Elucidating the Temporal Patterns of Gene Expression in the Inferred Regulatory Interactions of GmCOL1b in Glycine max.

The CONSTANS ( CO ) gene in Arabidopsis thaliana has a central role in photoperiodic regulation of flowering. However, the roles of CO genes in mediating flowering in soybeans ( Glycine max ) remain uncertain. We previously inferred regulatory interactions of a soybean CO homolog, GmCOL1b , using in-house RNA-seq data and the network inference algorithm package CausNet. Here, we identify potential GmCOL1b downstream genes and experimentally verify them by expressing GmCOL1b in soybean protoplast cells. Temporal expression patterns of these genes indicate the regulatory effects of GmCOL1b on the expression of the circadian clock genes GmLCL1 and GmLCL4 and the flowering regulator GmTEM1a .

Clarifying the Temporal Dynamics of the Circadian Clock and Flowering Gene Network Using Overexpression and Targeted Mutagenesis of Soybean EARLY FLOWERING 3-1 ( GmELF3-1 ).

With progressing climate fluctuations, an understanding of the molecular mechanisms of crop plants that regulate their flowering responses to environments is crucial. To achieve this goal, we aimed at clarifying the gene regulatory networks among the circadian clock and flowering genes in soybean ( Glycine max ). Based on our network inference approach , we hypothesize that GmELF3-1 , one of the Evening Complex (EC) gene homologs in soybean's circadian clock, may have an integrative role in transcriptional regulation of the circadian clock and flowering gene network. In this study, we verify GmELF3-1 ' s regulatory roles in its potential downstream genes by modulating the activity of GmELF3-1 using overexpression and CRISPR-Cas9 in soybean protoplasts. Our results indicate that GmELF3-1 may control the expression of the PRR genes in the circadian clock and the flowering gene GmCOL1a .

Experimental Verification of Inferred Regulatory Interactions of EARLY FLOWERING 3 ( GmELF3-1 ) in Glycine max.

Understanding the roles of evening complex (EC) genes in the circadian clock of plants can inform how diurnal transcriptional loops in the clock gene network function to regulate key physiological and developmental events, including flowering transition. Gene regulatory interactions among soybean's circadian clock and flowering genes were inferred using time-series RNA-seq data and the network inference algorithmic package CausNet. In this study, we seek to clarify the inferred regulatory interactions of the EC gene GmELF3-1. A gene expression analysis using soybean protoplasts as a transient model indicated regulatory roles of GmELF3-1 in expression of selected flowering genes.

Acute postoperative pain and dorsal root ganglia transcriptomic signatures following total knee arthroplasty (TKA) in rats: An experimental study.

Total knee arthroplasty (TKA) is the final treatment option for patients with advanced knee osteoarthritis (OA). Unfortunately, TKA surgery is accompanied by acute postoperative pain that is more severe than arthroplasty performed in other joints. Elucidating the molecular mechanisms specific to post-TKA pain necessitates an animal model that replicates clinical TKA procedures, induces acute postoperative pain, and leads to complete functional recovery. Here, we present a new preclinical TKA model in rats and report on functional and behavioral outcomes indicative of pain, analgesic efficacy, serum cytokine levels, and dorsal root ganglia (DRG) transcriptomes during the acute postoperative period. Following TKA, rats exhibited marked deficits in weight bearing that persisted for 28 days. Home cage locomotion, rearing, and gait were similarly impacted and recovered by day 14. Cytokine levels were elevated on postoperative days one and/or two. Treatment with morphine, ketorolac, or their combination improved weight bearing while gabapentin lacked efficacy. When TKA was performed in rats with OA, similar functional deficits and comparable recovery time courses were observed. Analysis of DRG transcriptomes revealed upregulation of transcripts linked to multiple molecular pathways including inflammation, MAPK signaling, and cytokine signaling and production. In summary, we developed a clinically relevant rat TKA model characterized by resolution of pain and functional recovery within five weeks and with pain-associated behavioral deficits that are partially alleviated by clinically administered analgesics, mirroring the postoperative experience of TKA patients.

Role of Heavy Metals in Diabetes: Mechanisms and Treatment Strategies.

Toxic metals affecting metabolic pathways have a broad range in the ecosystem from both natural and anthropogenic sources. Because of constant contamination from waste and untreated chemical effluents, their emissions have risen significantly over the last few decades, quickly gaining attention due to their crucial role in the development of several metabolic disorders, notably diabetes mellitus. Cadmium and arsenic not only spread widely in our atmosphere but are also linked to a wide range of health hazards. These are primarily accumulated in the liver, kidney, and pancreas once they reach the human body, where they have deleterious effects on the metabolism of glucose and its association with other metabolic pathways, particularly glycolysis, glycogenesis, and gluconeogenesis, by altering and impairing the specific activity of major enzymes. Impairment of hepatic glucose homeostasis plays a crucial role in diabetes mellitus pathogenesis. Impaired liver and kidney functions, as well as decreased pancreatic and muscle function, also contribute significantly to elevated levels of blood glucose. Heavy metals have the potential to cause changes in the conformation in these enzymes. They also impair hormonal balance by destroying the pancreas and adrenal glands. Such metals often facilitate the development of reactive oxygen species and inhibit antioxidant defense mechanisms, with multiple organs subsequently damaged. This review briefly discusses the involvement of heavy metals in metabolic disorders such as diabetes mellitus, the enzymes involved in this pathway, and glucose homeostasis.

Update on Diagnosis and Treatment of Adult Pulmonary Alveolar Proteinosis.

Pulmonary alveolar proteinosis (PAP) is a rare pulmonary surfactant homeostasis disorder resulting in buildup of lipo-proteinaceous material within the alveoli. PAP is classified as primary (autoimmune and hereditary), secondary, congenital and unclassifiable type based on the underlying pathogenesis. PAP has an insidious onset and can, in some cases, progress to severe respiratory failure. Diagnosis is often secured with bronchoalveolar lavage in the setting of classic imaging findings. Recent insights into genetic alterations and autoimmune mechanisms have provided newer diagnostics and treatment options. In this review, we discuss the etiopathogenesis, diagnosis and treatment options available and emerging for PAP.

Drug Induced Liver Injury Attributed to a Curcumin Supplement.

More severe reactions, higher acute liver failure rates, and higher recurrence rates on re-challenge occur with supplement-related Drug Induced Liver Injury (DILI) (Medina-Caliz et al., 2018). We report a case of curcumin-induced hepatocellular DILI in a 78-year old female admitted with jaundice, with a one-month latency. Extensive evaluation for alternative etiologies of hepatotoxicity was unremarkable. The Roussel Uclaf Causality Assessment Method (RUCAM) score of 6 for the supplement indicated a probable association (score >8: highly probable association). Peak levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were >20 times upper limit of normal. A 48% decrease in AST and ALT levels was observed 7 days after discontinuation of the supplement, and resolution of transaminitis was observed in 42 days. No re-challenge was performed. In conclusion, this case emphasizes the importance of recognizing curcumin supplements as DILI triggers. Furthermore, it reiterates the need for careful evaluation of herbal and dietary supplements (HDS) consumed by patients to identify potential DILI culprits, and to ultimately prevent DILI reactions with significant morbidity and mortality.

Frequency and features of nocturnal enuresis in Pakistani children aged 5 to 16 years based on ICCS criteria: a multi-center cross-sectional study from Karachi, Pakistan.

BACKGROUND: Nocturnal enuresis (NE) is a common symptom in children worldwide. International Children's Continence Society (ICCS) defines enuresis as either mono-symptomatic, NE with lower urinary tract symptoms and NE with co-morbid conditions. The objectives of this study were to determine the frequencies and types of NE and associated symptoms and conditions in children aged 5 to 16 years based on ICCS criteria. METHODS: A multi-center cross sectional study was conducted between November 2012 and December 2013 in the primary care clinics of four hospitals in Karachi. Children aged five to fifteen years were included through consecutive sampling. Informed consent was obtained from the parents and a pre-coded semi-structured questionnaire was used to obtain the information. Data was entered on SPSS version 20.0 and multivariable logistic regression analysis was used for data analysis. RESULTS: Out of 429 children aged between five and sixteen years, 243(56.9%) were boys and the remaining 186(43.1%) were girls. One hundred and eighty three children (43%) had nocturnal enuresis (NE). Forty four (10.3%), had mono-symptomatic NE, 57(31.1%) had associated lower urinary tract symptoms (NE-LUTS), whereas 30 (16.3%) had NE with a co-morbid condition. Fifty two (28.4%) NE's had at least one of both LUTS and a co-morbid condition. Out of the 246(57%) non-enuretic's, 31(12.6%) had a LUTS, 95(38.6%) had a co-morbid condition and 57(23.2%) had at least one of both LUTS and a co-morbid condition. The remaining 63 (25.6%) were symptom free. Increased voiding frequency, urgency, dysuria, suprapubic pain and daytime incontinence were the LUTS significantly associated with NE. Co-morbid conditions significantly associated with NE included constipation, congenital defects, developmental delay, and learning and sleep problems. CONCLUSION: Although NE can be an only symptom, it is often associated with lower urinary tract symptoms like dysuria, urgency, suprapubic pain, and daytime incontinence. Children presenting with NE often have co-morbid conditions like constipation, urinary tract infection, sleep disorders, and developmental delay. Many children presenting with these conditions as the primary complaint may also have NE. It should be addressed as unrecognized and untreated NE can cause additional morbidity and distress.

In silico analysis reveals widespread presence of three gene families, MAPK, MAPKK and MAPKKK, of the MAPK cascade from crop plants of Solanaceae in comparison to the distantly-related syntenic species from Rubiaceae, coffee.

Mitogen-activated protein kinases (MAPKs) are an important family of genes which play roles in vital plant processes, and they also help in coping against various kinds of environmental stresses including abiotic as well as biotic factors. The advancement of genomics calls for the annotation, identification, and detailed processing of the essential gene families in plants in order to provide insights into the importance of their central roles as well as for providing the basis for making their growth vigorous even under stressed conditions and, ultimately, to benefit from them by foreseeing the potential threats to their growth. In the current study, MAPK, MAPKK, and MAPKKK families of the MAPK cascade were identified and reported from five different agriculturally and economically important crop species of the Solanaceae and Rubiaceae families based on conserved signature motifs aligned throughout the members of the families under this gene superfamily. Genes reported from the species after strict filtering were: 89, tomato; 108, potato; 63, eggplant; 79, pepper; 64, coffee. These MAPKs were found to be randomly distributed throughout the genome on the chromosomes of the respective species. Various characteristics of the identified genes were studied including gene structure, gene and coding sequence length, protein length, isoelectric point, molecular weight, and subcellular localization. Moreover, maximum likelihood test of phylogeny was conducted on the retrieved sequences for the three MAPK cascade families to determine their homologous relationships which were also analyzed quantitatively by heat plots.

Quinoline derivatives: candidate drugs for a class B G-protein coupled receptor, the calcitonin gene-related peptide receptor, a cause of migraines.

Class B G-protein coupled receptors are involved in a wide variety of diseases and are a major focus in drug design. Migraines are a common problem, and one of their major causative agents is the class B G-protein coupled receptor, Calcitonin gene-related peptide (CGRP) receptor, a target for competitive drug discovery. The calcitonin receptor-like receptor generates complexes with a receptor activity-modifying protein, which determines the type of receptor protein formed. The CGRP receptor comprises a complex formed from the calcitonin receptor-like receptor and receptor activity-modifying protein 1. In this study, an in silico docking approach was used to target the calcitonin receptor-like receptor in the bound form with receptor activity-modifying protein 1 (CGRP receptor), as well as in the unbound form. In both cases, the resulting inhibitors bound to the same cavity of the calcitonin receptor-like receptor. The twelve evaluated compounds were competitive inhibitors and showed efficient inhibitory activity against the CGRP receptor and Calcitonin receptor-like receptor. The two studied quinoline derivatives demonstrated potentially ideal inhibitory activity in terms of binding interactions and low range nano-molar inhibition constants. These compounds could prove helpful in designing drugs for the effective treatment of migraines. We propose that quinoline derivatives possess inhibitory activity by disturbing CGRP binding in the trigeminovascular system and may be considered for further preclinical appraisal for the treatment of migraines.

Molecular docking studies of flavonoids for their inhibition pattern against ß-catenin and pharmacophore model generation from experimentally known flavonoids to fabricate more potent inhibitors for Wnt signaling pathway.

BACKGROUND: Canonical Wnt signaling plays a key role in tumor cell proliferation, which correlates with the accumulation of ß-catenin in cell due to inactivation of glycogen synthetase kinase-3 ß. However, uncontrolled expression of ß-catenin leads to fibromatosis, sarcoma and mesenchymal tumor formation. Recently, a number of polyphenolic compounds of naturally occurring flavonoid family have been screened for the inhibition of Wnt signaling. OBJECTIVE: Elucidation of the binding mode of inhibitors to ß-catenin, reporting more potent inhibitors for the disease-causing protein and designing a pharmacophore model based on naturally occurring compounds, flavonoids. MATERIALS AND METHODS: In this study, a comparative molecular docking analysis was performed to elucidate the binding mode of experimentally reported and unknown inhibitors. Based on the knowledge of geometry, binding affinity and drug score, we described a subset of novel inhibitors. RESULTS: The binding energy of known inhibitors (isorhamnetin, fisetin, genistein and silibinin) was observed in a range of -5.68 to -4.98 kcal/mol, while novel inhibitors (catechin, luteolin, coumestrol and ß-naphthoflavone) exhibited -6.50 to -5.22 kcal/mol. We observed good placement and strong interactions of selected compounds inside the binding pocket of ß-catenin. Moreover, flavonoid family members and T cell factors 4 (TCF4) compete for ß-catenin binding by sharing common binding residues. CONCLUSION: This study will largely help in understanding the molecular basis of ß-catenin/TCF4 inhibition through flavonoids by exploring their structural details. Finally, the novel inhibitors proposed in this study need further attention to uncover cancer treatment and with the generated pharmacophore model, more and potent ß-catenin inhibitors can be easily screened.

Warfarin Therapy: Survey of Patients' Knowledge of their Drug Regimen.

BACKGROUND: Warfarin is utilised for the treatment of thromboembolic disease. Its use demands a careful and continual monitoring given its narrow therapeutic index and potentially life-threatening complications. The aim of this study was to assess the extent of patients' knowledge of their warfarin therapy. METHODS: A total of 200 consecutive patients from a single community hospital completed an online survey questionnaire (www.eSurveysPro.com). Using the responses to the questionnaire, we recorded compliance to warfarin therapy, knowledge about drug interactions, adverse effects of warfarin therapy, complications, and resulting hospitalisation. RESULTS: We recruited 200 patients, 55% (109/200) women and 45% (91/200) men, among which 88% were compliant with their daily medication. Of the 200 patients, 56% were unaware of any potential drug interactions, 58% were unaware of any adverse effects, 27% had experienced adverse effects, 12% had been hospitalised because of adverse effects (33% of which were due to bleeding), and 65% kept a personal record of their international normalised ratio. CONCLUSION: Despite the high level of compliance, patient knowledge of warfarin therapy was low. Given the potential drug interactions and complexities involved with warfarin therapy, it is of high importance that medical professionals educate their patients and make them aware of any impending signs of emergent medical complications.

The effect of different environmental conditions on the encystation of Acanthamoeba castellanii belonging to the T4 genotype.

In this study, Acanthamoeba castellanii was cultivated under different stress conditions to induce possible encystation. The morphological and histological properties were analysed by light and electron microscopy as well as cyst-specific staining. The findings revealed that cysts prepared through liquid medium using higher osmolarity as a trigger (10% glucose with 50mM magnesium chloride for 72 h) are similar to cysts prepared using non-nutrient agar (nutrient deprivation as a trigger in plating assays for 14 days), as determined by SDS-resistance, cyst-specific Calcofluor white staining and transmission electron microscopy. Using liquid medium assay, A. castellanii encystation was studied by exposing trophozoites to media lacking growth ingredients (phosphate buffered saline or distilled water), inappropriate temperatures (4-45°C), pH (3-9), artificial light-dark cycles, 5% CO2, and microaerophilic conditions. Optimal encystation was observed when cells were incubated in PBS with 50mM MgCl2 and 10% glucose at 24-30°C at pH 7. Increasing temperature over 37°C or pH 9 adversely affected encystation, while light-dark cycles, 5% CO2 and microaerophilic conditions had no effect on encystation of A. castellanii. None of the aforementioned conditions had any effect on the viability of A. castellanii, as determined by Trypan blue exclusion assay. A complete knowledge of encystation in A. castellanii is crucial to our understanding of the biology of these ecologically and medically important organisms.