Cinnamaldehyde regulates H2O 2-induced skeletal muscle atrophy by ameliorating the proteolytic and antioxidant defense systems

Skeletal muscle atrophy/wasting is associated with impaired protein metabolism in diverse physiological and pathophysiological conditions. Elevated levels of reactive oxygen species (ROS), disturbed redox status, and weakened antioxidant defense system are the major contributing factors toward atrophy. Regulation of protein metabolism by controlling ROS levels and its associated catabolic pathways may help in treating atrophy and related clinical conditions. Although cinnamaldehyde (CNA) enjoys the established status of antioxidant and its role in ROS management is reported, impact of CNA on skeletal muscle atrophy and related pathways is still unexplored. In the current study, the impact of CNA on C2C12 myotubes and the possible protection of cultured cells from H ₂O ₂-induced atrophy is examined. Myotubes were treated with H ₂O ₂ in the presence and absence of CNA and the changes in the antioxidative, proteolytic systems, and mitochondrial functions were scored. Morphological analysis showed significant protective effects of CNA on length, diameter, and nuclei fusion index of myotubes. The evaluation of biochemical markers of atrophy; creatine kinase, lactate dehydrogenase, succinate dehydrogenase along with the study of muscle-specific structural protein (i.e., myosin heavy chain-fast [MHCf] type) showed significant protection of proteins by CNA. CNA pretreatment not only checked the activation of proteolytic systems (ubiquitin-proteasome E3-ligases [MuRF1/Atrogin1]), autophagy [Beclin1/LC3B], cathepsin L, calpain, caspase), but also prevented any alteration in the activities of antioxidative defense enzymes (catalase, glutathione- S-transferase, glutathione-peroxidase, superoxide dismutase, glutathione reductase). The results suggest that CNA protects myotubes from H ₂O ₂-induced atrophy by inhibiting/resisting the amendments in proteolytic systems and maintains cellular redox-balance.     

An aptasensor for rapid and sensitive detection of estrogen receptor alpha in human breast cancer

The analysis of estrogen receptor (ER) expression in breast carcinomas plays a crucial role in determining the endocrine responsiveness of tumors for systemic adjuvant therapy. Conventionally, the ER levels in breast carcinomas had been detected using the dextran-coated charcoal assay and radioimmunoassay, which are now substituted with safer and economic antibody-based assays such as immunohistochemistry (IHC) and enzyme-linked immunosorbent assay (ELISA). Despite a gold (Au) standard method, the IHC has been criticized for factors such as tissue fixation, antibody selection, and threshold staining for result interpretation that could falsify test accuracy and reproducibility. The quest for alternative methods of ER quantification in tissue samples paved the way for aptamer-based diagnostics. Previously, we have isolated a DNA aptamer against human ER alpha (ERα) using an in vitro evolution system. In this study, we developed an electrochemical sensor using the 76-nucleotide DNA ERα- aptamer for rapid, precise, and cost-effective detection of ERα expression in human breast cancer patients. The aptasensor was constructed by covalently immobilizing the thiolated ERα- aptamer onto a screen-printed Au electrode. Construction of aptasensors was confirmed through atomic force microscopy and differential pulse voltammetry measurements. A detection limit of 0.001 ng/ml was calculated for full-length ERα (66.2 kDa) in a detection time of 10 min. Analysis of the cancerous breast tissue samples using the ELISA and aptasensor methods enabled distinctive classification of samples into the categories of ER −ve, weak ER +ve, and strong ER +ve samples. The current change of this aptasensor lies within 5% after a storage of 60 days at 4°C. Further studies on a reasonably large sample size are required to realize the clinical potential of the sensor.     

Creation and validation of a reactor engineering model for multiphase red wine fermentations

Red wine fermentations are performed in the presence of grape skins and seeds to ensure the extraction of color and other phenolics. The presence of these solids results in two distinct phases in the fermentor, as the solids float to the top to form a “cap.” Modeling of red wine fermentation is, therefore, complex and must consider spatial heterogeneity to predict fermentation kinetics. We have developed a reactor-engineering model for red wine fermentations that includes the fundamentals of fermentation kinetics, heat transfer, diffusion, and compressible fluid flow. To develop the heat transfer component of the model, the heat transfer properties of grapes were experimentally determined as a function of fermentation progression. COMSOL was used to solve all components of the model simultaneously utilizing a finite element analysis approach. Predictions from this model were validated using prior experimental work. Model prediction and experimental data showed excellent agreement. The model was then used to predict spatial profiles of active yeast cell concentration and ethanol productivity, as well as liquid velocity profiles. Finally, the model was used to predict how these gradients would change with differences in initial bioavailable nitrogen concentration, a key parameter in predicting fermentation outcome in nitrogen-limited wine fermentations.     

Exploring sample cross-contamination in fish epidermal mucus

Cross-contamination of epidermal mucus was assessed at three sampling locations on the bodies of Pacific halibut Hippoglossus stenolepis by inducing contact between fish coated with labelled synthetic mucus and non-treated fish. Results indicate a positive relationship between sampling site exposure and sample contamination and that mucous sample cross-contamination can be mitigated by sampling in a location protected from external contact.     

Temperature‐dependence of minimum resource requirements alters competitive hierarchies in phytoplankton

Resource competition theory is a conceptual framework that provides mechanistic insights into competition and community assembly of species with different resource requirements. However, there has been little exploration of how resource requirements depend on other environmental factors, including temperature. Changes in resource requirements as influenced by environmental temperature would imply that climate warming can alter the outcomes of competition and community assembly. We experimentally demonstrate that environmental temperature alters the minimum light and nitrogen requirements – as well as other growth parameters – of six widespread phytoplankton species from distinct taxonomic groups. We found that species require the most nitrogen at the highest temperatures while light requirements tend to be lowest at intermediate temperatures, although there are substantial interspecific differences in the exact shape of this relationship. We also experimentally parameterize two competition models, which we use to illustrate how temperature, through its effects on species’ traits, alters competitive hierarchies in multispecies assemblages, determining community dynamics. Developing a mechanistic understanding of how temperature influences the ability to compete for limiting resources is a critical step towards improving forecasts of community dynamics under climate warming.     

Quality control and integration of genotypes from two calling pipelines for whole genome sequence data in the Alzheimer's disease sequencing project

The Alzheimer's Disease Sequencing Project (ADSP) performed whole genome sequencing (WGS) of 584 subjects from 111 multiplex families at three sequencing centers. Genotype calling of single nucleotide variants (SNVs) and insertion-deletion variants (indels) was performed centrally using GATK-HaplotypeCaller and Atlas V2. The ADSP Quality Control (QC) Working Group applied QC protocols to project-level variant call format files (VCFs) from each pipeline, and developed and implemented a novel protocol, termed “consensus calling,” to combine genotype calls from both pipelines into a single high-quality set. QC was applied to autosomal bi-allelic SNVs and indels, and included pipeline-recommended QC filters, variant-level QC, and sample-level QC. Low-quality variants or genotypes were excluded, and sample outliers were noted. Quality was assessed by examining Mendelian inconsistencies (MIs) among 67 parent-offspring pairs, and MIs were used to establish additional genotype-specific filters for GATK calls. After QC, 578 subjects remained. Pipeline-specific QC excluded ~12.0% of GATK and 14.5% of Atlas SNVs. Between pipelines, ~91% of SNV genotypes across all QCed variants were concordant; 4.23% and 4.56% of genotypes were exclusive to Atlas or GATK, respectively; the remaining ~0.01% of discordant genotypes were excluded. For indels, variant-level QC excluded ~36.8% of GATK and 35.3% of Atlas indels. Between pipelines, ~55.6% of indel genotypes were concordant; while 10.3% and 28.3% were exclusive to Atlas or GATK, respectively; and ~0.29% of discordant genotypes were. The final WGS consensus dataset contains 27,896,774 SNVs and 3,133,926 indels and is publicly available.     

Modulation of PDGF mediated osteoblast chemotaxis by leukemia inhibitory factor LIF

Cell migration is a key event in tissue repair and remodeling. PDGF, a growth factor for multiple target cells, has been shown to be a potent chemoattractant for a variety of mesenchymal cells. However, it is likely that PDGF-mediated cell migration will be influenced by other cytokines that can be produced during physiological and pathological conditions. Leukemia inhibitory factor (LIF), a cytokine that is produced by a variety of cells including osteoblasts, may promote bone formation, but the mechanism is not known. Since osteoblasts are responsible for laying down new matrix during skeletal remodeling, in this report we have examined whether PDGF or LIF influences the migration of osteoblasts. Among several cytokines and growth factors tested, only PDGF was able to elicit a major chemotactic (directed migration) and a minor chemokinetic (random-migration) response in osteoblasts. LIF alone was not active in either chemotaxis or chemokinesis but when included with PDGF it caused a reduction in chemokinesis. Further, pretreatment of osteoblasts with LIF caused an increase in PDGF-driven chemotaxis. Finally, osteoblasts exposed briefly to LIF synthesized a higher level of non-collagenous proteins upon further treatment with PDGF. These observations are consistent with a role for LIF in promoting bone formation, both by influencing directional migration of osteoblasts and in laying down new matrix. © 1996 Wiley-Liss, Inc.     

The effects of infant births on the sociosexual behavior and hormonal patterns of a cooperatively breeding primate (Cebuella pygmaea)

This study examined changes in the behavioral and hormonal patterns of cooperatively breeding pairs in a primate species with the passing of time and with specific reproductive events. We (1) compared patterns of sexual, agonistic, and affiliative behavior of newly paired pygmy marmosets with the same behavioral patterns immediately after the birth of their first set of infants; (2) determined if postpartum behavioral differences existed between pairs whose infants lived and those whose infants died; and (3) examined whether behavioral patterns changed over the course of ovarian cycles in parous pygmy marmosets as had been documented in nulliparous pairs. The behavior of pairs was recorded during daily half-hour focal samples for 60 days after pairing, and 30 or 60 days after the birth of infants for pairs whose litters died or lived, respectively. Daily urine samples from females during the study were analyzed for luteinizing hormone and pregnanediol glucuronide concentrations to determine dates of ovulation. The results indicated that males consistently altered their sexual behavior and olfactory monitoring of mates during periovulatory periods in the females' cycles both postpairing and postpartum, while similar rates of social and sexual behavior were maintained between the conditions. Sexual behavior occurred throughout the females' ovarian cycles. Peaks in sexual behavior during the periovulatory period in nulliparous pairs disappeared after the birth of infants. Pairs whose infants died showed higher rates of sexual behavior than pairs with surviving infants. Social and sexual behavior may function to maintain the relationship both during and outside of ovulation, especially with the loss of infants. © 1996 Wiley-Liss, Inc.     

Morphological development,cell number,and allocation of cells to trophectoderm and inner cell mass of in vitro fertilized and parthenogenetically developed buffalo embryos: The effect of IGF-I

The morphology and number of cells in the trophectoderm (TE) and inner cell mass (ICM) of buffalo blastocysts derived from in vitro fertilization and cultured in the presence or absence of insulin-like growth factor-I (IGF-I) were analyzed by differential fluorochrome staining technique. The total cell number (TCN), TE number, and ICM cell number were significantly higher in blastocysts developed in vitro in the presence of IGF-I as compared to blastocysts developed without IGF-I (P < 0.01). It was observed that the buffalo blastocyst took 5–9 days postfertilization to develop in vitro. In order to correlate the time required for blastocyst development and the allocation of cells to TE and ICM, blastocysts were designated as fast (developing on or before day 7) or slow (developing after day 7). The TCN, TE, and ICM cells of fast-developing blastocysts cultured in the presence of IGF-I were significantly higher than slow-developing blastocysts (P < 0.01). The blastocysts developed on day 6 had a mean total cell number 118.6 ± 21.4, which significantly decreased to 85.6 ± 17.4, 62.0 ± 14.5, and 17.0 ± 4.0 on days 7, 8, and 9, respectively (P < 0.05). Normal development of buffalo embryo showed that, on average, embryos reached compact morula stage at the earliest between days 4.5–5.5. Blastocysts developed, at the earliest, between days 5.0–6.0, and it took them, on average, 6.5 days to hatch from the zona pellucida. TCN, TE, and ICM increased three times from morula to blastocyst; however, the proportion of ICM to TCN remained the same, in both embryonic stages. TE approximately doubled in hatched blastocysts, as compared to unhatched blastocysts (P < 0.05). However, ICM cells were decreased. The time required for development of parthenogenetic blastocysts was observed to be greater as compared to in vitro fertilized (IVF) blastocysts. The total cell number of parthenogenetic blastocysts was 100.8 ± 11.3, including 59.2 ± 8.4 cells of TE and 42.1 ± 6.9 cells of ICM. © 1996 Wiley-Liss, Inc.     

Two independent approaches converge to the cloning of a new Leptosphaeria maculans avirulence effector gene,AvrLmS‐Lep2

Brassica napus (oilseed rape, canola) seedling resistance to Leptosphaeria maculans, the causal agent of blackleg (stem canker) disease, follows a gene‐for‐gene relationship. The avirulence genes AvrLmS and AvrLep2 were described to be perceived by the resistance genes RlmS and LepR2, respectively, present in B. napus ‘Surpass 400’. Here we report cloning of AvrLmS and AvrLep2 using two independent methods. AvrLmS was cloned using combined in vitro crossing between avirulent and virulent isolates with sequencing of DNA bulks from avirulent or virulent progeny (bulked segregant sequencing). AvrLep2 was cloned using a biparental cross of avirulent and virulent L. maculans isolates and a classical map‐based cloning approach. Taking these two approaches independently, we found that AvrLmS and AvrLep2 are the same gene. Complementation of virulent isolates with this gene confirmed its role in inducing resistance on Surpass 400, Topas‐LepR2, and an RlmS‐line. The gene, renamed AvrLmS‐Lep2, encodes a small cysteine‐rich protein of unknown function with an N‐terminal secretory signal peptide, which is a common feature of the majority of effectors from extracellular fungal plant pathogens. The AvrLmS‐Lep2/LepR2 interaction phenotype was found to vary from a typical hypersensitive response through intermediate resistance sometimes towards susceptibility, depending on the inoculation conditions. AvrLmS‐Lep2 was nevertheless sufficient to significantly slow the systemic growth of the pathogen and reduce the stem lesion size on plant genotypes with LepR2, indicating the potential efficiency of this resistance to control the disease in the field.