A computational framework for modeling cell–matrix interactions in soft biological tissues

Biomechanics and Modeling in Mechanobiology - Living soft tissues appear to promote the development and maintenance of a preferred mechanical state within a defined tolerance around a so-called set...     

Efficiency of direct and indirect shoot organogenesis in different genotypes of Rosa hybrida

To optimize indirect regeneration (IR) and direct regeneration (DR) in Rosa hybrida cv. Apollo different explant types and different concentrations of plant growth regulators were investigated. Among the different auxins studied and over all explant types, 10???M 2,4-dichlorophenoxyacetic acid (2,4-D) promoted the highest frequency of callus production for IR. The highest frequency of regeneration (60.8?%) was obtained when calli were transferred to Murashige and Skoog medium supplemented with 2.5???M thidiazuron (TDZ) and 2???M gibberellic acid. The highest frequency of regeneration (80.2?%) for DR was obtained from leaves cultured on the medium containing 10???M TDZ. The efficiency of IR and DR were compared in four different rose cultivars including ??Apollo??, ??Black Baccara??, ??Maroussia?? and ??Amanda??. The frequency of regeneration in all four cultivars was significantly higher in DR compared to IR. Also shoots regenerated by DR appeared earlier than the shoots regenerated by IR. The results of flow cytometry showed that the shoots derived from IR to DR were tetraploid like the original cultivars.     

Oryzalin-induced chromosome doubling in<Emphasis Type="Italic"> Rosa</Emphasis> and its effect on plant morphology and pollen viability

Shoot tips of the diploid rose Thérèse Bugnet were treated in vitro to oryzalin at concentrations of 5 and 15 M. Tetraploid shoots were obtained in highest frequencies (40%) after exposure to 5 M oryzalin for 14 days. Thin (1 mm) nodal sections were treated with 5 M oryzalin and the highest frequency of tetraploids (66%) was obtained after exposure for only 1 day. The shorter exposure times required to induce chromosome doubling in thin nodal sections is attributed to the more efficient delivery of oryzalin to the meristem. Tetraploids were obtained from four diploid roses and hexaploids from two triploid roses. Chromosome doubling was accompanied by increases in thickness and a darker green colouration of the leaves and, in all diploid to tetraploid and one triploid to hexaploid conversion, the breadth/length ratio of leaflets was significantly increased. Internodes were longer in tetraploids than diploids but significantly shorter in hexaploids than triploids. The number of petals per flower in the tetraploid form of Thérèse Bugnet was double that of the diploid. Significant increases in pollen viability accompanied chromosome doubling of all four diploids and one of the two triploids.Communicated by H. Nybom     

Analytical and Clinical Validation of a Digital Sequencing Panel for Quantitative,Highly Accurate Evaluation of Cell-Free Circulating Tumor DNA

Next-generation sequencing of cell-free circulating solid tumor DNA addresses two challenges in contemporary cancer care. First this method of massively parallel and deep sequencing enables assessment of a comprehensive panel of genomic targets from a single sample, and second, it obviates the need for repeat invasive tissue biopsies. Digital Sequencing^TM is a novel method for high-quality sequencing of circulating tumor DNA simultaneously across a comprehensive panel of over 50 cancer-related genes with a simple blood test. Here we report the analytic and clinical validation of the gene panel. Analytic sensitivity down to 0.1% mutant allele fraction is demonstrated via serial dilution studies of known samples. Near-perfect analytic specificity (> 99.9999%) enables complete coverage of many genes without the false positives typically seen with traditional sequencing assays at mutant allele frequencies or fractions below 5%. We compared digital sequencing of plasma-derived cell-free DNA to tissue-based sequencing on 165 consecutive matched samples from five outside centers in patients with stage III-IV solid tumor cancers. Clinical sensitivity of plasma-derived NGS was 85.0%, comparable to 80.7% sensitivity for tissue. The assay success rate on 1,000 consecutive samples in clinical practice was 99.8%. Digital sequencing of plasma-derived DNA is indicated in advanced cancer patients to prevent repeated invasive biopsies when the initial biopsy is inadequate, unobtainable for genomic testing, or uninformative, or when the patient’s cancer has progressed despite treatment. Its clinical utility is derived from reduction in the costs, complications and delays associated with invasive tissue biopsies for genomic testing.     

Ovarian response and pregnancy rate following different doses of eCG treatment in Chall ewes

The purpose of the study was to investigate the effects of different doses of equine chorionic gonadotropin (eCG) treatment on follicular development, ovulation and pregnancy rate during the breeding season in fat-tailed Chall ewes. Seventy-two cycling (62.5 ± 2.5 kg), multiparous Iranian Chall ewes were used in the trial. The ewes were randomly allocated to 6 groups (n = 12/group). Estrus was synchronized with the aid of controlled intravaginal drug release (CIDR) devices, inserted for 14 days. At the time of CIDR removal (day 14), the ewes received i.m. either 0 (control group, G0), 450 (G450), 550 (G550), 650 (G650), 750 (G750) or 850 (G850) IU eCG. Vasectomized rams were used to detect estrus in the ewes from 24 h after CIDR removal. Ovarian follicular activity was monitored with the aid of transrectal ultrasonography on the day of CIDR insertion (day 0) and daily from the day of eCG treatment (day 14), until estrus (day 16). During these days, blood samples were collected for the determination of plasma progesterone and estradiol concentrations. Laparoscopic intrauterine inseminations were conducted 54–60 h after CIDR removal. The number of CL's and pregnancy diagnosis was recorded using ultrasonography 7 and 54 days following AI, respectively. Half of ewes in control group and most of the ewes treated with eCG showed signs of estrus within 36 h of CIDR removal. The ewes in groups G750 and G850 recorded the highest number of large follicles at estrus and CL's 7 days later. The pregnancy rate in groups G550 (75.0%) and G650 (75.0%) was higher (P < 0.05) than that in the other groups. The ovarian response and estradiol concentration, as well as pregnancy rate showed that 550 or 650 IU eCG treatment is the most effective doses in improving the pregnancy rate in Iranian Chall ewes.     

Interaction of insulin with a triblock copolymer of PEG-(fumaric-sebacic acids)-PEG: thermodynamic and spectroscopic studies

A comparative study on the interaction of (PEG-co-P(FA/SC)-co-PEG) triblock copolymer with bovine and human insulins was carried out using isothermal titration calorimetry (ITC), circular dichroism (CD), and fluorescence spectroscopy. ITC data show that the copolymer has a low affinity for both proteins, with an association constant of about 7-9 x 10(3) M (-1). Results also show that binding is enthalpically driven, and disfavored by conformational entropy. CD spectroscopy studies reveal a small increase in the helical content and a decrease in beta-structure as well as random coil in both proteins. Acrylamide quenching experiments display reduced accessibility of tyrosines, while intrinsic fluorescence spectra show lower tyrosine emission. Furthermore, thermal unfolding experiments, studied by far-UV CD at 222 and 217 nm, demonstrate that upon interaction with the copolymer helix structure becomes less stable while the stability of beta-structure remains unchanged. Altogether, these observations indicate that (PEG-co-P(FA/SC)-co-PEG) triblock copolymer has similar effect(s) on both proteins.     

Impact of Growth Hormone (GH) Deficiency and GH Replacement upon Thymus Function in Adult Patients

Background

Despite age-related adipose involution, T cell generation in the thymus (thymopoiesis) is maintained beyond puberty in adults. In rodents, growth hormone (GH), insulin-like growth factor-1 (IGF-1), and GH secretagogues reverse age-related changes in thymus cytoarchitecture and increase thymopoiesis. GH administration also enhances thymic mass and function in HIV-infected patients. Until now, thymic function has not been investigated in adult GH deficiency (AGHD). The objective of this clinical study was to evaluate thymic function in AGHD, as well as the repercussion upon thymopoiesis of GH treatment for restoration of GH/IGF-1 physiological levels.

Methodology/Principal Findings

Twenty-two patients with documented AGHD were enrolled in this study. The following parameters were measured: plasma IGF-1 concentrations, signal-joint T-cell receptor excision circle (sjTREC) frequency, and sj/β TREC ratio. Analyses were performed at three time points: firstly on GH treatment at maintenance dose, secondly one month after GH withdrawal, and thirdly one month after GH resumption. After 1-month interruption of GH treatment, both plasma IGF-1 concentrations and sjTREC frequency were decreased (p<0.001). Decreases in IGF-1 and sjTREC levels were correlated (r = 0.61, p<0.01). There was also a decrease in intrathymic T cell proliferation as indicated by the reduced sj/β TREC ratio (p<0.01). One month after reintroduction of GH treatment, IGF-1 concentration and sjTREC frequency regained a level equivalent to the one before GH withdrawal. The sj/β TREC ratio also increased with GH resumption, but did not return to the level measured before GH withdrawal.

Conclusions

In patients with AGHD under GH treatment, GH withdrawal decreases thymic T cell output, as well as intrathymic T cell proliferation. These parameters of thymus function are completely or partially restored one month after GH resumption. These data indicate that the functional integrity of the somatotrope GH/IGF-1 axis is important for the maintenance of a normal thymus function in human adults.

Trial Registration

ClinicalTrials.gov NTC00601419     

Agroinfiltration: a rapid and reliable method to select suitable rose cultivars for blue flower production

Rose cultivars with blue flower color are among the most attractive breeding targets in floriculture. However, they are difficult to produce due to the low efficiency of transformation systems, interactive effects of hosts and vectors, and lengthy processes. In this study, agroinfiltration-mediated transient expression was investigated as a tool to assess the function of flower color genes and to determine appropriate host cultivars for stable transformation in Rosa hybrida. To induce delphinidin accumulation and consequently to produce blue hue, the petals of 30 rose cultivars were infiltrated with three different expression vectors namely pBIH-35S-CcF3′5′H, pBIH-35S-Del2 and pBIH-35S-Del8, harbouring different sets of flower color genes. The results obtained showed that the ectopic expression of the genes was only detected in three cultivars with dark pink petals (i.e. ‘Purple power’, ‘High & Mora’ and ‘Marina’) after 6–8 days. The high performance liquid chromatography analyses confirmed delphinidin accumulation in the infiltrated petals caused by transient expression of CcF3′5′H gene. Moreover, there were significant differences in the amounts of delphinidin among the three cultivars infiltrated with the three different expression vectors. More specifically, the highest delphinidin content was detected in the cultivar ‘Purple power’ (4.67 µg g^?1 FW), infiltrated with the pBIH-35S-Del2 vector. The expression of CcF3′5′H gene in the infiltrated petals was also confirmed by real time PCR. In conclusion and based on the findings of the present study, the agroinfiltration could be regarded as a reliable method to identify suitable rose cultivars in blue rose flower production programs.     

Intraventricular insulin potentiates the anorexic effect of corticotropin releasing hormone in rats

Intraventricular corticotropin releasing hormone (CRH) suppresses food intake and body weight as a stress response. Insulin, acting within the brain, also suppresses food intake and body weight, and this suppression is related to caloric homeostasis. We determined if increased insulin within the brain potentiates the anorexic effects of intraventricular CRH. Rats were food deprived for 17 h each day and then given 30-min access to Ensure. One-half received continuous third ventricular infusion of synthetic cerebrospinal fluid via osmotic minipumps, and one-half received insulin (0.6 mU/day). During the infusion, rats also received 0, 0.1, 1.0, or 5.0 microg of CRH into the lateral ventricle just before access to Ensure. Insulin alone had no effect on Ensure intake or body weight. CRH dose dependently reduced Ensure intake in both groups, and the reduction was greater in the insulin group. Hence, central insulin potentiated the ability of centrally administered CRH to suppress food intake. These findings suggest that stress-related influences over food intake, particularly those mediated via CRH, interact with relative adiposity as signaled to the brain by central insulin.