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Hepatocellular carcinoma (HCC) results in the abnormal regulation of cellular metabolic pathways. Constraint-based modeling approaches can be utilized to dissect metabolic reprogramming, enabling the identification of biomarkers and anticancer targets for diagnosis and treatment. In this study, two genome-scale metabolic models (GSMMs) were reconstructed by employing RNA sequencing expression patterns of hepatocellular carcinoma (HCC) and their healthy counterparts. An anticancer target discovery (ACTD) framework was integrated with the two models to identify HCC targets for anticancer treatment. The ACTD framework encompassed four fuzzy objectives to assess both the suppression of cancer cell growth and the minimization of side effects during treatment. The composition of a nutrient may significantly affect target identification. Within the ACTD framework, ten distinct nutrient media were utilized to assess nutrient uptake for identifying potential anticancer enzymes. The findings revealed the successful identification of target enzymes within the cholesterol biosynthetic pathway using a cholesterol-free cell culture medium. Conversely, target enzymes in the cholesterol biosynthetic pathway were not identified when the nutrient uptake included a cholesterol component. Moreover, the enzymes PGS1 and CRL1 were detected in all ten nutrient media. Additionally, the ACTD framework comprises dual-group representations of target combinations, pairing a single-target enzyme with an additional nutrient uptake reaction. Additionally, the enzymes PGS1 and CRL1 were identified across the ten-nutrient media. Furthermore, the ACTD framework encompasses two-group representations of target combinations involving the pairing of a single-target enzyme with an additional nutrient uptake reaction. Computational analysis unveiled that cell viability for all dual-target combinations exceeded that of their respective single-target enzymes. Consequently, integrating a target enzyme while adjusting an additional exchange reaction could efficiently mitigate cell proliferation rates and ATP production in the treated cancer cells. Nevertheless, most dual-target combinations led to lower side effects in contrast to their single-target counterparts. Additionally, differential expression of metabolites between cancer cells and their healthy counterparts were assessed via parsimonious flux variability analysis employing the GSMMs to pinpoint potential biomarkers. The variabilities of the fluxes and metabolite flow rates in cancer and healthy cells were classified into seven categories. Accordingly, two secretions and thirteen uptakes (including eight essential amino acids and two conditionally essential amino acids) were identified as potential biomarkers. The findings of this study indicated that cancer cells exhibit a higher uptake of amino acids compared with their healthy counterparts. [ABSTRACT FROM AUTHOR]

Background: The current treatment of osteogenesis imperfecta (OI) is imperfect. Our study thus delves into the potential of using Dickkopf-1 antisense (DKK1-AS) to treat OI. Methods: We analysed serum DKK1 levels and their correlation with lumbar spine and hip T-scores in OI patients. Comparative analyses were conducted involving bone marrow stromal cells (BMSCs) and bone tissues from wild-type mice, untreated OI mice, and OI mice treated with DKK1-ASor DKK1-sense (DKK1-S). Results: Significant inverse correlations were noted between serum DKK1 levels and lumbar spine (correlation coefficient = − 0.679, p = 0.043) as well as hip T-scores (correlation coefficient = − 0.689, p = 0.042) in OI patients. DKK1-AS improved bone mineral density (p = 0.002), trabecular bone volume/total volume fraction (p < 0.001), trabecular separation (p = 0.010), trabecular thickness (p = 0.001), trabecular number (p < 0.001), and cortical thickness (p < 0.001) in OI mice. DKK1-AS enhanced the transcription of collagen 1α1, osteocalcin, runx2, and osterix in BMSC from OI mice (all p < 0.001), resulting in a higher von Kossa-stained matrix area (p < 0.001) in ex vivo osteogenesis assays. DKK1-AS also reduced osteoclast numbers (p < 0.001), increased β-catenin and T-cell factor 4 immunostaining reactivity (both p < 0.001), enhanced mineral apposition rate and bone formation rate per bone surface (both p < 0.001), and decreased osteoclast area (p < 0.001) in OI mice. DKK1-AS upregulated osteoprotegerin and downregulated nuclear factor-kappa B ligand transcription (both p < 0.001). Bone tissues from OI mice treated with DKK1-AS exhibited significantly higher breaking force compared to untreated OI mice (p < 0.001). Conclusions: Our study elucidates that DKK1-AS has the capability to enhance bone mechanical properties, restore the transcription of osteogenic genes, promote osteogenesis, and inhibit osteoclastogenesis in OI mice. [ABSTRACT FROM AUTHOR]

In this paper, a fuzzy hierarchical optimization framework is proposed for identifying potential antiviral targets for treating severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in the heart. The proposed framework comprises four objectives for evaluating the elimination of viral biomass growth and the minimization of side effects during treatment. In the application of the framework, Dulbecco's modified eagle medium (DMEM) and Ham's medium were used as uptake nutrients on an antiviral target discovery platform. The prediction results from the framework reveal that most of the antiviral enzymes in the aforementioned media are involved in fatty acid metabolism and amino acid metabolism. However, six enzymes involved in cholesterol biosynthesis in Ham's medium and three enzymes involved in glycolysis in DMEM are unable to eliminate the growth of the SARS-CoV-2 biomass. Three enzymes involved in glycolysis, namely BPGM, GAPDH, and ENO1, in DMEM combine with the supplemental uptake of L-cysteine to increase the cell viability grade and metabolic deviation grade. Moreover, six enzymes involved in cholesterol biosynthesis reduce and fail to reduce viral biomass growth in a culture medium if a cholesterol uptake reaction does not occur and occurs in this medium, respectively. [ABSTRACT FROM AUTHOR]

Sclerostin (SOST) is a Wnt signaling inhibitor detrimental to osteogenic differentiation and bone mineral acquisition. While control of SOST action delays the pathogenesis of skeletal disorders, the effects of SOST vaccination on the estrogen deficiency-induced bone deterioration remain elusive. In this study, we generated a SOST-Fc fusion protein which was composed of a SOST peptide Pro-Asn-Ala-Ile-Gly along with an IgG Fc fragment. SOST-Fc vaccination increased serum anti-SOST antibody levels and reduced serum SOST concentrations in mice. In vitro, anti-SOST serum attenuated the SOST-induced inhibition of osteogenic gene expression in osteoblast cultures. Administration with SOST-Fc increased serum levels of bone formation marker osteocalcin and alleviated the ovariectomy escalation of serum resorption markers CTX-1 and TRAP5b concentrations. It remarkably lessened the estrogen deficiency-mediated deterioration of bone mineral density, morphometric characteristics of trabecular bone, and mechanical strength of femurs and lumbar spines. The SOST-Fc-treated skeletal tissue exhibited moderate responses to the adverse actions of ovariectomy to bone mineral accretion, osteoclast surface, trabecular separation, and fatty marrow histopathology. SOST-Fc treatment increased serum osteoclast-inhibitory factor osteoprotegrin levels in conjunction with strong Wnt3a, β-catenin, and TCF4 immunostaining in osteoblasts, whereas it weakened the estrogen deficiency enhancement of osteoclast-promoting factor receptor activator of nuclear factor-κB ligand. Taken together, blockade of SOST action by SOST-Fc vaccination sustains Wnt signaling, which harmonizes bone mineral accretion and resorption reactions and thereby ameliorates ovariectomy-induced bone loss. This study highlights SOST-Fc fusion protein as a new molecular therapeutic potential for preventing from osteoporotic disorders. [ABSTRACT FROM AUTHOR]

Cholestasis and hepatitis can cause continuous liver damage that may ultimately result in liver fibrosis. In a previous study, we demonstrated that microRNA-29a (miR-29a) protects against liver fibrosis. Toll-like receptor 2 (TLR2) and TLR4 are pattern recognition receptors of bacterial lipoprotein and lipopolysaccharide, both of which participate in activating hepatic stellate cells and liver fibrosis. The purpose of this study is to characterize the biological influence of miR-29a on TLR2 and TLR4 signaling in livers injured with bile duct ligation (BDL). We performed BDL on both miR-29a transgenic mice (miR-29aTg) and wild-type mice to induce cholestatic liver injury. Primary HSCs were transfected with a miR-29a mimic and inhibitor. In the wild-type mice, the BDL demonstrated significant α-smooth muscle actin fibrotic matrix formation and hepatic high mobility group box-1 expression. However, in the miR-29aTg mice, these factors were significantly reduced. Furthermore, miR-29a overexpression reduced the BDL exaggeration of TLR2, TLR4, MyD88, bromodomain-containing protein 4 (BRD4), phospho-p65 as well as proinflammatory cytokines, IL-1β, MCP-1, TGF-β, and TNF-α. In vitro, miR-29a mimic transfection reduced α-SMA, BRD4,TLR2, and TLR4 expressions in HSCs. This study provides new molecular insight into the ability of miR-29a to inhibit TLR2 and TLR4 signaling, which thus slows the progression of cholestatic liver deterioration. [ABSTRACT FROM AUTHOR]

Precision medicine considers an individual’s unique physiological characteristics as strongly influential in disease vulnerability and in response to specific therapies. Predicting an individual’s susceptibility to developing an illness, making an accurate diagnosis, maximizing therapeutic effects, and minimizing adverse effects for treatment are essential in precision medicine. We introduced model-based precision medicine optimization approaches, including pathogenesis, biomarker detection, and drug target discovery, for treating presynaptic dopamine overactivity. Three classes of one-hit and two-hit enzyme defects were detected as the causes of disease states by the optimization approach of pathogenesis. The cluster analysis and support vector machine was used to detect optimal biomarkers in order to discriminate the accurate etiology from three classes of disease states. Finally, the fuzzy decision-making method was employed to discover common and specific drug targets for each classified disease state. We observed that more accurate diagnoses achieved higher satisfaction grades and dosed fewer enzyme targets to treat the disease. Furthermore, satisfaction grades for common drugs were lower than for specific ones, but common drugs could simultaneously treat several disease states that had different etiologies. [ABSTRACT FROM AUTHOR]

Excess glucocorticoid administration impairs osteogenic activities, which raises the risk of osteoporotic disorders. Epigenetic methylation of DNA and histone regulates the lineage commitment of progenitor cells. This study was undertaken to delineate the actions of histone lysine demethylase 6a (UTX) with regard to the glucocorticoid impediment of osteogenic differentiation. Osteogenic progenitor cells responded to supraphysiological glucocorticoid by elevating CpG dinucleotide methylation proximal to transcription start sites within Runx2 and osterix promoters and Wnt inhibitor Dickkopf-1 (Dkk1) expression concomitant with low UTX expression. 5′-Aza-deoxycystidine demethylation of Runx2 and osterix promoters abolished the glucocorticoid inhibition of mineralized matrix accumulation. Gain of UTX function attenuated the glucocorticoid-induced loss of osteogenic differentiation, whereas UTX silencing escalated adipogenic gene expression and adipocyte formation. UTX sustained osteogenic gene transcription through maintaining its occupancy to Runx2 and osterix promoters. It also mitigated the trimethylation of histone 3 at lysine 27 (H3K27me3), which reduced H3K27me3 enrichment to Dkk1 promoter and thereby lowered Dkk1 transcription. Modulation of β-catenin and Dkk1 actions restored UTX signaling in glucocorticoid-stressed cells. In vivo, UTX inhibition by exogenous methylprednisolone and GSK-J4 administration, an effect that disturbed H3K27me3, β-catenin, Dkk1, Runx2, and osterix levels, exacerbated trabecular microarchitecture loss and marrow adiposity. Taken together, glucocorticoid reduction of UTX function hindered osteogenic differentiation. Epigenetic hypomethylation of osteogenic transcription factor promoters and H3K27 contributed to the UXT alleviation of Dkk1 transcription and osteogenesis in glucocorticoid-stressed osteogenic progenitor cells. Control of UTX action has an epigenetic perspective of curtailing glucocorticoid impairment of osteogenic differentiation and bone mass. Key messages: [ABSTRACT FROM AUTHOR]

Model-based optimization approaches are valuable in developing new drugs for human metabolic disorders. The core objective in most optimal drug designs is positive therapeutic effects. In this study, we considered the effects of therapeutic, adverse, and target variation simultaneously. A fuzzy optimization method was applied to formulate a multiobjective drug design problem for detecting enzyme targets in the presynaptic dopamine metabolic network to remedy two types of enzymopathies caused by deficiencies of vesicular monoamine transporter 2 (VMAT2) and tyrosine hydroxylase (TH). The fuzzy membership approach transforms a two-stage drug discovery problem into a unified decision-making problem. We developed a nested hybrid differential evolution algorithm to efficiently identify a set of potential drug targets. Furthermore, we also simulated the effects of current clinical drugs for Parkinson’s disease (PD) in this model and tried to clarify the possible causes of neurotoxic and neuroprotective effects. The optimal drug design could yield 100% satisfaction grade when both therapeutic effect and the number of targets were considered in the objective. This scenario required regulating one to three and one or two enzyme targets for 50%–95% and 50%–100% VMAT2 and TH deficiencies, respectively. However, their corresponding adverse and target variation effect grades were less satisfactory. For the most severe deficiencies of VMAT2 and TH, a compromise design could be obtained when the effects of therapeutic, adverse, and target variation were simultaneously applied to the optimal drug discovery problem. Such a trade-off design followed the no free lunch theorem for optimization; that is, a more serious dopamine deficiency required more enzyme targets and lower satisfaction grade. In addition, the therapeutic effects of current clinical medications for PD could be enhanced in combination with new enzyme targets. The increase of toxic metabolites after treatment might be the cause of neurotoxic effects of some current PD medications. [ABSTRACT FROM AUTHOR]

Crisp and fuzzy optimization approaches were applied to design an optimal temperature and pH control policy for a batch process of simultaneous saccharification and co-fermentation (SSCF) for ethanol production from lignocellulose, using the enzyme and recombinant strainZymomonas mobilisZM (pZB5). To determine an optimal temperature and pH control policy, we applied the Arrhenius relationship to each rate constant to express the temperature and pH effects in the kinetic model for both saccharification and fermentation. The goal of the optimal design was to determine the optimal temperature, pH value, initial lignocellulosic concentration, and fermentation time for maximizing the ethanol productivity under the constraints of the follow-up separation specifications. The interactive crisp and fuzzy optimization methods were applied to solve the trade-off optimization problems for obtaining a compromised design. The fuzzy goal attainment approach obtained a compromised design more flexibly than did the crisp optimization. We also compared the performances for batch and fed-batch SSCF, and used various composition proportions for the batch SSCF to determine a series of optimal designs for the fuzzy goal attainment problem. Batch SSCF was slightly more effective than fed-batch fermentation, and spruce exhibited the maximum productivity because of its higher cellulose and lower hemicellulose contents compared with those of other sources. [ABSTRACT FROM AUTHOR]

Total knee arthroplasty (TKA) inevitably perturbs the femoral medullary canal, which increases blood loss or morbidities associated with marrow embolization postoperatively. Computer navigation TKA reportedly minimizes medullary disturbance to alleviate perioperative blood loss. We performed a prospective comparative study, enrolling 87 patients with osteoarthritic knees from March 2011 to December 2011 in our hospital. The patients were separated into two groups, according to the surgeon they visited. Fifty-four patients underwent computer navigation TKAs and 33 had conventional TKAs. Levels of cell adhesion molecules (CAMs), including intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and platelet endothelial cellular adhesion molecule-1 (PECAM-1) in sera and hemovac drainage were measured by ELISA before and 24 hours after the surgery. We showed that patients receiving computer navigation TKAs had less blood loss and lower CAMs in serum and hemovac drainage after the operation. Less postoperative elevation of serum ICAM-1 (p=0.022) and PECAM-1 (p=0.003) from the preoperative baseline after the surgery was also noted. This study provides molecular evidence for the differential extent in vascular injury between conventional and navigation TKAs and sheds light on the possible benefits of computer navigation TKAs. Trial Registration: ClinicalTrials.gov [ABSTRACT FROM AUTHOR]

Motivation: Mathematical modeling and optimization have been used for detecting enzyme targets in human metabolic disorders. Such optimal drug design methods are generally differentiated as two stages, identification and decision-making, to find optimal targets. We developed a unified method named fuzzy equal metabolic adjustment to formulate an optimal enzyme target design problem for drug discovery. The optimization framework combines the identification of enzyme targets and a decision-making strategy simultaneously. The objectives of this algorithm include evaluations of the therapeutic effect of target enzymes, the adverse effects of drugs and the minimum effective dose (MED).Results: An existing generalized mass action system model of human uric acid (UA) metabolism was used to formulate the fuzzy optimization method for detecting two types of enzymopathies: hyperuricemia caused by phosphoribosylpyrophosphate synthetase (PRPPS) overactivity and Lesch–Nyhan syndrome. The fuzzy objectives were set so that the concentrations of the metabolites were as close as possible to the healthy levels. The target design included a diet control of ribose-5-phospahate (R5P). The diet control of R5P served as an extra remedy to reduce phosphate uptake entering the purine metabolic pathway, so that we could obtain a more satisfactory treatment than obtained for those without a diet control. Moreover, enhancing UA excretion resulted in an effective treatment of hyperuricemia caused by PRPPS overactivity. This result correlates with using probenecid and benbromazone, which are uricosuric agents present in current clinical medications. By contrast, the Lesch–Nyhan syndrome required at least three enzyme targets to cure hyperuricemia.Contact: chmfsw@ccu.edu.twSupplementary information: Supplementary data are available at Bioinformatics online. [ABSTRACT FROM AUTHOR]

Highlights: [•] A cell recycling two-tank system was used for continuous production of bioethanol. [•] Each tank fermentor equipped with a settler for recycling of flocculent yeast. [•] The system was effective for ethanol production at high dilution rates. [•] The productivity was tremendously enhanced compared with batch fermentation. [ABSTRACT FROM AUTHOR]

Non-alcoholic fatty liver disease (NAFLD), the most common cause of chronic liver disease, consists of fat deposited (steatosis) in the liver due to causes besides excessive alcohol use. The folding activity of heat shock protein 60 (HSP60) has been shown to protect mitochondria from proteotoxicity under various types of stress. In this study, we investigated whether HSP60 could ameliorate experimental high-fat diet (HFD)-induced obesity and hepatitis and explored the potential mechanism in mice. The results uncovered that HSP60 gain not only alleviated HFD-induced body weight gain, fat accumulation, and hepatocellular steatosis, but also glucose tolerance and insulin resistance according to intraperitoneal glucose tolerance testing and insulin tolerance testing in HSP60 transgenic (HSP60Tg) compared to wild-type (WT) mice by HFD. Furthermore, overexpression of HSP60 in the HFD group resulted in inhibited release of mitochondrial dsRNA (mt-dsRNA) compared to WT mice. In addition, overexpression of HSP60 also inhibited the activation of toll-like receptor 3 (TLR3), melanoma differentiation-associated gene 5 (MDA5), and phosphorylated-interferon regulatory factor 3 (p-IRF3), as well as inflammatory biomarkers such as mRNA of il-1β and il-6 expression in the liver in response to HFD. The in vitro study also confirmed that the addition of HSP-60 mimics in HepG2 cells led to upregulated expression level of HSP60 and restricted release of mt-dsRNA, as well as downregulated expression levels of TLR3, MDA5, and pIRF3. This study provides novel insight into a hepatoprotective effect, whereby HSP60 inhibits the release of dsRNA to repress the TLR3/MDA5/pIRF3 pathway in the context of NAFLD or hepatic inflammation. Therefore, HSP60 may serve as a possible therapeutic target for improving NAFLD. [ABSTRACT FROM AUTHOR]

Objective Excessive glucocorticoid treatment increases the incidence of osteopenia and osteonecrosis. MicroRNAs (miRNAs) reportedly target messenger RNA expression and regulate osteoblastogenesis and skeletal development. We undertook this study to investigate whether miR-29a regulates glucocorticoid-mediated bone loss. Methods Rats were given methylprednisolone, lentivirus-mediated miR-29a precursor, or lentivirus-mediated miR-29a inhibitor. Dual x-ray absorptiometry, micro-computed tomography, material testing, and enzyme-linked immunosorbent assay were performed to quantify bone mass, microarchitecture, peak load, and serum Dkk-1 levels. Differential miRNA expression profiles were detected using polymerase chain reaction arrays. The abundance of signaling molecules was assessed using immunoblotting. Results Glucocorticoid treatment induced loss of bone mineral density and trabecular microstructure in association with reduced miR-29a expression. Treatment with miR-29a precursor attenuated the adverse effects of glucocorticoid on bone mass, trabecular bone volume fraction, and biomechanical load-bearing capacity of bone tissue. Gain of miR-29a function alleviated the detrimental effects of glucocorticoid treatment on mineral acquisition and ex vivo osteoblast differentiation, and also reduced osteoclast surface, ex vivo osteoclast differentiation, and RANKL expression in bone microenvironments. Knockdown of miR-29a accelerated osteoclast resorption, cortical bone porosity, bone fragility, and loss of ex vivo osteogenic differentiation capacity. MicroRNA-29a regulated the abundance of Wnt signaling components (Wnt-3a, glycogen synthase kinase 3β, and β-catenin), the Wnt inhibitor Dkk-1, Akt, and phosphorylated ERK, and the expression of the osteogenic factors RUNX-2 and insulin-like growth factor 1 in bone tissue. Conclusion MicroRNA-29a signaling protected against glucocorticoid-induced disturbance of Wnt and Dkk-1 actions and improved osteoblast differentiation and mineral acquisition. Promotion of miR-29a signaling is an alternative strategy for alleviating glucocorticoid-induced bone deterioration. [ABSTRACT FROM AUTHOR]

Abstract: Excessive glucocorticoid administration accelerates osteoblast apoptosis and skeletal deterioration. Heat shock proteins (HSPs) regulate metabolic activities in osteoblastic cells. This study characterized the biological significance of HSP60 in glucocorticoid-induced bone loss. Rats were treated with glucocorticoid, HSP60 antisense oligonucleotides, or adenovirus-mediated HSP60 gene transfer. Bone mineral density, metaphyseal trabecular micro-architecture, and fragility were analyzed by dual X-ray absorptiometry, micro-computed tomography, and material testing, respectively. Differential proteomic profiles of bone tissue extracts were detected by bi-dimensional electrophoresis and mass spectrometry. Survival and proapoptotic signal transduction were quantified by immunoblotting. Glucocorticoid-treated rats had low bone mineral density and metaphyseal trabecular microstructure in association with downregulation of collagen 1α1 and HSP60 expressions in bone tissue. Gain of HSP60 function by adenovirus-mediated HSP60 gene transfer abrogated the deleterious effects of glucocorticoid treatment on bone mass, trabecular microstructure, and mechanical strength. Enhancement of HSP60 signaling attenuated the glucocorticoid-induced loss of trabecular bone volume, mineral acquisition reactions and osteoblast surface. HSP60 gene transfer activated ERK and Akt and reduced Bax and cytochrome c release, as well as caspase-3 cleavage, which attenuated the inhibitory effects of glucocorticoid treatment on osteoblast survival. Loss of HSP60 function by HSP60 antisense oligonucleotides accelerated mitochondrial apoptotic programs and osteoblast apoptosis. Knockdown of HSP60 induced loss of bone mass, micro-architecture integrity, and mechanical property. Taken together, loss of HSP60 signaling contributes to the glucocorticoid-induced enhancement of pro-apoptotic reactions, thereby accelerating osteoblast apoptosis and bone mass loss. Enhancement of HSP60 function is beneficial for protecting bone tissue against the glucocorticoid-induced inhibition of bone cell viability and bone formation. [Copyright &y& Elsevier]

Abstract: Computer-aided process/solvent design is introduced to find a feasible biocompatible solvent for an extractive fermentation and separation process. The designed biocompatible solvent serves as both the extractant for extractive fermentation and the entrainer for extractive distillation, to yield water-free ethanol. Several goals, such as maximizing production rate and extraction efficiency, and limiting solvent utilization, are simultaneously considered in the optimal solvent design problem. Thus, the design was formulated as a mixed-integer nonlinear optimization problem. A two-phase computational scheme was introduced to solve the problem. The mixed-integer hybrid differential evolution (MIHDE) algorithm was first applied to solve the problem in order to obtain a feasible solution. The feasible solution was then served as an initial starting point for the mixed-integer sequential quadratic programming (MISQP) solver to numerically confirm that the optimal design was achieved. We have compared the crisp and fuzzy approaches to the design problem. The fuzzy goal attainment approach is able to address goal trade-offs and yield an overall satisfactory grade for the problem. [ABSTRACT FROM AUTHOR]

Abstract: In this study, we considered a multi-stage integrated extractive fermentation with cell recycling for ethanol production using the genetically engineered Sacchromyces yeast 1400 (pLNH33), which can utilize glucose and xylose as carbon sources to produce ethanol. Each stage consists of a stirred-tank bioreactor, a cell settler and an extractor. A generalized mathematical model was formulated to express the multi-stage integrated process. The aim of the optimization problem was to obtain the maximum overall productivity and conversions subject to the interval inequality constraints for the residual glucose and xylose concentrations and the total sugar supply. A fuzzy goal attainment method was applied to the multiobjective problem in order to achieve the maximum satisfaction for all design requirements. From the computational results, the integrated extractive fermentation with cell recycling (involving the extraction of ethanol from the extractor in situ to alleviate product inhibition) led to an optimal overall productivity that was 8.0% higher than that obtained by the method of continuous fermentation with cell recycling, and about 13-fold higher than that obtained by the method of continuous fermentation without cell recycling. [Copyright &y& Elsevier]

Abstract: Intensive bone cell apoptosis contributes to osteonecrosis of femoral head (ONFH). Dickkopf-1 (DKK1) reportedly mediates various types of skeletal disorders. This study investigated whether DKK1 was linked to the occurrence of ONFH. Thirty-nine patients with various stages of ONFH were recruited. Bone specimens were harvested from 34 ONFH patients underwent hip arthroplasty, and from 10 femoral neck fracture patients. Bad, Bcl2 TNFα, DKK1, Wnt3a, LRP5, and Axin1 expressions were analyzed by quantitative RT-PCR and ELISA. Apoptotic cells were assayed using terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end-labelling (TUNEL). Primary bone-marrow mesenchymal cells were treated with DKK1 RNA interference and recombinant DKK1 protein. ONFH patients with the histories of being administrated corticosteroids and excessive alcohol consumption had significantly higher Bad and DKK1 mRNA expressions in bone tissue and DKK1 abundances in serum than femoral neck fracture patients. Bone cells adjacent to osteonecrotic bone displayed strong DKK1 immunoreactivity and TUNEL staining. Increased DKK1 expression in bone tissue and serum correlated with Bad expression and TUNEL staining. Serum DKK1 abundance correlated with the severity of ONFH. The DKK1 RNA interference and recombinant DKK1 protein regulated Bad expression and apoptosis of primary bone-marrow mesenchymal cells. Knock down of DKK1 reduced dexamethasone-induced apoptosis of mesenchymal cells. Taken together, promoted DKK1 expression was associated with bone cell apoptosis in the occurrence of ONFH patients with the histories of corticosteroid and alcohol intake and progression of ONFH. DKK1 expression in injured tissue provides new insight into ONFH pathogenesis. [Copyright &y& Elsevier]

Background: Mathematical modeling has been applied to the study and analysis of complex biological systems for a long time. Some processes in biological systems, such as the gene expression and feedback control in signal transduction networks, involve a time delay. These systems are represented as delay differential equation (DDE) models. Numerical sensitivity analysis of a DDE model by the direct method requires the solutions of model and sensitivity equations with time-delays. The major effort is the computation of Jacobian matrix when computing the solution of sensitivity equations. The computation of partial derivatives of complex equations either by the analytic method or by symbolic manipulation is time consuming, inconvenient, and prone to introduce human errors. To address this problem, an automatic approach to obtain the derivatives of complex functions efficiently and accurately is necessary. Results: We have proposed an efficient algorithm with an adaptive step size control to compute the solution and dynamic sensitivities of biological systems described by ordinal differential equations (ODEs). The adaptive direct-decoupled algorithm is extended to solve the solution and dynamic sensitivities of time-delay systems describing by DDEs. To save the human effort and avoid the human errors in the computation of partial derivatives, an automatic differentiation technique is embedded in the extended algorithm to evaluate the Jacobian matrix. The extended algorithm is implemented and applied to two realistic models with time-delays: the cardiovascular control system and the TNF-α signal transduction network. The results show that the extended algorithm is a good tool for dynamic sensitivity analysis on DDE models with less user intervention. Conclusions: By comparing with direct-coupled methods in theory, the extended algorithm is efficient, accurate, and easy to use for end users without programming background to do dynamic sensitivity analysis on complex biological systems with time-delays. [ABSTRACT FROM AUTHOR]

Recent studies have shown dysbiosis is associated with inflammatory bowel disease (IBD). However, trying to restore microbial diversity via fecal microbiota transplantation (FMT) or probiotic intervention fails to achieve clinical benefit in IBD patients. We performed a probiotic intervention on a simulated IBD murine model to clarify their relationship. IBD was simulated by the protocol of azoxymethane and dextran sodium sulfate (AOM/DSS) to set up a colitis and colitis-associated neoplasm model on BALB/c mice. A single probiotic intervention using Clostridium butyricum Miyairi (CBM) on AOM/DSS mice to clarify the role of probiotic in colitis, colitis-associated neoplasm, gut microbiota, and immune cytokines was performed. We found dysbiosis occurred in AOM/DSS mice. The CBM intervention on AOM/DSS mice failed to improve colitis and colitis-associated neoplasms but changed microbial composition and unexpectedly increased expression of proinflammatory IL-17A in rectal tissue. We hypothesized that the probiotic intervention caused dysbiosis. To clarify the result, we performed inverse FMT using feces from AOM/DSS mice to normal recipients to validate the pathogenic effect of dysbiosis from AOM/DSS mice and found mice on inverse FMT did develop colitis and colon neoplasms. We presumed the probiotic intervention to some extent caused dysbiosis as inverse FMT. The role of probiotics in IBD requires further elucidation. [ABSTRACT FROM AUTHOR]

Abstract: Problems of parameter estimation of nonlinear bioreaction systems are in general formulated as function optimization problems and are known to be frequently ill-conditioned and multimodal. While the optimization problems are defined on a large parameter search space, only few evolutionary algorithms are able to find a global solution to the large parameter search problem. In this study, a geometric mean mutation was embedded to hybrid differential evolution to replace a gene of the selected individual outside the assigned region. The replaced individuals were then applied to a differential mutation strategy to yield a perturbed individual. The benefit of using a large parameter search space to an inverse problem is to reduce the kinetic model complexity to yield a more compact formulation. Two inverse problems and twelve static benchmark problems with the large parameter search space are employed to illustrate the effectiveness of the proposed method. [Copyright &y& Elsevier]

Abstract: Aims: Long-term glucocorticoid administration is known to induce bone deterioration. Glycogen synthase kinase-3β (GSK-3β) signaling reportedly participates in bone remodeling. This study investigated whether GSK-3β inhibitor could regulate glucocorticoid-induced inhibition of osteoblast differentiation in vitro or bone mass in vivo. Main methods: MC3T3-E1 osteoblasts were treated with kinase-inactive GSK-3β mutant and 6-bromoindirubin-3′-oxim (BIO) and then exposed to 1µM dexamethasone. Survival and osteoblast differentiation of cell cultures were assessed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end-labeling, quantitative RT-PCR, and von Kossa staining. Mineral density, biomechanical properties and microenvironments of BIO- and glucocorticoid-treated rat bone tissues were analyzed using dual-energy X-ray absorptiometry, material testing, and histomorphometry, respectively. Key findings: Glucocorticoid decreased levels of phosphorylated Ser9-GSK-3β and β-catenin in osteoblast cultures. Kinase-inactive GSK-3β mutant and BIO treatments attenuated dexamethasone-induced inhibition of β-catenin, Runx2 abundance, and osteoblast differentiation but suppressed glucocorticoid-induced apoptosis of cell cultures. Exogenous BIO treatment alleviated methylprednisolone-induced impairment of mineral density, biomechanical strength, trabecular bone volume, osteoblast surface, and bone formation rate of rat bone tissue. BIO treatment also attenuated glucocorticoid-induced promotion of osteoclast surface and marrow adipocyte volume in bone tissue. Bone cells adjacent to glucocorticoid-stressed bone tissue displayed strong phosphorylated Ser9-GSK-3β and β-catenin immunostaining following BIO treatment. Significance: Inhibition of GSK-3β abrogated glucocorticoid-induced bone loss by increasing β-catenin- and Runx2-mediated osteoblast differentiation. Controlling GSK-3β signaling in bone cells may be a strategy for preventing glucocorticoid-induced osteopenia. [Copyright &y& Elsevier]

Purpose: To evaluate the effect and its mechanism of aminoguanidine (AG) on small-bowel protection after whole-abdominal irradiation (WAI) in rats. Methods and Materials: Male Sprague-Dawley rats (300–400 g) subjected to 12 Gy WAI were used for the study. Aminoguanidine at a dose of 50–800 mg/kg was administered by the gavage route 2 h before WAI. Mucosal damage of small bowel was evaluated by the grade of diarrhea and crypt survival; oxidative stress was determined by the level of 8-hydroxy 2′-deoxyguanosine (8-OHdG) with immunohistochemistry (IHC). Nitrosative stress was evaluated by the expression of inducible nitric oxide synthase (iNOS) and 3-nitrotyrosine (3-NT) with IHC, and systemic and portal vein NOx (nitrite + nitrate) levels were measured and compared with and without AG treatment after WAI. Results: Aminoguanidine showed a dose-dependent effect against WAI-induced diarrhea. Aminoguanidine at a dose of 400 mg/kg had the best protective effect, from 92% to 17% (p = 0.002). Aminoguanidine increased crypt survival from 23% to 46% (p = 0.003). It also significantly attenuated 8-OHdG expression but not 3-NT and iNOS expression at both 4 and 8 h after 12-Gy WAI. Aminoguanidine did not alter the portal vein NOx levels 4 and 8 h after 12-Gy WAI. Conclusion: Aminoguanidine has a radioprotective effect against radiation-induced small-bowel damage due to its antioxidant effect but not inhibition of nitric oxide production. Dietary AG may have a potentially protective effect on the small intestine of patients subjected to pelvic and abdominal radiotherapies. [Copyright &y& Elsevier]

Abstract: In this study, the computer-aided process/solvent design is introduced to find an optimal biocompatible solvent and to maximize the ethanol production rate simultaneously for the single- or double-stage extractive fermentation process with cell recycling. Such a process/solvent design problem is formulated as a mixed-integer nonlinear programming problem that is solved by mixed-integer hybrid differential evolution in order to obtain a global design. The double-stage process can use a smaller amount of fresh solvent to increase ethanol productivity compared with that of the single-stage process, but it will also decrease overall conversion. Comparing the case studies, the simultaneous process/solvent design could yield higher overall ethanol productivity than that of the process design. The maximum ethanol production rate for the double-stage extractive fermentation with cell recycling was about 10-fold higher than that of continuous fermentation and about twofold higher than that of continuous fermentation with cell recycling. [Copyright &y& Elsevier]

Abstract: Osteonecrosis of the femoral head (ONFH) is a skeletal disorder characterized by ischemic deterioration, bone marrow edema and eventually femoral head collapse. The systemic regulation of ONFH in adult patients has not been examined. Serum proteomic is an innovative tool that potentially detects simultaneous expressions of serum proteins in pathological contexts. We compared the serum proteome profiles of 11 adult patients with ONFH (3 females and 8 males) and 11 healthy volunteers (3 females and 8 males). The proteins in the aliquots of sera were subjected to isoelectric focusing, two-dimensional gel electrophoresis and silver staining. The protein spots were matched and quantified using an imaging analysis system. The differentially expressed protein spots were subjected to in-gel trypsin digestion. The peptide mass fingerprints were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF/TOF) and a bioinformation search. We found that ONFH patients showed significantly higher abundances of kininogen 1 variant, complement factor C3 precursor, and complement factor H and lower levels of antithrombin III chain B, apolipoprotein A--IV precursor, and gelsolin isoform α precursor. These proteins of interest were reported to modulate thrombotic/fibrinolytic reactions, oxidative stress, vessel injury, tissue necrosis or cell apoptosis in several tissue types under pathological contexts. Taken together, the occurrence of ONFH was associated with various serum protein expressions. Our high--throughput serum proteomic findings indicated that multiple pathological reactions presumably occurred in ONFH. [Copyright &y& Elsevier]

We evaluated whether proinflammatory cytokine expression and myofibroblast recruitment in subacromial bursa was linked to rotator cuff lesions with shoulder stiffness. We analyzed expressions of IL-1β, IL-6, and TNF-α in subacromial bursa and joint fluid collected from 14 patients with cuff tears with stiffness as a study group (Group I) and 14 patients with rotator cuff tears without shoulder stiffness as a control group (Group II) using real-time RT-PCR, immunohistochemistry, and ELISA. Myofibroblast apoptosis in subacromial bursa was analyzed using terminal deoxynucleotidyl transferase -mediated deoxyuridine triphosphate-biotin nick end-labeling (TUNEL) and α-smooth muscle actin immunofluorescence staining. Shoulder function was evaluated using the Constant score. Group I had higher mRNA expression ( p < 0.001) and immunoreactivities ( p < 0.001) of IL-1β. They also had higher levels of IL-1β, IL-6, and TNF-α in joint fluid. Increased IL-1β mRNA expression in the subacromial bursa and IL-1β levels in joint fluid were correlated with a preoperative deficit in shoulder motion ( p < 0.001) and preoperative Constant scores ( p < 0.001). Immunofluorescence observations showed that Group I subjects had more myofibroblasts ( p < 0.001) than Group II. In Group II, a significant correlation was found between apoptotic myofibroblasts and total myofibroblasts ( p = 0.002), but not in Group I ( p = 0.510). Increased expression of IL-1β and myofibroblast recruitment in the subacromial bursa in rotator cuff lesions are linked to shoulder stiffness. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:1090-1097, 2008 [ABSTRACT FROM AUTHOR]

In this study, crisp and fuzzy multiple-goal optimization approaches are respectively introduced to design an optimal biocompatible solvent to a two-stage extractive fermentation with cell recycling for ethanol production. When designing a biocompatible solvent for the extractive fermentation process, many issues, such as extractive efficiency, conversion, amount of solvent utilized and so on, have to be considered. An interactive multiple-goal design procedure is introduced to determine a trade-off result in order to satisfy such contradicted goals. Both approaches could be iterated to solve the interactive multiple-goal design problem in order to yield a trade-off result. However, the crisp optimization design is a tedious task that requires the designer to provide various pairs of the upper bounds for the design problem to obtain the corresponding solution. The fuzzy optimization approach is able to be trade-off several goals simultaneously and to yield the overall satisfactory grade for the product/process design problem. [Copyright &y& Elsevier]

In this study, crisp and flexible optimization approaches are, respectively, introduced to design an optimal biocompatible solvent for an extractive fermentation process. The optimal design problem is formulated as a mixed-integer nonlinear programming model in which performance requirements of the compounds are reflected in the objective and the constraints. In general, the requirements for the objective and constraints are not rigid; consequently, the flexible or fuzzy optimization approach is applied to soften the rigid requirement for maximization of the extraction efficiency and to consider the mass flow rate and biocompatibility of solvent as the softened inequality constraints to the solvent design problem. Having elicited the membership function for the objective function and the constraint, the optimal solvent design problem can be formulated as a flexible goal attainment problem. Mixed-integer hybrid differential evolution is applied to solve the problem in order to find a satisfactory design. [Copyright &y& Elsevier]

Abstract: We analyze the growth dynamics and production of an industrially interesting amino acid in a recombinant Escherichia coli strain, using in parallel two alternative modeling frameworks, namely S-systems and lin-log models. These models have identical steady-state solutions, but differ in their representation of transients. The models were parameterized with data from two bioprocesses that differed in their initial culture concentrations and then used to predict responses under yet a different initial culture regimen. Among the S-system models, several alternatives representing slightly different pathway structures, were tested. All were found to be capable of capturing the dynamics of all variables in the test systems and also of predicting the dynamic responses under new conditions. The lin-log models also captured the dynamics, but not as well as the S-system models. A probable reason for the inferior performance of lin-log models is their intrinsic property of not representing situations well where variable concentrations are moderately small. [Copyright &y& Elsevier]

Abstract: Dickkopf-1 (DKK1) has been found to act as a potent Wnt signaling-inhibitory factor for regulating skeletal disorders. We investigated whether modulation of DKK1 expression by end-capped phosphorothioate DKK1 antisense oligonucleotide could alter estrogen loss-induced bone loss. Ovariectomized or sham-operated rats were given 20 μg/kg/day DKK1 sense or antisense oligonucleotide or vehicle for 28 days. Femurs and tibiae were dissected to assess bone mass, biomechanical strength, immunohistochemistry and ex vivo osteoclast formation. We found that DKK1 antisense oligonucleotide significantly abrogated the suppressing effect of ovariectomy on weight, mineral content, mineral density and peak load of femurs. DKK1 antisense oligonucleotide treatment reduced ovariectomy promotion of ex vivo osteoclast differentiation of primary M-CSF-dependent bone marrow macrophages. Histomorphometric observation demonstrated that DKK1 antisense oligonucleotide treatment increased osteoblast number and impaired ovariectomy-promoted trabecular bone loss and osteoclast number in bone tissue. Osteoblastic cells adjacent to endosteum of trabecular bone and chondrocytes at calcified cartilage expressed intensive DKK1 and RANKL and weak OPG immunostaining in ovariectomized rat bone microenvironments. Osteogenic cells and chondral cells displayed weak DKK1, RANKL and OPG expression of bone tissue after DKK1 antisense oligonucleotide treatment. Taken together, attenuation of DKK1 expression in ovariectomized rat bone tissue alleviated loss of bone mass and biomechanical property. The regulatory action of DKK1 antisense oligonucleotide treatment on bone tissue appeared to suppress the promoting effect of estrogen deficiency on osteoclastogenesis-stimulatory factor RANKL expression and osteoclast differentiation. Control of DKK1 signaling can be used in the future as an alternative strategy for protecting estrogen deficiency induction of bone loss. [Copyright &y& Elsevier]

Abstract: A two-phase design approach is introduced to determine the optimal feed rate, fed glucose concentration and fermentation time to maximize protein productivity using recombinant Escherichia coli BL21 (pBAW2) strain. The first phase is applied to determine a primary S-system kinetic model using batch time-series data. Two runs were carried out in the second phase to achieve the maximum protein productivity for the fed-batch fermentation process. The computational results using the S-system kinetic model obtained from the second run are in better agreement with the experiments than those using the kinetic model obtained from batch time-series data. For cross-validation, two extra fed-batch experiments with different feed strategies were carried out for comparison with the optimal fed-batch result. From the experimental results, this approach could improve productivity by at least 3%. [Copyright &y& Elsevier]

Abstract: In this study, we have established a suitable model to describe the dynamic characteristic of an aspartase-overproducing Escherichia coli strain. A traditional Monod model and power-law models are respectively applied to describe the dynamic behaviors. When the model structure was selected, the parameter values of the model should be determined by the global/local optimization method. There are two major challenges, numerical integration for differential equations and finding global parameter values. In this study, we introduce hybrid differential evolution, which is a variant of genetic algorithms, to avoid obtaining a premature solution. In addition, we apply a modified collocation method to avoid numerical integration. The Monod''s model could only predict the growth characteristic of the recombinant E. coli, qualitatively. The S-system representation could suit for constructing the model structure of the microbial growth. [Copyright &y& Elsevier]

Abstract: Purpose. This study was performed to investigate the effect of shock wave treatment on the healing at tendon–bone interface in rabbits. Materials and methods. Thirty-six New Zealand White rabbits were used in this study. The anterior cruciate ligament was excised and replaced with the long digital extensor. The right knees (study group) were treated with 500 impulses of shock waves at 14kV, while the left knees (control group) received no shock waves. Histomorphological studies were performed in 24 rabbits at 1, 2, 4, 8, 12 and 24 weeks. Biomechanical studies were performed in 12 rabbits at 12 and 24 weeks. Results. There was significantly more trabecular bone around the tendons noted in the study group compared with the control group at different time intervals after 4 weeks (P &#60;0.05). The contacting between bone and tendon was significantly better in the study group than the control group after 8 weeks (P &#60;0.05). The tensile strength of the tendon–bone interface was significantly higher in the study group than the control group at 24 weeks (P =0.018), whereas similar modes of graft failure were noted between the two groups. Conclusion. Shock wave treatment significantly improves the healing rate of the tendon–bone interface in a bone tunnel in rabbits. The effect of shock waves appears to be time-dependent. [Copyright &y& Elsevier]

Vascular endothelial growth factor (VEGF) is an important regulator for angiogenesis and endochondral bone formation. Although low-intensity pulsed ultrasound (US) has been recently used for accelerating fracture healing, the effect of US stimulation on angiogenic factor production by osteoblasts remains undetermined. Here, we found that US elevation of VEGF-A expression in human osteoblasts to be mediated by nitric oxide (NO) and hypoxia-inducible factor-1α (HIF-1α). Human osteoblasts were treated with or without US stimulation (200 μs pulse, 1 kHz at 30 mW/cm2) for 20 min. Cells were subjected to assessment of VEGF-A expression, NO production, nitric oxide synthase (NOS) catalytic activities, and HIF-1α transactivation. Results showed that US significantly increased VEGF-A mRNA and protein levels in 6 h. US augmentation of VEGF level was transcriptionally mediated. Early inhibition of NO production, but not calcium or prostaglandin E2, significantly reduced US-enhanced VEGF-A levels. Osteoblasts responded to US treatment by increasing NO production, NOS catalytic activities, iNOS immunoexpression, nuclear HIF-1α activation, and binding to the VEGF-A promoter. Inhibition of NOS activity by N-nitro-l-arginine methyl ester (l-NAME) or blockade of guanylate cyclase activity by ODQ reduced US-augmented HIF-1α transactivation and VEGF-A levels. Conditioned medium harvested from US-treated osteoblasts promoted tube formation of human umbilical vein endothelial cells (HUVEC). Monoclonal VEGF-A antibody neutralization or l-NAME pretreatment reduced the promoting effect of conditioned medium on angiogenesis of HUVEC. Together, these findings show that NO plays an important role in mediating extracellular stimuli released by US and triggering intracellular response of osteoblasts to produce angiogenic factor after US treatment. [Copyright &y& Elsevier]

: BackgroundRecent studies have shown that platelets play an important role in the pathogenesis of reperfusion injury. Using an inferior epigastric artery skin flap as a flap ischemia/reperfusion (I/R) injury model, we investigated whether the administration of a nitric oxide (NO) donor, nitrosoglutathione (GSNO), could decrease platelet activation and modulate the NO synthase (NOS) activity of platelets and promote flap survival.: MethodsThirty minutes before flap reperfusion, normal saline (1 mL), nitrosoglutathione (GSNO 0.2, 0.6, 3 mg/kg), or NG-nitro-l-arginine-methyl ester (450 mg/kg) was injected intravenously in 10 rats, respectively. The p-selectin (CD62P) expression of platelet activation was detected by a flow cytometry. Immunohistochemical staining was performed to investigate the CD62P deposition on the microvasculature of the flap vessels. NOS isoform expression in the platelets was evaluated by Western blot. Tissue perfusion was monitored by using laser-Doppler flowmetry. Survival areas were assessed at 7 days postoperatively: ResultsAn optimal dose of GSNO (0.6 mg/kg), significantly decreased in CD62P expression on platelets (P < 0.001) and its deposition on the flap vessels, selectively suppressed iNOS induction of platelet, and significantly improved blood perfusion and the flap survival rate (59.8 ± 4.9% versus 22.1 ± 6.1%, P < 0.001). In contrast, the NO synthase inhibitor, NG-nitro-l-arginine methyl ester, although inhibiting iNOS expression of platelets, compromised platelet activation, tissue perfusion, and flap survival.: ConclusionThis study suggests that GSNO can appropriately donate NO to suppress platelet activation and platelet iNOS induction, resulting in less platelet activation, better blood perfusion, and flap survival after I/R injury. [Copyright &y& Elsevier]

Despite the success in clinical application, the exact mechanism of shock wave therapy remains unknown. We hypothesized that shock wave therapy induces the ingrowth of neovascularization and improves blood supply to the tissues. The purpose of this study was to investigate the effect of shock wave therapy on neovascularization at the tendon–bone junction. Fifty New Zealand white rabbits with body weight ranging from 2.5 to 3.5 kg were used in this study. The right limb (the study side) received shock wave therapy to the Achilles tendon near the insertion to bone. The left limb (the control side) received no shock wave therapy. Biopsies of the tendon–bone junction were performed in 0, 1, 4, 8 and 12 weeks. The number of neo-vessels was examined microscopically with hematoxylin–eosin stain. Neovascularization was confirmed by the angiogenic markers including vessel endothelial growth factor (VEGF) and endothelial nitric oxide synthase (eNOS) expressions and endothelial cell proliferation determined by proliferating cell nuclear antigen (PCNA) expression examined microscopically with immunohistochemical stains. The results showed that shock wave therapy produced a significantly higher number of neo-vessels and angiogenesis-related markers including eNOS, VEGF and PCNA than the control without shock wave treatment. The eNOS and VEGF began to rise in as early as one week and remained high for 8 weeks, then declined at 12 weeks; whereas the increases of PCNA and neo-vessels began at 4 weeks and persisted for 12 weeks. In conclusion, shock wave therapy induces the ingrowth of neovascularization associated with early release of angiogenesis-related markers at the Achilles tendon–bone junction in rabbits. The neovascularization may play a role to improve blood supply and tissue regeneration at the tendon–bone junction. [Copyright &y& Elsevier]

Study is aimed to elucidate the effect of porcine leukocyte antimicrobial protein on the proliferation of Staphylococcus aureus and Escherichia coli inoculated in ground meats. The antimicrobial proteins were isolated from porcine blood using 0.2 M sodium acetate extraction and cation-exchange column chromatography. Antimicrobial protein preparation consisted predominantly of 7.5 and 6 kDa molecules. Immunoblotting demonstrated that the 7.5 kDa molecule displayed defensin immuno-reactivity, a unique blood granulocyte antimicrobial protein. Both 7.5 and 6 kDa molecules could decrease apparent proliferation of test cultures. Sterilization (121 °C, 30 min) and proteolytic enzymes (pepsin, trypsin, and chymotrypsin) could significantly decrease bacteriocidal activities of antimicrobial protein preparation. Furthermore, antimicrobial protein preparation enhanced exposure of intercellular nucleotide of test cultures. Antimicrobial protein preparation inhibition of S. aureus and E. coli growth was concentration and time dependence. Adding 160 μg/g antimicrobial protein preparation to ground ham meat and sausage mince could significantly (P<0.05) hurdle viable colony formation of test cultures at 6 and 12 h, respectively. Taken together, porcine leukocyte antimicrobial protein preparation has potential to inhibit proliferation of S. aureus and E. coli inoculated in ground meat via induction of bacterolysis, suggesting this antimicrobial protein preparation be an alternative natural bio-preservation source for ground meat products. [Copyright &y& Elsevier]

In this study, we consider a two-stage fermenter with cell recycling including an extractor to produce lactic acid. When the extractor was omitted, the proposed process was simplified to two special processes. Under the same operating conditions, we compared the overall lactic acid productivity and glucose conversion for the three processes. The proposed fermentation process was more efficient than the other processes. To simultaneously obtain the maximum productivity and conversion, the problem is formulated as a fuzzy multiobjective optimization problem. The fuzzy decision is introduced to convert the multiobjective fuzzy optimization problem into the fuzzy goal attainment problem. Hybrid differential evolution is applied to solve the fuzzy goal attainment problem to obtain a global Pareto solution. [Copyright &y& Elsevier]

In this study, we have introduced a fuzzy decision-making approach to design a multi-objective optimal design problem of a multiproduct batch chemical plant. In the crisp decision-making approach, the designer is first to solve the multi-objective optimization problem, and then to check whether each optimal objective function value satisfies the rigid preference goal. In real world application, the preference goal for each objective function is an interval bound not a rigid value so that the problem becomes a fuzzy goal optimization problem. A monotonic increasing or decreasing membership function is used to define the degree of satisfaction for each objective function so that the fuzzy goal optimization problem is then represented as an augmented minimax problem formulated as mixed-integer nonlinear programming (MINLP) models. To obtain a unique solution, we have introduced the mixed-integer hybrid differential evolution (MIHDE) to solve the MINLP problems. The MIHDE can straightforwardly solve the problem without any mathematical model transformation. Two examples were used to illustrate the applicability of the proposed interactive algorithm. [Copyright &y& Elsevier]

Glucocorticoid provokes bone mass loss and fatty marrow, accelerating osteoporosis development. Bromodomain protein BRD4, an acetyl–histone-binding chromatin reader, regulates stem cell and tissue homeostasis. We uncovered that glucocorticoid inhibited acetyl Lys-9 at the histone 3 (H3K9ac)-binding Runx2 promoter and decreased osteogenic differentiation, whereas bromodomain protein 4 (BRD4) and adipocyte formation were upregulated in bone-marrow mesenchymal progenitor cells. BRD4 knockdown improved H3K9ac occupation at the Runx2 promoter and osteogenesis, but attenuated glucocorticoid-mediated adipocyte formation together with the unaffected H3K9ac-binding PPARγ2 promoter. BRD4 regulated epigenome related to fatty acid metabolism and the forkhead box P1 (Foxp1) pathway, which occupied the PPARγ2 promoter to modulate glucocorticoid-induced adipocytic activity. In vivo, BRD4 inhibitor JQ-1 treatment mitigated methylprednisolone-induced suppression of bone mass, trabecular microstructure, mineral acquisition, and osteogenic differentiation. Foxp1 signaling, marrow fat, and adipocyte formation in glucocorticoid-treated skeleton were reversed upon JQ-1 treatment. Taken together, glucocorticoid-induced H3K9 hypoacetylation augmented BRD4 action to Foxp1, which steered mesenchymal progenitor cells toward adipocytes at the cost of osteogenic differentiation in osteoporotic skeletons. BRD4 inhibition slowed bone mass loss and marrow adiposity. Collective investigations convey a new epigenetic insight into acetyl histone reader BRD4 control of osteogenesis and adipogenesis in skeleton, and highlight the remedial effects of the BRD4 inhibitor on glucocorticoid-induced osteoporosis. [ABSTRACT FROM AUTHOR]

Recent studies have found that microRNA-29a (miR-29a) levels are significantly lower in fibrotic livers, as shown with human liver cirrhosis. Such downregulation influences the activation of hepatic stellate cells (HSC). Phosphoinositide 3-kinase p85 alpha (PI3KP85α) is implicated in the regulation of proteostasis mitochondrial integrity and unfolded protein response (UPR) and apoptosis in hepatocytes. This study aimed to investigate the potential therapeutic role of miR-29a in a murine bile duct ligation (BDL)-cholestatic injury and liver fibrosis model. Mice were assigned to four groups: sham, BDL, BDL + scramble miRs, and BDL + miR-29a-mimic. Liver fibrosis and inflammation were assessed by histological staining and mRNA/protein expression of representative markers. Exogenous therapeutics of miR-29a in BDL-stressed mice significantly attenuated glutamic oxaloacetic transaminase (GOT)/glutamic-pyruvic transaminase (GPT) and liver fibrosis, and caused a significant downregulation in markers related to inflammation (IL-1β), fibrogenesis (TGF-β1, α-SMA, and COL1α1), autophagy (p62 and LC3B II), mitochondrial unfolded protein response (UPRmt; C/EBP homologous protein (CHOP), heat shock protein 60 (HSP60), and Lon protease-1 (LONP1, a mitochondrial protease), and PI3KP85α within the liver tissue. An in vitro luciferase reporter assay further confirmed that miR-29a mimic directly targets mRNA 3′ untranslated region (UTR) of PI3KP85α to suppress its expression in HepG2 cell line. Our data provide new insights that therapeutic miR-29a improves cholestasis-induced hepatic inflammation and fibrosis and proteotstasis via blocking PI3KP85α, highlighting the potential of miR-29a targeted therapy for liver injury. [ABSTRACT FROM AUTHOR]

MicroRNA-29 (miR-29) has been shown to play a critical role in reducing inflammation and fibrosis following liver injury. Non-alcoholic fatty liver disease (NAFLD) occurs when fat is deposited (steatosis) in the liver due to causes other than excessive alcohol use and is associated with liver fibrosis. In this study, we asked whether miR-29a could reduce experimental high fat diet (HFD)-induced obesity and liver fibrosis in mice. We performed systematical expression analyses of miR-29a transgenic mice (miR-29aTg mice) and wild-type littermates subjected to HFD-induced NAFLD. The results demonstrated that increased miR-29a not only alleviated HFD-induced body weight gain but also subcutaneous, visceral, and intestinal fat accumulation and hepatocellular steatosis in mice. Furthermore, hepatic tissue in the miR-29aTg mice displayed a weak fibrotic matrix concomitant with low fibrotic collagen1α1 expression within the affected tissues compared to the wild-type (WT) mice fed the HFD diet. Increased miR-29a signaling also resulted in the downregulation of expression of the epithelial mesenchymal transition-executing transcription factor snail, mesenchymal markers vimentin, and such pro-inflammation markers as il6 and mcp1 within the liver tissue. Meanwhile, miR-29aTg-HFD mice exhibited significantly lower levels of peroxisome proliferator-activated receptor γ (PPARγ), mitochondrial transcription factor A TFAM, and mitochondria DNA content in the liver than the WT-HFD mice. An in vitro luciferase reporter assay further confirmed that miR-29a mimic transfection reduced fatty acid translocase CD36 expression in HepG2 cells. Conclusion: Our data provide new insights that miR-29a can improve HDF-induced obesity, hepatocellular steatosis, and fibrosis, as well as highlight the role of miR-29a in regulation of NAFLD. [ABSTRACT FROM AUTHOR]

Aim. We wish to investigate the therapeutic potential of a single-session high-energy extracorporeal shock wave therapy (ESWT) on the rotator cuff lesions with shoulder stiffness. Patients and Methods. Thirty-seven patients afflicted with rotator cuff lesions with shoulder stiffness were randomized to receive either shockwave or sham treatment based on statistical randomization. In the shockwave group, we used Orthospec™ Extracorporeal Shock Wave Therapy 3000 impulse 24 kV (0.32 mJ/mm2) focused at two points as one session. The sham intervention entailed the use of the device in which the silicone pad was removed from the stand-off device. The visual analogue scale (VAS), muscle power of the shoulder, Constant and Murley score (CMS), and range of motion (ROM) of the shoulder were assessed for all patients. Ten milliliters of peripheral venous blood was obtained from every participant for the measurements of markers for inflammation, tissue regeneration, angiogenesis, and substance P before and at 1 week and 4 weeks after intervention. Results. The ESWT group has significantly better VAS, muscle power, CMS, and ROM at 6 and 12 months after intervention. No between-group differences were observed before as well as 1 and 4 weeks after intervention in the selected biomarkers. Conclusion. ESWT may be a good adjuvant for the treatment of rotator cuff lesions with shoulder stiffness. [ABSTRACT FROM AUTHOR]

Aim. We wish to investigate the therapeutic potential of a single-session high-energy extracorporeal shock wave therapy (ESWT) on the rotator cuff lesions with shoulder stiffness. Patients and Methods. Thirty-seven patients afflicted with rotator cuff lesions with shoulder stiffness were randomized to receive either shockwave or sham treatment based on statistical randomization. In the shockwave group, we used Orthospec™ Extracorporeal Shock Wave Therapy 3000 impulse 24 kV (0.32 mJ/mm2) focused at two points as one session. The sham intervention entailed the use of the device in which the silicone pad was removed from the stand-off device. The visual analogue scale (VAS), muscle power of the shoulder, Constant and Murley score (CMS), and range of motion (ROM) of the shoulder were assessed for all patients. Ten milliliters of peripheral venous blood was obtained from every participant for the measurements of markers for inflammation, tissue regeneration, angiogenesis, and substance P before and at 1 week and 4 weeks after intervention. Results. The ESWT group has significantly better VAS, muscle power, CMS, and ROM at 6 and 12 months after intervention. No between-group differences were observed before as well as 1 and 4 weeks after intervention in the selected biomarkers. Conclusion. ESWT may be a good adjuvant for the treatment of rotator cuff lesions with shoulder stiffness. [ABSTRACT FROM AUTHOR]

Spinal epidural fibrosis is one of the typical features attributable to failed back surgery syndrome, with excessive scar development in the dura and nerve roots. The microRNA-29 family (miR-29s) has been found to act as a fibrogenesis-inhibitory factor that reduces fibrotic matrix overproduction in various tissues. However, the mechanistic basis of miRNA-29a underlying the overabundant fibrotic matrix synthesis in spinal epidural scars post-laminectomy remained elusive. This study revealed that miR-29a attenuated lumbar laminectomy-induced fibrogenic activity, and epidural fibrotic matrix formation was significantly lessened in the transgenic mice (miR-29aTg) as compared with wild-type mice (WT). Moreover, miR-29aTg limits laminectomy-induced damage and has also been demonstrated to detect walking patterns, footprint distribution, and moving activity. Immunohistochemistry staining of epidural tissue showed that miR-29aTg was a remarkably weak signal of IL-6, TGF-β1, and DNA methyltransferase marker, Dnmt3b, compared to the wild-type mice. Taken together, these results have further strengthened the evidence that miR-29a epigenetic regulation reduces fibrotic matrix formation and spinal epidural fibrotic activity in surgery scars to preserve the integrity of the spinal cord core. This study elucidates and highlights the molecular mechanisms that reduce the incidence of spinal epidural fibrosis, eliminating the risk of gait abnormalities and pain associated with laminectomy. [ABSTRACT FROM AUTHOR]

Identifying essential targets in the genome-scale metabolic networks of cancer cells is a time-consuming process. The present study proposed a fuzzy hierarchical optimization framework for identifying essential genes, metabolites and reactions. On the basis of four objectives, the present study developed a framework for identifying essential targets that lead to cancer cell death and evaluating metabolic flux perturbations in normal cells that have been caused by cancer treatment. Through fuzzy set theory, a multiobjective optimization problem was converted into a trilevel maximizing decision-making (MDM) problem. We applied nested hybrid differential evolution to solve the trilevel MDM problem to identify essential targets in genome-scale metabolic models for five consensus molecular subtypes (CMSs) of colorectal cancer. We used various media to identify essential targets for each CMS and discovered that most targets affected all five CMSs and that some genes were CMS-specific. We obtained experimental data on the lethality of cancer cell lines from the DepMap database to validate the identified essential genes. The results reveal that most of the identified essential genes were compatible with the colorectal cancer cell lines obtained from DepMap and that these genes, with the exception of EBP, LSS, and SLC7A6, could generate a high level of cell death when knocked out. The identified essential genes were mostly involved in cholesterol biosynthesis, nucleotide metabolisms, and the glycerophospholipid biosynthetic pathway. The genes involved in the cholesterol biosynthetic pathway were also revealed to be determinable, if a cholesterol uptake reaction was not induced when the cells were in the culture medium. However, the genes involved in the cholesterol biosynthetic pathway became non-essential if such a reaction was induced. Furthermore, the essential gene CRLS1 was revealed as a medium-independent target for all CMSs. [ABSTRACT FROM AUTHOR]

Background: Despite advances in treatment, patients with refractory colorectal cancer (CRC) still have poor long-term survival, so there is a need for more effective therapeutic options. Methods: To evaluate the HDAC8 inhibition efficacy as a CRC treatment, we examined the effects of various HDAC8 inhibitors (HDAC8i), including BMX (NBM-T-L-BMX-OS01) in combination with temozolomide (TMZ) or other standard CRC drugs on p53 mutated HT29 cells, as well as wild-type p53 HCT116 and RKO cells. Results: We showed that HDAC8i with TMZ cotreatment resulted in HT29 arrest in the S and G2/M phase, whereas HCT116 and RKO arrest in the G0/G1 phase was accompanied by high sub-G1. Subsequently, this combination approach upregulated p53-mediated MGMT inhibition, leading to apoptosis. Furthermore, we observed the cotreatment also enabled triggering of cell senescence and decreased expression of stem cell biomarkers. Mechanistically, we found down-expression levels of β-catenin, cyclin D1 and c-Myc via GSK3β/β-catenin signaling. Intriguingly, autophagy also contributes to cell death under the opposite status of β-catenin/p62 axis, suggesting that there exists a negative feedback regulation between Wnt/β-catenin and autophagy. Consistently, the Gene Set Enrichment Analysis (GSEA) indicated both apoptotic and autophagy biomarkers in HT29 and RKO were upregulated after treating with BMX. Conclusions: BMX may act as a HDAC8 eraser and in combination with reframed-TMZ generates a remarkable synergic effect, providing a novel therapeutic target for various CRCs. 12mq7khsyzMPAFsnG-a-Nk Video Abstract [ABSTRACT FROM AUTHOR]

Parkinson disease (PD) is the second-most common neurodegenerative disease. The characteristic pathology of progressive dopaminergic neuronal loss in people with PD is associated with iron accumulation and is suggested to be driven in part by the novel cell death pathway, ferroptosis. A unique modality of cell death, ferroptosis is mediated by iron-dependent phospholipid peroxidation. The mechanisms of ferroptosis inhibitors enhance antioxidative capacity to counter the oxidative stress from lipid peroxidation, such as through the system xc−/glutathione (GSH)/glutathione peroxidase 4 (GPX4) axis and the coenzyme Q10 (CoQ10)/FSP1 pathway. Another means to reduce ferroptosis is with iron chelators. To date, there is no disease-modifying therapy to cure or slow PD progression, and a recent topic of research seeks to intervene with the development of PD via regulation of ferroptosis. In this review, we provide a discussion of different cell death pathways, the molecular mechanisms of ferroptosis, the role of ferroptosis in blood–brain barrier damage, updates on PD studies in ferroptosis, and the latest progress of pharmacological agents targeting ferroptosis for the intervention of PD in clinical trials. [ABSTRACT FROM AUTHOR]

A correction to the article "Construction of lentivirusmediated miR-29a precursor and inhibitor expression vectors in the June 2013 issue is presented.