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Objectives Exosomal Phospho-Tau-181(P-T181-tau), Total tau (T-tau), and amyloid-β peptide 42 (Aβ42) have been proved the capacity for the amnestic mild cognitive impairment (MCI) and the diagnosis of Alzheimer's disease (AD). This study aimed to explore the cognitive function and the levels of P-T181-tau, T-tau, and Aβ42 in neuronal-derived exosomes (NDEs) extracted from plasma in normal cognitive adults over 45 years old with olfactory dysfunction. Methods A cross-sectional survey of 29 participants aged over 45 was conducted. Plasma exosomes were isolated, precipitated, and enriched by immuno-absorption with anti- L1 cell adhesion molecule (L1CAM) antibody. NDEs were characterized by CD81, and extracted NDE protein (P-T181-tau, T-tau, and Aβ42) biomarkers were quantified by enzyme-linked immunosorbent assay (ELISAs). Olfactory performance was assessed by Sniffin' Sticks and cognitive performance was assessed by Montreal Cognitive Assessment (MoCA). Results There was no significant difference between adults with olfactory dysfunction and without olfactory dysfunction regarding the cognitive function as measured by MoCA and all the participants showed normal cognition. Adults with olfactory dysfunction showed a higher concentration of P-T181-tau in plasma NDEs than did adults without olfactory dysfunction (P = 0.034). Both the levels of P-T181-tau (r = - 0.553, P = 0.003) and T-tau (r = - 0.417, P = 0.034) negatively correlated with the odor identification scores. In addition, the level of T-tau negatively correlated with MoCA scores (r = - 0.597, P = 0.002). The levels of P-T181-tau (r = - 0.464, P = 0.022) and T-tau (r = - 0.438, P = 0.032) negatively correlated with the delayed recall scores. Conclusions This study demonstrated that cognition-related pathogenic proteins including P-T181-tau in plasma NDEs were significantly increased in adults over 45 years old with olfactory dysfunction before the occurrence of cognitive impairment. The impaired odor identification and the delayed recall function were highly associated with the increased levels of P-T181-tau and T-tau in plasma NDEs.

Soft-tissue sarcomas (STS) are rare malignancies showing lineage differentiation toward diverse mesenchymal tissues. Half of all high-grade STSs develop lung metastasis with a median survival of 15 months. Here, we used a genetically engineered mouse model that mimics undifferentiated pleomorphic sarcoma (UPS) to study the molecular mechanisms driving metastasis. High-grade sarcomas were generated with Cre recombinase technology using mice with conditional mutations in Kras and Trp53 (KP) genes. After amputation of the limb bearing the primary tumor, mice were followed for the development of lung metastasis. Using RNA-sequencing of matched primary KP tumors and lung metastases, we found that the long noncoding RNA (lncRNA) Nuclear Enriched Abundant Transcript 1 (Neat1) is significantly upregulated in lung metastases. Furthermore, NEAT1 RNA ISH of human UPS showed that NEAT1 is upregulated within a subset of lung metastases compared with paired primary UPS. Remarkably, CRISPR/Cas9-mediated knockout of Neat1 suppressed the ability of KP tumor cells to colonize the lungs. To gain insight into the underlying mechanisms by which the lncRNA Neat1 promotes sarcoma metastasis, we pulled down Neat1 RNA and used mass spectrometry to identify interacting proteins. Interestingly, most Neat1 interacting proteins are involved in RNA splicing regulation. In particular, KH-Type Splicing Regulatory Protein (KHSRP) interacts with Neat1 and is associated with poor prognosis of human STS. Moreover, depletion of KHSRP suppressed the ability of KP tumor cells to colonize the lungs. Collectively, these results suggest that Neat1 and its interacting proteins, which regulate RNA splicing, are involved in mediating sarcoma metastasis.Implications:Understanding that lncRNA NEAT1 promotes sarcoma metastasis, at least in part, through interacting with the RNA splicing regulator KHSRP may translate into new therapeutic approaches for sarcoma.

Background As fat and obesity play a vital role in the pathophysiology of metabolic dysfunction-associated fatty liver disease (MAFLD), this study aims to investigate the association between the fat mass and obesity-associated gene (FTO) and MAFLD. Methods Six SNPs (rs6499640, rs1421085, rs8050136, rs3751812, rs9939609 and rs9930506) within FTO were genotyped for 741 MAFLD patients (median age, 69.98; interquartile range, 66.55-75.93) and 825 healthy people (median age, 69.94; interquartile range, 66.39-75.64). Allele and genotype frequencies, pairwise linkage disequilibrium (LD) and haplotype analysis were calculated. Results BMI, waist circumference, systolic blood pressure, diastolic blood pressure, fasting plasma glucose, triglyceride, alanine transaminase, glutamyl transpeptidase and the prevalence of diabetes were found to be higher in the MAFLD individuals comparing to the control ones (P 0.8), and individuals with C-A-T-A haplotype had an elevated MAFLD risk (P =0.005). Conclusion The case-control study indicated that C variant of rs1421085, A variant of rs8050136, T variant of rs3751812 and A variant of rs9939609 are associated with elevated MAFLD risk in the older Chinese Han population.

During neocortical development, neural stem cells (NSCs) divide symmetrically to self-renew at the early stage and then divide asymmetrically to generate post-mitotic neurons. The molecular mechanisms regulating the balance between NSC self-renewal and neurogenesis are not fully understood. Using mouse in utero electroporation (IUE) technique and in vitro human NSC differentiation models including cerebral organoids (hCOs), we show here that regulator of cell cycle (RGCC) modulates NSC self-renewal and neuronal differentiation by affecting cell cycle regulation and spindle orientation. RGCC deficiency hampers normal cell cycle process and dysregulates the mitotic spindle, thus driving more cells to divide asymmetrically. These modulations diminish the NSC population and cause NSC pre-differentiation that eventually leads to brain developmental malformation in hCOs. We further show that RGCC might regulate NSC spindle orientation by affecting the organization of centrosome and microtubules. Our results demonstrate that RGCC is essential to maintain the NSC pool during cortical development and suggest that RGCC defects could have etiological roles in human brain malformations.

A label-free sensing strategy based on the enzyme-mimicking property of Glutathione-Ag nanoparticles (GSH-AgNPs) was demonstrated for colorimetric detection of vitamin B1 (VB1). Firstly, obvious blue color accompanied with an absorption peak at 652 nm was observed due to the high peroxidase-like activity of GSH-AgNPs towards 3,3′,5,5′-tetramethylbenzidine (TMB). Then, in the presence of VB1, the mimetic activity of GSH-AgNPs could be strongly restrained, evidenced as a promiment colorimetric change to colorless, which can be used to achieve the visualization detection VB1. Linear relationship between absorbance response and VB1 concentration from 0 to 0.2 µM were obtained. The detection limit was calculated as low as 40 nM. The inhibition reasons were thoroughly discussed. Considering the advantages of rapid response, easy procedure and high selectivity, the proposed method possesses potential application in environment and biological analysis for VB1 detection.

High quality is the main goal of today’s maize breeding and the investigation of grain quality traits would help to breed high-quality varieties in maize. In this study, genome-wide association studies in a set of 248 diverse inbred lines were performed with 83,057 single nucleotide polymorphisms (SNPs), and five grain quality traits were investigated in diverse environments for two years. The results showed that maize inbred lines showed substantial natural variations of grain quality and these traits showed high broad-sense heritability. A total of 49 SNPs were found to be significantly associated with grain quality traits. Among these SNPs, four co-localized sites were commonly detected by multiple traits. The candidate genes which were searched for can be classified into 11 biological processes, 13 cellular components, and 6 molecular functions. Finally, we found 29 grain quality-related genes. These genes and the SNPs identified in the study would offer essential information for high-quality varieties breeding programs in maize.

Flow has been widely studied in the field of positive psychology. However, little is known regarding its biological mechanism. This study aimed to ascertain flow-related gene loci. We investigated the association between flow and five single nucleotide polymorphisms associated with common mental disorders among a sample of 870 healthy 1st year students of Jining Medical University, Shandong Province, China. This study was approved by the Ethics Committee of Jining Medical University (approval No. JNMC-2016-KY-001) on June 1, 2016. rs11191454 demonstrated significant statistical association with flow after adjusting for age and gender (P = 0.004). The allele carriers achieved higher scores in all four dimensions of flow: merging of action and awareness, challenge-skill balance, sense of control, and clear goals. This biological research article indicates that rs11191454 in the arsenite methyltransferase (AS3MT) gene might be associated with flow in a Chinese Han population, and that might result from altered arsenic metabolism.

Background: Schizophrenia (SCZ) is a complex, heritable, and devastating psychiatric disorder. Mutations in the members of ABC transporters have been associated with psychiatric illnesses. Aims: In this study, we investigated whether 9 SNPs in ABCB1 (rs6946119, rs28401781, rs4148739, and rs3747802), ABCB6 (rs1109866, rs1109867, rs3731885, and rs3755047), and ABCG1 (rs182694) contribute to the risk of SCZ in a Han Chinese population. Methods: We conducted a case-control study in a Han Chinese population, involving 1,034 SCZ patients and 1,034 unrelated healthy controls to genotype 9 SNPs. Results: The analysis demonstrated that rs182694 of ABCG1 was significantly different between SCZ patients and controls as to allele (rs182694: p = 0.0070, χ2 = 7.27) and genotype frequencies (rs182694: p = 0.0013, χ2 = 13.35). Conclusions: Our findings support an association between ABCG1 polymorphism and SCZ in a Han Chinese population.

Background Mucormycosis is a rare fungal infection occurring chiefly in the lung or the rhino-orbital-cerebral compartment, particularly in patients with immunodeficiency or diabetes mellitus. Among Mucorales fungi, Rhizopus spp. are the most common cause of mucormycosis. Case presentation We report a case of pulmonary mucormycosis caused by Rhizopus microsporus in a young patient with diabetes but no other apparent risk factors. The diagnosis mainly relied on clinical manifestation, positive pulmonary tissue biopsy, and fungal culture. The patient was successfully treated with posaconazole oral suspension and remains asymptomatic at one-year follow-up. Conclusions Pulmonary mucormycosis is a life-threatening condition and posaconazole is an effective treatment for pulmonary mucormycosis caused by Rhizopus microspores.

T cell receptor (TCR) knockout is a critical step in producing universal chimeric antigen receptor T cells for cancer immunotherapy. A promising approach to achieving the knockout is to deliver the CRISPR/Cas9 system into cells using electrotransfer technology. However, clinical applications of the technology are currently limited by the low cell viability. In this study, we attempt to solve the problem by screening small molecule drugs with an immortalized human T cell line, Jurkat clone E6-1, for inhibition of apoptosis. The study identifies a few caspase inhibitors that could be used to simultaneously enhance the cell viability and the efficiency of plasmid DNA electrotransfer. Additionally, we show that the enhancement could be achieved through knockdown of caspase 3 expression in siRNA treated cells, suggesting that the cell death in electrotransfer experiments was caused mainly by caspase 3-dependent apoptosis. Finally, we investigated if the caspase inhibitors could improve TCR gene-editing with electrotransferred ribonucleoprotein, a complex of Cas9 protein and a T cell receptor-&alpha, constant (TRAC)-targeting single guide RNA (sgRNA). Our data showed that inhibition of caspases post electrotransfer could significantly increase cell viability without compromising the TCR disruption efficiency. These new findings can be used to improve non-viral T cell engineering.

Electrotransfection is a widely used method for delivering genes into cells with electric pulses. Although different hypotheses have been proposed, the mechanism of electrotransfection remains controversial. Previous studies have indicated that uptake and intracellular trafficking of plasmid DNA (pDNA) are mediated by endocytic pathways, but it is still unclear which pathways are directly involved in the delivery. To this end, the present study investigated the dependence of electrotransfection on macropinocytosis. Data from the study demonstrated that electric pulses induced cell membrane ruffling and actin cytoskeleton remodeling. Using fluorescently labeled pDNA and a macropinocytosis marker (i.e., dextran), the study showed that electrotransfected pDNA co-localized with dextran in intracellular vesicles. Furthermore, electrotransfection efficiency could be decreased significantly by reducing temperature or treatment of cells with a pharmacological inhibitor of Rac1 and could be altered by changing Rac1 activity. Taken together, the findings suggested that electrotransfection of pDNA involved Rac1-dependent macropinocytosis.

Venlafaxine is one of commonly prescribed antidepressants for major depressive disorder (MDD). Accumulated evidence implicates the involvement of glutamatergic receptors in the pathophysiology of MDD and antidepressant treatment. By using 193 MDD patients who have been taking venlafaxine for 6 weeks, we investigated whether single nucleotide polymorphisms (SNPs) in glutamate ionotropic receptor kainate type subunit 4 (GRIK4), glutamate ionotropic receptor AMPA type subunit 1 (GRIA1) and glutamate metabotropic receptor 7 (GRM7) were associated with treatment response. 14 SNPs were selected randomly depended on association studies. Efficacy of treatment was determined by 17-item of Hamilton Rating Scale. Allele and genotype frequencies were compared between responders and non-responders. After adjusting by the false discovery rate (FDR), rs6589847 and rs56275759 in GRIK4 and rs9870680 in GRM7 showed associating with venlafaxine treatment response at week 6. (FDR: P = .018, P = .042, and P = .040, respectively). Our results indicated that genetic variants in the GRIK4 and GRM7 may associate with the treatment response in MDD patients treated by venlafaxine.

The distal long arm of chromosome 22 (22q13.3) may harbor genes implicated in schizophrenia. This is evidenced by various genetic mapping studies. BRD1 and its neighboring gene ZBED4, both located within this region, have repeatedly been found to be associated with schizophrenia in the Caucasian population. In this study, we chose seven SNPs (two BRD1 SNPs, five ZBED4 SNPs) to carry out an association study between these two genes and schizophrenia in the Chinese population. However, no significant result was obtained, which was consistent with the Japanese population. Taken together, we could conclude that BRD1 and ZBED4 might be population specific in schizophrenia and may not account for a substantial proportion of genetic risk for schizophrenia in the Asian population.

Human dilated cardiomyopathy (DCM) is characterized by congestive heart failure and altered myocardial gene expression. Epigenetic changes, including DNA methylation, are implicated in the development of DCM but have not been studied extensively. Clinical human DCM and nonfailing control left ventricle samples were individually analyzed for DNA methylation and expressional changes. Expression microarrays were used to identify 393 overexpressed and 349 underexpressed genes in DCM (GEO accession number: GSE43435 ). Gene promoter microarrays were utilized for DNA methylation analysis, and the resulting data were analyzed by two different computational methods. In the first method, we utilized subtractive analysis of DNA methylation peak data to identify 158 gene promoters exhibiting DNA methylation changes that correlated with expression changes. In the second method, a two-stage approach combined a particle swarm optimization feature selection algorithm and a discriminant analysis via mixed integer programming classifier to identify differentially methylated gene promoters. This analysis identified 51 hypermethylated promoters and six hypomethylated promoters in DCM with 100% cross-validation accuracy in the group assignment. Generation of a composite list of genes identified by subtractive analysis and two-stage computation analysis revealed four genes that exhibited differential DNA methylation by both methods in addition to altered gene expression. Computationally identified genes ( AURKB, BTNL9, CLDN5, and TK1) define a central set of differentially methylated gene promoters that are important in classifying DCM. These genes have no previously reported role in DCM. This study documents that rigorous computational analysis applied to microarray analysis of healthy and diseased human heart samples helps to define clinically relevant DNA methylation and expressional changes in DCM.

The influence of the mowing agronomic measure on the quality of Saussurea involucrata was studied in Urumqi, Xinjiang using the method of combining biotechnology with botany, in order to reveal the effect of mowing strength on flavonoids and polysaccharide content in S. involucrata. The results show that S. involucrata has strong regeneration ability after mowing and it can be mown four times in a year. And the flavonoids content decreases while its polysaccharide content increases generally under different mowing times. In addition, the flavonoids content decreases in general, while its polysaccharide content reduces at first and then greatly increases with mowing frequency increases at the same study area. However, their flavonoids content meets medicinal quality standards of pharmacopoeia of the People's Republic of China in 2010 (flavonoid≥0.15％). For S. involucrate with longer growth period in whole life, it can get higher production of chemical composition by mowing. The results have important application value in production practice. It can provide a case of mowing scheme for the sustainable efficient use and quality control for the medicinal materials of S. involucrate.

Three-dimensional (3D) tumor models have been established in various microfluidic systems for drug delivery and resistance studies in vitro. However, one of the main drawbacks of these models is non-uniform distribution of cells, leaving regions with very low cell density within the 3D structures. As a result, molecular diffusion in the cell compartments is faster than that observed in solid tumors. To solve this problem, we developed a new technique for preparation of 3D tumor models in vitro. It was based on a microfluidic device containing three parallel channels separated by narrowly spaced posts. Tumor cells were loaded into the central channel at high density. To test the system, B16.F10 melanoma cells were perfusion-cultured overnight and the resulting 3D structure was characterized in terms of viability, density, and morphology of cells as well as transport properties of small fluorescent molecules. Immediately upon loading of tumor cells, the cell density was comparable to those observed in B16.F10 tumor tissues in vivo; and the viability of tumor cells was maintained through the overnight culture. The tumor model displayed low extracellular space and high resistance to diffusion of small molecules. For membrane-permeant molecules (e.g., Hoechst 33342), the rate of interstitial penetration was extremely slow, compared to membrane-impermeant molecules (e.g., sodium fluorescein). This versatile tumor model could be applied to in vitro studies of transport and cellular uptake of drugs and genes.

Pulsed electric fields can enhance interstitial transport of plasmid DNA (pDNA) in solid tumors. However, the extent of enhancement is still limited. To this end, the effects of cellular resistance to electric field-mediated gene delivery were investigated. The investigation used two tumor cell lines (4T1 (a murine mammary carcinoma) and B16.F10 (a metastatic subline of B16 murine melanoma)) either in suspensions or implanted in two in vivo models (dorsal skin-fold chamber (DSC) and hind leg). The volume fraction of cells was altered by pretreatment with a hyperosmotic mannitol solution (1 M). It was observed that the pretreatment reduced the volumes of 4T1 and B16.F10 cells, suspended in an agarose gel, by 50 and 46%, respectively, over a 20-min period, but did not cause significant changes ex vivo in volumes of hind-leg tumor tissues grown from the same cells in mice. The mannitol pretreatment in vivo improved electric field-mediated gene delivery in the hind-leg tumor models, in terms of reporter gene expression, but resulted in minimal enhancement in pDNA electrophoresis over a few microns distance in the DSC tumor models. These data demonstrated that hyperosmotic mannitol solution could effectively improve electric field-mediated gene delivery around individual cells in vivo by increasing the extracellular space.

Objective: To explore the therapeutic effect and mechanism of Gynostemma Pentaphyllum (GSM) on spleen-qi deficiency mice. Method: Mice were randomly divided into 4 groups: normal group, model group, Sijunzi decoction, Gynostemma Pentaphyllum group. The mice were given rhubarb decoction at the concentration of 1.56 g/kg by gavage for 5 days to establish spleen-qi deficiency mice model. The basic characteristics of weight, rectal temperature, stool, appetite and other characteristics of mice were observed. Swimming time, Water Maze Experiment, Open Field Test and serum immunoglobulin were detected. Result: Compared with model group, Gynostemma Pentaphyllum can significantly improve body weight of mice. Compared with model group, the swimming time of Gynostemma Pentaphyllum group was significantly increased on the fourth day, the vertical movement time and the number of swimming cycles, as well as the concentration of immunoglobulin IgG in the Gynostemma Pentaphyllum group was increased. Conclusion: Gynostemma Pentaphyllum has a certain effect on the overall treatment of spleen-qi deficiency mice. The therapeutic effect of Gynostemma Pentaphyllum on spleen-qi deficiency mice may be related to increase the immunoglobulin levels.

CRP3 is the muscle-specific form of the cysteine and glycine-rich protein family and plays an important role in myofiber differentiation. Here we isolated and characterized its coding gene CSRP3 from porcine muscle. Phylogenic analyses demonstrated that CSRP3 diverged first and is distinguished from two other members, CSRP1 and CSRP2. CSRP3 mRNA was up-regulated during the development of porcine embryonic skeletal muscle, indicating its potential importance in muscle growth. Genetic variant analyses detected multiple variations in an approximately 400 bp region covering exon 4 and its downstream intron, and two haplotypes were identified by sequencing. One of synonymous substitutions C1924T was used for linkage and association analyses. It was revealed that the substitution of C1924T had significant associations with firmness (P < 0.01), Lab Loin pH, Off Flavor Score and Water Holding Capacity (P < 0.05), and a suggestive effect (P < 0.1) on Flavor Score and Average Glycolytic Potential in a Berkshire × Yorkshire F2 population. The association analyses results agreed with the gene’s localization to a QTL region for meat quality traits on porcine chromosome 2p14-17 demonstrated by both linkage mapping and RH mapping. These results provide fundamental evidence for CSRP3 as a functional candidate gene affecting pig meat quality.

In addition to its established role in initiating the endocrine arm of the stress response, corticotropin-releasing factor (CRF) can act in the brain to modulate neural pathways that effect coordinated physiological and behavioral adjustments to stress. Although CRF is expressed in a set of interconnected limbic and autonomic cell groups implicated as primary sites of stress-related peptide action, most of these are lacking or impoverished in CRF receptor (CRFR) expression. Understanding the distribution of functional receptor expression has been hindered by the low resolution of ligand binding approaches and the lack of specific antisera, which have supported immunolocalizations at odds with analyses at the mRNA level. We have generated a transgenic mouse that reports expression of the principal, or type 1, CRFR (CRFR1). This mouse expresses GFP in a cellular distribution that largely mimics that of CRFR1 mRNA and is extensively colocalized with it in individual neurons. GFP-labeled cells display indices of activation (Fos induction) in response to central CRF injection. At the cellular level, GFP labeling marks somatic and proximal dendritic morphology with high resolution, and is also localized to axonal projections of at least some labeled cell groups. This includes a presence in synaptic inputs to central autonomic structures such as the central amygdalar nucleus, which is implicated as a stress-related site of CRF action, but lacks cellular CRFR1 expression. These findings validate a new tool for pursuing the role of central CRFR signaling in stress adaptation, and suggest means by which the pervasive ligand-receptor mismatch in this system may be reconciled.

Understanding mechanisms of formation of vascular networks under different experimental conditions is essential for improving treatment of angiogenesis-dependent diseases. To this end, we investigated the dose response of angiogenesis to basic fibroblast growth factor (bFGF) using the rat corneal pocket assay. The response was quantified, in terms of (i) the migration distance of vascular networks, (ii) the total vessel length, (iii) the distribution of the projected width of vessels, (iv) the distribution of the number of vessels, and (v) the distribution of vessel diameters. The quantification was based on new image analysis methods developed in the study. It was observed that the migration distance and the total vessel length increased by 82% and 199%, respectively, when the dose of bFGF was increased from 5 ng to 50 ng. The number and the diameter of vessels increased with the dose of bFGF as well. However, the last two parameters at a given dose of bFGF were approximately independent of the location in the middle region between the pellet and the limbus. These results provided useful information for understanding mechanisms of angiogenesis induced by bFGF and important data for validating a mathematical model of angiogenesis described in the second part of the study.

Previous data have demonstrated that doxorubicin (DOX) released from a lysolecithin-containing thermosensitive liposome (LTSL) can shut down blood flow in a human tumor xenograft (FaDu) in mice when the treatment is combined with hyperthermia (HT), suggesting that LTSL-DOX is a potential antivascular agent. To further understand mechanisms of the treatment, we investigated effects of LTSL-DOX (5 mg/kg body weight) plus HT (42 degrees C, 1 h) on microcirculation in another tumor (a murine mammary carcinoma, 4T07) implanted in mouse dorsal skin-fold chambers and dose responses of tumor (FaDu and 4T07) and endothelial cells to LTSL-DOX or free DOX with or without HT. We observed that LTSL-DOXHT could significantly reduce blood flow and microvascular density in 4T07 tumors. The antivascular efficacy of LTSLDOX- HT could be enhanced through increasing tumor microvascular permeability of liposomes by using platelet activating factor (PAF). We also observed that the dose responses of FaDu and 4T07 to DOX in vitro were similar to each other and could be enhanced by HT. Taken together, these data suggested that tumor microvascular permeability was more critical than the sensitivity of tumor cells to DOX in determining the antivascular efficacy of LTSL-DOX-HT treatment.

Engineering principles and new applications for the nascent field of synthetic biology are just beginning to be explored. Here, we report the engineering of four novel synthetic biological systems: (1) a Bacterial Dynamo, for generating electricity using modified magnetotactic bacteria on a microfabricated device; (2) Cancer StickyBots, for targeting and destroying tumour cells using engineered Escherichia coli cells; (3) Human Encryption, an information encoding, storage, and retrieval scheme for potential security and medical diagnostic applications; and (4) X-Verter, new strategies and tools for biological circuit design and BioBrick management. While each of these systems had distinct aims, they shared a common philosophy of rationally building useful and beneficial synthetic biological systems using fundamental engineering principles. They also demonstrated the potential usefulness of BioBricks and contributed to the Registry of Standard Biological Parts and synthetic biology community-at-large.

Endothelial cell (EC) aging and senescence are key events in atherogenesis and cardiovascular disease development. Age-associated changes in the local mechanical environment of blood vessels have also been linked to atherosclerosis. However, the extent to which cell senescence affects mechanical forces generated by the cell is unclear. In this study, we sought to determine whether EC senescence increases traction forces through age-associated changes in the glycocalyx and antioxidant regulator deacetylase Sirtuin1 (SIRT1), which is downregulated during aging. Traction forces were higher in cells that had undergone more population doublings and changes in traction force were associated with altered actin localization. Older cells also had increased actin filament thickness. Depletion of heparan sulfate in young ECs elevated traction forces and actin filament thickness, while addition of heparan sulfate to the surface of aged ECs by treatment with angiopoietin-1 had the opposite effect. While inhibition of SIRT1 had no significant effect on traction forces or actin organization for young cells, activation of SIRT1 did reduce traction forces and increase peripheral actin in aged ECs. These results show that EC senescence increases traction forces and alters actin localization through changes to SIRT1 and the glycocalyx.

GW5638 is a novel tissue-selective estrogen receptor (ER) modulator. Structurally, it is a derivative of tamoxifen that is known for its inhibitory effects on angiogenesis in an ER-independent manner. Therefore, it is possible that GW5638 has the same effects as tamoxifen on angiogenesis. To test this hypothesis, we used the rat cornea pocket assay and developed a new method that could precisely determine the total projected area of microvessels induced by basic fibroblast growth factor (bFGF) in the cornea. Animals in the study were treated with corn oil (control group), tamoxifen, or GW5638. After treatment, we observed that both GW5638 and tamoxifen could inhibit angiogenesis in the cornea (P

Recent studies have shown that up to 90% of viral vectors could disseminate to normal organs following intratumoral infusion. The amount of dissemination might be dependent on the infusion conditions. Therefore, we investigated the effects of infusion rate, volume, and dose on transgene expression in liver and tumor tissues after intratumoral infusion of an adenoviral vector encoding luciferase. Luciferase expression was determined through bioluminescence intensity measurement. We observed that the luciferase expression in the liver was independent of the infusion rate but increased with the infusion dose, whereas the luciferase expression in the tumor was a bell-shaped function of the infusion rate. The latter observation was consistent with the distribution pattern of Evans blue–labeled albumin after its solution was infused into tumors at the same infusion rates. We also observed that the infusion volume could affect luciferase expression in the tumor but not in the liver. These observations implied that virus dissemination was determined mainly by the infusion dose, whereas the amount of transgene expression in the tumor depended on the distribution volume of viral vectors in the tumor as well as the infusion dose. [Mol Cancer Ther 2006;5(2):362–6]

Systemic virus dissemination is a potential problem during local gene delivery in solid tumours. However, the kinetics and pathways of the dissemination have not been well characterised during the first 24 h after the infusion is started. To this end, we infused adenoviral vectors for luciferase or enhanced green fluorescence protein into three different tumour models in mice. During and/or after the infusion, we determined the amount of adenoviruses in the tumour, blood, and liver, and examined the transgene expression in the liver, lung, blood, and tumour. In addition, we intravenously injected tumour cells expressing luciferase and examined the biodistribution of these cells in the body. We observed transgene expression in the liver and tumour at 24 h after the infusion, but could not detect transgene expression in the blood and lung. The peak concentration of viral vectors in the plasma occurred during the intratumoral infusion. At 10 min after the infusion, few viral vectors remained in the blood and the ratio of copy numbers of adenoviruses between liver and tumour was2 in 80% andor = 10 in 40% of the mice. Most tumour cells injected intravenously accumulated in the lung within the first 24 h. Taken together, these data indicated that systemic virus dissemination occurred mainly during the first 10 min after the intratumoral infusion was started, and that the dissemination was due to infusion-induced convective transport of viral vectors into leaky tumour microvessels.

Interstitial transport of DNA is a rate-limiting step in electric field-mediated gene delivery in vivo. Interstitial transport of macromolecules, such as plasmid DNA, over a distance of several cell layers, is inefficient due to small diffusion coefficient and inadequate convection. Therefore, we explored electric field as a novel driving force for interstitial transport of plasmid DNA. In this study, agarose gels were used to mimic the interstitium in tissues as they had been well characterized and could be prepared reproducibly. We measured the electrophoretic movements of fluorescently labeled plasmid DNA in agarose gels with three different concentrations (1.0%, 2.0% and 3.0%) subjected to electric pulses at three different field strengths (100, 200 and 400 V/cm) and four different pulse durations (10, 50, 75, 99 ms). We observed that: (1) shorter pulses (10 ms) were not as efficient as longer pulses in facilitating plasmid transport through agarose gels; (2) plasmid electromobility reached a plateau at longer pulse durations; and (3) plasmid electromobility increased with applied electric energy, up to a threshold, in all three gels. These data suggested that both pulse strength and duration needed to be adequately high for efficient plasmid transport through extracellular matrix. We also found that electric field was better than concentration gradient of DNA as a driving force for interstitial transport of plasmid DNA.

The purpose of this report is to present an overview of the use of fluorescence imagingin vivo, with particular emphasis on oncology. It is important to note, however, that many of the methods described herein have been applied to the study of non-malignant tissues as well. Modern medicine and biology research has benefited greatly from an ever-expanding assortment of fluorescent markers and labels. These markers and labels have allowed investigators to observe the behavior and properties of cell and molecular entities of interest in the context of complicated biological systems such as a mammalian cell or a whole mouse. Methods developed to image fluorescence in whole mice have been valuable in studying patterns of tumor growth and metastases. Alternatively, more detailed information and a wide variety of endpoints can be obtained using “intravital” preparations. This review focuses on use of fluorescence imaging for intravital preparations. For detail on fluorescence imaging of whole animals, refer to reviews on this subject [1,2]. For oncologic applications, studies have focused primarily on window chamber preparations that allow for real-time visualization of tumor growth, vascularity, vascular responses to stimulation, vascular permeability, vascular orientation, flow instability, and the like. These endpoints have been used to show that there are functional differences between tumor and normal tissues with respect to these functions under baseline conditions and after therapeutic manipulation. Examples of some of these differences are provided in this review as a means to illustrate how they can be used.

The erroneous and unpredictable behavior of percutaneous glucose sensors just days following implantation has limited their clinical utility for diabetes management. Recent research has implicated the presence of adherent inflammatory cells as the key mitigating factor limiting sensor functionality in this period of days post-implantation. Here we present a novel in vitro platform to mimic the cell-embedded provisional matrix that forms adjacent to the sensor immediately after implantation for the focused investigation of the effects of early stage tissue response on sensor function. This biomimetic surrogate is formed by imbibing fibrin-based gels with physiological densities of inflammatory RAW 264.7 macrophages. When surrounding functional sensors, macrophage-embedded fibrin gels contribute to sensor signal declines that are similar in both shape and magnitude to those observed in previous whole blood and small animal studies. Signal decline in the presence of gels is both metabolically-mediated and sensitive to cell type and activation. Computational modeling of the experimental setup is also presented to validate the design by showing that the cellular glucose uptake parameters necessary to achieve such experimental declines align well with literature values. Together, these data suggest this in vitro provisional matrix surrogate may serve as an effective screening tool for testing the biocompatibility of future glucose sensor designs.

Apoptosis is a tightly controlled process in mammalian cells. It is important for embryogenesis, tissue homoeostasis, and cancer treatment. Apoptosis not only induces cell death, but also leads to the release of signals that promote rapid proliferation of surrounding cells through the Phoenix Rising (PR) pathway. To quantitatively understand the kinetics of interactions of different molecules in this pathway, we developed a mathematical model to simulate the effects of various changes in the PR pathway on the secretion of prostaglandin E2 (PGE2), a key factor for promoting cell proliferation. These changes include activation of caspase 3 (C3), caspase 7 (C7), and nuclear factor κB (NFκB). In addition, we simulated the effects of cyclooxygenase-2 (COX2) inhibition and C3 knockout on the level of secreted PGE2. The model predictions on PGE2 in MEF and 4T1 cells at 48 hours after 10-Gray radiation were quantitatively consistent with the experimental data in the literature. Compared to C7, the model predicted that C3 activation was more critical for PGE2 production. The model also predicted that PGE2 production could be significantly reduced when COX2 expression was blocked via either NFκB inactivation or treatment of cells with exogenous COX2 inhibitors, which led to a decrease in the rate of conversion from arachidonic acid to prostaglandin H2 in the PR pathway. In conclusion, the mathematical model developed in this study yielded new insights into the process of tissue regrowth stimulated by signals from apoptotic cells. In future studies, the model can be used for experimental data analysis and assisting development of novel strategies/drugs for improving cancer treatment or normal tissue regeneration., Author Summary Apoptosis, or programmed cell death, is known to be important for embryogenesis, tissue homoeostasis, and cancer treatment. Furthermore, researchers have recently observed that apoptosis may promote wound healing and tissue regeneration, and accelerate undesired solid tumor regrowth after chemotherapy/radiation therapy. Mechanisms of apoptosis-induced tissue regrowth are related to a molecular network discovered recently in our lab. To quantitatively understand the kinetics of interactions of different molecules in this network, we developed a mathematical model and validated it by comparing the simulation results to experimental data reported in previous studies. To gain new insights into the process of tissue regrowth after inducing apoptosis, we used the model to simulate the effects of radiation on the production of a key growth stimulating factor, PGE2, in apoptotic cells. Additionally, we simulated how PGE2 production could be altered when cells were treated with different inhibitors. We expect that the new mathematical model can be used in future studies to facilitate design of better approaches to cancer treatment or normal tissue regeneration.

The aims of the present study were to determine if melatonin exerts an effect on prolactin (PRL) secretion via the tuberoinfundibular dopaminergic (TIDA) neurons and if endogenous or exogenous melatonin has an entraining effect on the rhythmic changes of TIDA neuronal activity and PRL secretion. Melatonin given in the morning (10:00 h), dose- (0.01-1 mg/kg, ip) and time- (at 15 and 60 min, but not at 30 min) dependently stimulated TIDA neuronal activity in ovariectomized (OVX), estrogen-treated rats as determined by 3,4-dihydroxyphenylacetic acid (DOPAC) levels in the median eminence (ME). Serum PRL was concurrently inhibited by the injection. Melatonin administered in the afternoon (15:00 h) was even more effective in stimulating the lowered TIDA neuronal activity and inhibiting the increased PRL level than that given in the morning (10:00 h). S-20098, a melatonin agonist was also effective in stimulating the TIDA neurons. In contrast, S-20928, a putative melatonin antagonist, while it had no effect by itself, blocked the effect of S-20098. Although S-20928 failed to prevent melatonin's effect on ME DOPAC levels, six interspaced injections of S-20928, from 18:00 to 01:30 h, significantly blocked the increase of ME DOPAC levels at 03:00 h, indicating that the endogenous melatonin may play a role. We further used rats that received daily injection of melatonin (1 mg/kg, ip) at 18:00 h for 10 days and found that the injection augmented basal TIDA neuronal activity at 11:00 h and blunted the afternoon PRL surge. In all, melatonin can have an inhibitory effect on PRL secretion by stimulating the TIDA neurons, and it may help to entrain the circadian rhythms of both TIDA neuronal activity and PRL secretion.

Molecular charge is one of the main determinants of transvascular transport. There are, however, no data available on the effect of molecular charge on microvascular permeability of macromolecules in solid tumours. To this end, we measured tumour microvascular permeability to different proteins having similar size but different charge. Measurements were performed in the human colon adenocarcinoma LS174T transplanted in transparent dorsal skinfold chambers in severe combined immunodeficient (SCID) mice. Bovine serum albumin (BSA) and IgG were fluorescently labelled and were either cationized by conjugation with hexamethylenediamine or anionized by succinylation. The molecules were injected i.v. and the fluorescence in tumour tissue was quantified by intravital fluorescence microscopy. The fluorescence intensity and pharmacokinetic data were used to calculate the microvascular permeability. We found that tumour vascular permeability of cationized BSA (pI-range: 8.6-9.1) and IgG (pI: 8.6-9.3) was more than two-fold higher (4.25 and 4.65x10(-7) cm s(-1)) than that of the anionized BSA (pI approximately 2.0) and IgG (pI: 3.0-3.9; 1.11 and 1.93x10(-7) cm s(-1), respectively). Our results indicate that positively charged molecules extravasate faster in solid tumours compared to the similar-sized compounds with neutral or negative charges. However, the plasma clearance of cationic molecules was approximately 2x faster than that of anionic ones, indicating that the modification of proteins enhances drug delivery to normal organs as well. Therefore, caution should be exercised when such a strategy is used to improve drug and gene delivery to solid tumours.

The sequence of events that leads to tumor vessel regression and the functional characteristics of these vessels during hormone–ablation therapy are not known. This is because of the lack of an appropriate animal model and monitoring technology. By using in vivo microscopy and in situ molecular analysis of the androgen-dependent Shionogi carcinoma grown in severe combined immunodeficient mice, we show that castration of these mice leads to tumor regression and a concomitant decrease in vascular endothelial growth factor (VEGF) expression. Androgen withdrawal is known to induce apoptosis in Shionogi tumor cells. Surprisingly, tumor endothelial cells begin to undergo apoptosis before neoplastic cells, and rarefaction of tumor vessels precedes the decrease in tumor size. The regressing vessels begin to exhibit normal phenotype, i.e., lower diameter, tortuosity, vascular permeability, and leukocyte adhesion. Two weeks after castration, a second wave of angiogenesis and tumor growth begins with a concomitant increase in VEGF expression. Because human tumors often relapse following hormone–ablation therapy, our data suggest that these patients may benefit from combined anti-VEGF therapy.

Novel anti-neoplastic agents such as gene targeting vectors and encapsulated carriers are quite large (approximately 100–300 nm in diameter). An understanding of the functional size and physiological regulation of transvascular pathways is necessary to optimize delivery of these agents. Here we analyze the functional limits of transvascular transport and its modulation by the microenvironment. One human and five murine tumors including mammary and colorectal carcinomas, hepatoma, glioma, and sarcoma were implanted in the dorsal skin-fold chamber or cranial window, and the pore cutoff size, a functional measure of transvascular gap size, was determined. The microenvironment was modulated: ( i ) spatially, by growing tumors in subcutaneous or cranial locations and ( ii ) temporally, by inducing vascular regression in hormone-dependent tumors. Tumors grown subcutaneously exhibited a characteristic pore cutoff size ranging from 200 nm to 1.2 μm. This pore cutoff size was reduced in tumors grown in the cranium or in regressing tumors after hormone withdrawal. Vessels induced in basic fibroblast growth factor-containing gels had a pore cutoff size of 200 nm. Albumin permeability was independent of pore cutoff size. These results have three major implications for the delivery of therapeutic agents: ( i ) delivery may be less efficient in cranial tumors than in subcutaneous tumors, ( ii ) delivery may be reduced during tumor regression induced by hormonal ablation, and ( iii ) permeability to a molecule is independent of pore cutoff size as long as the diameter of the molecule is much less than the pore diameter.

Binding is crucial to the function of most biologically active molecules, but difficult to quantify directly in living tissue. To this end, fluorescence recovery after photobleaching was used to detect the immobilization of fluorescently labeled ligand caused by binding to receptors in vivo . Measurements of mAb affinity to target antigen within human tumor xenografts revealed a saturable binding isotherm, from which an in vivo carcinoembryonic antigen density of 0.56 nmol/g (5.0 × 10 5 /cell) and an association constant of K a ⩽ 4 × 10 7 M −1 were estimated. The present method can be adapted for in vivo studies of cell signaling, targeted drugs, gene therapy, and other processes involving receptor-ligand binding.

The hyperpermeability of tumor vessels to macromolecules, compared with normal vessels, is presumably due to vascular endothelial growth factor/vascular permeability factor (VEGF/VPF) released by neoplastic and/or host cells. In addition, VEGF/VPF is a potent angiogenic factor. Removal of this growth factor may reduce the permeability and inhibit tumor angiogenesis. To test these hypotheses, we transplanted a human glioblastoma (U87), a human colon adenocarcinoma (LS174T), and a human melanoma (P-MEL) into two locations in immunodeficient mice: the cranial window and the dorsal skinfold chamber. The mice bearing vascularized tumors were treated with a bolus (0.2 ml) of either a neutralizing antibody (A4.6.1) (492 μg/ml) against VEGF/VPF or PBS (control). We found that tumor vascular permeability to albumin in antibody-treated groups was lower than in the matched controls and that the effect of the antibody was time-dependent and influenced by the mode of injection. Tumor vascular permeability did not respond to i.p. injection of the antibody until 4 days posttreatment. However, the permeability was reduced within 6 h after i.v. injection of the same amount of antibody. In addition to the reduction in vascular permeability, the tumor vessels became smaller in diameter and less tortuous after antibody injections and eventually disappeared from the surface after four consecutive treatments in U87 tumors. These results demonstrate that tumor vascular permeability can be reduced by neutralization of endogenous VEGF/VPF and suggest that angiogenesis and the maintenance of integrity of tumor vessels require the presence of VEGF/VPF in the tissue microenvironment. The latter finding reveals a new mechanism of tumor vessel regression—i.e., blocking the interactions between VEGF/VPF and endothelial cells or inhibiting VEGF/VPF synthesis in solid tumors causes dramatic reduction in vessel diameter, which may block the passage of blood elements and thus lead to vascular regression.

We have characterized the effect of bone graft treatment by heating or freezing (with or without dimethyl sulfoxide (DMSO)). Tissue culture and dorsal skin-fold chambers in mice were used as sites to quantify the effect on angiogenesis, growth and calcification of neonatal femora. Fresh femora increased in both length and cartilage diameter (calcification in vivo only), but cryopreservation or heating abolished the increase in femoral dimensions. In vivo, femora of all experimental groups elicited an angiogenic response from the host tissue, which was most pronounced for fresh femora, weaker for DMSO-preserved frozen bone and poor for unprotected frozen bone and boiled femora. Freezing in the presence of a cryopreservative (DMSO) was found to preserve the angiogenic potential of frozen bone, whereas unprotected heating or freezing significantly impaired angiogenesis induction and growth potential.

Nonsteroidal anti-inflammatory agents have been used experimentally and clinically to suppress a variety of physiological events, including angiogenesis and formation of bone. The exact mechanisms by which indomethacin alters skeletal tissue generation are unknown, due in part to methodological limitations. By the use of an organ culture assay and an animal model using intravital microscopy in mice bearing dorsal skinfold chambers, the effect of indomethacin on growth and angiogenesis of neonatal femora was characterized over 16 days. In both assays, femora significantly elongated with time (P

P-selectin-mediated rolling is believed to be important in the recruitment of leukocytes to tissue after ischemia-reperfusion injury. The dorsal skin chamber was used to examine differences in the rolling and stable adhesion of circulating leukocytes in subcutaneous (SC) vessels of P-selectin-deficient and age-matched wild-type mice, both under basal conditions and after ischemia-reperfusion. Rolling in the postcapillary venules in SC tissue of P-selectin-deficient mice was significantly lower than that in wild-type mice under the basal conditions and post-ischemia-reperfusion (P < .05), but was not eliminated by the deletion of the P-selectin gene. No significant difference between P-selectin-deficient and wild-type mice in shear rate or leukocyte-endothelial adhesion was observed up to 24 hours after ischemia-reperfusion. These results show that P-selectin-mediated rolling is not a prerequisite for ischemia-reperfusion-induced leukocyte-endothelial adhesion in the skin.

PURPOSE The study was designed to determine trabecular meshwork (TM) stiffness and its relationship to outflow facility (C) in perfused normal human and porcine eyes. METHODS Human and porcine eyes were perfused at pressures of 10, 20, 30, and 40 mm Hg to determine C and how outflow resistance (R = 1/C) varied with the pressure. Following perfusions, TM tissue segments were dissected and stretched uniaxially to determine the circumferential bulk Young's modulus (E). The statistical significance of difference between different groups was evaluated using a two-tailed Student's t-test or Mann-Whitney U test. RESULTS A larger E correlated with a higher C measured at 10 and 20 mm Hg (P < 0.05), and a similar trend was observed at 30 and 40 mm Hg in human eyes (n = 7). Additionally, a higher C correlated to a lower variance of R, and a stiffer TM correlated to a lower variance of R in human eyes (P < 0.05). For porcine TM, E was inversely correlated to a cross-sectional area (P < 0.003, n = 11), and its value (24.9 and 1.5 kPa; geometric mean and geometric SE) was lower than E of human TM (515 ± 136 kPa; mean ± SE) (P < 0.01). C and variance of R were not significantly different between the species. CONCLUSIONS A higher circumferential stiffness of the TM correlated with a higher outflow facility and less IOP elevation-induced variation in outflow resistance in normal human eyes, but not in porcine eyes. For future studies, these correlations need to be evaluated in glaucomatous eyes to better understand normal and abnormal TM functions.

Simultaneous measurements of intravascular and interstitial oxygen partial pressure (PO2) in any tissue have not previously been reported, despite the importance of oxygen in health and in disease. This is due to the limitations of current techniques, both invasive and noninvasive. We have optically measured microscopic profiles of PO2 with high spatial resolution in subcutaneous tissue and transplanted tumors in mice by combining an oxygen-dependent phosphorescence quenching method and a transparent tissue preparation. The strengths of our approach include the ability to follow PO2 in the same location for several weeks and to relate these measurements to local blood flow and vascular architecture. Our results show that (i) PO2 values in blood vessels in well-vascularized regions of a human colon adenocarcinoma xenograft are comparable to those in surrounding arterioles and venules, (ii) carbogen (95% O2/5% CO2) breathing increases microvascular PO2 in tumors, and (iii) in unanesthetized and anesthetized mice PO2 drops to hypoxic values at < 200 microns from isolated vessels but drops by < 5 mmHg (1 mmHg = 133 Pa) in highly vascularized tumor regions. Our method should permit noninvasive evaluations of oxygen-modifying agents and offer further mechanistic information about tumor pathophysiology in tissue preparations where the surface of the tissue can be observed.

Human infections of H5N1 highly pathogenic avian influenza virus have continued to occur in China without corresponding outbreaks in poultry, and there is little conclusive evidence of the source of these infections. Seeking to identify the source of the human infections, we sequenced 31 H5N1 viruses isolated from humans in China (2005 to 2010). We found a number of viral genotypes, not all of which have similar known avian virus counterparts. Guided by patient questionnaire data, we also obtained environmental samples from live poultry markets and dwellings frequented by six individuals prior to disease onset (2008 and 2009). H5N1 viruses were isolated from 4 of the 6 live poultry markets sampled. In each case, the genetic sequences of the environmental and corresponding human isolates were highly similar, demonstrating a link between human infection and live poultry markets. Therefore, infection control measures in live poultry markets are likely to reduce human H5N1 infection in China.

Direct scarlet 4BE (DS-4BE) is a synthetic chemical dye and widely used in textile and dyeing industry. In this study, Penicillium sp. HS-DY08 was used as the original strain in the decoloraziation of DS-4BE and denitrification of nitrate and nitrite. Results indicated that HS-DY08 strain could decolorize DS-4BE dye in MSM medium and PDA media. Denitrification was obviously occurred in MSM medium containing nitrite or nitrate. Further study showed that HS-DY08 could synchronize decolorization and denitrification in MSM medium containing 30 mg/L DS-4BE dye and 100 mg/L nitrate. The decolorization and denitrification were completely finished within 6d, suggesting that HS-DY08 strain has practical significance and broad application prospect on the treatment of dye wastewater.

RNA interference (RNAi) is a fundamental mechanism of gene regulation and has been harnessed to produce a new class of drugs for treatment of various diseases. A key issue in these applications is how to effectively deliver RNAi therapeutics into target cells. This review is focused on advances in RNA delivery in vivo. To achieve it, novel strategies have been developed to enhance stability of RNA in cells and tissues, overcome barriers to transport of RNA or its carriers in the body, and reduce immunogenicity and cytotoxicity of treatment. Approaches to RNA delivery are divided into three categories in this review: biological, chemical, and physical. Advantages and disadvantages of each method are discussed. At present, effective delivery of RNAi therapeutics in vivo is still a challenge although significant advances have been made in this field.

The use of animal models in drug discovery studies presents issues with feasibility and ethical concerns. To address these limitations, in vitro tissue models have been developed to provide a means for systematic, repetitive, and quantitative investigation of drugs. By eliminating or reducing the need for animal subjects, these models can serve as platforms for more tightly controlled, high-throughput screening of drugs and for pharmacokinetic and pharmacodynamic analyses of drugs. The focus of this review is three-dimensional (3D) tissue models that can capture cell-cell and cell-matrix interactions. Compared to the 2D culture of cell monolayers, 3D models more closely mimic native tissues since the cellular microenvironment established in the 3D models often plays a significant role in disease progression and cellular responses to drugs. A growing body of research has been published in the literature, which highlights the benefits of the 3D in vitro models of various tissues. This review provides an overview of some successful 3D in vitro models that have been developed to mimic liver, breast, cardiac, muscle, bone, and corneal tissues as well as malignant tissues in solid tumors.

To further investigate the effect of a combination of genistein with survivin of RNA interference on the proliferation and apoptosis of DU-145 cells, the effect of genistein on the proliferation of DU-145 cells was detected by the MTT method and cytometry, and the apoptosis of cells was observed with fluorescence microscopy. In order to test combined genistein with transfection of small interfering RNA (siRNA) against survivin, a survivin siRNA plasmid was constructed and transfected into DU-145 cells. Genistein inhibited proliferation and induced apoptosis of cancerous DU-145 and Hela cells, whereas genistein had minimal effects for normal L-O2 cells. The stable transfected cell lines of DU-145, knockdown survivin by siRNA, displayed stronger apoptotic than untransfected DU-145, the transfected cell of DU-145 treated with genistein demonstrated the inhibition of proliferation and induction of apoptosis significantly; it showed genistein synergistic effect with RNAi in survivin for inhibition of prostate cancer cells.

Gene therapy has a great potential in cancer treatment. However, the efficacy of cancer gene therapy is currently limited by the lack of a safe and efficient means to deliver therapeutic genes into the nucleus of tumor cells. One method under investigation for improving local gene delivery is based on the use of pulsed electric field. Despite repeated demonstration of its effectiveness in vivo, the underlying mechanisms behind electric field-mediated gene delivery remain largely unknown. Without a thorough understanding of these mechanisms, it will be difficult to further advance the gene delivery. In this review, the electric field-mediated gene delivery in solid tumors will be examined by following individual transport processes that must occur in vivo for a successful gene transfer. The topics of examination include: (i) major barriers for gene delivery in the body, (ii) distribution of electric fields at both cell and tissue levels during the application of external fields, and (iii) electric field-induced transport of genes across each of the barriers. Through this approach, the review summarizes what is known about the mechanisms behind electric field-mediated gene delivery and what require further investigations in future studies. © 2007 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 97:691–711, 2008

Pathogenic bacteria such as Listeria and Yersinia gain initial entry by binding to host target cells and stimulating their internalization. Bacterial uptake entails successive, increasingly strong associations between receptors on the surface of bacteria and hosts. Even with genetically identical cells grown in the same environment, there are vast differences in the number of bacteria entering any given cell. To gain insight into this variability, we examined uptake dynamics of Escherichia coli engineered to express the invasin surface receptor from Yersinia, which enables uptake via mammalian host β1-integrins. Surprisingly, we found that the uptake probability of a single bacterium follows a simple power-law dependence on the concentration of integrins. Furthermore, the value of a power-law parameter depends on the particular host-bacterium pair but not on bacterial concentration. This power-law captures the complex, variable processes underlying bacterial invasion while also enabling differentiation of cell lines., Author Summary Uptake of bacteria by mammalian cells is highly variable within a population of host cells and between host cell types. A detailed but unwieldy mechanistic model describing individual host-pathogen receptor binding events is captured by a simple power-law dependence on the concentration of the host receptors. The power-law parameters capture characteristics of the host-bacterium pair interaction and can differentiate host cell lines. This study has important implications for understanding the accuracy and precision of therapeutics employing receptor-mediated transport of materials to mammalian hosts.