New two-stage pH combined with dissolved oxygen control strategy for cyclic ß-1,2 glucans synthesis.

On the basis of a novel two-stage pH combined with dissolved oxygen (DO) control strategy in fed-batch fermentation, this research addresses the influence of pH on cyclic ß-1,2-glucans (CßGs) biosynthesis and melanin accumulation during the production of CßGs by Rhizobium radiobacter ATCC 13,333. Under these optimal fermentation conditions, the maximum cell concentration and CßGs concentration in a 7-L stirred-tank fermenter were 7.94 g L-1 and 3.12 g L-1, which were the maximum production reported for R. radiobacter. The melanin concentration of the fermentation broth was maintained at a low level, which was beneficial to the subsequent separation and purification of the CßGs. In addition, a neutral extracellular oligosaccharide (COGs-1) purified by the two-stage pH combined with DO control strategy fermentation medium was structurally characterized. Structural analyses indicated that COGs-1 was a family of unbranched cyclic oligosaccharides composed of only ß-1,2-linked D-glucopyranose residues with degree of polymerization between 17 and 23, namely CßGs. This research provides a reliable source of CßGs and structural basis for further studies of biological activity and function. KEY POINTS: • A two-stage pH combined with DO control strategy was proposed for CßGs production and melanin biosynthesis by Rhizobium radiobacter. • The final extracellular CßGs production reached 3.12 g L-1, which was the highest achieved by Rhizobium radiobacter. • The existence of CßGs could be detected by TLC quickly and accurately.

MicroRNA-21 as a diagnostic marker for hepatocellular carcinoma: A systematic review and meta-analysis.

BACKGROUND: MicroRNA-21 (miR-21) is one of the oncogenic miRNAs which may be a potential diagnostic biomarker for hepatocellular carcinoma (HCC). METHODS: We systematically searched Medline, Embase, the Cochrane Library, ISI Web of Knowledge, Scopus from inception to August 15, 2018, and reference lists of identified primary studies. Two independent investigators extracted patient and study characteristics. The sensitivity and specificity of microRNA-21 for HCC detection and were analyzed with a random effect model. The area under summary receiver operating characteristic curve (AUC) was used to estimate overall test performance. RESULTS: A total of 515 HCC patients, and 338 healthy or chronic hepatitis controls from six published studies were enrolled in this meta-analysis. All articles were published in English with moderate-to-high quality. The overall pooled sensitivity and specificity were 85.2% (73.3% to 88.4%) and 79.2% (68.4% to 87.0%), respectively. The AUC area was 0.89 (95% CI: 0.85-0.91). The studies had moderate heterogeneity (I2=70.11%). None of the subgroups investigated-ethnicity, controls, sample source-could account for the heterogeneity. CONCLUSION: MiR-21 is a helpful biomarker for early diagnosis of HCC. Nevertheless, the results of the test must be interpreted carefully in the context of medical history, erological tests and imaging examinations for HCC surveillance.

MicroRNA-195 chemosensitizes colon cancer cells to the chemotherapeutic drug doxorubicin by targeting the first binding site of BCL2L2 mRNA.

The mechanisms underlying doxorubicin (Dox) resistance in colon cancer cells are not fully understood. MicroRNA (miRNA) play important roles in tumorigenesis and drug resistance. However, the relationship between miRNA and Dox resistance in colon cancer cells has not been previously explored. In this study, we utilized microRNA array and real-time PCR to verify that miR-127, miR-195, miR-22, miR-137 were significantly down-regulated, while miR-21, miR-592 were up-regulated in both HT29/DOX and LOVO/DOX cell lines. In vitro cell viability assay showed that knockdown of miR-195 in HT29 and LOVO cells caused a marked inhibition of Dox-induced cytotoxicity. Moreover, we explored that miR-195 is involved in repression of BCL2L2 expression through targeting its 3'-untranslated region, especially the first binding site within its mRNA. Furthermore, down-regulation of miR-195 conferred DOX resistance in parental cells and reduced cell apoptosis activity, while over-expression of miR-195 sensitized resistant cells to DOX and enhanced cell apoptosis activity, all of which can be partly rescued by BCL2L2 siRNA and cDNA expression. These results may have implications for therapeutic strategies aiming to overcome colon cancer cell resistance to Dox.

Reversible reactivity by optic nerve astrocytes.

Reactive astrocytes are typically studied in models that cause irreversible mechanical damage to axons, neuronal cell bodies, and glia. Here, we evaluated the response of astrocytes in the optic nerve head to a subtle injury induced by a brief, mild elevation of the intraocular pressure. Astrocytes demonstrated reactive remodeling that peaked at three days, showing hypertrophy, process retraction, and simplification of their shape. This was not accompanied by any significant changes in the gene expression profile. At no time was there discernible damage to the optic axons, as evidenced by electron microscopy and normal anterograde and retrograde transport. Remarkably, the morphological remodeling was reversible. These findings underscore the plastic nature of reactivity. They show that reactivity can resolve fully if the insult is removed, and suggest that reactivity per se is not necessarily deleterious to axons. This reaction may represent very early events in the sequence that eventually leads to glial scarring.

Endogenous EGF maintains Sertoli germ cell anchoring junction integrity and is required for early recovery from acute testicular ischemia/reperfusion injury.

Administration of exogenous epidermal growth factor (EGF) improves testicular injury after acute ischemia-reperfusion (IR) stress, but the molecular basis is poorly understood. The role of endogenous EGF in testicular recovery and the underlying intracellular signaling pathways involved were herein investigated. In mice, testicular IR injury significantly enhanced the expression level of endogenous Egf at the very beginning of reperfusion. Expression of EGF receptor (Egfr (ErbB1)) was accordingly upregulated 3  h after reperfusion. Deprivation of majority of circulated EGF by sialoadenectomy aggravated testicular detriment (especially in pachytene spermatocytes), enhanced germ cell apoptosis, and thereafter resulted in impaired meiotic differentiation after IR insult. Mechanistically, endogenous EGF signaling appeared to be indispensable for the proper maintenance of Sertoli germ cells anchoring junction dynamics during the early testicular recovery. We also provided the in vitro evidences in a well-established rat Sertoli germ cell co-cultures model that the pro-survival effect of endogenous EGF on germ cells in response to testicular IR insult is mediated, at least in part, via the phosphatidylinositol 3-kinase/pAkt pathway. Collectively, our results suggest that the augment of endogenous EGF during the early testicular recovery may act on top of an endocrinous cascade orchestrating the intimate interactions between Sertoli cells and germ cells and may operate as indispensable defensive mechanism in response to testicular IR stress. Future studies in this field would shed light on this complicated pathogenesis.

Expression and localization of α2A-adrenergic receptor in the rat post-natal developing cochlea.

Lots of adrenergic receptors (ARs) are widely present across the auditory pathways and are positioned to affect auditory and vestibular functions. However, noradrenergic regulation in the cochlea has not been well characterized. In this study, a rat model of noise-induced hearing loss was developed to investigate the expression of α2A-adrenergic receptor (AR) after acoustic trauma, then, we investigated the expression of α2A-AR in the developing rat cochlea using immunofluorescence, qRT-PCR, and Western blotting. We found that the expression of α2A-AR significantly increased in rats exposed to noise compared with controls. Immunofluorescence analysis demonstrated that α2A-AR is localized on hair cells (HCs), spiral ganglion neurons (SGNs), and the stria vascularis (SV) in the postnatal developing cochlea from post-natal day (P) 0 to P28. Furthermore, we observed α2A-AR mRNA reached a maximum level at P14 and P28 when compared with P0, while no significant differences in α2A-AR protein levels at the various stages when compared with P0. This study provides direct evidence for the expression of α2A-AR in HCs, SGNs, and the SV of the cochlea, indicating that norepinephrine might play a vital role in hearing function within the cochlea through α2A-AR.

Trimethylamine-N-oxide: a potential biomarker and therapeutic target in ischemic stroke.

Ischemic stroke is by far the most common cerebrovascular disease and a major burden to the global economy and public health. Trimethylamine-N-oxide (TMAO), a small molecule compound produced by the metabolism of intestinal microorganisms, is reportedly associated with the risk of stroke, as well as the severity and prognosis of stroke; however, this conclusion remains contentious. This article reviews the production of TMAO, TMAO's relationship with different etiological types of ischemic stroke, and the possibility of reducing TMAO levels to improve the prognosis of ischemic stroke.

Calcineurin activation causes retinal ganglion cell degeneration.

PURPOSE: We previously reported that calcineurin, a Ca(2+)/calmodulin-dependent serine/threonine phosphatase, is activated and proposed that it participates in retinal ganglion cell (RGC) apoptosis in two rodent ocular hypertension models. In this study, we tested whether calcineurin activation by itself, even in the absence of ocular hypertension, is sufficient to cause RGC degeneration. METHODS: We compared RGC and optic nerve morphology after adeno-associated virus serotype 2 (AAV2)-mediated transduction of RGCs with constitutively active calcineurin (CaNCA) or unactivated, wild-type calcineurin (CaNwt). Retinas and optic nerves were harvested 7-16 weeks after injection of the AAV into mouse vitreous. In flatmounted retinas, the transduced RGCs were identified with immunohistochemistry. The morphology of the RGCs was revealed by immunostaining for neurofilament SMI32 or by using GFP-M transgenic mice. A modified Sholl analysis was applied to analyze the RGC dendritic morphology. Optic nerve damage was assessed with optic nerve grading according to the Morrison standard. RESULTS: CaNwt and CaNCA were highly expressed in the injected eyes. Compared to the CaNwt-expressing RGCs, the CaNCA-expressing RGCs had smaller somas, smaller dendritic field areas, shorter total dendrite lengths, and simpler dendritic branching patterns. At 16 weeks, the CaNCA-expressing eyes had greater optic nerve damage than the CaNwt-expressing eyes. CONCLUSIONS: Calcineurin activation is sufficient to cause RGC dendritic degeneration and optic nerve damage. These data support the hypothesis that calcineurin activation is an important mediator of RGC degeneration, and are consistent with the hypothesis that calcineurin activation may contribute to RGC neurodegeneration in glaucoma.

Inhalation of hydrogen gas attenuates ouabain-induced auditory neuropathy in gerbils.

AIM: Auditory neuropathy (AN) is a hearing disorder characterized by abnormal auditory nerve function with preservation of normal cochlear hair cells. This study was designed to investigate whether treatment with molecular hydrogen (H(2)), which can remedy damage in various organs via reducing oxidative stress, inflammation and apoptosis, is beneficial to ouabain-induced AN in gerbils. METHODS: AN model was made by local application of ouabain (1 mmol/L, 20 mL) to the round window membrane in male Mongolian gerbils. H(2) treatment was given twice by exposing the animals to H(2) (1%, 2%, and 4%) for 60 min at 1 h and 6 h after ouabain application. Before and 7 d after ouabain application, the hearing status of the animals was evaluated using the auditory brainstem response (ABR) approach, the hear cell function was evaluated with distortion product otoacoustic emissions (DPOAE). Seven days after ouabain application, the changes in the cochleae, especially the spiral ganglion neurons (SGNs), were morphologically studied. TUNEL staining and immunofluorescent staining for activated caspase-3 were used to assess the apoptosis of SGNs. RESULTS: Treatment with H(2) (2% and 4%) markedly attenuated the click and tone burst-evoked ABR threshold shift at 4, 8, and 16 kHz in ouabain-exposed animals. Neither local ouabain application, nor H(2) treatment changed the amplitude of DPOAE at 4, 8, and 16 kHz. Morphological study showed that treatment with H(2) (2%) significantly alleviated SGN damage and attenuated the loss of SGN density for each turn of cochlea in ouabain-exposed animals. Furthermore, ouabain caused significantly higher numbers of apoptotic SGNs in the cochlea, which was significantly attenuated by the H(2) treatment. However, ouabain did not change the morphology of cochlear hair cells. CONCLUSION: The results demonstrate that H(2) treatment is beneficial to ouabain-induced AN via reducing apoptosis. Thus, H(2) might be a potential agent for treating hearing impairment in AN patients.

Enhanced solubility of curcumin by complexation with fermented cyclic ß-1,2-glucans.

Curcumin (CUR) is a low-solubility polyphenolic compound with many physiological functions. Cyclic ß-1,2-glucans (cyclosophoraoses [Cys]), which contain rings of different sizes with degrees of polymerization ranging from 17 to 23, were obtained from Rhizobium radiobacter ATCC 1333, a soil microorganism. The complexation ability and solubility enhancement of cyclic ß-1,2-glucans with insoluble curcumin were investigated. Phase-solubility analysis revealed that the stoichiometric ratio of the inclusion complexes was 1:1. The stability constant of Cys was 930 M-1, which was 7.68 times that of α-cyclodextrin (α-CD) and 2.09 times that of ß-cyclodextrin (ß-CD). The characteristics of the curcumin/Cys inclusion complexes were successfully determined by using Fourier transform infrared (FTIR) spectrometry, differential scanning calorimetry (DSC), nuclear magnetic resonance (1H NMR) spectroscopy, and scanning electron microscopy (SEM). Moreover, a 1:1 molecular model of the curcumin/Cys inclusion complexes was established through molecular docking analysis. These findings indicated that cyclic ß-1,2-glucans successfully formed complexes with curcumin, which suggested that they could be used as solubility-increasing agents. To the best of our knowledge, this is the first report in which curcumin has been embedded into cyclic ß-1,2-glucans resulting in an increase in its aqueous solubility.

WITHDRAWN: Reprint of: Mechanisms of retinal ganglion cell injury and defense in glaucoma.

The Publisher regrets that this article is an accidental duplication of an article that has already been published, doi:10.1016/j.exer.2010.04.002. The duplicate article has therefore been withdrawn.

Facies-Controlled Geostatistical Porosity Model for Estimation of the Groundwater Potential Area in Hongliu Coalmine, Ordos Basin, China.

Accurate and reliable evaluations of potential groundwater areas are of significance in the hydrogeological assessments of coalfields because water inrush disasters may be caused by unclear groundwater potential. A three-dimensional geological model of porosity based on deterministic modeling and a facies-controlled method are used to determine the groundwater potential of the coal measure aquifer. The modeling processes are as follows: based on the interlayer and discontinuity (faults) data extracted from boreholes and geological maps, an integrated sequence framework model is developed. Using the results of sedimentary microfacies identification and the method of deterministic modeling, a sedimentary microfacies model is successfully established. Finally, based on facies-controlled and sequential Gaussian methods, an effective porosity model is established that can predict the groundwater potential. The predicted results show that sandstones sedimented in channel, point bar, and batture environments possess high effective porosity and strong groundwater potential; however, the sandstones sedimented in interdistributary bays, flood plains, and sand sheets possess low effective porosity. Model validation was performed based on the hydrological pumping test data collected from observation boreholes, drainage water inflow data from dewatered boreholes in the tunnel around workface, and the mine water inflow in tunnels and the workfaces. The validation analysis results show that the effective porosity and sedimentary facies were correlated with the actual flux. The predicted results are consistent with the actual flux data, validating the predicted model.

Fabricating a wettable microwells array onto a nitrogen plasma-treated ITO substrate: high-throughput fluorimetric platform for selective sensing of ammonia in blood using polymer-stabilized NH2-MIL-125.

A high-throughput and selective fluorimetric platform has been constructed for the analysis of ammonia in blood by using a polymer-stabilized metal-organic framework (MOF) of porous NH2-MIL-125, which was coated onto a wettable microwells array constructed on an indium tin oxide (ITO) substrate. It was found that the nitrogen plasma treatment for the ITO substrate could create a super-hydrophilic interface that combined with the hydrophobic pattern yielded a wettable microwells array, enabling the condensation-based enrichment of targets from the sample droplets. Moreover, the NH2-MIL-125 MOF encapsulated using polymers could be firmly coated onto the microwells to act as fluorescent probes for sensing NH3 with enhanced responses. In addition, the use of the polymer polyvinyl pyrrolidone could protect and stabilize the crystals of NH2-MIL-125 probe in aqueous media, revealing the improved hydrophilicity and significantly depressed signal background. The as-developed fluorimetric platform, containing a MOF-coated microwells array, can enable the detection of ammonia in blood, with concentrations ranging linearly from 0.10 to 300 µM. More importantly, this plasma treatment-based fabrication route may hold promise for designing different wettable microwells arrays for the high-throughput detection of multiple targets in the fields of biomedical analysis and environmental monitoring.

Mechanisms of retinal ganglion cell injury and defense in glaucoma.

Glaucoma is a disease in which retinal ganglion cells (RGCs) die leading ultimately to blindness. Over the past decade and a half, information has begun to emerge regarding specific molecular responses of the retina to conditions of elevated intraocular pressure (IOP). It is now clear that the state of the RGC in glaucoma depends on a balance of pro-survival and pro-death pathways in the retina and details of these responses are still being worked out. In this review, we will discuss the evidence supporting the involvement of specific apoptotic cascades as well as the insults that trigger RGC apoptosis. In addition, we will present evidence supporting the existence of endogenous protective mechanisms as well as exogenous neuroprotective strategies.

CircFLNA Acts as a Sponge of miR-646 to Facilitate the Proliferation, Metastasis, Glycolysis, and Apoptosis Inhibition of Gastric Cancer by Targeting PFKFB2.

BACKGROUND: Many studies have confirmed that circular (circRNA) is involved in the development of gastric cancer (GC). However, the role of circFLNA in the progression of GC remains unclear. METHODS: Quantitative real-time PCR (qRT-PCR) was used to measure the relative expression of circFLNA, microRNA (miR)-646 and 6-phosphofructo-2-kinase/fructose-2, 6-biphosphatase 2 (PFKFB2). Cell counting kit 8 (CCK8) assay, transwell assay and flow cytometry were performed to determine the proliferation, migration, invasion and apoptosis of cells, respectively. GC tumor xenograft models were built to confirm the function of circFLNA silencing on GC tumor growth in vivo. Furthermore, the lactate production, glucose consumption, ATP level and glucose uptake were detected to assess the glycolysis of cells. Then, the interaction between miR-646 and circFLNA or PFKFB2 was confirmed using dual-luciferase reporter assay. RNA immunoprecipitation (RIP) assay was used to verify the interaction between miR-646 and circFLNA further. In addition, Western blot (WB) analysis was employed to detect the relative protein expression of PFKFB2. RESULTS: Our results found that circFLNA was upregulated in GC tissues and cells. Silencing of circFLNA could suppress the proliferation, migration, invasion, glycolysis, and enhance the apoptosis of GC cells. Also, circFLNA knockdown reduced GC tumor volume and weight in vivo. Further experiments revealed that circFLNA could sponge miR-646, and miR-646 could target PFKFB2. The rescue experiments indicated that miR-646 inhibitor could reverse the suppressive effect of circFLNA silencing on GC progression, and PFKFB2 overexpression also could invert the inhibition effect of miR-646 on GC progression. CONCLUSION: Our data concluded that circFLNA played a pro-cancer role in GC, which suggested that circFLNA might be a potential biomarker for GC treatment.

Combined effect of recombinant human adenovirus p53 and curcumin in the treatment of liver cancer.

The development of an effective therapeutic intervention for liver cancer is a worldwide challenge that remains to be adequately addressed. Of note, TP53, which encodes the p53 protein, is an important tumor suppressor gene, 61% of TP53 is functionally inactivated in liver cancer. Recombinant human adenovirus p53 (rAd-p53) is the first commercial product that has been used for gene therapy. In the present study, the combined mechanistic effects of rAd-p53 and curcumin, a naturally occurring compound with previously reported anti-inflammatory, antioxidant and anti-cancer properties, were assessed in liver cancer cells, using HepG2 cells as the model cell line. The administration of either curcumin or rAd-p53 promoted apoptosis, suppressed epithelial-mesenchymal transition (EMT) and blocked G2/M phase progression in HepG2 cells, which were potentiated further when both agents were applied together. Combined rAd-p53 and curcumin treatment resulted in higher p53 (P<0.01) and p21 (P<0.01) expression compared with rAd-p53 or curcumin were added alone, suggesting an additive effect on TP53 expression. Additionally, curcumin and rAd-p53 were demonstrated to regulate the activation of mitogen-activated protein kinases (MAPKs) ERK1/2, p38 MAPK and JNK. These results indicated that the combination of rAd-p53 with curcumin synergistically potentiates apoptosis and inhibit EMT compared with either rAd-p53 or curcumin treatment alone via the regulation of TP53 regulation. Mechanistically, this effect on TP53 expression may involve the ERK1/2, p38 MAPK and JNK signaling pathways. The current study provides new insights that can potentially advance the development of therapeutic strategies for liver cancer treatment.

[Effect of histone acetylation on osteogenic differentiation of periodontal ligament stem cells derived from periodontitis tissue].

Epigenetics is defined as a change in gene expression without the alteration of the genetic sequence. Such a change would be inherited by offspring. Histone acetylation is a type of epigenetics. Existing studies proposed that chronic periodontitis is related to epigenetic modification. In this review, we summarised the influence of chronic periodontitis on periodontal ligament stem cells by histone acetylation.

Sirt3 confers protection against acrolein-induced oxidative stress in cochlear nucleus neurons.

Acrolein is a ubiquitous dietary and environmental pollutant, which can also be generated endogenously during cellular stress. However, the molecular mechanisms underlying acrolein-induced neurotoxicity, especially in ototoxicity conditions, have not been fully determined. In this study, we investigated the mechanisms on acrolein-induced toxicity in primary cultured cochlear nucleus neurons with focus on Sirt3, a mitochondrial deacetylase. We found that acrolein treatment induced neuronal injury and programmed cell death (PCD) in a dose dependent manner in cochlear nucleus neurons, which was accompanied by increased intracellular reactive oxygen species (ROS) generation and lipid peroxidation. Acrolein exposure also significantly reduced the mitochondrial membrane potential (MMP) levels, promoted cytochrome c release and decreased mitochondrial ATP production. In addition, increased ER tracker fluorescence and activation of ER stress factors were observed after acrolein treatment, and the ER stress inhibitors were shown to attenuate acrolein-induced toxicity in cochlear nucleus neurons. The results of western blot and RT-PCR showed that acrolein markedly decreased the expression of Sirt3 at both mRNA and protein levels, and reduced the activity of downstream mitochondrial enzymes. Furthermore, overexpression of Sirt3 by lentivirus transfection partially prevented acrolein-induced neuronal injury in cochlear nucleus neurons. These results demonstrated that acrolein induces mitochondrial dysfunction and ER stress in cochlear nucleus neurons, and Sirt3 acts as an endogenous protective factor in acrolein-induced ototoxicity.

Smad3/4 Binding to Promoter II of P450arom So As to Regulate Aromatase Expression in Endometriosis.

Activin A can stimulate aromatase P450 (P450arom) expression in eutopic endometrial stromal cells (ESCs) of endometriosis by activin type I receptor-Smad pathway. In order to identify Smad3/4 binding to P450arom promoter II that mediates activin A response in ESCs, polymerase chain reaction (PCR) products of a serial truncated deletion of the P450arom promoter II region between -904 and +87 bp were inserted into the pGL3-basic vector to generate the promoter reporter plasmids. Luciferase reporter plasmids were cotransfected into cells with or without activin A (25 ng/mL). The pGL3 -705/+87 revealed a luciferase activity similar to pGL3 -904/+87, whereas progressive truncation to position -464/+87 and -192/+87, the luciferase activity was significant variation. Chromatin immunoprecipitation assay and Smad4-small interfering RNA (siRNA) testify that Smad3/4 binds to the activin A-responsive aromatase promoter in ESCs. Chromatin immunoprecipitation assay-PCR assay demonstrated anti-Smad3 antibody complexes could interact with the amplified DNA of the activin A-responsive P450arom promoter. Mutations of the binding site (-141/-138 bp, -165/-162 bp) in P450arom promoter II significantly reduced promoter activity of activin A fold-induction to 26% and 28%, respectively. We cotransfected pGL3 -705/+87 with control siRNA and Smad4-siRNA into ESCs in the presence of activin A. Luciferase analysis showed that Smad4-siRNA abolished increased promoter activity of activin A-induced P450arom expression. The effect of activin A on the p-Smad3 accumulation in the cytoplasm and nucleus was significantly abrogated following the pretreatment of ESCs with Smad4-siRNA. In conclusion, activated Smad3 proteins can bind to P450arom promoter -705/+87 bp region, responsive to activin A in ESCs, which can promote P450arom transcription.