Publications by authors named "Sajjad Ahmad"

214 Publications

Immuno-Informatics Analysis of Pakistan-Based HCV Subtype-3a for Chimeric Polypeptide Vaccine Design.

Vaccines (Basel) 2021 Mar 21;9(3). Epub 2021 Mar 21.

King Edward Medical University, Lahore 54000, Pakistan.

Hepatitis C virus (HCV) causes chronic and acute hepatitis infections. As there is extreme variability in the HCV genome, no approved HCV vaccine has been available so far. An effective polypeptide vaccine based on the functionally conserved epitopes will be greatly helpful in curing disease. For this purpose, an immuno-informatics study is performed based on the published HCV subtype-3a from Pakistan. First, the virus genome was translated to a polyprotein followed by a subsequent prediction of T-cell epitopes. Non-allergenic, IFN-γ producer, and antigenic epitopes were shortlisted, including 5 HTL epitopes and 4 CTL, which were linked to the final vaccine by GPGPG and AAY linkers, respectively. Beta defensin was included as an adjuvant through the EAAAK linker to improve the immunogenicity of the polypeptide. To ensure its safety and immunogenicity profile, antigenicity, allergenicity, and various physiochemical attributes of the polypeptide were evaluated. Molecular docking was conducted between TLR4 and vaccine to evaluate the binding affinity and molecular interactions. For stability assessment and binding of the vaccine-TLR4 docked complex, molecular dynamics (MD) simulation and MMGBSA binding free-energy analyses were conducted. Finally, the candidate vaccine was cloned in silico to ensure its effectiveness. The current vaccine requires future experimental confirmation to validate its effectiveness. The vaccine construct produced might be useful in providing immune protection against HCV-related infections.
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http://dx.doi.org/10.3390/vaccines9030293DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8004085PMC
March 2021

Structure-Based Virtual Screening Identifies Multiple Stable Binding Sites at the RecA Domains of SARS-CoV-2 Helicase Enzyme.

Molecules 2021 Mar 7;26(5). Epub 2021 Mar 7.

Department of Biology, College of Science, United Arab Emirates University, Al Ain 15551, United Arab Emirates.

With the emergence and global spread of the COVID-19 pandemic, the scientific community worldwide has focused on search for new therapeutic strategies against this disease. One such critical approach is targeting proteins such as helicases that regulate most of the SARS-CoV-2 RNA metabolism. The purpose of the current study was to predict a library of phytochemicals derived from diverse plant families with high binding affinity to SARS-CoV-2 helicase (Nsp13) enzyme. High throughput virtual screening of the Medicinal Plant Database for Drug Design (MPD3) database was performed on SARS-CoV-2 helicase using AutoDock Vina. Nilotinib, with a docking value of -9.6 kcal/mol, was chosen as a reference molecule. A compound (PubChem CID: 110143421, ZINC database ID: ZINC257223845, eMolecules: 43290531) was screened as the best binder (binding energy of -10.2 kcal/mol on average) to the enzyme by using repeated docking runs in the screening process. On inspection, the compound was disclosed to show different binding sites of the triangular pockets collectively formed by Rec1A, Rec2A, and 1B domains and a stalk domain at the base. The molecule is often bound to the ATP binding site (referred to as binding site 2) of the helicase enzyme. The compound was further discovered to fulfill drug-likeness and lead-likeness criteria, have good physicochemical and pharmacokinetics properties, and to be non-toxic. Molecular dynamic simulation analysis of the control/lead compound complexes demonstrated the formation of stable complexes with good intermolecular binding affinity. Lastly, affirmation of the docking simulation studies was accomplished by estimating the binding free energy by MMPB/GBSA technique. Taken together, these findings present further in silco investigation of plant-derived lead compounds to effectively address COVID-19.
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http://dx.doi.org/10.3390/molecules26051446DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7962107PMC
March 2021

Synthesis of Michael Adducts as Key Building Blocks for Potential Analgesic Drugs: In vitro, in vivo and in silico Explorations.

Drug Des Devel Ther 2021 23;15:1299-1313. Epub 2021 Mar 23.

Department of Chemistry, COMSATS University Islamabad, Abbottabad Campus, Abbottabad, 22060, Pakistan.

Background: Organocatalytic asymmetric Michael addition is a strong approach for C-C bond formation. The objective of the study is to design molecules by exploiting the efficiency of Michael Adducts. We proceeded with the synthesis of Michael adducts by tailoring the substitution pattern on maleimide and trans-β-nitro styrene as Michael acceptors. The synthesized compounds were evaluated for dual cyclooxygenases (COX) and lipoxygenase (LOX) inhibition.

Methods: The compounds (4, 9-11) were synthesized through Michael additions. The cyclooxygenases (COX-1 and 2) and lipoxygenase (5-LOX) assays were used for in vitro evaluations of compounds. After the acute toxicity studies, the in vivo analgesic potential was determined with acetic acid induced writhing, tail immersion, and formalin tests. Furthermore, the possible roles of adrenergic and dopaminergic receptors were also studied. Extensive computational studies were performed to get a better understanding regarding the binding of this compound with protein target.

Results: Four Michael adducts (4, 9-11) were synthesized. Compound 4 was obtained in enantio- and diastereopure form. The stereopure compound 4 showed encouraging COX-1 and-2 inhibitions with IC values of 128 and 65 μM with SI of 1.94. Benzyl derivative 11 showed excellent COX-2 inhibition with the IC value of 5.79 μM and SI value 7.96. Compounds 4 and 11 showed good results in in vivo models of analgesia like acetic acid test, tail immersion, and formalin tests. Our compounds were not active in dopaminergic and adrenergic pathways and so were acting centrally. Through extensive computational studies, we computed binding energies, and pharmacokinetic predictions.

Conclusion: Our findings conclude that our synthesized Michael products (pyrrolidinedione 4 and nitroalkane 11) can be potent centrally acting analgesics. Our in silico predictions suggested that the compounds have excellent pharmacokinetic properties. It is concluded here that dual inhibition of COX/LOX pathways provides a convincing step towards the discovery of safe lead analgesic molecules.
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http://dx.doi.org/10.2147/DDDT.S292826DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8001115PMC
March 2021

Pituitary infundibular epidermoid cyst: a rare cause of hypopituitarism.

BMJ Case Rep 2021 Mar 24;14(3). Epub 2021 Mar 24.

Department of Endocrinology, University of Wales College of Medicine, Cardiff, UK.

A 53-year-old man presented with 6 months history of weight loss associated with nausea, fatigue, dizziness and headache. On arrival he was in adrenal crisis. Biochemistry revealed anterior hypopituitarism with low cortisol, thyroxine, testosterone and a slightly raised prolactin. He was commenced on steroids, thyroxine and testosterone. MRI pituitary gland was reported to have a 9.4 mm microadenoma. Cabergoline was started for a possible microprolactinoma. Follow-up MRI showed increase in the size of complex cystic lesion causing chiasmal compression raising a possibility of craniopharyngioma. Visual fields assessment was normal. In view of the rapid enlargement, to protect vision and obtain a tissue diagnosis he underwent endoscopic trans-sphenoidal surgery. A cystic lesion was noted intraoperatively originating from pituitary stalk with intrasellar and suprasellar extension. It was filled with white caseous material and fluid. Histology revealed epidermoid cyst. His headache resolved postoperatively.
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http://dx.doi.org/10.1136/bcr-2020-241065DOI Listing
March 2021

Self-reported knowledge, attitudes, and practice of final year dental students in relation to child abuse: a multi-centre study.

Int J Paediatr Dent 2021 Mar 17. Epub 2021 Mar 17.

Discipline Lead Paediatric Dentistry, School of Dentistry, The University of Queensland, Australia.

Background: The number of child abuse cases is increasing worldwide therefore it is important to educate individuals having contact with children about it. This includes dentists who play a pivotal role in detecting and reporting these cases.

Aim: To identify and compare the final year dental student's knowledge, attitudes and practice in relation to child abuse.

Design: A 38-item and four part online questionnaire was distributed to students of 11 dental schools in 10 countries. SPSS and GraphPad Prism were used for data analysis. The levels of statistical significance were determined using a chi-square test. P ≤ 0.05 was considered to be statistically significant.

Results: A total of 660 students completed the survey. Fifty-six percent of the students received formal training on child abuse, and 86% wanted additional training. The knowledge of child abuse was significantly higher in Australia, USA and Jordan compared to other countries. The internet (60.3%) was widely used as an information source for child abuse.

Conclusions: The study showed that dental students lack knowledge and experience in recognising and reporting child abuse. Most respondents indicated a desire for additional training, therefore dental schools should review what they are currently teaching and make changes as appropriate.
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http://dx.doi.org/10.1111/ipd.12781DOI Listing
March 2021

In Silico Validation, Fabrication and Evaluation of Nano-Liposomes of Bistorta amplexicaulis Extract for Improved Anticancer Activity Against Hepatoma Cell Line (HepG2).

Curr Drug Deliv 2021 Mar 16. Epub 2021 Mar 16.

Department of Biochemistry Pir Mehr Ali Shah University of Arid Agriculture Rawalpindi, 46300. Pakistan.

Background: Bistorta amplexicaulis of the genus Polygonum (Polygonaceae) has been reported for its antioxidant and anticancer activities. However, the low cellular uptake of the compounds in its extract limit their therapeutic application.

Objectives: The present study was aimed at developing a nanoliposomal carrier system for B. amplexicaulis extracts for its improved cellular uptake thus resulting in enhanced anticancer activity.

Methods: Ultra Pressure Liquid Chromatography (UPLC) was used to identify major compounds in the plant extract. Nanoliposomes (NLs) were prepared using a thin-film rehydration method using DPPC, PEG2000DSPE and cholesterol, followed by characterization through several parameters. In vitro screening was performed against breast cancer cell line (MCF-7) and Hepatocellular carcinoma cell line (HepG-2) using MTT-assay. Raw extract and nanoliposomes were tested on Human Umbilical Vein Endothelial Cells (HUVEC). Moreover, molecular docking was performed to validate the data obtained through wet lab.

Results: The UHPLC method identified gallic acid, caffeic acid, chlorogenic acid and catechin as the major compounds in the extract. The NLs with size ranging between 140-155 nm, zeta potential -16.9 to -19.8 mV and good polydispersity index of < 0.1 were prepared, with a high encapsulation efficiency of 81%. The MTT assay showed significantly (p > 0.05) high uptake and cytotoxicity of NLs as compared to the plant extract. Moreover, a reduced toxicity against HUVEC cells was observed as compared to the extract. Also, the docking of identified compounds suggested a favorable interaction to the SH2 domain of both STAT3 and STAT5.

Conclusion: Overall, the results suggested NLs as a potential platform that could be developed for the improved intracellular delivery of plant extract thus increasing their therapeutic outcomes.
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http://dx.doi.org/10.2174/1567201818666210316113640DOI Listing
March 2021

Low uptake of COVID-19 prevention behaviours and high socioeconomic impact of lockdown measures in South Asia: Evidence from a large-scale multi-country surveillance programme.

SSM Popul Health 2021 Mar 13;13:100751. Epub 2021 Feb 13.

Lee Kong Chian School of Medicine, Nanyang Technological University, Singapore.

Background: South Asia has become a major epicentre of the COVID-19 pandemic. Understanding South Asians' awareness, attitudes and experiences of early measures for the prevention of COVID-19 is key to improving the effectiveness and mitigating the social and economic impacts of pandemic responses at a critical time for the Region.

Methods: We assessed the knowledge, behaviours, health and socio-economic circumstances of 29,809 adult men and women, at 93 locations across four South Asian countries. Data were collected during the national lockdowns implemented from March to July 2020, and compared with data collected prior to the pandemic as part of an ongoing prospective surveillance initiative.

Results: Participants were 61% female, mean age 45.1 years. Almost half had one or more chronic disease, including diabetes (16%), hypertension (23%) or obesity (16%). Knowledge of the primary COVID-19 symptoms and transmission routes was high, but access to hygiene and personal protection resources was low (running water 63%, hand sanitisers 53%, paper tissues 48%). Key preventive measures were not widely adopted. Knowledge, access to, and uptake of COVID-19 prevention measures were low amongst people from disadvantaged socio-economic groups. Fifteen percent of people receiving treatment for chronic diseases reported loss of access to long-term medications; 40% reported symptoms suggestive of anxiety or depression. The prevalence of unemployment rose from 9.3% to 39.4% (P < 0.001), and household income fell by 52% (P < 0.001) during the lockdown. Younger people and those from less affluent socio-economic groups were most severely impacted. Sedentary time increased by 32% and inadequate fruit and vegetable intake increased by 10% (P < 0.001 for both), while tobacco and alcohol consumption dropped by 41% and 80%, respectively (P < 0.001), during the lockdown.

Conclusions: Our results identified important knowledge, access and uptake barriers to the prevention of COVID-19 in South Asia, and demonstrated major adverse impacts of the pandemic on chronic disease treatment, mental health, health-related behaviours, employment and household finances. We found important sociodemographic differences for impact, suggesting a widening of existing inequalities. Our findings underscore the need for immediate large-scale action to close gaps in knowledge and access to essential resources for prevention, along with measures to safeguard economic production and mitigate socio-economic impacts on the young and the poor.
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http://dx.doi.org/10.1016/j.ssmph.2021.100751DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7902538PMC
March 2021

Biomaterials for corneal endothelial cell culture and tissue engineering.

J Tissue Eng 2021 Jan-Dec;12:2041731421990536. Epub 2021 Feb 16.

Department of Eye and Vision Science, Institute of Ageing and Chronic Disease, University of Liverpool, Liverpool, UK.

The corneal endothelium is the posterior monolayer of cells that are responsible for maintaining overall transparency of the avascular corneal tissue via pump function. These cells are non-regenerative in vivo and therefore, approximately 40% of corneal transplants undertaken worldwide are a result of damage or dysfunction of endothelial cells. The number of available corneal donor tissues is limited worldwide, hence, cultivation of human corneal endothelial cells (hCECs) in vitro has been attempted in order to produce tissue engineered corneal endothelial grafts. Researchers have attempted to recreate the current gold standard treatment of replacing the endothelial layer with accompanying Descemet's membrane or a small portion of stroma as support with tissue engineering strategies using various substrates of both biologically derived and synthetic origin. Here we review the potential biomaterials that are currently in development to support the transplantation of a cultured monolayer of hCECs.
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http://dx.doi.org/10.1177/2041731421990536DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7894589PMC
February 2021

Rational design of potent anti-COVID-19 main protease drugs: An extensive multi-spectrum in silico approach.

J Mol Liq 2021 May 12;330:115636. Epub 2021 Feb 12.

Foundation University Medical College, Foundation University Islamabad, Islambad, Pakistan.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) as a novel coronavirus and the etiological agent of global pandemic coronavirus disease (COVID-19) requires quick development of potential therapeutic strategies. Computer aided drug design approaches are highly efficient in identifying promising drug candidates among an available pool of biological active antivirals with safe pharmacokinetics. The main protease (M) enzyme of SARS-CoV-2 is considered key in virus production and its crystal structures are available at excellent resolution. This marks the enzyme as a good starting receptor to conduct an extensive structure-based virtual screening (SBVS) of ASINEX antiviral library for the purpose of uncovering valuable hits against SARS-CoV-2 M. A compound hit (BBB_26580140) was stand out in the screening process, as opposed to the control, as a potential inhibitor of SARS-CoV-2 M based on a combined approach of SBVS, drug likeness and lead likeness annotations, pharmacokinetics, molecular dynamics (MD) simulations, and end point MM-PBSA binding free energy methods. The lead was further used in ligand-based similarity search (LBSS) that found 33 similar compounds from the ChEMBL database. A set of three compounds (SCHEMBL12616233, SCHEMBL18616095, and SCHEMBL20148701), based on their binding affinity for M, was selected and analyzed using extensive MD simulation, hydrogen bond profiling, MM-PBSA, and WaterSwap binding free energy techniques. The compounds conformation with M show good stability after initial within active cavity moves, a rich intermolecular network of chemical interactions, and reliable relative and absolute binding free energies. Findings of the study suggested the use of BBB_26580140 lead and its similar analogs to be explored in vivo which might pave the path for rational drug discovery against SARS-CoV-2 M.
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http://dx.doi.org/10.1016/j.molliq.2021.115636DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7879066PMC
May 2021

Cross-coupling reactions towards the synthesis of natural products.

Mol Divers 2021 Feb 20. Epub 2021 Feb 20.

Department of Chemistry, University of Management and Technology, Lahore, Pakistan.

Cross-coupling reactions are powerful synthetic tools for the formation of remarkable building blocks of many naturally occurring molecules, polymers and biologically active compounds. These reactions have brought potent transformations in chemical and pharmaceutical disciplines. In this review, we have focused on the use of cross-coupling reactions such as Suzuki, Negishi, Heck, Sonogashira and Stille in the total synthesis of some natural products of recent years (2016-2020). A short introduction of mentioned cross-coupling reactions along with highlighted aspects of natural products has been stated in separate sections. Additionally, few examples of natural products via incorporation of more than one type of cross-coupling reaction have also been added to demonstrate the importance of these reactions in organic synthesis.
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http://dx.doi.org/10.1007/s11030-021-10195-6DOI Listing
February 2021

SARS-CoV-2: An Update on Genomics, Risk Assessment, Potential Therapeutics and Vaccine Development.

Int J Environ Res Public Health 2021 02 8;18(4). Epub 2021 Feb 8.

College of Life Science and Technology, Guangxi University, Nanning 530004, China.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a great threat to public health, being a causative pathogen of a deadly coronavirus disease (COVID-19). It has spread to more than 200 countries and infected millions of individuals globally. Although SARS-CoV-2 has structural/genomic similarities with the previously reported SARS-CoV and MERS-CoV, the specific mutations in its genome make it a novel virus. Available therapeutic strategies failed to control this virus. Despite strict standard operating procedures (SOPs), SARS-CoV-2 has spread globally and it is mutating gradually as well. Diligent efforts, special care, and awareness are needed to reduce transmission among susceptible masses particularly elder people, children, and health care workers. In this review, we highlighted the basic genome organization and structure of SARS-CoV-2. Its transmission dynamics, symptoms, and associated risk factors are discussed. This review also presents the latest mutations identified in its genome, the potential therapeutic options being used, and a brief explanation of vaccine development efforts against COVID-19. The effort will not only help readers to understand the deadly SARS-CoV-2 virus but also provide updated information to researchers for their research work.
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http://dx.doi.org/10.3390/ijerph18041626DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7915969PMC
February 2021

Synthesis, characterization and advanced sustainable applications of titanium dioxide nanoparticles: A review.

Ecotoxicol Environ Saf 2021 Apr 6;212:111978. Epub 2021 Feb 6.

Universiti Teknologi, Malaysia.

Nanotechnology is capturing great interest worldwide due to their stirring applications in various fields. Among nanoparticles (NPs), titanium dioxide (TiO) NPs have been widely used in daily life and can be synthesized through various physical, chemical, and green methods. Green synthesis is a non-toxic, cost-effective, and eco-friendly route for the synthesis of NPs. Plenty of work has been reported on the green, chemical, physical and biological synthesis of TiO NPs and these NPs can be characterized through high tech. instruments. In the present review, dense data have been presented on the comparative synthesis of TiO NPs with different characteristics and their wide range of applications. Among the TiO NPs synthesis techniques, the green methods have been proven to be efficient than chemical synthesis methods because of the less use of precursors, time-effectiveness, and energy-efficiency during the green synthesis procedures. Moreover, this review describes the types of plants (shrubs, herbs and trees), microorganisms (bacteria, fungi and algae), biological derivatives (proteins, peptides, and starches) employed for the synthesis of TiO NPs. The TiO NPs can be effectively used for the treatment of polluted water and positively affected the plant physiology especially under abiotic stresses but the response varied with types, size, shapes, doses, duration of exposure, metal species along with other factors. This review also highlights the regulating features and future standpoints for the measurable enrichment in TiO NPs product and perspectives of TiO NPs reliable application.
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http://dx.doi.org/10.1016/j.ecoenv.2021.111978DOI Listing
April 2021

Human leukocyte antigen B*0702 is protective against ocular Stevens-Johnson syndrome/toxic epidermal necrolysis in the UK population.

Sci Rep 2021 Feb 3;11(1):2928. Epub 2021 Feb 3.

Academic Unit of Ophthalmology, Birmingham and Midland Eye Centre, Institute of Inflammation and Ageing, College of Medical and Dental Sciences, University of Birmingham, Dudley Road, Birmingham, B18 7QU, UK.

Stevens-Johnson Syndrome and Toxic Epidermal Necrolysis (SJS/TEN) are part of a disease continuum of vesiculobullous mucocutaneous reactions affecting the skin and mucous membranes including the ocular surface. Manifestations of disease range from mild dry eye to progressive conjunctival cicatrisation, limbal epithelial stem cell failure and corneal blindness. In Far Eastern and South East Asian populations where SJS/TEN is prevalent, numerous human leukocyte antigen (HLA) gene variants at the A, B and C loci have been identified as risk factors for developing SJS/TEN with severe ocular complications (SOC). By contrast, the incidence of SJS/TEN with SOC in European countries is relatively low. To date, ocular SJS/TEN risk altering alleles have not been widely investigated in European populations. In this study, we analysed the association of HLA -A, -B and -C alleles with SJS/TEN in 33 patients residing in the UK with age matched controls. The data showed statistically significant novel negative allele association with HLA-B*0702 and a trend with HLA-C*0702 in the patient group, indicating these alleles are protective. Further characterisation of protective and risk alleles in other ethnic groups is required to fully elucidate the putative role of these alleles in the susceptibility of SJS/TEN with or without severe ocular complications in patients in the UK.
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http://dx.doi.org/10.1038/s41598-021-82400-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7859395PMC
February 2021

Computational Determination of Potential Multiprotein Targeting Natural Compounds for Rational Drug Design Against SARS-COV-2.

Molecules 2021 Jan 28;26(3). Epub 2021 Jan 28.

Key Laboratory of Ministry of Education for Medicinal Plant Resource and Natural Pharmaceutical Chemistry, Shaanxi Normal University, Xi'an 710062, China.

SARS-CoV-2 caused the current COVID-19 pandemic and there is an urgent need to explore effective therapeutics that can inhibit enzymes that are imperative in virus reproduction. To this end, we computationally investigated the MPD3 phytochemical database along with the pool of reported natural antiviral compounds with potential to be used as anti-SARS-CoV-2. The docking results demonstrated glycyrrhizin followed by azadirachtanin, mycophenolic acid, kushenol-w and 6-azauridine, as potential candidates. Glycyrrhizin depicted very stable binding mode to the active pocket of the Mpro (binding energy, -8.7 kcal/mol), PLpro (binding energy, -7.9 kcal/mol), and Nucleocapsid (binding energy, -7.9 kcal/mol) enzymes. This compound showed binding with several key residues that are critical to natural substrate binding and functionality to all the receptors. To test docking prediction, the compound with each receptor was subjected to molecular dynamics simulation to characterize the molecule stability and decipher its possible mechanism of binding. Each complex concludes that the receptor dynamics are stable (Mpro (mean RMSD, 0.93 Å), PLpro (mean RMSD, 0.96 Å), and Nucleocapsid (mean RMSD, 3.48 Å)). Moreover, binding free energy analyses such as MMGB/PBSA and WaterSwap were run over selected trajectory snapshots to affirm intermolecular affinity in the complexes. Glycyrrhizin was rescored to form strong affinity complexes with the virus enzymes: Mpro (MMGBSA, -24.42 kcal/mol and MMPBSA, -10.80 kcal/mol), PLpro (MMGBSA, -48.69 kcal/mol and MMPBSA, -38.17 kcal/mol) and Nucleocapsid (MMGBSA, -30.05 kcal/mol and MMPBSA, -25.95 kcal/mol), were dominated mainly by vigorous van der Waals energy. Further affirmation was achieved by WaterSwap absolute binding free energy that concluded all the complexes in good equilibrium and stability (Mpro (mean, -22.44 kcal/mol), PLpro (mean, -25.46 kcal/mol), and Nucleocapsid (mean, -23.30 kcal/mol)). These promising findings substantially advance our understanding of how natural compounds could be shaped to counter SARS-CoV-2 infection.
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http://dx.doi.org/10.3390/molecules26030674DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7865386PMC
January 2021

Proteome wide vaccine targets prioritization and designing of antigenic vaccine candidate to trigger the host immune response against the Mycoplasma genitalium infection.

Microb Pathog 2021 Mar 29;152:104771. Epub 2021 Jan 29.

Department of Bioinformatics and Biological Statistics, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, PR China; State Key Laboratory of Microbial Metabolism, Shanghai-Islamabad-Belgrade Joint Innovation Centre on Antibacterial Resistances, Joint International Research Laboratory of Metabolic & Developmental Sciences and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200030, PR China; Peng Cheng Laboratory, Vanke Cloud City Phase I Building 8, Xili Street, Nashan District, Shenzhen, Guangdong, 518055, PR China. Electronic address:

Mycoplasma genitalium is a small size, sexually transmitted bacterial pathogen that causes urethritis in males and cervicitis in females. Being resistant to antibiotics, difficulty in diagnosis, treatment, and control of this cosmopolitan infection, vaccination is the alternating method for its effective management. Herein, this study was conducted to computationally design a multi-epitope vaccine to boost host immune responses against M. genitalium. To achieve the study aim, immunoinformatics approaches were applied to the said pathogen's proteomics sequence data. B and T cell epitopes were projected from the three shortlisted vaccine proteins; MG014, MG015, Hmw3MG317. The final vaccine ensemble comprises cytotoxic and helper T cell epitopes fused through appropriate linkers. The epitopes peptide is then liked to an adjuvant for efficient recognition and processing by the host immune system. The various physicochemical parameters such as allergenicity, antigenicity, theoretical pI, GRAVY, and molecular weight of the vaccine were checked and found safe and effective to be used in post-experimental studies. The stability and binding affinity of the vaccine with the TLR1/2 heterodimer were ensured by performing molecular docking. The best-docked complex was considered, ranked top having the lowest binding energy and strong intermolecular binding and stability. Finally, the vaccine constructs better expression was obtained by in silico cloning into the pET28a (+) vector in Escherichia coli K-12 strain, and immune simulation validated the immune response. In a nutshell, all these approaches lead to developing a multi-epitope vaccine that possessed the ability to induce cellular and antibody-mediated immune responses against the pathogen used.
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http://dx.doi.org/10.1016/j.micpath.2021.104771DOI Listing
March 2021

species; a potential source of anti-SARS-CoV-2 main protease inhibitors.

J Biomol Struct Dyn 2021 Jan 15:1-11. Epub 2021 Jan 15.

NUMS Department of Biological Sciences, National University of Medical Sciences, Rawalpindi, Pakistan.

The COVID-19 being a preconized global pandemic by the World Health Organization needs persuasive immediate research for possible medications. The present study was carried out with a specific aim to computationally evaluate and identify compounds derived from species as the plausible inhibitors against 3-chymotrypsin-like main protease (3CLpro) or main protease (M), which is a key enzyme in the life-cycle of coronavirus. The compounds were isolated from the crude extracts of species. Among the isolated compounds, novel inhibitory leads were identified using techniques. Molecular docking revealed that stigmasterol (-8.3 kcal/mol), chondrillasterol (-7.9 kcal/mol) and hexadecnoic acid (-6.9 kcal/mol)) among others bind in the substrate-binding pocket and also interacted with the catalytic dyad of the 3-CLpro. Further evaluation using 50 ns molecular dynamic simulation and MMPB-GBSA indicated that among the top three docking hits, hexadecanoic acid was found to be the most promising anti-COVID-19 lead against the main protease. Hexadecanoic acid might serve as a potent anti-SARS-CoV-2 compound to combat COVID-19, however, and validation and optimization is needed. Communicated by Ramaswamy H. Sarma.
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http://dx.doi.org/10.1080/07391102.2021.1873188DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7814571PMC
January 2021

Spinal epidural lipomatosis: a rare association of Cushing's disease.

Endocrinol Diabetes Metab Case Rep 2020 Oct 19;2020. Epub 2020 Oct 19.

GIM/Diabetes & Endocrinology, University Hospital of Wales, Cardiff, UK.

Summary: Excess cortisol is associated with hypertrophy and redistribution of adipose tissue leading to central obesity which is classically seen in Cushing's syndrome. Abnormal accumulation of fatty tissue in the spinal canal is most commonly associated with chronic steroid therapy and rarely reported with endogenous Cushing's syndrome. Herein, we describe a case of spinal epidural lipomatosis (SEL) associated with Cushing's disease. A 17-year-old man was referred with lower limb weakness, weight gain, multiple stretch marks, back pain and loss of height. He had clinical and biochemical features of Cushing's syndrome. MRI and Inferior Petrosal Sinus Sampling (IPSS) confirmed a pituitary adenoma as the source. On day 1 post trans-sphenoidal adenectomy he developed spastic paraparesis with a sensory deficit to the level of T5. MRI spine showed increased fat deposition in the spinal canal from T2 to T9 consistent with a diagnosis of SEL. He was managed conservatively and made a good recovery following restoration of eucortisolism and a period of rehabilitation.

Learning Points: SEL is a serious complication of glucocorticoid excess and should be considered in any patient presenting with new lower limb neurological symptoms associated with hypercortisolism. It is important to distinguish symptomatic SEL from cortisol-induced proximal myopathy by good history and clinical examination. MRI of the spine is the gold standard investigation for making a diagnosis of SEL. Restoration of eucortisolism can lead to resolution of fat accumulation and good neurological outcome.
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http://dx.doi.org/10.1530/EDM-20-0111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7576635PMC
October 2020

Risk assessment of potentially toxic metal(loid)s in Vigna radiata L. under wastewater and freshwater irrigation.

Chemosphere 2021 Feb 27;265:129124. Epub 2020 Nov 27.

Department of Environmental Sciences, COMSATS University Islamabad, Vehari Campus, 61100, Pakistan.

Depending on the use and management, wastewater generation can be a severe environmental dilemma or a potential source. Proper application and management of municipal water (MW) in agriculture could be its sustainable use. Until now, there is rare data about the combined application of wastewater and freshwater in agriculture that could be considered as sustainable water management strategy. Also, plant (oxidative) stress responses to wastewater application have been rarely investigated. Here, we elucidated the influence of MW irrigation, diluted with canal water (CW) and groundwater (tap water; TW), on Vigna radiata to evaluate (i) the accumulation of potentially toxic metal(loid)s (PTMs; arsenic, copper, cadmium, iron, manganese, lead, nickel, zinc) in different plant tissues, (ii) biochemical modifications in plants, (iii) relative compartmentation of PTMs inside plant, and (iv) PTMs-induced health risk. Results revealed that the soil-plant transfer of PTMs and physiological changes in V. radiata varied depending on the irrigation water type. Noticeably, plants sequestered most of the PTM contents in roots (average 64%) and less were translocated to plant shoots. All the irrigation treatments provoked oxidative stress in V. radiata with high production of hydrogen peroxide, followed by an oxidation of membrane lipids and a decrease in chlorophyll content. The estimated cancer risk and hazard quotients values revealed a potential risk to human health (HQ: 2.2-108.8, CR: 0.0002-0.664), especially for arsenic, cadmium and lead. The integrated risk estimated from PTMs highlighted the unsuitability of all the treatments for crop irrigation. Therefore, in areas with high PTM levels in MW and freshwaters their mixed use is not an ideal management practice. Conclusively, this study helps to strictly monitor the quality of irrigation water before applying to crops and develop a suitable management and remediation strategy.
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http://dx.doi.org/10.1016/j.chemosphere.2020.129124DOI Listing
February 2021

Vaginal Microbiome-Based Bacterial Signatures for Predicting the Severity of Cervical Intraepithelial Neoplasia.

Diagnostics (Basel) 2020 Nov 26;10(12). Epub 2020 Nov 26.

Department of Obstetrics and Gynecology, School of Medicine, Kyungpook National University, Daegu 41404, Korea.

Although emerging evidence revealed that the gut microbiome served as a tool and as biomarkers for predicting and detecting specific cancer or illness, it is yet unknown if vaginal microbiome-derived bacterial markers can be used as a predictive model to predict the severity of CIN. In this study, we sequenced V3 region of 16S rRNA gene on vaginal swab samples from 66 participants (24 CIN 1-, 42 CIN 2+ patients) and investigated the taxonomic composition. The vaginal microbial diversity was not significantly different between the CIN 1- and CIN 2+ groups. However, we observed dominant type (16.7%), which does not belong to conventional community state type (CST). Moreover, a minimal set of 33 bacterial species was identified to maximally differentiate CIN 2+ from CIN 1- in a random forest model, which can distinguish CIN 2+ from CIN 1- (area under the curve (AUC) = 0.952). Among the 33 bacterial species, was selected as the most impactful predictor in our model. This finding suggests that the random forest model is able to predict the severity of CIN and vaginal microbiome may play a role as biomarker.
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http://dx.doi.org/10.3390/diagnostics10121013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7761147PMC
November 2020

Cd induced biphasic response in soil alkaline phosphatase and changed soil bacterial community composition: The role of background Cd contamination and time as additional factors.

Sci Total Environ 2021 Feb 16;757:143771. Epub 2020 Nov 16.

College of Biological and Environment, Nanjing Forestry University, Nanjing, Jiangsu, China; Co-Innovation Center for the Sustainable Forestry in Southern Jiangsu Province, Nanjing, Jiangsu, China; National Positioning Observation Station of Hung-tse Lake Wetland Ecosystem in Jiangsu Province, Nanjing, Jiangsu, China. Electronic address:

Hormesis is an intriguing phenomenon characterized by low-dose stimulation and high-dose inhibition. The hormetic phenomena have been frequently reported in the past decades, but the researches on the biphasic responses of soil enzymes are still limited. The main objective of this study is to explore dose response of alkaline phosphatase (ALP) to Cd (0, 0.003, 0.03, 0.3, 3.0 and 30 mg/kg) in the presence of different levels of background Cd contamination (bulk soil with no added Cd, BS; low background Cd, LB; medium background Cd, MB; and high background Cd, HB). ALP activity at 0.003-0.3 mg Cd/kg was 13-39% higher than that of the control (0 mg Cd/kg) for HB after 7 d. Similarly, the enzyme activities at 0.003-0.03 mg Cd/kg were 2-25% and 14-17% higher than those of the controls for MB and HB after 60 d. After 90 d, ALP activities at 0.3-3.0 mg Cd/kg increased by 11-17% for LB. The dose-response curves had the shape of an inverted U, showing biphasic responses at days 7 (HB), 60 (MB and HB) and 90 (LB). After 60 days of exposure, total operational taxonomic units (OTU) numbers and unique species exposed to Cd stress displayed hormetic-response curve for MB. The relative abundances of Agrobacterium, Salinimicrobiums, Bacilllus, and Oceanobacillus displayed significantly positive correlations with ALP activity. This suggested that bacterial communities potentially contribute to ALP's hormesis. This study further provides new insights into the ecological mechanisms of pollutant-induced hormesis, and substantially contributes to the ecological risk assessment of Cd pollution.
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http://dx.doi.org/10.1016/j.scitotenv.2020.143771DOI Listing
February 2021

A computational study to disclose potential drugs and vaccine ensemble for COVID-19 conundrum.

J Mol Liq 2021 Feb 10;324:114734. Epub 2020 Nov 10.

Foundation University Medical College, Foundation University Islamabad, DHA-I, Islamabad 44000, Pakistan.

The nucleocapsid (N) protein of SARS-COV-2, a virus responsible for the current COVID-19 pandemic, is considered a potential candidate for the design of new drugs and vaccines. The protein is central to several critical events in virus production, with its highly druggable nature and rich antigenic determinants making it an excellent anti-viral biomolecule. Docking-based virtual screening using the Asinex anti-viral library identified binding of drug molecules at three specific positions: loop 1 region, loop 2 region and β-sheet core pockets, the loop 2 region being the most common binding and stable site for the bulk of the molecules. In parallel, the protein was characterized by vaccine design perspective and harboured three potential B cell-derived T cell epitopes: PINTNSSPD, GVPINTNSS, and DHIGTRNPA. The epitopes are highly antigenic, virulent, non-allergic, non-toxic, bind with good affinity to the highly prevalent DRB*0101 allele and show an average population coverage of 95.04%. A multi-epitope vaccine ensemble which was 83 amino acids long was created. This was highly immunogenic, robust in generating both humoral and cellular immune responses, thermally stable, and had good physicochemical properties that could be easily analyzed in in vivo and in vitro studies. Conformational dynamics of both drug and vaccine ensemble with respect to the receptors are energetically stable, shedding light on favourable conformation and chemical interactions. These facts were validated by subjecting the complexes to relative and absolute binding free energy methods of MMGB/PBSA and WaterSwap. A strong agreement on the system stability was disclosed that supported ligand high affinity potential for the receptors. Collectively, this work sought to provide preliminary experimental data of existing anti-viral drugs as a possible therapy for COVID-19 infections and a new peptide-based vaccine for protection against this pandemic virus.
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http://dx.doi.org/10.1016/j.molliq.2020.114734DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7654302PMC
February 2021

Rational Drug Design for PqsA Enzyme: An Guided Study to Block Biofilm Formation.

Front Mol Biosci 2020 15;7:577316. Epub 2020 Oct 15.

84 Heukseok-ro, Dongjak-gu, Department of Biomedical Engineering, Chung-Ang University, Seoul, South Korea.

is an opportunistic gram-negative bacterium implicated in acute and chronic nosocomial infections and a leading cause of patient mortality. Such infections occur owing to biofilm formation that confers multidrug resistance and enhanced pathogenesis to the bacterium. In this study, we used a rational drug design strategy to inhibit the quorum signaling system of by designing potent inhibitory lead molecules against anthranilate-CoA ligase enzyme encoded by the gene. This enzyme produces autoinducers for cell-to-cell communication, which result in biofilm formation, and thus plays a pivotal role in the virulence of . A library of potential drug molecules was prepared by performing ligand-based screening using an available set of enzyme inhibitors. Subsequently, structure-based virtual screening was performed to identify compounds showing the best binding conformation with the target enzyme and forming a stable complex. The two hit compounds interact with the binding site of the enzyme through multiple short-range hydrophilic and hydrophobic interactions. Molecular dynamic simulation and MM-PBSA/GBSA results to calculate the affinity and stability of the hit compounds with the PqsA enzyme further confirmed their strong interactions. The hit compounds might be useful in tackling the resistant phenotypes of this pathogen.
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http://dx.doi.org/10.3389/fmolb.2020.577316DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7593710PMC
October 2020

Towards a novel peptide vaccine for Middle East respiratory syndrome coronavirus and its possible use against pandemic COVID-19.

J Mol Liq 2021 Feb 6;324:114706. Epub 2020 Nov 6.

Department of Biological Sciences, National University of Medical Sciences, the Mall, Rawalpindi 46000, Pakistan.

Middle East respiratory syndrome coronavirus (MERS-CoV) is an emerging health concern due to its high mortality rate of 35%. At present, no vaccine is available to protect against MERS-CoV infections. Therefore, an in silico search for potential antigenic epitopes in the non-redundant proteome of MERS-CoV was performed herein. First, a subtractive proteome-based approach was employed to look for the surface exposed and host non-homologous proteins. Following, immunoinformatics analysis was performed to predict antigenic B and T cell epitopes that were used in the design of a multi-epitopes peptide. Molecular docking study was carried out to predict vaccine construct affinity of binding to Toll-like receptor 3 (TLR3) and understand its binding conformation to extract ideas about its processing by the host immune system. We identified membrane protein, envelope small membrane protein, non-structural protein ORF3, non-structural protein ORF5, and spike glycoprotein as potential candidates for subunit vaccine designing. The designed multi-epitope peptide then linked to β-defensin adjuvant is showing high antigenicity. Further, the sequence of the designed vaccine construct is optimized for maximum expression in the expression system. A rich pattern of hydrogen and hydrophobic interactions of the construct was observed with the TLR3 allowing stable binding of the construct at the docked site as predicted by the molecular dynamics simulation and MM-PBSA binding energies. We expect that the panel of subunit vaccine candidates and the designed vaccine construct could be highly effective in immunizing populations from infections caused by MERS-CoV and could possible applied on the current pandemic COVID-19.
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http://dx.doi.org/10.1016/j.molliq.2020.114706DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7644433PMC
February 2021

A combine approach of chemical synthesis, biological evaluation and structural dynamics studies revealed thiazole substituted arylamine derivatives as potent FabH enzyme inhibitors.

Bioorg Chem 2020 12 24;105:104426. Epub 2020 Oct 24.

Department of Chemistry, Quaid-i-Azam University, Islamabad 45320, Pakistan. Electronic address:

Bacterial FabH enzyme is a broad-spectrum antimicrobial target and can be used in the design of novel antibiotics. This study reports chemical synthesis of thiazole based amine compounds as FabH inhibitors, followed by biological evaluation, and computational drug designing analysis with ultimate objective to guide further biological optimization of the identified hits. The compounds were synthesized through Pd-PEPPSI catalyzed cross coupling strategy for the Buchwald-Hartwig amination of thiazole-substituted aryl bromide. Pd-PEPPSI pre-catalysts were utilized for the cross couple with the diverse range of functionalized electron-deficient and electron-rich anilines and aliphatic amines. The thiazole based heteroaryl bromide coupling was found to be challenging and only specialized Pd-PEPPSI-IPr and Pd-PEPPSI-IPent catalysts were found to be effective providing the coupling product yield in the range of 78% to 99%. Biological investigation depicted compound 3f to be effective against Bacillus subtilis, Staphylococcus aureus, Staphylococcus epidermis, and Escherichia coli with mean + standard deviation value of 9.6 ± 0.4, 11.6 ± 0.4, 15.6 ± 0.4, and 11.6 ± 0.4, respectively. This compound is also active against free radicals with EC value of 39.45 µg/ml. Comparative docking predictions unravel the 3f binding mode at FabH active tunnel as such to block complete access for the natural substrate and involved balanced hydrogen and hydrophobic interactions. FabH-3f complex dynamics in solution found the docked conformation between the protein and compound of higher stability with mean carbon alpha deviation of 1.87 Å and mean residual deviation of 0.88 Å. Intermolecular interactions analysis depicted Asn274 from FabH active pocket to be significant in compound holding and strengthening of interaction as the simulation progresses. This was supported further by radial distribution function (RDF) and axial frequency distribution (AFD) that demonstrated the high distribution of compound atoms in close proximity of Asn274 residue and decrease in interaction distance. Further, the docking and simulation findings were validated through MMPB/GBSA methods that complements the compound affinity for the said target. In a nutshell, the identified hit could be subjected to structure, biological and pharmacokinetic optimization for development of effective FabH inhibitors.
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http://dx.doi.org/10.1016/j.bioorg.2020.104426DOI Listing
December 2020

En-face analysis of the human limbal lymphatic vasculature.

Exp Eye Res 2020 12 28;201:108278. Epub 2020 Sep 28.

Department Ophthalmology, Medical University of Innsbruck, Austria. Electronic address:

Purpose: To describe the location and morphometric characteristics of the human limbal lymphatic vasculature and its relation to the marginal corneal vascular arcades (MCA).

Methods: Ex vivo confocal microscopic (CM) imaging and immunofluorescence double staining for CD-31 and D2-40 of histological en-face sections using 12 preserved human cadaveric corneoscleral discs were performed, followed by a semi-automated morphometric analysis of the two-dimensional vascular network architecture.

Results: Ex vivo CM confirmed the presence of 2 distinct vascular networks. The haematic limbal vascular complex (HLVC) extended further into the cornea, forming typical MCAs. The lymphatic limbal vascular complex (LLVC) was peripheral from the termination of Bowman's layer and was also found to be peripheral to and deeper than the HLVC. LLVC and HLVC were significantly different with respect to vessel diameter, segment length and wall thickness.

Conclusion: The lymphatic vasculature of the human corneoscleral limbal region displays specific morphometric features that allow its differentiation from haematic vessels using CM.
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http://dx.doi.org/10.1016/j.exer.2020.108278DOI Listing
December 2020

Preloaded Descemet Membrane Endothelial Keratoplasty Grafts With Endothelium Outward: A Cross-Country Validation Study of the DMEK Rapid Device.

Cornea 2020 Sep 16. Epub 2020 Sep 16.

International Center for Ocular Physiopathology, Fondazione Banca degli Occhi del Veneto Onlus, Venice, Italy.

Purpose: To validate the "Descemet membrane endothelial keratoplasty (DMEK) Rapid" device for the cross-country transportation of preloaded DMEK grafts preserved with endothelium outward.

Methods: DMEK grafts were stripped and loaded in the DMEK Rapid device with tissue culture medium (TCM) or transport medium (TM) with endothelium outward. The device was mounted in a 40-mL flask and preserved for 4 days on a rocker to simulate transportation (study A, n = 24) or shipped in the TM from Italy to the United Kingdom (study B, n = 9) and evaluated within 72 hours. All the tissues were stained with Alizarin red. Viability of the cells was checked postsimulations and posttransportation and was confirmed using live/dead staining. Expression of tight junction proteins was evaluated.

Results: In study A, the endothelial cell loss observed from the TCM group was 20.8% (±5.2) compared with 19.5% (±6.7) from the TM group (P = 0.41) after transport simulation. Alizarin red showed minimal uncovered areas in both groups. There were no statistical differences in viability between the TM (80.83%) and TCM groups (78.83%). In study B, 12.9% (±7.8) endothelial cell loss was observed after transporting the tissues from Italy to the United Kingdom with no significant difference between prestrip and posttransportation (P = 0.05). Alizarin red staining did not show any uncovered area. Live/dead analysis showed 85.16% cell viability after transportation. zonula occludens-1 (ZO-1) was expressed in all tissues.

Conclusions: The DMEK Rapid device is safe for preloading and shipping DMEK grafts internationally with endothelium outward within 72 hours when preserved in the transport media.
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http://dx.doi.org/10.1097/ICO.0000000000002493DOI Listing
September 2020

Synthetic media for preservation of corneal tissues deemed for endothelial keratoplasty and endothelial cell culture.

Acta Ophthalmol 2020 Sep 10. Epub 2020 Sep 10.

International Center for Ocular Physiopathology, Fondazione Banca degli Occhi del Veneto Onlus, Venice, Italy.

Purpose: To compare the difference between various endothelial graft preparation methods and endothelial cell culture from tissues that are preserved in serum-based and synthetic medium.

Methods: In a randomized masked study, the tissues (n = 64) were preserved in Cornea Max (serum-based) and Cornea Syn (synthetic) series for 36 days at their respective preservation conditions. Following organ culture, corneal tissues (n = 48) were used to prepareDescemet stripping automated endothelial keratoplasty (DSAEK), preloaded ultra-thin (UT) -DSAEK, prestripped Descemet membrane endothelial keratoplasty (DMEK), free-floating DMEK, and preloaded DMEK with endothelium inward and outward grafts. These tissues were preserved for another 4days at room temperature in dextran supplemented media following which they were subjected to trypan blue, alizarin red, live/dead and Zonula Occludens-1 (ZO-1) staining. A separate set of tissues (n = 16) from both the series was used for human corneal endothelial cell (HCEnC) culture. At confluence, the proliferation and cell doubling rate was calculated and the cultured cells were subjected to live/dead, ZO-1, 2A12 and Ki-67 staining. Mann-Whitney test was performed with p < 0.05 deemed statistically significant.

Results: After preparation and preservation of the tissues for endothelial keratoplasty, alizarin red showed standard endothelial morphology from both the groups. Endothelial cell loss, hexagonality and uncovered areas did not show statistically significant differences (p > 0.05) between both groups. For HCEnC, cell doubling rate was 4.7 days (p > 0.05). All the antibodies were expressed in both the groups. Hexagonality, polymorphism, cell area, viable/dead cells and Ki-67 positivity were not statistically significant (p > 0.05).

Conclusions: Complete synthetic organ culture series is safe and advantageous for carrying out advanced endothelial keratoplasty graft preparation procedures and for HCEnC culture as it is free from animal or animal-derived products.
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http://dx.doi.org/10.1111/aos.14583DOI Listing
September 2020

Human corneal endothelial cells from older donors can be cultured and passaged on cell-derived extracellular matrix.

Acta Ophthalmol 2020 Sep 10. Epub 2020 Sep 10.

Institute of Ophthalmology, University College London, London, UK.

Purpose: To investigate the effect of culturing human corneal endothelial cells (HCEnCs) from older donors on extracellular matrix (ECM) derived from human corneal endothelial cell line (HCEC-12).

Methods: HCEC-12 cells were cultured on lab-tek chamber slides for 9 days. Upon confluence, the cells were ruptured using ammonium hydroxide leaving the released ECM on the slide surface which was visualized using scanning electron microscope (SEM). HCEnCs from old aged donor tissues (n = 40) were isolated and cultured on either fibronectin-collagen (FNC) or HCEC-12 ECM at passage (P) 0. At subsequent passages (P1 and P2), cells were sub-cultured on FNC and ECM separately. Live/dead analysis and tight junction using ZO-1 staining were used to record percentage viability and morphological changes. The protein composition of HCEC-12 ECM was then analysed using liquid chromatography-mass spectrometry.

Results: SEM images showed long fibrillar-like structures and a fully laid ECM upon confluence. HCEnCs cultured from older donor tissues on this ECM showed significantly better proliferation and morphometric characteristics at subsequent passages. Out of 1307 proteins found from the HCEC-12 derived ECM, 93 proteins were evaluated to be matrix oriented out of which 20 proteins were exclusively found to be corneal endothelial specific.

Conclusions: ECM derived from HCEC-12 retains protein and growth factors that stimulate the growth of HCEnCs. As the current clinical trials are from younger donors that are not available routinely for cell culture, HCEnCs from older donors can be cultured on whole ECM and passaged successfully.
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http://dx.doi.org/10.1111/aos.14614DOI Listing
September 2020

Trend and abrupt change analysis in water quality of Urmia Lake in comparison with changes in lake water level.

Environ Monit Assess 2020 Sep 7;192(10):623. Epub 2020 Sep 7.

Department of Civil and Environmental Engineering, University of Nevada, Las Vegas, NV, USA.

Urmia Lake, as the largest lake in Iran borders, has a special role in the ecosystem of the region. The water level in this lake declines in recent year remarkably, so monitoring the lake water quality is important from an environmental view. In this research, the changes in the qualitative variables of the lake water (including electrical conductivity (EC), pH, total dissolved solids (TDS), and sodium adsorption ratio (SAR)) are compared with the changes in the lake's water level based on the Mann-Kendall nonparametric test. Further, abrupt change points in the time series of quality variables were detected by the Pettitt test. Studies were carried out on samples collected from five different stations during 2005-2015. The results showed that the water level of Urmia Lake had a significant decreasing trend and also, except for TDS, the other investigated quality variables had negative trends during the studied period. It was observed that in general, the values of the Z statistic in the stations located in the eastern part of the lake were more than the stations located in the western part, and also the stations located in the northern parts had a higher trend than those in the south of the bridge.
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http://dx.doi.org/10.1007/s10661-020-08577-8DOI Listing
September 2020