L respectively, there was not significant difference (P > 0.05). After application of 18beta-GA concentration > or = 100 micrmol/L, the C(input), G(input) and R(input) of the single smooth muscle cells was very close.

CONCLUSIONGap junctional coupling is enhanced in the SHR cerebral arterial smooth muscle cells. 18beta-GA concentration-dependent inhibits Wistar rat's and SHR cerebral arteriolar gap junctions between arterial smooth muscle cells. The inhibitory potency is similar between the two different rats. When 18beta-GA concentration is > or = 100 micromol/L, it can completely block gap junctions between arteriolar smooth muscle cells.

Animals ; Cerebral Arteries ; cytology ; Gap Junctions ; drug effects ; Glycyrrhetinic Acid ; analogs & derivatives ; pharmacology ; Male ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; drug effects ; Patch-Clamp Techniques ; Rats ; Rats, Inbred SHR ; Rats, Wistar

OBJECTIVETo investigate the effect of Src kinase inhibitor PP2 on intercellular communication of gap junction in breast cancer cells.

METHODSCultured breast cancer Hs578T cells were treated with various concentrations of pp2 (0,1,2,4,8,16,32 μmol/L) for 24h. Cell growth was determined by MTT assay; dye spread in Hs578T cells was measured by Parachute assay; and the expression of Src kinase in Hs578T cells was detected by Western blot.

RESULTSMTT assay showed that the survive rate of Hs578T cells treated with PP2 (1 ≊ 8 μmol/L) was 98% ± 3% ≊ 94 % ± 4%. Parachute assay showed that compared to control group the standard normalized dye spread rates of Hs578T cells treated with 1,2,4 and 8 μmol/L PP2 were 1.60 ± 0.08,2.00 ± 0.05,2.20 ± 0.05 and 2.70 ± 0.09,respectively (all P<0.01). Moreover,compared to control group at the same time points,the standard normalized dye spread of Hs578T cells treated with 8 μmol/L PP2 for 6,12 and 24 h were 1.4 ± 0.05,1.7 ± 0.06,and 2.2 ± 0.07,respectively (all P<0.01). Western blot showed that the expression ratios of Src kinase/β-actin of Hs578T cells treated with 1,2,4 and 8 μmol/L PP2 for 24 h were 0.93 ± 0.02,0.70 ± 0.09,0.66 ± 0.09 and 0.36 ± 0.10,which were significantly inhibited compared with control group (P<0.05 or 0.01). And the expression ratio of Src kinase/β-actin of Hs578T cells treated with 8 μmol/L PP2 for 6,12 and 24h was 0.82 ± 0.03,0.66 ± 0.08 and 0.59 ±0.09, which were all inhibited significantly compared to control group (P<0.01).

CONCLUSIONPP2 enhances the gap junction function in breast cancer Hs578T cells, which is probably related to the inhibition of Src kinase.

Breast Neoplasms ; pathology ; Cell Line, Tumor ; Female ; Gap Junctions ; drug effects ; Humans ; Pyrimidines ; administration & dosage ; pharmacology ; src-Family Kinases ; metabolism

OBJECTIVETo investigate the effect of valproate acid sodium (VPA) on gap junction intercellular communication in breast cancer Hs578T cells and explore the mechanism.

METHODSMTT assay was used to detect the cytotoxicity of VPA on Hs578T cells, and parachute assay was used to detect the effect of VPA on dye spread of the cells. Western blotting was employed to detect the expression changes of Cx43 total protein in VPA-treated Hs578T cells. The effect of VPA on the expression of Cx43 on the surface of Hs578T cells was examined with immunofluorescence assay.

RESULTSMTT assay showed no obvious cytotoxicity of VPA on Hs578T cells at the concentrations below 10 mmol/L. VPA below 5 mmol/L obviously increased the gap junction function in Hs578T cells (P<0.01), and significantly enhanced the expression of Cx43 total protein (P<0.01) and Cx43 expression on the surface of Hs578T cells (P<0.01).

CONCLUSIONVPA can obviously increase the gap junction function in Hs578T cells possibly by enhancing Cx43 total protein expression and Cx43 protein expression on the surface of Hs578T cells.

Breast Neoplasms ; metabolism ; Cell Communication ; drug effects ; Cell Line, Tumor ; Connexin 43 ; metabolism ; Female ; Gap Junctions ; drug effects ; Humans ; Valproic Acid ; pharmacology