Burkholderia pseudomallei BipD modulates host mitophagy to evade killing.

Mitophagy is critical for mitochondrial quality control and function to clear damaged mitochondria. Here, we found that Burkholderia pseudomallei maneuvered host mitophagy for its intracellular survival through the type III secretion system needle tip protein BipD. We identified BipD, interacting with BTB-containing proteins KLHL9 and KLHL13 by binding to the Back and Kelch domains, recruited NEDD8 family RING E3 ligase CUL3 in response to B. pseudomallei infection. Although evidently not involved in regulation of infectious diseases, KLHL9/KLHL13/CUL3 E3 ligase complex was essential for BipD-dependent ubiquitination of mitochondria in mouse macrophages. Mechanistically, we discovered the inner mitochondrial membrane IMMT via host ubiquitome profiling as a substrate of KLHL9/KLHL13/CUL3 complex. Notably, K63-linked ubiquitination of IMMT K211 was required for initiating host mitophagy, thereby reducing mitochondrial ROS production. Here, we show a unique mechanism used by bacterial pathogens that hijacks host mitophagy for their survival.

High-Temperature Chlorination of Nickel Oxide Using Calcium Chloride.

Attempts have been made to extract nickel from ores and nickel-containing wastes using the chlorination method. However, the use of gaseous chlorinating agents is limited due to their toxicity. High-temperature chlorination of nickel oxide using calcium chloride is analyzed in this study. The volatilization percentage is positively correlated to temperature and CaCl2 dosage and negatively correlated to oxygen partial pressure. The apparent activation energy is calculated to be 142.91 kJ/mol, between 1173 K and 1323 K, which suggests that the high-temperature chlorination of nickel oxide using calcium chloride is controlled by a chemical reaction.

Citrus-derived DHCP inhibits mitochondrial complex II to enhance TRAIL sensitivity via ROS-induced DR5 upregulation.

Heat-modified citrus pectin, a water-soluble indigestible polysaccharide fiber derived from citrus fruits and modified by temperature treatment, has been reported to exhibit anticancer effects. However, the bioactive fractions and their mechanisms remain unclear. In this current study, we isolated an active compound, trans-4,5-dihydroxy-2-cyclopentene-l-one (DHCP), from heat-treated citrus pectin, and found that is induces cell death in colon cancer cells via induction of mitochondrial ROS. On the molecular level, DHCP triggers ROS production by inhibiting the activity of succinate ubiquinone reductase (SQR) in mitochondrial complex II. Furthermore, cytotoxicity, apoptotic activity, and activation of caspase cascades were determined in HCT116 and HT-29 cell-based systems, the results indicated that DHCP enhances the sensitivity of cancer cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), with DHCP-induced ROS accounting for the synergistic effect between DHCP and TRAIL. Furthermore, the combination of DHCP and TRAIL inhibits the growth of HCT116 and HT-29 xenografts synergistically. ROS significantly increases the expression of TRAIL death receptor 5 (DR5) via the p53 and C/EBP homologous protein pathways. Collectively, our findings indicate that DHCP has a favorable toxicity profile and is a new TRAIL sensitizer that shows promise in the development of pectin-based pharmaceuticals, nutraceuticals, and dietary agents aimed at combating human colon cancer.

Superhydrophobic metal organic framework doped polycarbonate porous monolith for efficient selective removal oil from water.

A series of superhydrophobic polycarbonate porous monoliths modified with metal organic framework (Z8/PC) were firstly fabricated through a facile thermally impacted non-solvent induced phase separation method for efficient selective oil/water separation. The performance of the monoliths on oil/water separation was evaluated in terms of selectivity, equilibrium adsorption capacity, corrosion resistance, kinetics, and circulation. The results showed that the use of ZIF-8 significantly compensated for the shortage of pure monolith. Compared with pure PC monolith, the hydrophobic angle of the Z8/PC-2 monolith promoted from 136.18° to 154.25° due to the micro-nano flower surface. Meanwhile, the Z8/PC-2 monolith displayed a more intricate and continuous interconnected 3D hierarchical micro-nano structure, which possessed the monolith a higher specific surface area of 146.84 m2 g-1 and porosity of 89.5%. What's more, more superior oil/water separation abilities of Z8/PC-2 monolith were manifested by the selective removal of oil or organic solvent from water within 30s, high equilibrium adsorption capacity, and excellent corrosion resistance. In addition, the ten-cycle regeneration of porous monoliths via centrifugation or evaporation displayed additional attractiveness. Therefore, porous Z8/PC monolith will be a promising candidate for the efficient selective oil/water separation of oil spills and organic solvents.

A 3-O-methylated heterogalactan from Pleurotus eryngii activates macrophages.

Mushroom-derived polysaccharides exhibit various biological activities owing to their diverse structural features. Here, we purified a 3-O-methylated heterogalactan (WPEP-N-b, Mw 21.4 kDa) from the fruiting bodies of Pleurotus eryngii. WPEP-N-b is composed primarily of galactose (43.8%), mannose (39.3%), methyl-galactose (11.7%) and glucose (9.2%) residues, with the main chain being composed of α-1,6-linked D-Galp and 3-O-Me-D-Galp, branched at O-2 with single t-ß-D-Manp as major the side chain. ß-1,6-D-Glcp residues are present as minor components either in side-chains or backbone. WPEP-N-b increases macrophage phagocytosis and secretion of NO, TNF-α, IL-6 and IL-1ß. Mechanistic studies demonstrate that WPEP-N-b promotes the degradation of IκB-α, and enhances phosphorylation of MAPKs and the NF-κB p65 subunit. Our results also indicate that this polysaccharide activates RAW264.7 cells via MAPK and NF-κB signaling pathways and the Toll-like receptor 2(TLR2). These results increase our understanding as to how mushroom-derived polysaccharides modulate the immunologic process.

Polysaccharide structure and immunological relationships of RG-I pectin from the bee pollen of Nelumbo nucifera.

From the bee pollen of Nelumbo nucifera, we used hot water extraction and chromatographic fractionation on DEAE-cellulose and Sepharose CL-6B to purify polysaccharides (WNPP) to homogeneity. WNPP-2-RG (3.8 × 105 Da) consisted primarily of Rha (11.5%), GalA (12.0%), Gal (41.2%) and Ara (29.7%). Structure analysis showed that this polysaccharide is a RG-I type pectin containing AG-I and AG-II side chains comprised of 1,5-L-Ara (25.6%), t-D-Gal (12.0%), and 1,6-D-Gal (18.3%) as the primary linkage types. Immunological activity assays demonstrated that WNPP-2-RG increased lymphocyte proliferation and macrophage phagocytosis, and induced production of NO. Compared to WNPP-2-RG, its hydrolysates (RG-8H-P) had no effect on lymphocyte proliferation, although they did promote macrophage phagocytosis and decreased NO production. Our results suggest that Gal side chain residues may play a crucial role in macrophage phagocytosis, whereas Ara is unimportant for NO production and neither Gal nor Ara impact lymphocyte proliferation.

Structural characterization and macrophage activation of a hetero-galactan isolated from Flammulina velutipes.

We isolated and purified a new polysaccharide (WFVP-N-b1) with a molecular weight of 20 kDa from Flammulina velutipes. Results showed that WFVP-N-b1 is composed of an α (1 → 6)-linked D-galactan backbone and branched at the O-2 of its Galp residues by an α-D-(1 → 6)-linked Manp attached to t-ß-D-Glcp or t-α-D-Fucp side chains. WFVP-N-b1 can significantly induce cytokines secretion and release of toxic molecules. On a cellular level, WFVP-N-b1 is recognized by Toll-like receptor 4 (TLR4). Thereby, the hetero-galactan increased the phosphorylation of mitogen-activated protein kinases (MAPKs) and Akt, promoted degradation of IκB-α and the nuclear translocation of the NF-κB p65 subunit. Importantly, our results indicate that WFVP-N-b1 activated macrophage is mediated by autophagy, as blockade of WFVP-N-b1-induced autophagy by Baf-A1 significantly decreases macrophage activation. This is the first report that hetero-galactan-induced macrophage activation is mediated by autophagy. Collectively, WFVP-N-b1 activated RAW264.7 cells through MAPKs, autophagy, and Akt/NF-κB signaling pathways via TLR4 receptor.

Aurora-A regulates autophagy through the Akt pathway in human prostate cancer.

BACKGROUND: Aurora A kinase is frequently overexpressed in a variety of tumor types, including the prostate. However, the function of Aurora A in autophagy in prostate cancer has not been investigated. Here, we aimed to study the functioning mechanism and autophagy associated signaling pathways of Aurora A in prostate cancer. METHODS: To investigate the biological function of Aurora A, down-regulation of Aurora A was performed followed by functional testing assays. Immunohistochemistry was used to detect the expression of Aurora A in human prostate cancer specimens. CCK8, Transwell, flow cytometric analysis and measurement of tumor formation in nude mice were performed to test the effects of Aurora A down-regulation in vivo and in vitro. Signaling pathway analysis was performed by using Western blot. Autophagy activity was measured by monitoring the expression levels of LC3-II. RESULTS: Aurora A overexpression was significantly higher in human prostate cancer specimens than in BPH. Furthermore, Aurora A knockdown inhibited the proliferation of prostate cancer cells by suppressing the Akt pathway, indicating that Akt is a novel Aurora A substrate in prostate cancer. Additionally, Aurora A down-regulation prompts autophagy in prostate cancer cells. Most importantly, Aurora A ablation almost fully abrogates tumorigenesis in nude mice, suggesting that Aurora A is a key oncogenic effector in prostate cancer. CONCLUSIONS: Taken together, our data suggest that Aurora-A plays an important role in the suppression of autophagy by inhibiting the phosphorylation of Akt, which in turn prevents autophagy-induced apoptosis in prostate cancer.

Immunomodulatory effects of Hericium erinaceus derived polysaccharides are mediated by intestinal immunology.

This study was aimed at investigating the immunomodulating activity of Hericium erinaceus polysaccharide (HEP) in mice, by assessing splenic lymphocyte proliferation (cell-mediated immunity), serum hemolysin levels (humoral immunity), phagocytic capacity of peritoneal cavity phagocytes (macrophage phagocytosis), and NK cell activity. ELISA of immunoglobulin A (SIgA) in the lamina propria, and western blotting of small intestinal proteins were also performed to gain insight into the mechanism by which HEP affects the intestinal immune system. Here, we report that HEP improves immune function by functionally enhancing cell-mediated and humoral immunity, macrophage phagocytosis, and NK cell activity. In addition, HEP was found to upregulate the secretion of SIgA and activate the MAPK and AKT cellular signaling pathways in the intestine. In conclusion, all these results allow us to postulate that the immunomodulatory effects of HEP are most likely attributed to the effective regulation of intestinal mucosal immune activity.

Multiple approaches to assess pectin binding to galectin-3.

Although several approaches have been used to evaluate binding of carbohydrates to lectins, results are not always comparable, especially with larger polysaccharides. Here, we quantitatively assessed and compared binding of pectin-derived polysaccharides to galectin-3 (Gal-3) using five methods: surface plasmon resonance (SPR), bio-layer interferometry (BLI), fluorescence polarization (FP), competitive fluorescence-linked immunosorbance (cFLISA), and the well-known cell-based hemagglutination assay (G3H). Our studies revealed that whereas Gal-3-pectin binding parameters determined by SPR and BLI were comparable and correlated with inhibitory potencies from the G3H assay, results using FP and cFLISA assays were highly variable and depended greatly on the probe and mass of the polysaccharide. In the cFLISA assay, for example, pectins showed no inhibition when using the DTAF-labeled asialofetuin probe, but did when using a DTAF-labeled pectin probe. And the FP approach with the DTAF-lactose probe did not work on polysaccharides and large galactan chains, although it did work well with smaller galactans. Nevertheless, even though results derived from all of these methods are in general agreement, derived KD, IC50, and MIC values do differ. Our results reflect the variability using various techniques and therefore will be useful to investigators who are developing pectin-derived Gal-3 antagonists as anti-cancer agents.

In vivo anticancer and immunomodulating activities of mannogalactoglucan-type polysaccharides from Lentinus edodes (Berkeley) Singer.

There is considerable interest in the potential of mushrooms in modulating the immune system and/or suppressing tumor growth. Among the studied bioactive compounds in mushrooms, polysaccharides are the most important. Nontoxic fungal polysaccharides have a more important role in immunomodulating and antitumor activities which are related to their effects to act of immune effecter cells such as lymphocytes, macrophages, dendritic cells, and natural killer cells involved in the innate and adaptive immunity. Two mannogalactoglucan-type polysaccharides (WPLE-N-2 and WPLE-A0.5-2), purified from the fruiting bodies of Lentinus edodes, were evaluated for their effects on the cellular immune response of Sarcoma 180 (S-180)-bearing mice. Mice were treated with 100 mg/kg body weight of the polysaccharides for 10 days. Significant tumor regressions of the polysaccharide groups' mice were observed compared to the control group. These polysaccharides could induce an increase in nitrite oxide (NO) production in peritoneal macrophages, significantly increase macrophage phagocytosis of tumor-bearing mice and augment concanavalin (ConA) and lipopolysaccharide (LPS)-induced splenocytes proliferation. Our results indicated that immunomodulating activity occurred through host mediation in response to lymphocyte proliferation, macrophage phagocytosis and induction of NO production while the antitumor activity occurred through direct cytotoxicity. Our findings suggest that mannogalactoglucan-type polysaccharides from L. edodes can be explored as novel potential immunostimulants. Our research provides essential data to a better understanding of L. edodes bioactive compounds, especially polysaccharides. Our results also confirm the key role of ß-linkages in the antitumor and immunomodulating effects of polysaccharides.

Increased expression of ESCO1 is correlated with poor patient survival and its role in human bladder cancer.

There is increasing evidence suggesting that establishment of sister chromatid cohesion N-acetyltransferase 1 (ESCO1) was involved in tumorigenesis. However, its role in bladder cancer remains unclear. In this study, we aimed to study the clinical correlation and biological significance of ESCO1 in bladder cancer. Our results showed that ESCO1 was significantly over-expressed in bladder cancer tissues compared with that in adjacent normal tissues. And, increased ESCO1 expression was significantly associated with higher grade (P < 0.001), higher tumor stage (P = 0.014), and multifocality (P = 0.042). Kaplan-Meier analysis and Cox proportional hazards model were performed to determine the prognostic significance of ESCO1, and the results showed that ESCO1 is a useful prognostic marker for bladder cancer patients. Moreover, we found that ESCO1 knockdown inhibited the growth, migration, and invasion of bladder cancer cells. In conclusion, our findings indicated that ESCO1 may play an important role in human bladder cancer, and ESCO1 might serve as a novel target and prognosis factor for human bladder cancer.

PACE4 regulates apoptosis in human prostate cancer cells via endoplasmic reticulum stress and mitochondrial signaling pathways.

BACKGROUND: PACE4 is a proprotein convertase capable of processing numerous substrates involved in tumor growth, invasion, and metastasis. However, the precise role of PACE4 during prostate cancer cell apoptosis has not been reported. METHODS: In the present study, human prostate cancer cell lines DU145, LNCaP, and PC3 were transfected with PACE4 small interfering (si)RNA to investigate the underlying mechanisms of apoptosis. RESULTS: We revealed that PACE4 siRNA exhibited antitumor activity by inducing apoptosis, as determined by Cell Counting Kit-8 (CCK-8), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltet-razolium bromide (MTT) assay, cell cycle analysis, Hoechst staining, caspase-3/7 activity, and western blot analysis. In addition, PACE4 siRNA significantly increased the ratio of Bax/Bcl-2, which led to the release of cytochrome c. Moreover, PACE4 siRNA also induced endoplasmic reticulum stress by increasing the expression of GRP78, GRP94, p-PERK, and p-eIF2α. The ratio of Bax/Bcl-2 and GRP78 were also increased in PACE4 gene knockdown prostate cancer cells compared with the control cells. CONCLUSION: These data demonstrate that PACE4 siRNA may exert its antitumor activity through mitochondrial and endoplasmic reticulum stress signaling pathways, indicating it may be a novel therapeutic target for prostate cancer.

The role of tumor suppressor gene SOX11 in prostate cancer.

SOX genes play an important role in a number of developmental processes. The transcription factor SOX11 is one of the members of the SOX family emerging as important transcriptional regulators. The aim of this study was to investigate the role of SOX11 in prostate cancer (PCa) and its expression pattern and clinical significance. The gene expression of SOX11 in human PCa tissues compared with benign prostate hyperplasia (BPH) tissues was detected using real-time quantitative reverse transcriptase-polymerase chain reaction (QRT-PCR) analysis and immunohositochemistry. SOX11 overexpression cell model was used to examine the role of SOX11 in cell growth and metastasis in vitro. The results showed that the positive rate of SOX11 staining was 16.67 % (10/60) in cases of prostatic carcinoma and 81.67 % (49/60) in cases of BPH, and the difference of SOX11 expression between PCa and BPH was statistically significant (P < 0.001). SOX11 mRNA level was lowly expressed in PCa cell lines compared to RWPE-1. SOX11 overexpression suppresses PCa cell migration and invasion. In conclusion, our findings demonstrate that SOX11 could suppress cell proliferation, migration, and invasion of PCa in vitro.

Long-term follow-up on the effects of sigmoid-rectal pouch for urinary diversion.

PURPOSE: The aim of this study was to investigate the long-term clinical effects of sigmoidrectal pouch for urinary diversion. MATERIALS AND METHODS: A total of 45 patients, including 40 males and 5 females, underwent sigmoid-rectal pouch procedure. The patients aged from 38 to 70 years with a mean age of 59 years. The postoperative follow-up ranged from 6 months to 19 years with an average of 6 years. Postoperative continence and voiding were analyzed, urinary reservoir pressure was measured and the complications of upper urinary tract were determined. The index of quality of life (QoL) in the International Prostate Symptom Score (IPSS) was used to evaluate the degree of satisfaction to urinate. RESULTS: Forty patients had slight incontinence in the early postoperative stage and could control urination well 30 days postoperatively. The volume of pouch was 270-600 mL with an average of 375 mL. The basic pressure during filling period was 6-20 cmH2O with an average 15 cmH2O, the maximum filling pressure was 15-30 cmH2O with an average 26 cmH2O. The compliance of sigmoid-rectal pouch was fine with an average of 30 (range 18-40) mL/ cmH2O. There were no severe complications such as hyperchloremic acidosis or retrograde pyelonephritis. Six patients had slight hydronephrosis. The index of QoL were 0-2 in 20 patients, 3 in five patients and 4 in two patients. CONCLUSION: The sigmoid-rectal pouch operation was simple and acceptable by surgeons and patients. It may be an ideal urinary diversion for patients with muscle-invasive bladder cancer, especially for patients on whom urethrectomy should be done.

Mouse senile amyloid fibrils deposited in skeletal muscle exhibit amyloidosis-enhancing activity.

Amyloidosis describes a group of protein folding diseases in which amyloid proteins are abnormally deposited in organs and/or tissues as fine fibrils. Mouse senile amyloidosis is a disorder in which apolipoprotein A-II (apoA-II) deposits as amyloid fibrils (AApoAII) and can be transmitted from one animal to another both by the feces and milk excreted by mice with amyloidosis. Thus, mouse AApoAII amyloidosis has been demonstrated to be a "transmissible disease". In this study, to further characterize the transmissibility of amyloidosis, AApoAII amyloid fibrils were injected into transgenic Apoa2(c)Tg(+/-) and normal R1.P1-Apoa2(c) mice to induce AApoAII systemic amyloidosis. Two months later, AApoAII amyloid deposits were found in the skeletal muscles of amyloid-affected mice, primarily in the blood vessels and in the interstitial tissues surrounding muscle fibers. When amyloid fibrils extracted from the skeletal muscles were subjected to Western blot analysis, apoA-II was detected. Amyloid fibril fractions isolated from the muscles not only demonstrated the structure of amyloid fibrils but could also induce amyloidosis in young mice depending on its fibril conformation. These findings present a possible pathogenesis of amyloidosis: transmission of amyloid fibril conformation through muscle, and shed new light on the etiology involved in amyloid disorders.

Amyloid fibrils formed by selective N-, C-terminal sequences of mouse apolipoprotein A-II.

In mice, amyloidogenic type C apolipoprotein A-II (apoA-II) forms amyloid fibrils in age-associated amyloidosis. To understand the mechanism of amyloid fibril formation by apoA-II, we examined the polymerization of synthetic partial peptides of apoA-II in vitro. None of the partial apoA-II peptides polymerized into amyloid fibrils when tested as a single species mixture. We found a unique mechanism in which N- and C-terminal peptides associated into amyloid fibrils in a 1:1 ratio at pH 2.5. The 11-residue amino acid sequence (6-16), which is a common sequence of type B apoA-II and type C apoA-II proteins in amyloidosis-resistant mice and amyloidosis-susceptible mice, respectively, was critical for polymerization into amyloid fibrils. The 18-residue-long amino acid sequence (48-65) is also necessary for nucleation, but not for the extension phase. These findings suggest that there may be different mechanisms underlying the nucleation and extension phases of apoA-II amyloid fibril formation. We also found that amino acid substitutions between type B apoA-II (Pro5, Val38) and type C apoA-II (Gln5, Ala38) did not affect either phase. The strategy of using synthetic partial peptides of amyloidogenic proteins in vitro is a useful system for understanding amyloid fibril formation and for the development of novel therapies.

Fecal transmission of AA amyloidosis in the cheetah contributes to high incidence of disease.

AA amyloidosis is one of the principal causes of morbidity and mortality in captive cheetahs (Acinonyx jubatus), which are in danger of extinction, but little is known about the underlying mechanisms. Given the transmissible characteristics of AA amyloidosis, transmission between captive cheetahs may be a possible mechanism involved in the high incidence of AA amyloidosis. In this study of animals with AA amyloidosis, we found that cheetah feces contained AA amyloid fibrils that were different from those of the liver with regard to molecular weight and shape and had greater transmissibility. The infectious activity of fecal AA amyloid fibrils was reduced or abolished by the protein denaturants 6 M guanidine.HCl and formic acid or by AA immunodepletion. Thus, we propose that feces are a vehicle of transmission that may accelerate AA amyloidosis in captive cheetah populations. These results provide a pathogenesis for AA amyloidosis and suggest possible measures for rescuing cheetahs from extinction.

Cross-seeding and cross-competition in mouse apolipoprotein A-II amyloid fibrils and protein A amyloid fibrils.

Murine senile [apolipoprotein A-II amyloid (AApoAII)] and reactive [protein A amyloid (AA)] amyloidosis are reported to be transmissible diseases via a seeding mechanism similar to that observed in the prion-associated disorders, although de novo amyloidogenesis and the progression of AApoAII or AA amyloidosis remain unclear. We examined the effect of co-injection of AApoAII and AA fibrils and multiple inflammatory stimuli in R1.P1-Apoa2(c) mice with the amyloidogenic Apoa2(c) allele. Both AApoAII and AA amyloidosis could be induced in this system, but the two types of amyloid fibrils preferentially promote the formation of the same type of fibrils while inhibiting the formation of the other. Furthermore, we demonstrate that AA or AApoAII amyloidosis could be cross-seeded by predeposited AApoAII or AA fibrils and that the predeposited amyloid fibrils were degraded when the fibril formation was reduced or stopped. In addition, a large proportion of the two amyloid fibrils colocalized during the formation of new fibrils in the spleen and liver. Thus, we propose that AApoAII and AA can both cross-seed and cross-compete with regard to amyloid formation, depending on the stage of amyloidogenesis. These results will aid in the clarification of the mechanisms of pathogenesis and progression of amyloid disorders.