Immunostimulatory and anti-tumor activity of a water-soluble polysaccharide from <Emphasis Type="Italic">Phellinus baumii</Emphasis> mycelia

A 17-kDa water-soluble polysaccharide (PB) was isolated and purified from Phellinus baumii using the DEAE-Sephadex A-50 and LPLC-Sephadex G-75 methods. Exposure of RAW264.7 macrophages to PB resulted in a significant increase of the cellular proliferation rate, nitric oxide production and expression levels of the IL-1β, IL-18, IL-6, IL-12p35 and IL-12p40 genes. An MTT assay indicated that PB markedly suppressed the proliferation of HepG₂ human liver cancer cells in a dose-dependent manner. Cell cycle analysis demonstrated that PB caused cell cycle arrest at the S phase, and 400 μg/ml of PB induced apoptotic cell death in HepG₂ cells after 48 h. The results suggested that PB inhibited the proliferation of HepG₂ cells by inducing cell cycle arrest at S phase, leading to apoptosis. In summary, our data indicate that the PB exerts immunoregulatory and anti-tumor activities in vitro.     

Daxx过表达对氧化应激诱导的肝肿瘤细胞凋亡的影响

死亡结构域相关蛋白Daxx可以敏化多种肿瘤细胞的凋亡过程,但对于肝肿瘤细胞株HepG2的影响未见报道．为了研究Daxx增加肝HepG2细胞对药物敏感性的影响及机制,为开发药物新的药理作用提供理论依据,分别转染pEGFP-C1和pEGFP-C1-Daxx这两个载体到HepG2细胞．实验分组如下：（1）正常对照组（未转染细胞组）;（2）pEGFP-C1空载体转染组（HepG2/GFP细胞）;（3）pEGFP-C1-Daxx表达载体转染组（nepG2/GFP-Daxx细胞）．筛选稳定细胞株,用逆转录聚合酶链反应检测mRNA的表达;用过氧化氢孵育24h诱导细胞凋亡,采用MTT法和流式细胞术检测细胞凋亡率,Western blot检测蛋白质的表达．经G418筛选稳定的细胞运用RT-PCR技术分析其mRNA,结果显示,转染绿色荧光蛋白Daxx表达载体的细胞Daxx的mRNA明显上调：用荧光显微镜观察到Daxx蛋白主要定位于细胞核．用过氧化氢诱导HepG2细胞凋亡,观察到过氧化氢呈浓度依赖性地抑制HepG2细胞活性．正常对照细胞、HepG2／GFP、HepG2／GFP-Daxx 3组细胞的IC50值分别是0．72、0．76、0．49mmol／L．并且运用流式细胞仪检测到HepG2／GFP-Daxx组细胞凋亡率明显高于转染空载体质粒组与未转染组（（42．9&#177;8．42）vs（27．3&#177;6．38）or（28．5&#177;4．71））．提示HepG2/GFP-Daxx细胞对过氧化氢的反应性较未转染细胞和HepG2/GFP敏感．还运用Western-blot检测到活化的caspase3在Daxx转染组细胞表达最强,达到（204．66&#177;19．68）％,而未转染和HepG2/GFP组细胞分别是（100&#177;3．1）％、（107．39&#177;20．1）％,进一步说明了Daxx可以增加HepG2细胞对于过氧化氢的敏感性．同时,观察到过氧化氢处理24h后,Daxx转染组细胞磷酸化的JNK表达明显高于空载体转染组和未转染细胞组．上述结果表明：a．Daxx可以增加肝HepG2细胞对过氧化氢诱导的细胞凋亡敏感性;b．Daxx蛋白敏化过氧化氢诱导的HepG2细胞凋亡可能与协同增加JNK活性有关．     

The anti-tumor effect of human monocyte-derived dendritic cells loaded with HSV-TK/GCV induced dying cells

Herpes simplex virus thymidine kinase (HSV-TK) gene and dendritic cells (DC) have been used as the pioneering in cancer therapy. HSV-TK gene can induce apoptosis and necrosis in tumor cells in the presence of the non-toxic prodrug ganciclovir (GCV). We investigated the anti-tumor effect of DC vaccination by introducing dying cells from HSV-TK gene treatment as an adjuvant. HepG₂-TK cell line was established by transfecting human hepatoma cell line HepG₂ (HLA-A₂ positive) with HSV-TK gene. Dying tumor cells were generated by culturing HepG₂-TK cells with GCV. After engulfed dying cells efficiently, immature DCs (imDC) derived from human monocytes were fully matured and elicited marked proliferation and cytotoxicity against HLA matched HepG₂ cells in autologous peripheral blood mononuclear cells (PBMC). It also implied that HepG₂ specific CTLs played an important role in the cytotoxicity which was primarily depended on Th1 responses. Given the feasibility of inducing dying cells by HSV-TK/GCV in vivo, our results suggest an effective method in clinical human hepatocellular carcinoma (HCC) treatment by an in vitro model of applying HSV-TK gene modified human tumor cells integrated with DC vaccination.     

ClC-3 chloride channel prevents apoptosis induced by hydrogen peroxide in basilar artery smooth muscle cells through mitochondria dependent pathway

ClC-3 Cl⁻ channel plays an important role in cell volume regulation and cell cycle. In vascular smooth muscle cells, we have found that ClC-3 was involved in ET-1 induced cell proliferation. The present study was designed to further investigate the role of ClC-3 Cl⁻ channel in H₂O₂-induced apoptosis and its underlying mechanisms in rat basilar arterial smooth muscle cell (BASMCs). By using ClC-3 cDNA and small interference RNA (siRNA) transfection strategy, it was found that overexpression of ClC-3 significantly decreased the apoptotic rate of H₂O₂-treated BASMCs and increased the cell viability, whereas silencing of ClC-3 with siRNA produced opposite effects and increased the apoptotic rate. ClC-3 overexpression decreased cytochrome C release and caspase-3 activation, and increased both the stability of mitochondrial membrane potential and the ratio of Bcl-2/Bax, whereas silencing of ClC-3 produced opposite effect. Furthermore, we demonstrated that overexpression of ClC-3 attenuated, whereas silencing of ClC-3 facilitated, the degradation of LaminA, one of the structural matrix proteins, in BASMCs. Our data suggest that ClC-3 Cl⁻ channel can modulate H₂O₂-induced apoptosis in BASMCs via the intrinsic, mitochondrial pathway.     

15-Deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) Induces Cell Death Through Caspase-independent Mechanism in A172 Human Glioma Cells

15-Deoxy-^Δ12,14-prostaglandin J₂ (15d-PGJ₂) is a naturally occurring cyclopentenone metabolite of prostaglandin D₂ (PGD₂) and is known as a specific potent ligand for the peroxisome proliferators activator receptor-γ (PPARγ). 15d-PGJ₂ inhibits cell growth and induces apoptosis in a number of different cancer cells. However, the underlying mechanism by which 15d-PGJ₂ induces cell death remains to be defined. The present study was undertaken to determine the effect of 15d-PGJ₂ on cell death in A172 human glioma cells. 15d-PGJ₂ caused reactive oxygen species (ROS) generation. 15d-PGJ₂-induced ROS production and cell death were prevented by the antioxidant N-acetylcysteine. Activation of mitogen-activated protein kinases (MAPK) was not observed in cells treated with 15d-PGJ₂ and inhibitors of MAPK subfamilies also were not effective in preventing 15d-PGJ₂-induced cell death. 15d-PGJ₂ treatment caused mitochondrial dysfunction, as evidenced by depolarization of mitochondrial membrane potential. 15d-PGJ₂ induced caspase activation at 24 h of treatment, but the 15d-PGJ₂-induced cell death was not prevented by caspase inhibitors. The antiapoptotic protein XIAP levels and release of apoptosis inducing factor (AIF) into the cytosol were not altered by 15d-PGJ₂ treatment. Taken together, these findings indicate that 15d-PGJ₂ triggers cell death through a caspase-independent mechanism and ROS production and disruption of mitochondrial membrane potential play an important role in the 15d-PGJ₂-induced cell death in A172 human glioma cells.     

Induction of mitochondrial biogenesis protects against caspase-dependent and caspase-independent apoptosis in L6 myoblasts

Apoptotic signaling plays an important role in skeletal muscle degradation, atrophy, and dysfunction. Mitochondria are central executers of apoptosis by directly participating in caspase-dependent and caspase-independent cell death signaling. Given the important apoptotic role of mitochondria, altering mitochondrial content could influence apoptosis. Therefore, we examined the direct effect of modest, but physiological increases in mitochondrial biogenesis and content on skeletal muscle apoptosis using a cell culture approach. Treatment of L6 myoblasts with SNAP or AICAR (5 h/day for 5 days) significantly increased PGC-1, AIF, cytochrome c, and MnSOD protein content as well as MitoTracker staining. Following induction of mitochondrial biogenesis, L6 myoblasts displayed decreased sensitivity to apoptotic cell death as well as reduced caspase-3 and caspase-9 activation following exposure to staurosporine (STS) and C2-ceramide. L6 myoblasts with higher mitochondrial content also exhibited reduced apoptosis and AIF release following exposure to hydrogen peroxide (H₂O₂). Analysis of several key apoptosis regulatory proteins (ARC, Bax, Bcl-2, XIAP), antioxidant proteins (catalase, MnSOD, CuZnSOD), and reactive oxygen species (ROS) measures (DCF and MitoSOX fluorescence) revealed that these mechanisms were not responsible for the observed cellular protection. However, myoblasts with higher mitochondrial content were less sensitive to Ca^2 +-induced mitochondrial permeability transition pore formation (mPTP) and mitochondrial membrane depolarization. Collectively, these data demonstrate that increased mitochondrial content at physiological levels provides protection against apoptotic cell death by decreasing caspase-dependent and caspase-independent signaling through influencing mitochondrial Ca^2 +-mediated apoptotic events. Therefore, increasing mitochondrial biogenesis/content may represent a potential therapeutic approach in skeletal muscle disorders displaying increased apoptosis.     

Induction of apoptosis by Polygonatum odoratum lectin and its molecular mechanisms in murine fibrosarcoma L929 cells

Background

The Galanthus nivalis agglutinin (GNA)-related lectins have been reported to bear antiproliferative and apoptosis-inducing activities in cancer cells; however, the precise mechanisms by which GNA-related lectins induce cell death are still only rudimentarily understood.

Methods

In the present study, Polygonatum odoratum lectin (designated POL), a mannose-binding specific GNA-related lectin, possessed a remarkable antiproliferative activity toward murine fibrosarcoma L929 cells. And, this lectin induced L929 cell apoptosis in a caspase-dependent manner. In addition, POL treatment increased the levels of FasL and Fas-Associated protein with Death Domain (FADD) proteins and resulted in caspase-8 activation. Also, POL treatment caused mitochondrial transmembrane potential collapse and cytochrome c release, leading to activations of caspase-9 and caspase-3. Moreover, POL treatment enhanced tumor necrosis factor α (TNFα)-induced L929 cell apoptosis.

Results

Our data demonstrate for the first time that this lectin induces apoptosis through both death-receptor and mitochondrial pathways, as well as amplifies TNFα-induced L929 cell apoptosis.

General significance

These inspiring findings would provide new molecular basis for further understanding cell death mechanisms of the Galanthus nivalis agglutinin (GNA)-related lectins in future cancer investigations.     

Anti-Hepatoma Activity of a Novel Compound Glaucocalyxin H In Vivo and In Vitro

Glaucocalyxin H (GLH) is a new compound isolated from a traditional Chinese medical herb Isodon japonica var. glaucocalyx which has been used for folk medicine. This study was carried out for the first time to investigate the potential role of GLH in anti-hepatoma activity and underlying mechanisms in it. GLH could inhibit the growth of tumor in mice and induce HepG₂ cells to death as assessed by the tumor reduction assay, toxic assay, morphological change, and survival rate assay. Many antitumor drugs originated from plants could inhibit the growth of tumor by inducing cells to apoptosis. The morphological changes of HepG₂ cells treated with different concentrations of GLH under fluorescence and electron microscope and apoptotic rates were detected to verify its effect on apoptosis. As shown in the study, GLH could induce HepG₂ cells to apoptosis in a dose-dependent manner. Bcl₂ and Bax proteins played important roles in apoptosis and the disequilibrium between Bcl₂ and Bax might result in apoptosis. The expression of Bax protein was upregulated and Bcl₂ protein was downregulated in HepG₂ cells treated with GLH assessed by Western blotting, and they were in a dose-dependent manner. Taken together, GLH can inhibit the growth of hepatoma cells in vivo and in vitro by inducing cell apoptosis due to the decreased Bcl₂ and increased Bax proteins suggesting that GLH could be a potential candidate as an anti-hepatoma agent for the therapeutic treatment of hepatoma.

Electronic supplementary material

The online version of this article (doi:10.1208/s12249-014-0227-3) contains supplementary material, which is available to authorized users.KEY WORDS: apoptosis, Glaucocalyxin H, hepatoma, HepG₂ cell     

CoCl2 induces apoptosis through the mitochondria- and death receptor-mediated pathway in the mouse embryonic stem cells

Embryonic hypoxia/ischemia is a major cause of a poor fetal outcome and future neonatal and adult handicaps. However, biochemical cellular events in mouse embryonic stem (mES) cells during hypoxia remains unclear. This study investigated the underlying mechanism of apoptosis in mES cells under CoCl₂-induced hypoxic/ischemic conditions. CoCl₂ enhanced the expression of hypoxia-inducible factor-1α (HIF-1α) and the accumulation of reactive oxygen species in mES cells. The CoCl₂-treated mES cells showed a decrease in cell viability as well as typical apoptotic changes, cell shrinkage, chromatin condensation, and nuclear fragmentation and an extended G₂/M phase of the cell cycle. CoCl₂ augmented the release of cytochrome c into the cytosol from the mitochondria with a concomitant loss of the mitochondrial transmembrane potential (ΔΨ_m) and upregulated the voltage-dependent anion channel. In addition, CoCl₂-induced caspase-3, -8, and -9 activation and upregulation of p53 level, whereas downregulated Bcl-2 and Bcl-xL, a member of the anti-apoptotic Bcl-2 family in mES cells. Furthermore, CoCl₂ led to the upregulation of Fas and Fas-ligand, which are the death receptor assemblies, as well as the cleavage of Bid in mES cells. These results suggest that CoCl₂ induces apoptosis through both mitochondria- and death receptor-mediated pathways that are regulated by the Bcl-2 family in mES cells.     

Reactions of 2-thiopyridone and related N-, S- and C-methylated derivatives with [Rh2Cl2(CO)4]: crystal and molecular structure of fac-[Rh(MeC5H3NS)3] containing 6-methyl-2-thiopyridonato ligands

[Rh₂Cl₂(CO)₄] reacts with the ligands L (2-pyridone, 2-thiopyridone, and the isomers 6-methyl-2-thiopyridone, 2-methylmercaptopyridine, and N-methylthiopyridone) to give initially, when L/Rh = 1, the bridged-cleaved compounds cis- [RhCl(CO)₂L]. Further additions of 2-methyl- mercaptopyridine, N-methylthiopyridone, or 2-pyridone caused no further change, but 2- thiopyridone and 6-methyl-2-thiopyridone gave new cis-dicarbonyl species (L/Rh = 2) and eventually monocarbonyl species (L/Rh > 3). All these solutions are air-sensitive and air oxidation of a solution of [Rh₂Cl₂(CO)₄] with an excess of 6-methyl-2- thiopyridone gave fac-[Rh(MeC₅H₃NS)₃] the X-ray structure of which shows three equivalent chelating 6-methyl-2-thiopyridonato ligands.     

Role of ERK in Hydrogen Peroxide-Induced Cell Death of Human Glioma Cells

Oxidative stress is known to induce cell death in a wide variety of cell types, apparently by modulating intracellular signaling pathways. Activation of extracellular signal-regulated kinase (ERK) in oxidative stress remains controversial. In some cellular systems, the ERK activation is associated with protection against oxidative stress, while in other system, the ERK activation is involved in apoptotic cell death. The present study was undertaken to examine the role of ERK activation in H₂O₂-induced cell death of human glioma (A172) cells. H₂O₂ resulted in a time- and dose-dependent cell death, which was largely attributed to apoptosis. H₂O₂ treatment caused marked sustained activation of ERK. The ERK activation and cell death induced by H₂O₂ was prevented by catalase, the hydrogen peroxide scavenger, and U0126, an inhibitor of ERK upstream kinase MEK1/2. Transient transfection with constitutive active MEK1, an upstream activator of ERK1/2, increased H₂O₂-induced cell death, whereas transfection with dominant-negative mutants of MEK1 decreased the cell death. The ERK activation and cell death caused by H₂O₂ was inhibited by antioxidants (N-acetylcysteine and trolox), Ras inhibitor, and suramin. H₂O₂ produced depolarization of mitochondrial membrane potential and its effect was prevented by catalase and U0126. Taken together, these findings suggest that growth factor receptor/Ras/MEK/ERK signaling pathway plays an active role in mediating H₂O₂-induced apoptosis of human glioma cells and functions upstream of mitochondria-dependent pathway to initiate the apoptotic signal.     

The involvement of mitochondria and the caspase-9 activation pathway in rituximab-induced apoptosis in FL cells

Despite the wide use of anti-CD20 antibody rituximab in the cancer treatment of B cell malignancies, the signalling pathways of CD20-induced apoptosis are still not understood. By using dominant negative (DN)-caspase-9 overexpressing follicular lymphoma cells we demonstrated that the activation of caspase-9 was essential for rituximab-mediated apoptosis. The death receptor pathway mediated by caspase-8 activation was not involved in rituximab-mediated apoptosis since overexpression of FLIP_short or FLIP_long proteins, inhibitors of caspase-8 activation, could not inhibit rituximab-induced apoptosis. However, the death receptor pathway activation by anti-Fas antibodies showed an additive effect on rituximab-induced apoptosis. The stabilisation of the mitochondrial outer membrane by Bcl-x_L overexpression inhibited cell death, showing the important role of mitochondria in rituximab-induced apoptosis. Interestingly, the rituximab-induced release of cytochrome c and collapse of mitochondrial membrane potential were regulated by caspase-9. We suggest that caspase-9 and downstream caspases may feed back to mitochondria to amplify mitochondrial disruption during intrinsic apoptosis.     

Protective effect of augmenter of liver regeneration on hydrogen peroxide-induced apoptosis in SH-SY5Y human neuroblastoma cells

Background: Hydrogen peroxide, as other reactive oxygen species (ROS) produced during redox processes, induces lipid membrane peroxidation and protein degeneration causing cell apoptosis. ROS are recently considered as messengers in cell signalling processes, which, through reversible protein disulphide bridges formation, activate regulatory factors of cell proliferation and apoptosis. Disulphide bridges formation is catalysed by sulphydryl oxidase enzymes.

Aim: The neuroprotective effect of ALR protein (Alrp), a sulphydryl oxidase enzyme, on H₂O₂-induced apoptosis in SH-SY5Y cells has been evaluated.

Methods: Cell viability, flow cytometric evaluation of apoptotic cells, fluorescent changes of nuclear morphology, immunocytochemistry Alrp detection, Western blot evaluation of mitochondrial cyt c release and mitochondrial swelling were determined.

Results: Alrp prevents the H₂O₂-induced cell viability loss, apoptotic cell death and mitochondrial swelling in SH-SY5Y cells in culture.

Conclusions: The data demonstrate that Alrp improves SH-SY5Y cells survival in H₂O₂-induced apoptosis. It is speculated that this effect could be related to the Alrp enzymatic activity.     

Prostaglandin A2 activates intrinsic apoptotic pathway by direct interaction with mitochondria in HL-60 cells

HL-60 cells treated by prostaglandin (PG) A₂ showed characteristics of apoptosis such as accumulation of hypodiploid and annexin V positive cells, condensed and fragmented nuclei, cytochrome c (Cyt C) release from mitochondria and activation of caspase-1, -2, -3, -7 and -9. PGA₂-induced cell death was rescued by inhibitors of caspase-9 and -3, but PGA₂-induced Cyt C release was not prevented by caspase inhibitors. During Cyt C release by PGA₂, mitochondrial transmembrane potential was maintained and mitochondrial permeability transition pore was not formed. In addition, anti-apoptotic BCL-2 family proteins like BCL-2 and BCL-XL, and ROS scavengers including ascorbic acid and 2,2,6,6-tetramethyl-1-piperidinyloxy were not able to inhibit Cyt C release as well as apoptosis by PGA₂. Finally, it was shown that PGA₂-induced Cyt C release in vitro from purified mitochondria in the absence of cytosolic components. Furthermore, thiol-containing compounds such as N-acetylcysteine, l-cysteine and monothioglycerol prevented Cyt C release, and hence induction of apoptosis. Taken together, these results suggest that PGA₂ activates intrinsic apoptotic pathway by directly stimulating mitochondrial outer membrane permeabilization to release Cyt C, in which thiol-reactivity of PGA₂ plays a pivotal role.     

Ginsenoside-Rd potentiates apoptosis induced by hydrogen peroxide in basilar artery smooth muscle cells through the mitochondrial pathway

Our previous studies showed that ginsenoside-Rd, a purified component from Panax notoginseng, inhibited cell proliferation and reversed basilar artery remodeling. The aim of this study was to investigate whether ginsenoside- Rd influences H₂O₂-induced apoptosis in basilar artery smooth muscle cells (BASMCs). The results showed that ginsenoside-Rd significantly potentiated H₂O₂-induced cell death and cell apoptosis. This resulted in a concentration-dependent reduction of the cell viability. Ginsenoside-Rd further increased cytochrome C release and caspase-9/caspase-3 activations, and reduced the stability of mitochondrial membrane potential (MMP) and the ratio of Bcl-2/Bax. Cyclosporine A, an inhibitor of mitochondrial-permeability transition, inhibited alteration of mitochondrial permeability induced by H₂O₂ and reversed the effect of ginsenoside-Rd on MMP. Our data strongly suggest that ginsenoside-Rd potentiated H₂O₂-induced apoptosis of BASMCs through the mitochondria-dependent pathway.     

High Bcl-2/Bax ratio in Walker tumor cells protects mitochondria but does not prevent H<Subscript>2</Subscript>O<Subscript>2</Subscript>-induced apoptosis via calcineurin pathways

It has been previously shown that Walker 256 tumor cells express a high content of the anti-apoptotic protein Bcl-2 which protects mitochondria against the damaging effects of Ca²⁺. In the present study, we analyze H₂O₂-induced apoptotic death in two different types of tumor cells: Walker 256 and SCC-25. Treatment with H₂O₂ (4mM) increased reactive oxygen species generation and the concentration of cytosolic free Ca²⁺. These alterations preceded apoptosis in both cell lines. In Walker cells, which show a high Bcl-2/Bax ratio, apoptosis was dependent on calcineurin activation and independent of changes in mitochondrial membrane potential (Δ < eqid1 > _m), as well as cytochrome c release. In contrast, in SCC-25 cells, which show a lower Bcl-2/Bax ratio, apoptosis was preceded by a decrease in Δ < eqid2 > _m, mitochondrial permeability transition, and cytochrome c release. Caspase-3 activation occurred in both cell lines. The data suggest that although the high Bcl-2/Bax ratio protected the mitochondria of Walker cells from oxidative stress, it was not sufficient to prevent apoptosis through calcineurin pathways.     

Apoptosis of mesenchymal stem cells induced by hydrogen peroxide concerns both endoplasmic reticulum stress and mitochondrial death pathway through regulation of caspases,p38 and JNK

Poor survival of mesenchymal stem cells (MSCs) compromised the efficacy of stem cell therapy for myocardial infarction. The increase of exogenous reactive oxygen species (ROS) in infracted heart is one of the important factors that challenged the survival of donor MSCs. In the study we aimed to evaluate the effect of oxidative stress on the cell death of MSCs and investigate its mechanisms in order to help with the identification of new biological compounds to reduce donor cells damage. Apoptosis of MSCs were evaluated with Hoechst 33342 staining and flow cytometry analysis. The mitochondrial membrane potential of MSCs was analyzed with JC‐1 staining. Signaling pathways involved in H₂O₂ induced apoptosis were analyzed with Western blot. H₂O₂ induced apoptosis of MSCs in a dose‐ and time‐dependent manner. H₂O₂ induced apoptosis of MSCs via both endoplasmic reticulum (ER) and mitochondrial pathways rather than extrinsic apoptosis pathway. H₂O₂ caused transient rather than sustained activation of p38 and JNK with no effect on ERK1/2 pathway. P38 was involved in the regulation of early apoptosis of MSCs while JNK was involved in the late apoptosis. P38 directed both ER stress and mitochondria death pathway in the early apoptosis. In conclusion, exogenous ROS was a major factor to induce apoptosis of MSCs. Both ER stress and mitochondria death pathway were involved in the apoptosis of MSCs. H₂O₂ activated p38 that directed the above two pathways in the regulation of early apoptosis of MSCs while JNK was involved in the late apoptosis of MSCs. J. Cell. Biochem. 111: 967–978, 2010. © 2010 Wiley‐Liss, Inc.     

Protective effects of methyl gallate on H2O2-induced apoptosis in PC12 cells

Neurodegenerative disorders are a class of diseases that have been linked to apoptosis induced by elevated levels of reactive oxygen species (ROS). ROS activates the apoptotic cascade through mitochondrial dysfunction and damage to lipids, proteins and DNA. Recently, fruit and tea-derived polyphenols have been found to be beneficial in decreasing oxidative stress and increasing overall health. Further, polyphenols including epigallocatechin gallate (EGCG) have been reported to inhibit apoptotic signaling and increase neural cell survival. In an effort to better understand the beneficial properties associated with polyphenol consumption, the aim of this study was to explore the neuroprotective effects of EGCG, methyl gallate (MG), gallic acid (GA) and N-acetylcysteine (NAC) on H₂O₂-induced apoptosis in PC12 cells and elucidate potential protective mechanisms. Cell viability data demonstrates that MG and NAC pre-treatments significantly increase viability of H₂O₂-stressed cells, while pre-treatments with EGCG and GA exacerbates stress. Quantitation of apoptosis and mitochondrial membrane potential shows that MG pre-treatment prevents mitochondria depolarization, however does not inhibit apoptosis and is thus evidence that MG can inhibit mitochondria-mediated apoptosis. Subsequent analysis of DNA degradation and caspase activation reveals that MG inhibits activation of caspase 9 and has a partial inhibitory effect on DNA degradation. These findings confirm the involvement of both intrinsic and extrinsic apoptotic pathways in H₂O₂-induced apoptosis and suggest that MG may have potential therapeutic properties against mitochondria-mediated apoptosis.     

Synthesis, characterization, and C-H/C-C cleavage reactions of two rhodium-trispyrazolylborate dihydrides

Treatment of Tp′Rh(PMe₃)Cl₂ and Tp′Rh(CNCH₂CMe₃)Cl₂ with Cp₂ZrH₂ produces Tp′Rh(PMe₃)H₂ and Tp′Rh(CNCH₂CMe₃)H₂, respectively, in excellent yield. Photolysis of benzene solutions of each dihydride complex generates hydrogen and the fragment [Tp′Rh(L)] which inserts into the solvent C-H bond. The phosphine dihydride has also been shown to be a catalyst for the hydrogenation of biphenylene, showing a capability to cleave C-C bonds. Reductive elimination of benzene from Tp′Rh(PMe₃)PhH is nearly 250 times slower than from Cp*Rh(PMe₃)PhH.     

Resveratrol-induced mitochondrial dysfunction and apoptosis are associated with Ca2+ and mCICR-mediated MPT activation in HepG2 cells

Resveratrol, a natural polyphenolic antioxidant, has been reported to possess the cancer chemopreventive potential in wide range by means of triggering tumor cells apoptosis through various pathways. It induced apoptosis through the activation of the mitochondrial pathway in some kinds of cells. In the present reports, we showed that resveratrol-induced HepG2 cell apoptosis and mitochondrial dysfunction was dependent on the induction of the mitochondrial permeability transition (MPT), because resveratrol caused the collapse of the mitochondrial membrane potential (ΔΨ_m) with the concomitant release of cytochrome c (Cyt.c). In addition, resveratrol induced a rapid and sustained elevation of intracellular [Ca²⁺], which compromised the mitochondrial ΔΨ_m and triggered the process of HepG2 cell apoptosis. In permeabilized HepG2 cells, we further demonstrated that the effect of the resveratrol was indeed synergistic with that of Ca²⁺ and Ca²⁺ is necessary for resveratrol-induced MPT opening. Calcium-induced calcium release from mitochondria (mCICR) played a key role in mitochondrial dysfunction and cell apoptosis: (1) mCICR inhibitor, ruthenium red (RR), prevent MPT opening and Cyt.c release; and (2) RR attenuated resveratrol-induced HepG2 cell apoptotic death. Furthermore, resveratrol promotes MPT opening by lowering Ca²⁺-threshold. These data suggest modifying mCICR and Ca²⁺ threshold to modulate MPT opening may be a potential target to control cell apoptosis induced by resveratrol. Xuemei Tian—Foundation item: Chinese National Natural Science Foundation (No.30300455).