Connexin32 activates necroptosis through Src-mediated inhibition of caspase 8 in hepatocellular carcinoma.

Necroptosis is an alternative form of programmed cell death that generally occurs under apoptosis-deficient conditions. Our previous work showed that connexin32 (Cx32) promotes the malignant progress of hepatocellular carcinoma (HCC) by enhancing the ability of resisting apoptosis in vivo and in vitro. Whether triggering necroptosis is a promising strategy to eliminate the apoptosis-resistant HCC cells with high Cx32 expression remains unknown. In this study, we found that Cx32 expression was positively correlated with the expression of necroptosis protein biomarkers in human HCC specimens, cell lines, and a xenograft model. Treatment with shikonin, a well-used necroptosis inducer, markedly caused necroptosis in HCC cells. Interestingly, overexpressed Cx32 exacerbated shikonin-induced necroptosis, but downregulation of Cx32 alleviated necroptosis in vitro and in vivo. Mechanistically, Cx32 was found to bind to Src and promote Src-mediated caspase 8 phosphorylation and inactivation, which ultimately reduced the activated caspase 8-mediated proteolysis of receptor-interacting serine-threonine protein kinase 1/3, the key molecule for necroptosis activation. In conclusion, we showed that Cx32 contributed to the activation of necroptosis in HCC cells through binding to Src and then mediating the inactivation of caspase 8. The present study suggested that necroptosis inducers could be more favorable than apoptosis inducers to eliminate HCC cells with high expression of Cx32.

Inhibition of the ATR-DNAPKcs-RB axis drives G1/S-phase transition and sensitizes triple-negative breast cancer (TNBC) to DNA holliday junctions.

Targeting the DNA damage response (DDR) is a promising strategy in oncotherapy, as most tumor cells are sensitive to excess damage due to their repair defects. Ataxia telangiectasia mutated and RAD3-related protein (ATR) is a damage response signal transduction sensor, and its therapeutic potential in tumor cells needs to be precisely investigated. Herein, we identified a new axis that could be targeted by ATR inhibitors to decrease the DNA-dependent protein kinase catalytic subunit (DNAPKcs), downregulate the expression of the retinoblastoma (RB), and drive G1/S-phase transition. Four-way DNA Holliday junctions (FJs) assembled in this process could trigger S-phase arrest and induce lethal chromosome damage in RB-positive triple-negative breast cancer (TNBC) cells. Furthermore, these unrepaired junctions also exerted toxic effects to RB-deficient TNBC cells when the homologous recombination repair (HRR) was inhibited. This study proposes a precise strategy for treating TNBC by targeting the DDR and extends our understanding of ATR and HJ in tumor treatment.

The gap junction inhibitor INI-0602 attenuates mechanical allodynia and depression-like behaviors induced by spared nerve injury in rats.

Gap junctions (GJs) are novel molecular targets for pain therapeutics due to their pain-promoting function. INI-0602, a new GJ inhibitor, exerts a neuroprotective role, while its role in neuropathic pain is unclear. The objective was to investigate the analgesic role and mechanisms of INI-0602 in neuropathic pain induced by spared nerve injury (SNI), and whether INI-0602 attenuated pain-induced depression-like behaviors. Rats were randomly assigned to saline treatment groups (sham+NS and SNI+NS) or INI-0602 treatment groups (sham+INI-0602 and SNI+INI-0602). The von Frey test was used to assess pain behavior, and the sucrose preference test, the forced swimming test, and the tail suspension test were used to assess depression-like behaviors. Gap junction intercellular communication (GJIC) was measured by parachute assay. Western blots were used to determine the protein expression. In vitro, INI-0602 significantly suppressed GJIC by decreasing connexin43 and connexin32 expression. In vivo, INI-0602 significantly suppressed mechanical allodynia during initiation (7 days after SNI) and the maintenance phase (21 days after SNI) and simultaneously attenuated accompanying depression-like behaviors. Furthermore, INI-0602 markedly suppressed the activation of astrocytes and microglia on days 7 and 21 by reducing GJIC. Finally, INI-0602 reversed the changes in the brain-derived neurotrophic factor and Nr2b subunits of the N-methyl-D-aspartate receptor in SNI rats, suggesting that these effects of INI-0602 were related to its analgesic effect. Our findings demonstrated that blocking GJs with INI-0602 attenuated mechanical pain hypersensitivity and related depression-like behaviors in SNI rats by reducing glial activation.

Gut Barrier Dysfunction Induced by Aggressive Fluid Resuscitation in Severe Acute Pancreatitis is Alleviated by Necroptosis Inhibition in Rats.

Fluid resuscitation is the first-line antishock treatment in severe acute pancreatitis (SAP). Currently, although mentions of complications related to aggressive fluid resuscitation are very frequent, a lack of proper handling of complications remains. One of the most important complications is intestinal barrier injury, including intestinal ischemia-reperfusion injury following aggressive fluid resuscitation. Once injured, the intestinal barrier may serve as the source of additional diseases, including systemic inflammatory response syndrome and multiple organ dysfunction syndrome, which aggravate SAP. This study focused on the underlying mechanisms of gut barrier dysfunction in rats induced by aggressive fluid resuscitation in SAP. This study further indicated the important role of necroptosis in intestinal barrier injury which could be relieved by using necroptosis-specific inhibitor Nec-1 before aggressive fluid resuscitation, thus reducing intestinal barrier damage. We also found pancreas damage after intestinal ischemia/reperfusion challenge and indicated the effects of high mobility group protein B1 in the vicious cycle between SAP and intestinal barrier damage.