Does Microbiome Contribute to Longevity? Compositional and Functional Differences in Gut Microbiota in Chinese Long-Living (>90 Years) and Elderly (65-74 Years) Adults.

The study of longevity and its determinants has been revitalized with the rise of microbiome scholarship. The gut microbiota have been established to play essential protective, metabolic, and physiological roles in human health and disease. The gut dysbiosis has been identified as an important factor contributing to the development of multiple diseases. Accordingly, it is reasonable to hypothesize that the gut microbiota of long-living individuals have healthy antiaging-associated gut microbes, which, by extension, might provide specific molecular targets for antiaging treatments and interventions. In the present study, we compared the gut microbiota of Chinese individuals in two different age groups, long-living adults (aged over 90 years) and elderly adults (aged 65-74 years) who were free of major diseases. We found significantly lower relative abundances of bacteria in the genera Sutterella and Megamonas in the long-living individuals. Furthermore, we established that while biological processes such as autophagy (GO:0006914) and telomere maintenance through semiconservative replication (GO:0032201) were enhanced in the long-living group, response to lipopolysaccharide (GO:0032496), nicotinamide adenine dinucleotide oxidation (GO:0006116), and S-adenosyl methionine metabolism (GO:0046500) were weakened. Moreover, the two groups were found to differ with respect to amino acid metabolism. We suggest that these compositional and functional differences in the gut microbiota may potentially be associated with mechanisms that contribute to determining longevity or aging.

METTL3 regulates N6-methyladenosine modification of ANGPTL3 mRNA and potentiates malignant progression of stomach adenocarcinoma.

BACKGROUND: N6-methyladenosine (m6A) is associated with mammalian mRNA biogenesis, decay, translation and metabolism, and also contributes greatly to gastrointestinal tumor formation and development. Therefore, the specific mechanisms and signaling pathways mediated by methyltransferase-like 3 (METTL3), which catalyzes the formation of m6A chemical labeling in stomach adenocarcinoma (STAD), are still worth exploring. METHODS: Quantitative real-time PCR (qRT-PCR) was constructed to detect the expression of METTL3 in gastric cancer cell lines and patient tissues. The biological function of METTL3 was investigated in vitro/in vivo by Cell Counting Kit-8, colony formation assay, Transwell assay and nude mouse tumorigenesis assay. Based on the LinkedOmics database, the genes co-expressed with METTL3 in the TCGA STAD cohort were analyzed to clarify the downstream targets of METTL3. Methylated RNA immunoprecipitation-qPCR (MeRIP-qPCR) and RNA stability analysis were employed to explore the mechanism of METTL3 in gastric cancer progression. RESULTS: We analyzed TCGA data and found that METTL3 was frequently elevated in STAD, and demonstrated that METTL3 was present at high levels in clinical STAD tissues and cells. High METTL3 expression was more likely to have advanced TNM tumors and distant metastasis. On the other hand, METTL3 silencing effectively impeded the higher oncogenic capacity of AGS and HGC27 cells in vivo and in vitro, as reflected by slowed cell growth and diminished migration and invasion capacities. Continued mining of the TCGA dataset identified the co-expression of angiopoietin-like 3 (ANGPTL3) and METTL3 in STAD. Lower level of ANGPTL3 was related to increased level of METTL3 in STAD samples and shorter survival times in STAD patients. ANGPTL3 enrichment limited the growth and metastasis of STAD cells. Besides, ANGPTL3 mRNA levels could be decreased by METTL3-dominated m6A modifications, a result derived from a combination of MeRIP-qPCR and RNA half-life experiments. Importantly, the inhibitory effect of METTL3 silencing on cancer could be reversed to some extent by ANGPTL3 inhibition. CONCLUSIONS: Overall, our findings suggested that METTL3 functioned an oncogenic role in STAD by reducing ANGPTL3 expression in an m6A-dependent manner. The discovery of the METTL3-ANGPTL3 axis and its effect on STAD tumor growth will contribute to further studies on the mechanisms of gastric adenocarcinoma development.

Angiopoietin-like protein 3 promotes colorectal cancer progression and liver metastasis partly via the mitogen-activated protein kinase 14 pathway.

Colorectal cancer (CRC) remains one of the most common malignancies worldwide, and liver metastasis represents a considerable challenge during CRC treatment. Aberrant expression of angiopoietin-like protein 3 (ANGPTL3) has been reported in several human cancer types. However, the function and mechanism of ANGPTL3 in CRC remain unclear. In this study, we first explored ANGPTL3 expression profiles in CRC datasets from ONCOMINE and in local samples from patients with CRC. We then elucidated the function of ANGPTL3 via knockdown and overexpression experiments. Bioinformatic analyses were performed to investigate the biological function and associated molecular mechanisms of ANGPTL3 in CRC oncogenesis and development. Finally, a xenograft model of liver metastasis was used to determine the role of ANGPTL3 in CRC metastasis. Our findings indicated that ANGPTL3 expression was upregulated in human CRC tissues, with high ANGPTL3 expression significantly correlated with poor survival of patients with CRC. ANGPTL3 overexpression promoted the proliferation and migration of CRC cells partially through mitogen-activated protein kinase 14 (MAPK14), while ANGPTL3 silencing had the opposite effect. Moreover, ANGPTL3 downregulation suppressed tumor growth and liver metastasis in xenograft mice. Collectively, the results presented here indicate that ANGPTL3 promotes cell proliferation and liver metastasis partly via MAPK14, suggesting that ANGPTL3 plays a tumor-promoting role in CRC progression and thus may represent a therapeutic target for CRC treatment.

The Role of Plant Progesterone in Regulating Growth, Development, and Biotic/Abiotic Stress Responses.

Progesterone is a steroid hormone that performs important functions in mammals. However, studies on its physiological functions in plants have gradually increased in recent years. Therefore, this review summarizes the regulatory functions of progesterone on plant growth and development, as well as its response to stress. Moreover, the plant metabolic processes of progesterone are also discussed. Overall, progesterone is ubiquitous in plants and can regulate numerous plant physiological processes at low concentrations. Since progesterone shares similar characteristics with plant hormones, it is expected to become a candidate for plant hormone. However, most of the current research on progesterone in plants is limited to the physiological level, and more molecular level research is needed to clarify progesterone signaling pathways.

Coxsackievirus A2 Leads to Heart Injury in a Neonatal Mouse Model.

Coxsackievirus A2 (CVA2) has emerged as an active pathogen that has been implicated in hand, foot, and mouth disease (HFMD) and herpangina outbreaks worldwide. It has been reported that severe cases with CVA2 infection develop into heart injury, which may be one of the causes of death. However, the mechanisms of CVA2-induced heart injury have not been well understood. In this study, we used a neonatal mouse model of CVA2 to investigate the possible mechanisms of heart injury. We detected CVA2 replication and apoptosis in heart tissues from infected mice. The activity of total aspartate transaminase (AST) and lactate dehydrogenase (LDH) was notably increased in heart tissues from infected mice. CVA2 infection also led to the disruption of cell-matrix interactions in heart tissues, including the increases of matrix metalloproteinase (MMP)3, MMP8, MMP9, connective tissue growth factor (CTGF) and tissue inhibitors of metalloproteinases (TIMP)4. Infiltrating leukocytes (CD45+ and CD11b+ cells) were observed in heart tissues of infected mice. Correspondingly, the expression levels of inflammatory cytokines in tissue lysates of hearts, including tumor necrosis factor alpha (TNF-α), interleukin-1beta (IL-1ß), IL6 and monocyte chemoattractant protein-1 (MCP-1) were significantly elevated in CVA2 infected mice. Inflammatory signal pathways in heart tissues, including phosphatidylinositol 3-kinase (PI3K)-AKT, mitogen-activated protein kinases (MAPK) and nuclear factor kappa B (NF-κB), were also activated after infection. In summary, CVA2 infection leads to heart injury in a neonatal mouse model, which might be related to viral replication, increased expression levels of MMP-related enzymes and excessive inflammatory responses.

CPNE1 Enhances Colorectal Cancer Cell Growth, Glycolysis, and Drug Resistance Through Regulating the AKT-GLUT1/HK2 Pathway.

INTRODUCTION: Colorectal cancer (CRC) is a major cause of cancer-related mortality worldwide. Copines-1 (CPNE1) has been shown to be overexpressed in various cancers; however, the role of CPNE1 in CRC remains unknown. Therefore, it is of great importance to elucidate the role of CPNE1 in CRC and its underlying mechanism of action. METHODS: CPNE1 expression in CRC tissues was measured by quantitative real-time PCR and immunohistochemical (IHC) staining. CPNE1 was knocked down (KD) or overexpressed using small inferring RNAs or lentiviral transduction in CRC cells. The proliferation, apoptosis, glycolysis, and mitochondrial respiration of CRC cells were assessed by cell counting kit-8, flow cytometry, and Xfe24 extracellular flux analyzer assays, respectively. The role of CPNE1 in tumor growth and chemoresistance was further confirmed in xenograft and patient-derived tumor xenograft models, respectively. RESULTS: CPNE1 mRNA and protein were upregulated in CRC tissues. CPNE1 promoted proliferation, inhibited apoptosis, increased mitochondrial respiration, enhanced aerobic glycolysis by activating AKT signaling, upregulated glucose transporter 1 (GLUT1) and hexokinase 2 (HK2), and downregulated the production of cleaved Caspase-3 (c-Caspase 3). CPNE1 also contributed to chemoresistance in CRC cells. CPNE1 KD inhibited tumor growth and increased the sensitivity of tumors to oxaliplatin in vivo. CONCLUSION: CPNE1 promotes CRC progression by activating the AKT-GLUT1/HK2 cascade and enhances chemoresistance.

Effect and mechanism of the long noncoding RNA MALAT1 on retinal neovascularization in retinopathy of prematurity.

AIMS: The most typical pathological manifestation of retinopathy of prematurity (ROP) is Retinal neovascularization (RNV). Long noncoding RNA (lncRNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) has been reported to mediate angiogenesis. Our experiment aimed to research the effect and mechanism of the MALAT1 on RNV in ROP. MAIN METHODS: C57 mice was used to establish oxygen-introduced retinopathy (OIR), and divided into control, hyperoxia, hyperoxia control siRNA, and hyperoxia MALAT1 siRNA groups. KEY FINDINGS: It was shown that MALAT1 mRNA was high expressed in the retinas of OIR mice. Further studies revealed that after intravitreal injection of MALAT1 siRNA, the degree of retinopathy was significantly reduced compared with OIR group. In addition, the protein and mRNA expression levels of CCN1, AKT and VEGF were significantly decreased. This was accompanied by a decrease in inflammatory genes including IL-1ß, IL-6, and TNF-α compared with the hyperoxia control siRNA mice. SIGNIFICANCE: The result suggested that MALAT1 may be involved in the process of RNV in ROP and MALAT1 siRNA may be a promising agent for the treatment of ROP by inhibiting RNV.

CircNOL10 Acts as a Sponge of miR-135a/b-5p in Suppressing Colorectal Cancer Progression via Regulating KLF9.

BACKGROUND: Circular RNAs (circRNAs) have been documented as key regulators during progression of malignant human cancer, including colorectal cancer (CRC). However, the underlying molecular mechanisms of circNOL10 in CRC remain unclear. METHODS: The real-time quantitative polymerase chain reaction was used to quantify the expression of circNOL10, miR-135a-5p, miR-135b-5p, and Krüppel-like factor 9 (KLF9). Kaplan-Meier curve was employed to assess the relationship between survival time of CRC patients and expression level of circNOL10. Cell ability of proliferation was measured by Cell Counting Kit8 and colony formation assays. Cell-cycle analysis was performed using flow cytometry assay. In addition, migration and invasion of CRC cell were examined with transwell analysis. The protein expression level was measured with Western blot assay. The interaction relationship of different molecules was analyzed by bioinformatics database and confirmed by dual-luciferase reporter, RNA immunoprecipitation, and RNA pulldown assay. The functional role of circNOL10 in vivo was determined by xenograft experiment. RESULTS: CircNOL10 was decreased in CRC tissues and cells and was associated with poor outcomes. Gain-of-functional experiment revealed that overexpression of circNOL10 constrained proliferation, cell-cycle progression, migration, and invasion of CRC cells, which was abolished by overexpression of miR-135a-5p or miR-135b-5p. Additionally, miR-135a-5p and miR-135b-5p, targets of circNOL10, regulated KLF9 expression in a negative feedback. Consistently, the results of xenograft experiment suggested that overexpression of circNOL10 inhibited tumor growth in vivo. CONCLUSION: In summary, our results showed that circNOL10 impeded CRC development by mediating proliferation, cell cycle, migration, and invasion by sponging miR-135a-5p and miR-135b-5p, which provided new understanding for CRC treatment.

FGF9 promotes cisplatin resistance in colorectal cancer via regulation of Wnt/ß-catenin signaling pathway.

Development of cisplatin resistance in colorectal cancer is largely caused by dysregulation of signaling pathways, including the Wnt/ß-catenin signaling pathway, in cancer cells. Further investigation into the molecular mechanism of chemoresistance could improve outcomes for patients with colorectal cancer. The present study determined that fibroblast growth factor 9 (FGF9) was overexpressed in tumor tissues compared with normal tissues from patients with colorectal cancer. Using the colorectal cancer cell line LoVo, transfection of recombinant FGF9 decreased cisplatin-induced cell apoptosis whilst FGF9 silencing increased cisplatin-induced apoptosis. Western blot analysis and reverse transcription-quantitative polymerase chain reaction demonstrated that FGF9 decreased adenomatous polyposis coli (APC) mRNA and protein expression and contributed to activation of the Wnt/ß-catenin signaling pathway. Notably, an increase in FGF9 and ß-catenin protein expression and a decrease in APC protein expression was observed in the established LoVo cisplatin resistant cell line (LoVo/cisplatin). Silencing of FGF9 reversed cisplatin resistance of LoVo/cisplatin cells. In conclusion, the present findings suggested that FGF9 activated the Wnt signaling pathway and was a mediator of cisplatin resistance in colorectal cancer.

Nuclear factor κB -inducing kinase is a diagnostic marker of gastric cancer.

Nuclear factor-κB-inducing kinase (NIK) is a new regulator of nuclear factor-κB signaling, which plays an important role in tumorigenesis. This study aimed to examine the expression of NIK in gastric cancer and investigate its clinical significance.Tumor issues were collected from 80 gastric cancer patients who received surgery and the diagnosis was confirmed by postoperative pathological analysis. The expression of NIK in gastric cancer tissues and adjacent normal mucosa was detected by immunohistochemical analysis. The associations between NIK expression and clinicopathological features of the patients were further analyzed.NIK expression was significantly higher in gastric cancer tissues than in adjacent normal tissues (P < .05). Furthermore, NIK expression showed significant association with UICC stage, T status, and differentiation, but not with age and gender of gastric cancer patients.NIK is overexpressed in gastric cancer and is a potential diagnostic marker of gastric cancer.

Postnatal development of mice with combined genetic depletions of lamin A/C, emerin and lamina-associated polypeptide 1.

Mutations in LMNA encoding lamin A/C and EMD encoding emerin cause cardiomyopathy and muscular dystrophy. Lmna null mice develop these disorders and have a lifespan of 7-8 weeks. Emd null mice show no overt pathology and have normal skeletal muscle but with regeneration defects. We generated mice with germline deletions of both Lmna and Emd to determine the effects of combined loss of the encoded proteins. Mice without lamin A/C and emerin are born at the expected Mendelian ratio, are grossly normal at birth but have shorter lifespans than those lacking only lamin A/C. However, there are no major differences between these mice with regards to left ventricular function, heart ultrastructure or electrocardiographic parameters except for slower heart rates in the mice lacking both lamin A/C and emerin. Skeletal muscle is similarly affected in both of these mice. Lmna+/- mice also lacking emerin live to at least 1 year and have no significant differences in growth, heart or skeletal muscle compared to Lmna+/- mice. Deletion of the mouse gene encoding lamina-associated protein 1 leads to prenatal death; however, mice with heterozygous deletion of this gene lacking both lamin A/C and emerin are born at the expected Mendelian ratio but had a shorter lifespan than those only lacking lamin A/C and emerin. These results show that mice with combined deficiencies of three interacting nuclear envelope proteins have normal embryonic development and that early postnatal defects are primarily driven by loss of lamin A/C or lamina-associated polypeptide 1 rather than emerin.

Imbalanced nucleocytoskeletal connections create common polarity defects in progeria and physiological aging.

Studies of the accelerated aging disorder Hutchinson-Gilford progeria syndrome (HGPS) can potentially reveal cellular defects associated with physiological aging. HGPS results from expression and abnormal nuclear envelope association of a farnesylated, truncated variant of prelamin A called "progerin." We surveyed the diffusional mobilities of nuclear membrane proteins to identify proximal effects of progerin expression. The mobilities of three proteins-SUN2, nesprin-2G, and emerin-were reduced in fibroblasts from children with HGPS compared with those in normal fibroblasts. These proteins function together in nuclear movement and centrosome orientation in fibroblasts polarizing for migration. Both processes were impaired in fibroblasts from children with HGPS and in NIH 3T3 fibroblasts expressing progerin, but were restored by inhibiting protein farnesylation. Progerin affected both the coupling of the nucleus to actin cables and the oriented flow of the cables necessary for nuclear movement and centrosome orientation. Progerin overexpression increased levels of SUN1, which couples the nucleus to microtubules through nesprin-2G and dynein, and microtubule association with the nucleus. Reducing microtubule-nuclear connections through SUN1 depletion or dynein inhibition rescued the polarity defects. Nuclear movement and centrosome orientation were also defective in fibroblasts from normal individuals over 60 y, and both defects were rescued by reducing the increased level of SUN1 in these cells or inhibiting dynein. Our results identify imbalanced nuclear engagement of the cytoskeleton (microtubules: high; actin filaments: low) as the basis for intrinsic cell polarity defects in HGPS and physiological aging and suggest that rebalancing the connections can ameliorate the defects.

TorsinA controls TAN line assembly and the retrograde flow of dorsal perinuclear actin cables during rearward nuclear movement.

The nucleus is positioned toward the rear of most migratory cells. In fibroblasts and myoblasts polarizing for migration, retrograde actin flow moves the nucleus rearward, resulting in the orientation of the centrosome in the direction of migration. In this study, we report that the nuclear envelope-localized AAA+ (ATPase associated with various cellular activities) torsinA (TA) and its activator, the inner nuclear membrane protein lamina-associated polypeptide 1 (LAP1), are required for rearward nuclear movement during centrosome orientation in migrating fibroblasts. Both TA and LAP1 contributed to the assembly of transmembrane actin-associated nuclear (TAN) lines, which couple the nucleus to dorsal perinuclear actin cables undergoing retrograde flow. In addition, TA localized to TAN lines and was necessary for the proper mobility of EGFP-mini-nesprin-2G, a functional TAN line reporter construct, within the nuclear envelope. Furthermore, TA and LAP1 were indispensable for the retrograde flow of dorsal perinuclear actin cables, supporting the recently proposed function for the nucleus in spatially organizing actin flow and cytoplasmic polarity. Collectively, these results identify TA as a key regulator of actin-dependent rearward nuclear movement during centrosome orientation.

Lamina-associated polypeptide 1 is dispensable for embryonic myogenesis but required for postnatal skeletal muscle growth.

Lamina-associated polypeptide 1 (LAP1) is an integral protein of the inner nuclear membrane that has been implicated in striated muscle maintenance. Mutations in its gene have been linked to muscular dystrophy and cardiomyopathy. As germline deletion of the gene encoding LAP1 is perinatal lethal, we explored its potential role in myogenic differentiation and development by generating a conditional knockout mouse in which the protein is depleted from muscle progenitors at embryonic day 8.5 (Myf5-Lap1CKO mice). Although cultured myoblasts lacking LAP1 demonstrated defective terminal differentiation and altered expression of muscle regulatory factors, embryonic myogenesis and formation of skeletal muscle occurred in both mice with a Lap1 germline deletion and Myf5-Lap1CKO mice. However, skeletal muscle fibres were hypotrophic and their nuclei were morphologically abnormal with a wider perinuclear space than normal myonuclei. Myf5-Lap1CKO mouse skeletal muscle contained fewer satellite cells than normal and these cells had evidence of reduced myogenic potential. Abnormalities in signalling pathways required for postnatal hypertrophic growth were also observed in skeletal muscles of these mice. Our results demonstrate that early embryonic depletion of LAP1 does not impair myogenesis but that it is necessary for postnatal skeletal muscle growth.

Protein-Bound Polysaccharide from Corbicula fluminea Inhibits Cell Growth in MCF-7 and MDA-MB-231 Human Breast Cancer Cells.

A novel protein-bound polysaccharide, CFPS-1, isolated from Corbicula fluminea, is composed predominantly of mannose (Man) and glucose (Glc) in a molar ratio of 3.1:12.7. The polysaccharide, with an average molecular weight of about 283 kDa, also contains 10.8% protein. Atomic force microscopy, high-performance liquid chromatography, Fourier transform infrared spectroscopy, gas chromatography/mass spectrometry, and nuclear magnetic resonance spectroscopy analyses revealed that CFPS-1 has a backbone of 1,6-linked and 1,4,6-linked-α-D-Glc, which is terminated with a 1-linked-α-D-Man residue at the O-4 position of 1,4,6-linked-α-D-Glc, in a molar ratio of 3:1:1. Preliminary in vitro bioactivity tests revealed that CFPS-1 effectively and dose-dependently inhibits human breast cancer MCF-7 and MDA-MB-231 cell growth, with an IC50 of 243 ± 6.79 and 1142 ± 14.84 µg/mL, respectively. In MCF-7, CFPS-1 produced a significant up-regulation of p53, p21, Bax and cleaved caspase-7 and down-regulation of Cdk4, cyclin D1, Bcl-2 and caspase-7. These effects resulted in cell cycle blockade at the S-phase and apoptosis induction. In contrast, in MDA-MB-231, with limited degree of change in cell cycle distribution, CFPS-1 increases the proportion of cells in apoptotic sub-G1 phase executed by down-regulation of Bcl-2 and caspase-7 and up-regulation of Bax and cleaved caspase-7. This study extends our understanding of the anticancer mechanism of C. fluminea protein-bound polysaccharide.

Perfluorooctane Sulfonate Induces Autophagy-Dependent Apoptosis through Spinster 1-Mediated lysosomal-Mitochondrial Axis and Impaired Mitophagy.

Lysosomal membrane permeabilization (LMP) and subsequently impaired autophagosome degradation was induced in HepG2 cells after treatment with perfluorooctane sulfonate (PFOS) for 24 h in our previous studies. We found that treatment of HepG2 cells with PFOS-induced autophagosome formation at earlier stage (6 h) of treatment in this study. The autophagosome formation inhibitor 3-methyladenine (3-MA) was able to relieve PFOS-induced LMP and release of cathepsin D in HepG2 cells. Knockdown of Spinster 1, a lysosomal membrane permease, attenuated PFOS-induced LMP in HepG2 cells. We proposed that Spinster 1 might work as a specific molecule that linked autophagy with LMP. PFOS-induced collapse of mitochondrial transmembrane potential was cathepsin D and autophagy dependent. Addition of 3-MA relieved PFOS-induced apoptosis, which was evidenced by Hoechst assay, AV/PI staining and caspase-3 activity assay. Inhibition of autophagosome formation by Atg5 siRNA attenuated PFOS-induced apoptosis. Treatment of HepG2 cells with PFOS for 24 h impaired mitophagy, as evidenced by an increase of cells with giant mitochondria and impairment of colocalization of PINK1 with light chain 3. In summary, we report that PFOS induces autophagy-dependent apoptosis in HepG2 cells through the lysosomal-mitochondrial axis and impairment of mitophagy, suggesting that autophagy is a primary target for PFOS toxicity. These findings provide new mechanistic insights into PFOS-induced hepatotoxicity.

A mutation abolishing the ZMPSTE24 cleavage site in prelamin A causes a progeroid disorder.

In 1994 in the Journal of Cell Science, Hennekes and Nigg reported that changing valine to arginine at the endoproteolytic cleavage site in chicken prelamin A abolishes its conversion to lamin A. The consequences of this mutation in an organism have remained unknown. We now report that the corresponding mutation in a human subject leads to accumulation of prelamin A and causes a progeroid disorder. Next generation sequencing of the subject and her parents' exomes identified a de novo mutation in the lamin A/C gene (LMNA) that resulted in a leucine to arginine amino acid substitution at residue 647 in prelamin A. The subject's fibroblasts accumulated prelamin A, a farnesylated protein, which led to an increased percentage of cultured cells with morphologically abnormal nuclei. Treatment with a protein farnesyltransferase inhibitor improved abnormal nuclear morphology. This case demonstrates that accumulation of prelamin A, independent of the loss of function of ZMPSTE24 metallopeptidase that catalyzes processing of prelamin A, can cause a progeroid disorder and that a cell biology assay could be used in precision medicine to identify a potential therapy.

Citreoviridin Induces Autophagy-Dependent Apoptosis through Lysosomal-Mitochondrial Axis in Human Liver HepG2 Cells.

Citreoviridin (CIT) is a mycotoxin derived from fungal species in moldy cereals. In our previous study, we reported that CIT stimulated autophagosome formation in human liver HepG2 cells. Here, we aimed to explore the relationship of autophagy with lysosomal membrane permeabilization and apoptosis in CIT-treated cells. Our data showed that CIT increased the expression of LC3-II, an autophagosome biomarker, from the early stage of treatment (6 h). After treatment with CIT for 12 h, lysosomal membrane permeabilization occurred, followed by the release of cathepsin D in HepG2 cells. Inhibition of autophagosome formation with siRNA against Atg5 attenuated CIT-induced lysosomal membrane permeabilization. In addition, CIT induced collapse of mitochondrial transmembrane potential as assessed by JC-1 staining. Furthermore, caspase-3 activity assay showed that CIT induced apoptosis in HepG2 cells. Inhibition of autophagosome formation attenuated CIT-induced apoptosis, indicating that CIT-induced apoptosis was autophagy-dependent. Cathepsin D inhibitor, pepstatin A, relieved CIT-induced apoptosis as well, suggesting the involvement of the lysosomal-mitochondrial axis in CIT-induced apoptosis. Taken together, our data demonstrated that CIT induced autophagy-dependent apoptosis through the lysosomal-mitochondrial axis in HepG2 cells. The study thus provides essential mechanistic insight, and suggests clues for the effective management and treatment of CIT-related diseases.

Citreoviridin induces ROS-dependent autophagic cell death in human liver HepG2 cells.

Citreoviridin (CIT) is one of toxic mycotoxins derived from fungal species in moldy cereals. Whether CIT exerts hepatotoxicity and the precise molecular mechanisms of CIT hepatotoxicity are not completely elucidated. In this study, the inhibitor of autophagosome formation, 3-methyladenine, protected the cells against CIT cytotoxicity, and the autophagy stimulator rapamycin further decreased the cell viability of CIT-treated HepG2 cells. Knockdown of Atg5 with Atg5 siRNA alleviated CIT-induced cell death. These finding suggested the hypothesis that autophagic cell death contributed to CIT-induced cytotoxicity in HepG2 cells. CIT increased the autophagosome number in HepG2 cells observed under a transmission electron microscope, and this effect was confirmed by the elevated LC3-II levels detected through Western blot. Reduction of P62 protein levels and the result of LC3 turnover assay indicated that the accumulation of autophagosomes in the CIT-treated HepG2 cells was due to increased formation rather than impaired degradation. The pretreatment of HepG2 cells with the ROS inhibitor NAC reduced autophagosome formation and reversed the CIT cytotoxicity, indicating that CIT-induced autophagic cell death was ROS-dependent. In summary, ROS-dependent autophagic cell death of HpeG2 cells described in this study may help to elucidate the underlying mechanism of CIT cytotoxicity.

Lamina-associated polypeptide-1 interacts with the muscular dystrophy protein emerin and is essential for skeletal muscle maintenance.

X-linked Emery-Dreifuss muscular dystrophy is caused by loss of function of emerin, an integral protein of the inner nuclear membrane. Yet emerin null mice are essentially normal, suggesting the existence of a critical compensating factor. We show that the lamina-associated polypeptide1 (LAP1) interacts with emerin. Conditional deletion of LAP1 from striated muscle causes muscular dystrophy; this pathology is worsened in the absence of emerin. LAP1 levels are significantly higher in mouse than human skeletal muscle, and reducing LAP1 by approximately half in mice also induces muscle abnormalities in emerin null mice. Conditional deletion of LAP1 from hepatocytes yields mice that exhibit normal liver function and are indistinguishable from littermate controls. These results establish that LAP1 interacts physically and functionally with emerin and plays an essential and selective role in skeletal muscle maintenance. They also highlight how dissecting differences between mouse and human phenotypes can provide fundamental insights into disease mechanisms.