The effect of modification of DNA interference on myostatin gene expression in mice.

Myostatin is a known negative regulator of muscle tissue growth. Thus, an inhibitor of myostatin may be therapeutically useful as an anabolic agent for the muscle tissue. A promising gene-silencing approach for gene therapy is DNA interference (DNAi), a sequence that is complementary to the promoter region of a target gene. To confer resistance to nuclease digestion, several modifications such as methylphosphonate or phosphorothioate have been proposed, wherein a nonbridging oxygen atom in the oligonucleotide phosphate backbone is replaced by sulphur. The aim of the present study was to assess the effectiveness of the DNAi molecule with phosphorothioate (PS) and without phosphorothioate (WPS) modification for inhibition of myostatin gene expression in mice. Eighteen four-week-old male BALB/c mice were randomly divided into three groups: DNAi-PS (n = 6), DNAi-WPS (n = 6) and control (n = 6). Intraperitoneal injections of DNAi (10 mg/kg) were given once a week, and mice body weights were measured weekly and sacrificed after three weeks. The expression of myostatin was assessed using real-time quantitative polymerace chain reaction. For histological evaluation, the skeletal muscle tissue was dissected from the biceps. The results were analysed by a t-test. Results demonstrated that administration of DNAi intraperitoneally with modification could suppress myostatin expression by up to 70%. Leg weight and histological analysis proved that chemically modified DNAi significantly suppressed the myostatin gene in mice. Overall, the results on DNA-induced gene silencing by antisense DNA oligonucleotides in animals can provide insight into the treatment of inherited diseases.

Rumen-protected l-carnitine supplementation during mating period altered metabolic status and reproductive performance of ewes.

Current study hypothesized that dietary l-carnitine (LC) inclusion during the mating period ameliorates both metabolic status and reproductive performance of ewes. Seventy Baluchi ewes (52 ± 4.2 kg of bodyweight and 18 ± 6 months old of age) were enrolled in this study. Animals were randomly allocated into two dietary treatments, control (only basal diet) or basal diet plus supplementation with a rumen-protected LC (Carneon 20 Rumin-pro; 20% LC; Kaesler Nutrition GmbH) at the rate of 10 g/head/day from 21 days before until 35 days after introducing rams to the ewes (MP). Feed intake was monitored by subtracting the ort from feed offered. Blood sample collection was conducted on Days -10, +10 and +20 relative to MP. Pregnancy was confirmed on Day 30 post-MP. Feed intake of the ewes in the LC group was higher than the control (p < 0.05). LC supplementation increased the cholesterol concentration in the ewes (p < 0.05). Blood urea concentration of animals in the LC group was significantly lower than the control (p < 0.05). The mRNA expression of toll-like receptor 4 was evidently lower in animals supplemented with LC than the control (p < 0.05). Both lambing and fecundity rates in the LC group tended to be higher compared with the control. LC supplementation showed potential to alter certain metabolites in the ewes. A tendency for higher lambing rate may partly be driven by dams efficient energy partitioning to support foetal growth and maintaining pregnancy.

Identification of selection signatures in Capra hircus and Capra aegagrus in Iran.

Identification of selection signatures may provide a better understanding of domestication process and candidate genes contributing to this process. In this study, two populations of domestic and wild goats from Iran were analyzed to identify selection signatures. RSB, iHS, and XP-EHH statistics were used in order to identify robust selection signatures in the goat genome. Genotype data of domestic and wild goats from the NextGen project was used. The data was related to 18 Capra aegagrus (wild goat) and 20 Capra hircus (domestic goat) from Iran. The iHS method indicated 675 and 441 selection signatures in C. aegagrus and C. hircus, respectively. RSB and XP-EHH methods showed about 370 and 447 selection signatures in C. aegagrus and C. hircus, respectively. These selection signatures were mainly associated with milk production, fleece trait, mammary epithelial cells, reproduction, and immune system.

The immunomodulatory effects of lactoferrin and its derived peptides on NF-κB signaling pathway: A systematic review and meta-analysis.

BACKGROUND: Lactoferrin is a versatile protein with important modulatory functions in inflammation and immune response. This glycoprotein can bind and sequester iron and LPS, thereby intervening in certain signaling pathways and biological processes. In the present meta-analysis, we aimed to pool experimental data regarding the immunomodulatory effects of lactoferrin and its derived peptides on the NF-κB signaling pathway. MATERIALS: We searched PubMed, Google Scholar, and Web of Science databases and obtained all related articles published before April 2022. Finally, 25 eligible studies were selected, and their reports were analyzed. METHODS: We used Review Manager Version 5.2 to compute the standardized mean difference (SMD) and its 95% confidence interval. In addition, the source of heterogeneity was explored using meta-regression and sensitivity analysis. The symmetry of the funnel plot and Egger's test were also used to evaluate publication bias utilizing Comprehensive Meta-Analysis Version 2. RESULTS: Comparing the group of cells and animals exposed to lipopolysaccharide alone with the group that received pretreatment with lactoferrin and its derivatives, we observed significant reductions in TNF-α, IL-1 beta, and IL-6 levels by 8.73 pg/mL, 2.21 pg/mL, and 3.24 pg/mL, respectively, in the second group. Additionally, IKK-ß, p-IκB, and NF-κB (p65) levels were significantly lower by 7.37-fold, 15.02-fold, and 3.88-fold, respectively, in various cells and tissues. CONCLUSION: Based on the results of this meta-analysis, lactoferrin and its derived peptides can be considered potent prophylactic and therapeutic candidates against inflammation-associated diseases by targeting the NF-kB pathway.

Differential Expression of RNAseq Imprinted Genes from Bovine Females Before and After Puberty.

The productivity of beef cows depends on early reproduction traits such as puberty and has an economic impact on the efficiency of production system. Imprinted genes modulate many important endocrine processes such as growth, the onset of puberty and maternal reproductive and behavior. The role of imprinted genes in puberty is a challenging subject since they show the reciprocal role of maternal and paternal genomes in progeny. Although, there are evidences of the involvement of imprint genes in puberty in human, the role of this type of genes in the onset of puberty in cattle has not been studied yet. Here we examined the expression of 27 imprinted genes in pre and post puberty in a bovine model to find differentially expressed imprinted genes in maternal-paternal purebreds and reciprocal crosses across eight tissues and discussed the task of these genes in this crucial process of development and in onset of puberty. DLK1 and MKRN3 that previously described as cause of the central precocious puberty (CPP) in human were differentially expressed in this study. Functional annotation analysis of differentially imprinted genes in different tissues showed significant biological processes of cellular response to growth factor stimulus, response to growth factor, response to parathyroid hormone, developmental growth and the importance of alternative splicing. The results of this study have implications in understanding the role of imprinted genes in the onset of puberty in cattle.

Effects of adding poly-histidine tag on stability, antimicrobial activity and safety of recombinant buforin I expressed in periplasmic space of Escherichia coli.

The lack of cost-effective methods for producing antimicrobial peptides has made it impossible to use their high potential as a new and powerful class of antimicrobial agents. In recent years, extensive research has been conducted to decrease the cost of recombinant proteins production through microorganisms, transgenic animals, and plants. Well-known genetic and physiological characteristics, short-term proliferation, and ease of manipulation make E. coli expression system a valuable host for recombinant proteins production. Expression in periplasmic space is recommended to reduce the inherently destructive behavior of antimicrobial peptides against the expressing microorganism and to decline susceptibility to proteolytic degradation. In this study, a pET-based expression system was used to express buforin I at E. coli periplasmic space, and its antimicrobial, hemolytic, and cell toxicity activities as well as structural stability were evaluated. The hemolysis activity and cytotoxicity of His-tagged buforin I were negligible and its antimicrobial activity did not show a significant difference compared to synthetic buforin I. In addition, in silico investigating of stability of native and His-tagged buforin I showed that RMSF, RMSD and Rg curves had followed a similar trend during 150 ns simulation. Furthermore, evaluating the modelled structures, FTIR and X-ray methods of both peptides indicated an insignificant structural difference. It was concluded that the recombinant buforin I could be a viable alternative to some currently used antibiotics by successfully expressing it in the pET-based expression system.

Asymmetric growth-limiting development of the female conceptus.

Introduction: Sex differences in prenatal growth may contribute to sex-dependent programming effects on postnatal phenotype. Methods: We integrated for the first time phenotypic, histomorphological, clinico-chemical, endocrine and gene expression analyses in a single species, the bovine conceptus at mid-gestation. Results: We demonstrate that by mid-gestation, before the onset of accelerated growth, the female conceptus displays asymmetric lower growth compared to males. Female fetuses were smaller with lower ponderal index and organ weights than males. However, their brain:body weight, brain:liver weight and heart:body weight ratios were higher than in males, indicating brain and heart 'sparing'. The female placenta weighed less and had lower volumes of trophoblast and fetal connective tissue than the male placenta. Female umbilical cord vessel diameters were smaller, and female-specific relationships of body weight and brain:liver weight ratios with cord vessel diameters indicated that the umbilico-placental vascular system creates a growth-limiting environment where blood flow is redistributed to protect brain and heart growth. Clinico-chemical indicators of liver perfusion support this female-specific growth-limiting phenotype, while lower insulin-like growth factor 2 (IGF2) gene expression in brain and heart, and lower circulating IGF2, implicate female-specific modulation of key endocrine mediators by nutrient supply. Conclusion: This mode of female development may increase resilience to environmental perturbations in utero and contribute to sex-bias in programming outcomes including susceptibility to non-communicable diseases.

Investigating the Probiotic Properties and Antimicrobial Activity of Lactic Acid Bacteria Isolated from an Iranian Fermented Dairy Product, Kashk.

In the present study, kashk samples were collected from two regions of Iran, the Fars (Abadeh) and Razavi Khorasan (Kalat) provinces. Fifteen bacteria were isolated and physiological and biochemical assays were performed. After identification to the genus level, eight isolates were identified as lactic acid bacteria (LAB) and subjected to molecular identification and probiotic properties assays. The results revealed that the isolates were Enterococcus faecium KKP 3772 (KF1), Enterococcus faecium C1 (KF2), Pediococcus pentosaceus H11 (KF3), Pediococcus pentosaceus VNK-1 (KK4), Lactococcus lactis RSg (KK1), Enterococcus faecalis P190052 (KK2), Enterococcus mundtii CECT972T (KK3), and Lactiplantibacillus plantarum PM411 (KK5). Only the numbers of L. lactis RSg (KK1) and Lpb. Plantarum PM411 (KK5) decreased to below 9 Log CFU/mL after acidic conditions (pH = 3) and showed weak antibacterial activity. Enterococcus mundtii CECT972T (KK3) and E. faecium C1(KF2) were highly susceptible to bile salts, while P. pentosaceus VNK-1 (KK4) and P. pentosaceus H11 (KF3) showed the highest resistance. All of the isolates were resistant to tetracycline and sensitive to chloramphenicol and gentamicin. The antimicrobial activity of P. pentosaceus VNK-1 (KK4) and P. pentosaceus H11 (KF3) was higher than other isolates and consequently, their inhibition zones were larger. The adhesion capabilities of LAB isolates to intestinal epithelial cells were evaluated by examining the auto-aggregation factor and cell surface hydrophobicity. The highest and lowest cell surface hydrophobicity and auto-aggregation were obtained from P. pentosaceus VNK-1 (KK4) and E. mundtii CECT972T (KK3), respectively. In general, P. pentosaceus VNK-1 (KK4) and P. pentosaceus H11 (KF3) have shown better probiotic properties as compared to other isolates.

Interactive effect of exercise training and growth hormone administration on histopathological and functional assessment of the liver in male Wistar rats.

BACKGROUND: Abuse of growth hormone (GH) is expanding in exercising populations due to its lipolytic and anabolic actions. The purpose of this study was to examine the interactive effect of exercise training and GH administration on histopathological and functional assessment in the liver of male Wistar rats. METHODS: Forty-eight male Wistar rats were randomly divided into six groups including control + saline group (CS), GH injection group (GI), resistance training + saline group (RS), aerobic training + saline group (AS), resistance training + GH injection group (RG), aerobic training + GH injection group (AG). All groups were injected with either saline or GH 1 h before each training session. RT and AT were performed five days/week for a total of 8-weeks. At the end of the study, blood samples and liver tissue samples were taken to evaluate circulating AST, ALT, and ALP enzymes, as well as albumin protein. Histopathology of liver tissue was performed via qualitative microscopic evaluation. RESULTS: Microscopic evaluation of liver tissue did not show any histopathologic changes. All the groups administered with GH showed a significant increase in ALT, ALP, and albumin protein (P<0.05). However, AST enzyme concentrations increased significantly only in the RG group (P=0.022). In addition, neither RS nor the AS groups showed significant AST, ALT, and ALP changes, but serum albumin concentration significantly increased in the AS group (P=0.033). CONCLUSIONS: The elevation of liver enzymes showed that GH administration with or without exercise training might cause severe liver damage.

Developing novel liquid biopsy by selective capture of viral RNA on magnetic beads to detect COVID-19.

Objectives: Early, specific, and sensitive detection methods of COVID-19 are essential for force stopping its worldwide infection. Although CT images of the lung and/or viral RNA extraction followed by real-time reverse-transcriptase-polymerase chain reaction (rRT-PCR) are widely used; they have some limitations. Here, we developed a highly sensitive magnetic bead-based viral RNA extraction assay followed by rRT-PCR. Materials and Methods: Case group included oropharyngeal/nasopharyngeal and blood samples from 30 patients diagnosed positive by PCR test for COVID-19 and control group included 30 same samples from COVID-19 negative PCR test individuals. RNA was extracted, using viral RNA extraction kit as well as using our hand-made capture bead-based technique. A one-step cDNA synthesis and Real Time PCR was conducted. A two-step comparison of the different viral RNA extraction methods for oropharyngeal/nasopharyngeal and blood samples was performed. Student t-test was applied with a P<0.05 considered statistically significant. Results: In the case group, all 30 mucosal samples extracted either with viral RNA extraction kit or with beads-based assay were COVID-19 positive although in the latter category, Cqs were much lower. Although 43% of plasma samples extracted by bead-based method were found to be positive but no plasma samples extracted with column-based kit were detected positive by Real Time PCR. Conclusion: Bead-based RNA extraction method can reduce RNA loss by its single-tube performance and enhance the test sensitivity. It is also more sensitive to lower viral loads as shown in the detection of blood samples and the lower Cqs of mucosal samples.

Differential expression analysis of genes and long non-coding RNAs associated with KRAS mutation in colorectal cancer cells.

KRAS mutation is responsible for 40-50% of colorectal cancers (CRCs). RNA-seq data and bioinformatics methods were used to analyze the transcriptional profiles of KRAS mutant (mtKRAS) in comparison with the wild-type (wtKRAS) cell lines, followed by in-silico and quantitative real-time PCR (qPCR) validations. Gene set enrichment analysis showed overrepresentation of KRAS signaling as an oncogenic signature in mtKRAS. Gene ontology and pathway analyses on 600 differentially-expressed genes (DEGs) indicated their major involvement in the cancer-associated signal transduction pathways. Significant hub genes were identified through analyzing PPI network, with the highest node degree for PTPRC. The evaluation of the interaction between co-expressed DEGs and lncRNAs revealed 12 differentially-expressed lncRNAs which potentially regulate the genes majorly enriched in Rap1 and RAS signaling pathways. The results of the qPCR showed the overexpression of PPARG and PTGS2, and downregulation of PTPRC in mtKRAS cells compared to the wtKRAS one, which confirming the outputs of RNA-seq analysis. Further, significant upregualtion of miR-23b was observed in wtKRAS cells. The comparison between the expression level of hub genes and TFs with expression data of CRC tissue samples deposited in TCGA databank confirmed them as distinct biomarkers for the discrimination of normal and tumor patient samples. Survival analysis revealed the significant prognostic value for some of the hub genes, TFs, and lncRNAs. The results of the present study can extend the vision on the molecular mechanisms involved in KRAS-driven CRC pathogenesis.

MS-HRM protocol: a simple and low-cost approach for technical validation of next-generation methylation sequencing data.

DNA methylation is a fundamental epigenetic process and have a critical role in many biological processes. The study of DNA methylation at a large scale of genomic levels is widely conducted by several techniques that are next-generation sequencing (NGS)-based methods. Methylome data revealed by DNA methylation next-generation sequencing (mNGS), should be always verified by another technique which they usually have a high cost. In this study, we offered a low-cost approach to corroborate the mNGS data. In this regard, mNGS was performed on 6 colorectal cancer (case group) and 6 healthy individual colon tissue (control group) samples. An R-script detected differentially methylated regions (DMRs), was further validated by high resolution melting (MS-HRM) analysis. After analyzing the data, the algorithm found 194 DMRs. Two locations with the highest level of methylation difference were verified by MS-HRM, which their results were in accordance with the mNGS. Therefore, in the present study, we suggested MS-HRM as a simple, accurate and low-cost method, useful for confirming methylation sequencing results.

Intestinal changes and immune responses during Clostridium perfringens-induced necrotic enteritis in broiler chickens.

Clostridium perfringens-induced necrotic enteritis (NE) is an economically important disease of broiler chickens. The present study evaluated the effect of C. perfringens on the intestinal histomorphometry, enteric microbial colonization, and host immune responses using 3 experimental NE reproduction methods. The experimental groups consisted of 1) unchallenged Control diet (corn-soybean meal), 2) Control diet + Eimera inoculation at d 11 followed by C. perfringens challenge at d 15 (ECp), 3) Wheat-based diet + C. perfringens challenge (WCp), and 4) Wheat-based diet + Eimeria inoculation followed by C. perfringens challenge (WECp). The results showed that chickens receiving ECp and WECp had reduced (P < 0.05) bird performance coupled with enteric gross lesions and epithelial damage at d 17 and 24 of age compared to unchallenged control birds. These ECp and WECp administered birds also had increased (P < 0.05) ileal colonization by clostridia and E. coli at d 17 and 24, while the resident Lactobacillus counts were reduced (P < 0.05) at d 24 of age. Furthermore, at d 24, jejunal transcription of IL-6, IL-10, annexin-A1 and IL-2 genes was upregulated (P < 0.05) in the ECp group, whereas the transcription of TNF receptor associated factor (TRAF)-3 gene was increased (P < 0.05) in WECp treated birds when compared to unchallenged control group. Additionally, stimulation of chicken splenocytes and cecal tonsilocytes with virulent C. perfringens bacilli or their secretory proteins resulted in a higher (P < 0.05) frequency of T cells and their upregulation of MHC-II molecule, as determined by flow cytometry. These findings suggest that C. perfringens, while inducing epithelial damage and changes in microbiota, can also trigger host immune responses. Furthermore, NE reproduction methods using coccidia with or without the wheat-based dietary predisposition seem to facilitate an optimal NE reproduction in broiler chickens and thus, may provide better avenues for future C. perfringens research.

Beneficial worm allies warn plants of parasite attack below-ground and reduce above-ground herbivore preference and performance.

Antagonistic interactions among different functional guilds of nematodes have been recognized for quite some time, but the underlying explanatory mechanisms are unclear. We investigated responses of tomato (Solanum lycopersicum) to two functional guilds of nematodes-plant parasite (Meloidogyne javanica) and entomopathogens (Heterorhabditis bacteriophora, Steinernema feltiae below-ground, and S. carpocapsae)-as well as a leaf mining insect (Tuta absoluta) above-ground. Our results indicate that entomopathogenic nematodes (EPNs): (1) reduced root knot nematode (RKN) infestation below-ground, (2) reduced herbivore (T. absoluta) host preference and performance above-ground, and (3) induced overlapping plant defence responses by rapidly activating polyphenol oxidase and guaiacol peroxidase activity in roots, but simultaneously suppressing this activity in above-ground tissues. Concurrently, we investigated potential plant signalling mechanisms underlying these interactions using transcriptome analyses. We found that both entomopathogens and plant parasites triggered immune responses in plant roots with shared gene expression. Secondary metabolite transcripts induced in response to the two nematode functional guilds were generally overlapping and showed an analogous profile of regulation. Likewise, we show that EPNs modulate plant defence against RKN invasion, in part, by suppressing active expression of antioxidant enzymes. Inoculations of roots with EPN triggered an immune response in tomato via upregulated phenylpropanoid metabolism and synthesis of protease inhibitors in plant tissues, which may explain decreased egg laying and developmental performance exhibited by herbivores on EPN-inoculated plants. Furthermore, changes induced in the volatile organic compound-related transcriptome indicated that M. javanica and/or S. carpocapsae inoculation of plants triggered both direct and indirect defences. Our results support the hypothesis that plants "mistake" subterranean EPNs for parasites, and these otherwise beneficial worms activate a battery of plant defences associated with systemic acquired resistance and/or induced systemic resistance with concomitant antagonistic effects on temporally co-occurring subterranean plant pathogenic nematodes and terrestrial herbivores.

Improvement of the performance of anticancer peptides using a drug repositioning pipeline.

The use of anticancer peptides (ACPs) as an alternative/complementary strategy to conventional chemotherapy treatments has been shown to decrease drug resistance and/or severe side effects. However, the efficacy of the positively-charged ACP is inhibited by elevated levels of negatively-charged cell-surface components which trap the peptides and prevent their contact with the cell membrane. Consequently, this decreases ACP-mediated membrane pore formation and cell lysis. Negatively-charged heparan sulphate (HS) and chondroitin sulphate (CS) have been shown to inhibit the cytotoxic effect of ACPs. In this study, we propose a strategy to promote the broad utilization of ACPs. In this context, we developed a drug repositioning pipeline to analyse transcriptomics data generated for four different cancer cell lines (A549, HEPG2, HT29, and MCF7) treated with hundreds of drugs in the LINCS L1000 project. Based on previous studies identifying genes modulating levels of the glycosaminoglycans (GAGs) HS and CS at the cell surface, our analysis aimed at identifying drugs inhibiting genes correlated with high HS and CS levels. As a result, we identified six chemicals as likely repositionable drugs with the potential to enhance the performance of ACPs. The codes in R and Python programming languages are publicly available in https://github.com/ElyasMo/ACPs_HS_HSPGs_CS. As a conclusion, these six drugs are highlighted as excellent targets for synergistic studies with ACPs aimed at lowering the costs associated with ACP-treatment.

KRAS-related long noncoding RNAs in human cancers.

KRAS is one of the most widely prevalent proto-oncogenes in human cancers. The constitutively active KRAS oncoprotein contributes to both tumor onset and cancer development by promoting cell proliferation and anchorage-independent growth in a MAPK pathway-dependent manner. The expression of microRNAs (miRNAs) and the KRAS oncogene are known to be dysregulated in various cancers, while long noncoding RNAs (lncRNAs) can act as regulators of the miRNAs targeting KRAS oncogene in different cancers and have gradually become a focus of research in recent years. In this review article, we summarize recent advances in the research on lncRNAs that have sponging effects on KRAS-targeting miRNAs as crucial mediators of KRAS expression in different cell types and organs. A deeper understanding of lncRNA function in KRAS-driven cancers is of major fundamental importance and will provide a valuable clinical tool for the diagnosis, prognosis, and eventual treatment of cancers.

Buforin I an alternative to conventional antibiotics: Evaluation of the antimicrobial properties, stability, and safety.

Cationic antimicrobial peptides are being developed as a promising class of antimicrobial sub-stances. The introduction of a new antibiotic component requires a comprehensive study of its properties so that it can be relied upon to continue laboratory procedures and clinical trials on laboratory animals or human volunteers. Antimicrobial activity of buforin I was evaluated against 15 of the most important pathogenic bacterial and fungal strains. This was followed by assessing anti-biofilm activity, time-dependent inhibitory, thermal stability, plas-ma stability, hemolysis, and cytotoxic activities. The range of obtained MICs was between 4 and 16 µg/mL. The most resistant and most sensitive microbial strains were S. salivarius and C. perfringens, respectively. Buforin I not only inhibited biofilm formation, but also showed a high biofilm radiation activity. Buforin I was stable in human plasma and also at different temperatures including 40, 60, and 80 °C. Although no significant anti-cancer properties were observed for buforin I, the lack of cytotoxicity as well as the lack of hemolytic activity confirm its safety. The high therapeutic index indicated that buforin I has a considerable pharmaceutical potential and can be a reasonable candidate to replace antibiotics or administered in combination with antibiotics to increase the effectiveness as well as reduce the dose of antibiotics.

Antibacterial effects assessment on some livestock pathogens, thermal stability and proposing a probable reason for different levels of activity of thanatin.

There is a continuing need to prevent the increasing use of common antibiotic and find the replacement to combat the drug/antibiotic resistant bacteria such as antimicrobial peptides (AMPs) such as thanatin peptide. In this study, recombinant thanatin peptide was expressed in the HEK293 cell line. Then the antimicrobial properties of this peptide on some poultry and farm animal's pathogen strains were assessed. The thermal-stability of thanatin was predicted in various temperatures through in silico analysis. Afterwards, according to Minimum Inhibitory Concentration (MIC) results, Escherichia coli and Pseudomonas aeruginosa were chosen to test the hypothesis of LptA/LptD-thanatin interaction, computationally. Relative amino acid sequences and crystallography structures were retrieved and missed tertiary structures were predicted. The interaction of thanatin with LptA and LptD of Escherichia coli and Pseudomonas aeruginosa were analyzed subsequently. The antibacterial activity of thanatin peptide was evaluated between 6.25 and 100 µg/mL using minimum inhibitory concentration. Also, the amounts of minimum bactericidal concentrations (MBC) were between 12.5 and 200 µg/mL. The bioinformatics analysis followed by the in vitro assessment, demonstrated that thanatin would be thermally stable in the body temperature of poultry and farm animals. Thanatin could penetrate to the outer membrane domain of LptD in Escherichia coli and it could block the transition path of this protein while the entrance of LptD in Pseudomonas aeruginosa was blocked for thanatin by extra residues in comparison with Escherichia coli LptD. In addition, the quality of interaction, with regard to the number and distance of interactions which leads to higher binding energy for thanatin and LptD of Escherichia coli was much better than Pseudomonas aeruginosa. But the site and quality of interaction for thanatin and LptA was almost the same for Escherichia coli and Pseudomonas aeruginosa. Accordingly, thanatin can prevent the assembly of LptA periplasmic bridge in both pathogens. The antibacterial and thermal stability of the thanatin peptide suggested that thanatin peptide might serve as a natural alternative instead of common antibiotics in the veterinary medicine. The outcome of this in silico study supports the MIC results. Therefore, a probable reason for different level of activity of thanatin against Escherichia coli and Pseudomonas aeruginosa might be the quality of LptA/LptD-thanatin interaction.

A DNA methylation panel for high performance detection of colorectal cancer.

One of the most promising ways to diagnose cancer especially colorectal cancer (CRC) is to trace its epigenetic events. In this article, a discovery step for detection of methylated DNA markers (MDMs) was performed using SureSelectXT Methyl-Seq in CRC case and control groups in addition to several methylation profiling datasets (GSE48684, GSE53051, GSE77718, GSE101764, and GSE42752). In silico validation of MDMs in colorectal and other cancers was conducted by Lnc2met. MethyLight assay was run on 40 and 47 case and control formalin-fixed paraffin-embedded tissues, respectively and the performance of selected genes were classified by support vector machine (SVM). As a result, 180 regions were identified among all common genes. In addition to SEPT9 and SFRP2, the best three MDM regions were selected from SLC30A10, AKR1B1 and GALNT14. Based on all assays, the best performance was accomplished by SEPT9/AKR1B1 with 98% sensitivity, 99% specificity, 125 positive likelihood ratio, 0.02 negative likelihood ratio and 5074 diagnostic odds ratio. Our results indicate that the AKR1B1/SEPT9 methylation panel detects CRC with a higher performance than SEPT9 methylation, which is a commercial diagnostic test for CRC. However, the creation of a clinically valuable test derived from this study requires performance evaluation in liquid biopsies.

Peripartal Rumen-Protected L-Carnitine Manipulates the Productive and Blood Metabolic Responses in High-Producing Holstein Dairy Cows.

This study aimed to monitor the effect of including rumen-protected L-carnitine (Carneon 20 Rumin-Pro, Kaesler Nutrition GmbH, Cuxhaven, Germany) in the transition diet on the productive and metabolic responses of multiparous high-producing Holstein dairy cows. Thirty-two multiparous cows were allocated in a completely randomized design to receive the same diet plus 60 g fat prill containing 85% palmitic acid (control, n = 16) or 100 g rumen-protected L-carnitine (RLC, n = 16); at 28 days before expected calving until 28 days in milk (DIM). Fat prill was included in the control diet to balance the palmitic acid content of both experimental diets. Milk production over the 28 DIM for the control and RLC groups was 46.5 and 47.7 kg, respectively. Milk fat content tended to increase upon rumen-protected L-carnitine inclusion (p = 0.1). Cows fed rumen-protected L-carnitine had higher fat- and energy-corrected milk compared with the control group. Pre- and post-partum administration of L-carnitine decreased both high- and low-density lipoprotein concentrations in peripheral blood of post-partum cows. The results of this study indicated that the concentration of triglycerides and beta-hydroxybutyrate was not significantly different between the groups, whereas the blood non-esterified fatty acid concentration was markedly decreased in cows supplemented with L-carnitine. Animals in the RLC group had a significant (p < 0.05) lower blood haptoglobin concentration at 7 and 14 DIM than the control. Animals in the RLC group had a lower concentration of blood enzymes than those of the control group. The mRNA abundance of Toll-like receptors 4, cluster of differentiation 14, and myeloid differential protein 2 did not significantly change upon the supplementation of L-carnitine in the transition diet. In summary, the dietary inclusion of RLC improved dairy cow's performance during the early lactation period. Greater production, at least in part, is driven by improved energy utilization efficiency and enhanced metabolic status in animals during the periparturient period.