Deadly liaisons: fatal attraction between CCN matricellular proteins and the tumor necrosis factor family of cytokines

Recent studies have revealed an unexpected synergism between two seemingly unrelated protein families: CCN matricellular proteins and the tumor necrosis factor (TNF) family of cytokines. CCN proteins are dynamically expressed at sites of injury repair and inflammation, where TNF cytokines are also expressed. Although TNFα is an apoptotic inducer in some cancer cells, it activates NFκB to promote survival and proliferation in normal cells, and its cytotoxicity requires inhibition of de novo protein synthesis or NFκB signaling. The presence of CCN1, CCN2, or CCN3 overrides this requirement and unmasks the apoptotic potential of TNFα, thus converting TNFα from a proliferation-promoting protein into an apoptotic inducer. These CCN proteins also enhance the cytotoxicity of other TNF cytokines, including LTα, FasL, and TRAIL. Mechanistically, CCNs function through integrin α₆β₁ and the heparan sulfate proteoglycan (HSPG) syndecan-4 to induce reactive oxygen species (ROS) accumulation, which is essential for apoptotic synergism. Mutant CCN1 proteins defective for binding α₆β₁-HSPGs are unable to induce ROS or apoptotic synergism with TNF cytokines. Further, knockin mice that express an α₆β₁-HSPG-binding defective CCN1 are blunted in TNFα- and Fas-mediated apoptosis, indicating that CCN1 is a physiologic regulator of these processes. These findings implicate CCN proteins as contextual regulators of the inflammatory response by dictating or enhancing the cytotoxicity of TNFα and related cytokines.     

OPA1 cleavage depends on decreased mitochondrial ATP level and bivalent metals

OPA1, an intra-mitochondrial dynamin GTPase, is a key actor of outer and inner mitochondrial membrane dynamic. OPA1 amino-terminal cleavage by PARL and m-AAA proteases was recently proposed to participate to the mitochondrial network dynamic in a DeltaPsi(m)-dependent way, and to apoptosis. Here, by an in vitro approach combining the use of purified mitochondrial fractions and mitochondrial targeting drugs, we intended to identify the central stimulus responsible for OPA1 cleavage. We confirm that apoptosis induction and PTPore opening, as well as DeltaPsi(m) dissipation induce OPA1 cleavage. Nevertheless, our experiments evidenced that decreased mitochondrial ATP levels, either generated by apoptosis induction, DeltaPsi(m) dissipation or inhibition of ATP synthase, is the common and crucial stimulus that controls OPA1 processing. In addition, we report that ectopic iron addition activates OPA1 cleavage, whereas zinc inhibits this process. These results suggest that the ATP-dependent OPA1 processing plays a central role in correlating the energetic metabolism to mitochondrial dynamic and might be involved in the pathophysiology of diseases associated to excess of iron or depletion of zinc and ATP.     

Propionibacterium acnes induces acute TNFα-mediated apoptosis of hepatocytes followed by inflammatory T-cell-mediated granulomatous hepatitis in mice

The CD3+/TCR+ T-cell-mediated hepatic inflammation induced byPropionibacterium acnes could be divided into an acute and a chronic phase. The acute phase occurred within 72 h after injection and displayed hepatic apoptosis. Anti-TNF antibody inhibited both theP. acnes-induced hepatic apoptosis and lymphocyte infiltration seen in this phase, indicating the involvement of this cytokine. Thereafter, a chronic phase was manifested from days 7 to 14 after injection. It was characterized as granulomatous inflammation admixed with apoptosis of infiltrating lymphocytes and some hepatocytes. Immunohistochemical staining showed that the infiltrating lymphocytes displayed TNF, TNF type I receptor and a variety of cytokines including IL-1, IL-4, IL-6, IL-10, IFN or IL-12. Interestingly, in naive mice, the arteries in the liver constitutively expressed IFN. Its expression appeared to be substantially increased at 48 h, decreased at 72 h, and increased again on day 14 afterP. acnes injection. Furthermore, Fas or FasL was only detected on the lymphocytes within the granuloma. We conclude thatP. acnes can induce a TNF-mediated acute hepatic apoptosis which subsequently progress to a T-cell-mediated granulomatous hepatitis with increased expression of multiple cytokines and Fas/FasL.     

Effect of H. pylori on the expression of TRAIL, FasL and their receptor subtypes in human gastric epithelial cells and their role in apoptosis

BACKGROUND AND AIMS: In the human stomach expression of TNF-related apoptosis inducing ligand (TRAIL) and its receptors and the modulatory role of Helicobacter pylori are not well described. Therefore, we investigated the effect of H. pylori on the expression of TRAIL, FasL and their receptors (TRAIL-R1-R4, Fas) in gastric epithelial cells and examined their role in apoptosis. MATERIALS AND METHODS: mRNA and protein expression of TRAIL, FasL and their receptors were analyzed in human gastric epithelial cells using RT-PCR, Western blot, and immunohistochemistry. Gastric epithelial cells were incubated with FasL, TRAIL and/or H. pylori, and effects on expression, cell viability and epithelial apoptosis were monitored. Apoptosis was analyzed by histone ELISA, DAPI staining and immunohistochemistry. RESULTS: TRAIL, FasL and their receptor subtypes were expressed in human gastric mucosa, gastric epithelial cell primary cultures and gastric cancer cells. TRAIL, FasL and H. pylori caused a time- and concentration-dependent induction of DNA fragmentation in gastric cancer cells with synergistic effects. In addition, H. pylori caused a selective up-regulation of TRAIL, TRAIL-R1 and Fas mRNA and protein expression in gastric cancer cells. CONCLUSIONS: Next to FasL and Fas, TRAIL and all of its receptor subtypes are expressed in the human stomach and differentially modulated by H. pylori. TRAIL, FasL and H. pylori show complex interaction mediating apoptosis in human gastric epithelial cells. These findings might be important for the understanding of gastric epithelial cell kinetics in patients with H. pylori infection.     

Role of apoptosis and Fas/FasL system in the oogenesis of the spotted ray Torpedo marmorata

In the present paper we investigated the role played by apoptosis during oogenesis in the cartilaginous fish Torpedo marmorata. TEM, TUNEL and immunohistochemical techniques were employed to specifically reveal morphological and biochemical hallmarks of apoptosis in specimens from birth to sexual maturity. Data obtained demonstrate that apoptosis occurs in prefollicular oocyte selection, in maintaining the homeostasis of granulosa in healthy growing oocyte and in resorbing atretic follicles. In this respect, the involvement of apoptosis in Torpedo marmorata oogenesis closely parallels that found in mammals, thus confirming that strategies of germ cell selection among vertebrates have been evolutionarily preserved.     

The apoptotic effect of intercalating agents on HPV-negative cervical cancer C-33A cells

Summary. Cervical cancer is one of the leading causes of female cancer death worldwide with about 500,000 deaths per year. Both mitomycin C and cisplatin are alkylating agents, which bind and intercalate DNA, and thus used as anti-cancer drugs. In these studies, we focused on investigating the apoptotic effects of intercalating agents on HPV-negative cervical cancer C-33A cells. Accordingly, C-33A cells were treated with carboplatin, mitomycin C or cisplatin. Cell cycle analysis revealed that treatment with mitomycin C and cisplatin but not with carboplatin resulted in apoptosis. Both mitomycin C and cisplatin induced apoptosis in C-33A cells via caspase-8 and -3 processing in a Fas/FasL-dependent manner and also suppressed IL-18 expression, while they down-regulated IκB expression and up-regulated p65 expression. These results suggest that both mitomycin C and cisplatin induce apoptosis, not only via the caspase-8 and -3 dependent Fas/FasL pathway, but also via the regulation of NF-κB activity and IL-18 expression in HPV-negative cervical cancer C-33A cells.     

Genetic polymorphisms of Fas (CD95) and Fas ligand (CD178) influence the rise in CD4+ T cell count after antiretroviral therapy in drug-naïve HIV-positive patients

Fas and Fas ligand (FasL) are the main genes that control cell death in the immune system. Indeed, they are crucial for the regulation of T lymphocyte homeostasis because they can influence cell proliferation. A strong debate exists on the importance of Fas/FasL system during HIV infection, which is characterized by the loss of CD4+ T cells directly, or indirectly, caused by the virus. To investigate whether the genetic background of the host plays a role in the immunoreconstitution, we studied the influence of different Fas and FasL polymorphisms on CD4+ T lymphocyte count and plasma viral load following initiation of highly active antiretroviral therapy (HAART) in drug-naïve HIV+ patients. We studied 131 individuals, who were compared to 136 healthy donors. Statistical analysis was performed by using X ² test, Fischer's Exact Test, and analysis for repeated measurements. The group of HIV+ patients had an unexpected lower frequency of FasLnt169 polymorphism (delT allele) than healthy controls (p=0.039). We then observed no significant differences in the immune reconstitution, in terms of CD4+ T cell increase, when the influence of single alleles of the gene Fas or FasL was considered. However, the combination of some polymorphisms of Fas or FasL significantly influenced CD4+ T cell production and viral load decrease, showing that these genes can play a role in the immunoreconstitution triggered by antiretroviral therapy.