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The aberrant activation of signaling pathways contributes to cancer cells with metabolic reprogramming. Thus, targeting signaling modulators is considered a potential therapeutic strategy for cancer. Subcellular fractionation, coimmunoprecipitation, biochemical analysis, and gene manipulation experiments revealed that decreasing the interaction of kirsten rat sarcoma viral oncogene homolog (KRAS) with p110α in lipid rafts with the use of naringenin (NGN), a citrus flavonoid, causes lipid raft-associated phosphatidylinositol 3-kinase (PI3K)−GTP-ras-related C3 botulinum toxin substrate 1 (Rac1)−protein kinase B (Akt)-regulated metabolic dysfunction of glycolysis and mitochondrial oxidative phosphorylation (OXPHOS), leading to apoptosis in human nasopharyngeal carcinoma (NPC) cells. The use of lethal-7g (let-7g) mimic and let-7g inhibitor confirmed that elevated let-7g resulted in a decrease in KRAS expression, which attenuated the PI3K−Rac1−Akt−BCL-2/BCL-xL-modulated mitochondrial energy metabolic functions. Increased let-7g depends on the suppression of the RNA-specificity of monocyte chemoattractant protein-induced protein-1 (MCPIP1) ribonuclease since NGN specifically blocks the degradation of pre-let-7g by NPC cell-derived immunoprecipitated MCPIP1. Converging lines of evidence indicate that the inhibition of MCPIP1 by NGN leads to let-7g upregulation, suppressing oncogenic KRAS-modulated PI3K–Rac1–Akt signaling and thereby impeding the metabolic activities of aerobic glycolysis and mitochondrial OXPHOS.

Tricalcium phosphate (TCP) has gained attention due to its interconnected porous structures which promote fibrovascular invasion and bony replacement. Moreover, when gelatin is added and crosslinked with genipin (GGT), TCP exhibits robust biocompatibility and stability, making it an excellent bone substitute. In this study, we incorporated emodin and lumbrokinase (LK) into GGT to develop an antibacterial biomaterial. Emodin, derived from various plants, possesses antibacterial and anti-inflammatory properties. LK comprises proteolytic enzymes extracted from the earthworm Lumbricus rubellus and exhibits fibrinolytic activity, enabling it to dissolve biofilms. Additionally, LK stimulates osteoblast activity while inhibiting osteoclast differentiation. GGT was combined with emodin and lumbrokinase to produce the GGTELK composite. The biomedical effects of GGTELK were assessed through in vitro assays and an ex vivo bone defect model. The GGTELK composite demonstrated antibacterial properties, inhibiting the growth of S. aureus and reducing biofilm formation. Moreover, it exhibited anti-inflammatory effects by reducing the secretion of IL-6 in both in vivo cell experiments and the ex vivo model. Therefore, the GGTELK composite, with its stability, efficient degradation, biocompatibility, and anti-inflammatory function, is expected to serve as an ideal bone substitute.

Targeting the activities of endoplasmic reticulum (ER)–mitochondrial-dependent metabolic reprogramming is considered one of the most promising strategies for cancer treatment. Here, we present biochemical subcellular fractionation, coimmunoprecipitation, gene manipulation, and pharmacologic evidence that induction of mitochondria-localized phospho (p)-cyclin dependent kinase 1 (CDK1) (Thr 161)–cyclin B1 complexes by apigenin in nasopharyngeal carcinoma (NPC) cells impairs the ER–mitochondrial bioenergetics and redox regulation of calcium (Ca++) homeostasis through suppressing the B cell lymphoma 2 (BCL-2)/BCL-2/B-cell lymphoma-extra large (BCL-xL)-modulated anti-apoptotic and metabolic functions. Using a specific inducer, inhibitor, or short hairpin RNA for acid sphingomyelinase (ASM) demonstrated that enhanced lipid raft-associated ASM activity confers alteration of the lipid composition of lipid raft membranes, which leads to perturbation of protein trafficking, and induces formation of p110α free p85α–unphosphorylated phosphatase and tensin homolog deleted from chromosome 10 complexes in the lipid raft membranes, causing disruption of phosphatidylinositol 3-kinase (PI3K)−protein kinase B (Akt)−GTP-ras-related C3 botulinum toxin substrate 1 (Rac1)-mediated signaling, thus triggering the p-CDK1 (Thr 161))–cyclin B1-mediated BCL-2 (Thr 69/Ser 87)/BCL-xL (Ser 62) phosphorylation and accompanying impairment of ER–mitochondria-regulated bioenergetic, redox, and Ca++ homeostasis. Inhibition of apigenin-induced reactive oxygen species (ROS) generation by a ROS scavenger N-acetyl-L-cysteine blocked the lipid raft membrane localization and activation of ASM and formation of ceramide-enriched lipid raft membranes, returned PI3K−Akt−GTP-Rac1-modulated CDK1–cyclin B1 activity, and subsequently restored the BCL-2/BCL-xL-regulated ER–mitochondrial bioenergetic activity. Thus, this study reveals a novel molecular mechanism of the pro-apoptotic activity of ASM controlled by oxidative stress to modulate the ER–mitochondrial bioenergetic metabolism, as well as suggests the disruption of CDK1–cyclin B1-mediated BCL-2/BCL-xL oncogenic activity by triggering oxidative stress−ASM-induced PI3K−Akt−GTP-Rac1 inactivation as a therapeutic approach for NPC.

The specialized cholesterol/sphingolipid-rich membrane domains termed lipid rafts are highly dynamic in the cancer cells, which rapidly assemble effector molecules to form a sorting platform essential for oncogenic signaling transduction in response to extra- or intracellular stimuli. Density-based membrane flotation, subcellular fractionation, cell surface biotinylation, and co-immunoprecipitation analyses of bichalcone analog ((E)-1-(4-Hydroxy-3-((4-(4-((E)-3-(pyridin-3-yl)acryloyl)phenyl)piperazin-1-yl)methyl)phenyl)-3-(pyridin-3-yl)prop-2-en-1-one (TSWU-BR4)-treated cancer cells showed dissociation between GRP78 and p85α conferring the recruitment of PTEN to lipid raft membranes associated with p85α. Ectopic expression of GRP78 could overcome induction of lipid raft membrane-associated p85α−unphosphorylated PTEN complex formation and suppression of GRP78−PI3K−Akt−GTP-Rac1-mediated and GRP78-regulated PERK−Nrf2 antioxidant pathway and cancer cell invasion by TSWU-BR4. Using specific inducer, inhibitor, or short hairpin RNA for ASM demonstrated that induction of the lipid raft membrane localization and activation of ASM by TSWU-BR4 is responsible for perturbing homeostasis of cholesterol and ceramide levels in the lipid raft and ER membranes, leading to alteration of GRP78 membrane trafficking and subsequently inducing p85α−unphosphorylated PTEN complex formation, causing disruption of GRP78−PI3K−Akt−GTP-Rac1-mediated signal and ER membrane-associated GRP78-regulated oxidative stress balance, thus inhibiting cancer cell invasion. The involvement of the enrichment of ceramide to lipid raft membranes in inhibition of NF-κB-mediated MMP-2 expression was confirmed through attenuation of NF-κB activation using C2-ceramide, NF-κB specific inhibitors, ectopic expression of NF-κB p65, MMP-2 promoter-driven luciferase, and NF-κB-dependent reporter genes. In conclusion, localization of ASM in the lipid raft membranes by TSWU-BR4 is a key event for initiating formation of ceramide-enriched lipid raft membrane platforms, which causes delocalization of GRP78 from the lipid raft and ER membranes to the cytosol and formation of p85α−unphosphorylated PTEN complexes to attenuate the GRP78-regulated oxidative stress balance and GRP78−p85α−Akt−GTP-Rac1−NF-κB−MMP-2-mediated cancer cell invasion.

Induction of the generation of endoplasmic reticulum (ER) calcium (Ca++)-mediated reactive oxygen species (ROS) by gallic acid (GA) has been implicated in the mitochondrial apoptotic death of human oral cancer (OC) cells, but the molecular mechanism by which GA causes ER Ca++ release of OC cells to undergo cell death remains unclear. Here, we report that GA-induced phosphorylation of B-cell lymphoma 2 (BCL-2)-interacting killer (BIK) (threonine (Thr) 33/Serine (Ser) 35) and p53 (Ser 15 and Ser 392), Bcl-2-associated x protein (BAX)/BCL-2 antagonist killer 1 (BAK) oligomerization on the ER and mitochondria, rising of cytosolic Ca++ and ROS, cytochrome c (Cyt c) release from the mitochondria, Ψm loss, and apoptosis were suppressed in cells co-treated with a specific inhibitor of casein kinase II (CK II) (4,5,6,7-tetrabromobenzotriazole). Small interfering RNA (siRNA)-mediated suppression of BIK inhibited GA-induced oligomeric complex of BAX/BAK in the ER and mitochondria, increase of cytosolic Ca++ and ROS, and apoptosis, but did not attenuate the increase in the level of Ser 15-phosphated p53 induced by GA. Blockade of p53 expression by short hairpin RNA suppressed BAX/BAK oligomerization and ER Ca++–ROS-associated apoptosis induced by GA but did not affect GA-induced phospho-BIK (Thr 33/Ser 35) levels. Induction of mitochondrial Cyt c release and ROS generation, increased cytosolic Ca++ level, and apoptosis by GA was attenuated by expression of the BAX or BAK siRNA. Over-expression of BCL-2 (but not BCL-XL) inhibited formation of ER oligomeric BAX/BAK by GA. Our results demonstrated that activation of the CK II by GA is required for the BIK-mediated ROS-dependent apoptotic activity of ER-associated BAX/BAK.

Background/Aim: Matrix metalloproteinase-2 (MMP-2) has been implicated in the pathogenesis of breast cancer (BC). However, there is limited research on the role of MMP-2 genotypes in BC risk. This study aimed to investigate the associations between two MMP-2 promoter polymorphisms, rs243865 and rs2285053, and BC risk. Materials and Methods: MMP-2 genotypes were analyzed using PCR-based RFLP methodology in a cohort comprising 1,232 BC cases and 1,232 controls. Results: Genotypic frequencies of MMP-2 rs243865 and rs2285053 in controls were consistent with Hardy-Weinberg equilibrium (p=0.3702 and 0.2036, respectively). There were no significant differences in the distribution of rs243865 and rs2285053 genotypes between BC cases and controls (p for trend=0.1602 and 0.2170, respectively). Variant genotypes at rs243865 and rs2285053 appeared to confer a protective effect, although not statistically significant (all p>0.05). Similarly, the variant T allele at rs243865 and rs2285053 showed a non-significant trend towards decreased BC risk (OR=0.84 and 0.89, 95%CI=0.69-1.02 and 0.78-1.02, p=0.0811 and 0.1043, respectively). There was no interaction observed between MMP-2 rs243865 or rs2285053 genotypes and age. Stratified analysis did not reveal significant associations between MMP-2 rs243865 or rs2285053 genotypes and triple-negative breast cancer (TNBC) (p=0.6458 and 0.8745, respectively). Among both TNBC and non-TNBC cases, none of the variant genotypes at rs243865 or rs2285053 showed significant associations with TNBC (all p>0.05). Conclusion: MMP-2 rs243865 and rs2285053 genotypes appear to have a minimal impact on individual susceptibility to BC or TNBC. [ABSTRACT FROM AUTHOR]

Induction of the generation of endoplasmic reticulum (ER) calcium (Ca++)-mediated reactive oxygen species (ROS) by gallic acid (GA) has been implicated in the mitochondrial apoptotic death of human oral cancer (OC) cells, but the molecular mechanism by which GA causes ER Ca++ release of OC cells to undergo cell death remains unclear. Here, we report that GA-induced phosphorylation of B-cell lymphoma 2 (BCL-2)-interacting killer (BIK) (threonine (Thr) 33/Serine (Ser) 35) and p53 (Ser 15 and Ser 392), Bcl-2-associated x protein (BAX)/BCL-2 antagonist killer 1 (BAK) oligomerization on the ER andmitochondria, rising of cytosolic Ca++ and ROS, cytochrome c (Cyt c) release from the mitochondria, ψm loss, and apoptosis were suppressed in cells co-treated with a specific inhibitor of casein kinase II (CK II) (4,5,6,7-tetrabromobenzotriazole). Small interfering RNA (siRNA)-mediated suppression of BIK inhibited GA-induced oligomeric complex of BAX/BAK in the ER and mitochondria, increase of cytosolic Ca++ and ROS, and apoptosis, but did not attenuate the increase in the level of Ser 15-phosphated p53 induced by GA. Blockade of p53 expression by short hairpin RNA suppressed BAX/BAK oligomerization and ER Ca++--ROS-associated apoptosis induced by GA but did not affect GA-induced phospho-BIK (Thr 33/Ser 35) levels. Induction of mitochondrial Cyt c release and ROS generation, increased cytosolic Ca++ level, and apoptosis by GA was attenuated by expression of the BAX or BAK siRNA. Over-expression of BCL-2 (but not BCL-XL) inhibited formation of ER oligomeric BAX/BAK by GA. Our results demonstrated that activation of the CK II by GA is required for the BIK-mediated ROS-dependent apoptotic activity of ER-associated BAX/BAK. [ABSTRACT FROM AUTHOR]

Mice deficient in hepatocyte nuclear factor 1 α (HNF-1α) develop dwarfism, liver dysfunction, and type 2 diabetes mellitus. Liver dysfunction in HNF-1α-null mice includes severe hepatic glycogen accumulation and dyslipidemia. The liver dysfunction may appear as soon as 2 weeks after birth. Since the HNF-1α-null mice become diabetic 2 weeks after birth, the early onset of the liver dysfunction is unlikely to be due to the diabetic status of the mice. More likely, it is due directly to the deficiency of HNF-1α in liver. Although the HNF-1α-null mice have an average life span of 1 year, the severe liver phenotype has thwarted attempts to study the pathogenesis of maturity-onset diabetes of the young type 3 (MODY3) and to examine therapeutic strategies for diabetes prevention and treatment in these mice. To circumvent this problem, we have generated a new Hnf-1α mutant mouse line, Hnf-1α[sup kin/kin], using gene targeting to inactivate the Hnf-1α gene and at the same time, to incorporate the Cre-loxP DNA recombination system into the locus for later revival of the Hnf-1α gene in tissues by tissue-specifically expressed Cre recombinase. The Hnf-1α[sup kin/kin] mice in which the expression of HNF-1α was inactivated in germ line cells were indistinguishable from the HNF-1α-null mice with regard to both the diabetes and liver phenotypes. Intriguingly, when the inactivated Hnf-1α gene was revived in liver (hepatic Hnf-1α revived) by the Cre recombinase driven by an albumin promoter, the Hnf-1α[sup kin/kin] mice, although severely diabetic, grew normally and did not develop any of the liver dysfunctions. In addition, we showed that the expression of numerous genes in pancreas, including a marker gene for pancreas injury, was affected by liver dysfunction but not by the deficiency of HNF-1α in pancreas. Thus, our hepatic-Hnf-1α-revived mice may serve as a useful... [ABSTRACT FROM AUTHOR]