The breast cancer susceptibility gene BRCA2 is required for the maintenance of telomere homeostasis.

Inactivating mutations in the breast cancer susceptibility gene BRCA2 cause gross chromosomal rearrangements. Chromosome structural instability in the absence of BRCA2 is thought to result from defective homology-directed DNA repair. Here, we show that BRCA2 links the fidelity of telomere maintenance with genetic integrity. Absence of BRCA2 resulted in signs of dysfunctional telomeres, such as telomere shortening, erosions, and end fusions in proliferating mouse fibroblasts. BRCA2 localized to the telomeres in S phase in an ATR-dependent manner, and its absence resulted in the accumulation of common fragile sites, particularly at the G-rich lagging strand, and increased the telomere sister chromatid exchange in unchallenged cells. The incidence of common fragile sites and telomere sister chromatid exchange increased markedly after treatment with replication inhibitors. Congruently, telomere-induced foci were frequently observed in the absence of Brca2, denoting activation of the DNA damage response and abnormal chromosome end joining. These telomere end fusions constituted a significant portion of chromosome aberrations in Brca2-deficient cells. Our results suggest that BRCA2 is required for telomere homeostasis and may be particularly important for the replication of G-rich telomeric lagging strands.

Methylalpinumisoflavone inhibits lipopolysaccharide-induced inflammation in microglial cells by the NF-kappaB and MAPK signaling pathway.

Neuroinflammation is chronic inflammation within the brain that is attributed to prolonged activation of microglial cells and results in neurodegenerative events, such as neuronal dysfunction and neuronal loss. Therefore, suppression of neuroinflammation would theoretically slow progression of neurodegenerative disease. In this study, we investigated the anti-inflammatory effects of 4'-O-methylalpinumisoflavone (methylalpinumisoflavone), isolated from Cudrania tricuspidata, against LPS-induced microglial activation in BV2 cells. Exposure of BV2 cells to LPS (0.5 µg/mL) significantly increased production of pro-inflammatory mediators, including NO, PGE(2), and pro-inflammatory cytokines. Conversely, pre-treatment with methylalpinumisoflavone (10 and 20 µg/mL) prior to treatment with LPS resulted in a significant decrease of LPS-induced production of pro-inflammatory mediators in a dose-dependent manner. In addition, reduction of pro-inflammatory mediators by treatment with methylalpinumisoflavone prior to treatment with LPS was accompanied by a decrease in translocation of NF-κB p50 and p65 from the cytoplasm to the nucleus and by a decrease in activation of mitogen-activated protein kinases (MAPKs), such as ERK1/2 and JNK. Taken together, these results suggest that methylalpinumisoflavone suppressed LPS-induced microglial activation and production of pro-inflammatory mediators by decreasing NF-κB signaling and by phosphorylation of MAPKs. These results suggest the potential of methylalpinumisoflavone as an anti-inflammatory drug candidate.

Heat-Killed Lactiplantibacillus plantarum LRCC5314 Mitigates the Effects of Stress-Related Type 2 Diabetes in Mice via Gut Microbiome Modulation.

The incidence of stress-related type 2 diabetes (stress-T2D), which is aggravated by physiological stress, is increasing annually. The effects of Lactobacillus, a key component of probiotics, have been widely studied in diabetes; however, studies on the effects of postbiotics are still limited. Here, we aimed to examine the mechanism through which heat-killed Lactiplantibacillus plantarum LRCC5314 (HK-LRCC5314) alleviates stress-T2D in a cold-induced stress-T2D C57BL/6 mouse model. HK-LRCC5314 markedly decreased body weight gain, adipose tissue (neck, subcutaneous, and epididymal) weight, and fasting glucose levels. In the adipose tissue, mRNA expression levels of stress-T2D associated factors (NPY, Y2R, GLUT4, adiponectin, and leptin) and pro-inflammatory factors (TNF-α, IL-6, and CCL-2) were also altered. Furthermore, HK-LRCC5314 increased the abundance of Barnesiella, Alistipes, and butyrate-producing bacteria, including Akkermansia, in feces and decreased the abundance of Ruminococcus, Dorea, and Clostridium. Thus, these findings suggest that HK-LRCC5314 exerts protective effects against stress-T2D via gut microbiome modulation, suggesting its potential as a supplement for managing stress-T2D.

Brca2 abrogation engages with the alternative lengthening of telomeres via break-induced replication.

A subset of cancer cells maintains their telomeres without telomerase through the recombination-based alternative lengthening of telomeres (ALT) pathway. Currently, it is not yet clear in what context ALT is induced and how the pathway choice is made. Here, we show that abrogation of Brca2 reinforces break-induced replication (BIR) and engages with ALT pathway. Brca2 depletion in telomerase-null mouse cells alleviated the growth defect, accompanied by telomere elongation, suggesting the induction of ALT. We also found that Brca2-depleted telomerase-null cells exhibited dynamic clustering of telomeres from G2 phase in Promyelocytic Nuclear (PML) bodies. For Brca2-deficient ALT induction, Rad51 was dispensable but Mre11 and Rad52 were required. Congruently, conservative telomeric DNA synthesis was apparent in mitosis, indicating that the absence of Brca2 directed towards Rad52-mediated BIR. Collectively, we propose that Brca2 abrogation can instigate ALT tumourigenesis through the induction of BIR. This study implies that inhibitors of BIR may be useful for BRCA2-associated ALT-type cancers. Assessing ALT features may be considered for the tailored therapy of BRCA2-associated cancers.

Curcumin protected PC12 cells against beta-amyloid-induced toxicity through the inhibition of oxidative damage and tau hyperphosphorylation.

One of the pathological hallmarks of Alzheimer's disease is the progressive accumulation of beta-amyloid (Abeta) in the form of senile plaques, and Abeta insult to neuronal cells has been identified as one of the major causes of the onset of the disease. Curcumin, the major and most active antioxidant of Curcuma longa, protects neuronal cells against Abeta-induced toxicity. Therefore, in this study, we investigated the neuroprotective mechanisms by which curcumin acts against Abeta (25-35)-induced toxicity in PC12 cells. Following the exposure of PC12 cells to 10 microM Abeta (25-35) for 24h, significant increases in the level of antioxidant enzymes, and DNA damage were observed, and these increases were accompanied by a decrease in cell viability, and an increase in intracellular calcium levels and tau hyperphosphorylation. In addition, pretreatment of PC12 cells with 10 microg/ml curcumin for 1h significantly reversed the effect of Abeta, by decreasing the oxidative stress, and DNA damage induced by Abeta, as well as attenuating the elevation of intracellular calcium levels and tau hyperphosphorylation induced by Abeta. Taken together, these data indicate that curucmin protected PC12 cells against Abeta-induced neurotoxicity through the inhibition of oxidative damage, intracellular calcium influx, and tau hyperphosphorylation.

Panax ginseng effects on DNA damage, CYP1A1 expression and histopathological changes in testes of rats exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin.

The effects of Panax ginseng extracts on DNA damage, expression of cytochrome P450 (CYP) 1A1 and reproductive toxicity were evaluated in the testis of rats exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxinthe (TCDD). Fifty rats were divided into five groups according to treatment with 2,3,7,8-TCDD and P. ginseng extracts. Single cell gel electrophoresis assays were performed to evaluate DNA damage that occurred in the lymphocytes of rats. Histological changes in the seminiferous tubules of the testis were determined using Johnsen's scoring system and Real Time-PCR was performed to evaluate the mRNA expression of CYP1A1. Significant pathological effects were observed in the 2,3,7,8-TCDD treated rats including a reduced seminiferous tubular diameter, an increased number of damaged tubules (maturation arrest, eosinophilic degeneration and spermatid giant cells) and increased Johnsen's score. DNA damage and the expression of CYP1A1 mRNA were significantly increased in rat testes. There were no significant differences between the control and animals treated with P. ginseng extracts. However, a significantly decreased level of DNA damage, decreased CYP1A1 expression and reduced pathological effects were observed in the 2,3,7,8-TCDD with P. ginseng extracts treated groups when compared with the TCDD treated group. In summary, our study demonstrates that 2,3,7,8-TCDD induces the pathological and genotoxical damage in rat testes, while P. ginseng extract treatment exhibits a therapeutic capacity to reduce these effects via reduction of CYP1A1 mRNA.

Formaldehyde exposure induces airway inflammation by increasing eosinophil infiltrations through the regulation of reactive oxygen species production.

Formaldehyde (FA) is a well-known cytotoxic irritant to the airways, but the mechanism of airway inflammation due to FA has not been clarified. In the present study, C57BL/6 mice were exposed to two concentrations (5 and 10ppm) of FA for 6h/day, 5days/week, for 2 weeks. The FA-exposed mice had much higher number of CCR3(+) eosinophils than control mice, and showed upregulated gene expression of CC-chemokine receptor-3 (CCR3), eotaxin and intercellular adhesion molecules-1 (ICAM-1) as well as an increased expression of proinflammatory and Th2 cytokines, such as interleukin (IL)-1ß, IL-4 and IL-5. In addition, FA exposure revealed a considerable increase in the serum levels of IgG1, IgG3, IgA and IgE compared to controls. Histopathological analysis of the lung tissues demonstrated eosinophils and mononuclear cell infiltration of the alveolar cell walls and alveolar spaces. Gene expression of thioredoxin (TRX), redox-regulating antioxidant proteins, was markedly suppressed in FA-exposed mice, and thereby intracellular ROS levels were increased along with increased FA concentration. These results were consistent with an increase in the number of CCR3-expressing eosinophils, and indicate that FA-induced ROS was generated from eosinophils recruited to the inflammatory sites of the airways.

Paradoxical delay of senescence upon depletion of BRCA2 in telomerase-deficient worms.

BRCA2 is a multifunctional tumor suppressor involved in homologous recombination (HR), mitotic checkpoint regulation, and telomere homeostasis. Absence of Brca2 in mice results in progressive shortening of telomeres and senescence, yet cells are prone to neoplastic transformation with elongated telomeres, suggesting that BRCA2 has positive and negative effects on telomere length regulation along the path to tumorigenesis. Using Caenorhabditis elegans as a model, we show here that depletion of BRC-2, an ortholog of BRCA2, paradoxically delays senescence in telomerase-deficient mutant worms. Telomerase-deficient worms (trt-1) exhibit early replication senescence due to short telomeres. It should be noted that worms mutated in brc-2 are not viable as well due to massive genotoxic insults. However, when BRC-2 is depleted by RNA interference in trt-1 mutant worms, the number of generations is unexpectedly increased with telomere length maintained, compared to telomerase mutants. Interestingly, depletion of other HR genes such as rad-51 and rad-54 exhibited similar effects. In worms doubly deficient of telomerase and brc-2, rad-51, or rad-54, extra telomeric C-circles were generated, suggesting that abrogation of HR induces an alteration in telomere environment favorable to illegitimate telomere maintenance when telomerase is absent. Collectively, absence of BRC-2 in telomerase-deficient background first leads to telomere shortening, followed by an induction of an as-yet-unknown telomere maintenance pathway, resulting in delay of senescence. The results have implications in the understanding of dysfunctional BRCA2-associated tumorigenesis.

Isolation, sequencing, and characterization of the cytochrome bo operon from Vitreoscilla.

The entire operon encoding the sodium pumping cytochrome bo from the bacterium Vitreoscilla was isolated and sequenced, and this sequence was analyzed by blast and hydropathy plots. There are fairly similar phylogenetic relationships which apply to all five proteins, but overall greater similarity to members of the gamma subdivision than the beta subdivision of the Proteobacteria. Hydropathy plots of all five Cyo proteins show near identity with those of the corresponding E. coli subunits, indicating that the similarity extends from sequence to structure. The operon appears to have a typical Shine-Dalgarno sequence, an E. coli-like promoter, and several possible binding sites for regulatory proteins. The Vitreoscilla Cyo B subunit (the probable Na+ pump) is almost identical to E. coli Cyo B at 18 key amino acids; thus, there are no obvious changes in Vitreoscilla Cyo B that hint at the details of its Na+ pumping ability.

Gene expression profiling in lung tissues from rats exposed to formaldehyde.

Formaldehyde is a ubiquitous toxic organic compound recently classified as a carcinogen by the International Agency for Research on Cancer and one of the major factors causing sick building syndrome. In this study, we have investigated the effects of formaldehyde on mRNA expression in rat lung tissues by applying genomics. Rats were exposed to ambient air and two different concentrations of formaldehyde (0, 5, 10 ppm) for 2 weeks at 6 h/day and 5 days/week in an inhalation chamber. Malondialdehyde (MDA) assay and carbonyl spectrometric assay were conducted to determine lipid peroxidation and protein oxidation levels and Comet assays were used for genotoxicity evaluation. Level of MDA, carbonyl insertion and DNA damage in the lungs of rats exposed to FA were found to be dose dependently increased. Gene expression was evaluated by using a bio-array hybridization analysis. A total of 21 (2 up- and 19 down-regulated) genes were identified as biomarkers for formaldehyde effects. Several differentiated gene groups were found. Genes involved in apoptosis, immunity, metabolism, signal transduction, transportation, coagulation and oncogenesis were found to be up- and down-regulated. Among these genes, the mRNA expressions of cytochrome P450, hydroxymethylbilane synthase, glutathione reductase, carbonic anhydrase 2, natriuretic peptide receptor 3, lysosomal associated protein transmembrane 5, regulator of G-protein signaling 3, olfactomedin related ER localized protein, and poly (ADP-ribose) polymerase-1 were confirmed by quantitative RT-PCR analysis. In summary, the MDA lipid peroxidation and the carbonyl protein oxidation assays showed that cytotoxic effects increased with increasing formaldehyde levels. Genomic analysis showed that 21 genes were up- or down-regulated. Of these genes, nine were confirmed by quantitative RT-PCR and could be potential biomarkers for human diseases associated with formaldehyde exposure.